Title: Effect of leg immersion in mild warm carbonated water on skin and muscle blood flow

Authors: Ogoh S, Washio T, Suzuki K, Ikeda K, Hori T, Olesen ND, Muraoka Y.

Journal: Physiol Rep. 2018 Sep;6(18):e13859. doi: 10.14814/phy2.13859. PMID: 30221833; PMCID: PMC6139710.

Link to full text: Effect of leg immersion in mild warm carbonated water on skin and muscle blood flow

Abstract: Leg immersion in carbonated water improves endothelial-mediated vasodilator function and decreases arterial stiffness but the mechanism underlying this effect remains poorly defined. We hypothesized that carbonated water immersion increases muscle blood flow. To test this hypothesis, 10 men (age 21 ± 0 years; mean ± SD) underwent lower leg immersion in tap or carbonated water at 38°C. We evaluated gastrocnemius muscle oxyhemoglobin concentration and tissue oxygenation index using near-infrared spectroscopy, skin blood flow by laser Doppler flowmetry, and popliteal artery (PA) blood flow by duplex ultrasound. Immersion in carbonated, but not tap water elevated PA (from 38 ± 14 to 83 ± 31 mL/min; P < 0.001) and skin blood flow (by 779 ± 312%, P < 0.001). In contrast, lower leg immersion elevated oxyhemoglobin concentration and tissue oxygenation index with no effect of carbonation (P = 0.529 and P = 0.495). In addition, the change in PA blood flow in response to immersion in carbonated water correlated with those of skin blood flow (P = 0.005) but not oxyhemoglobin concentration (P = 0.765) and tissue oxygenation index (P = 0.136) while no relations was found for tap water immersion. These findings indicate that water carbonation has minimal effect on muscle blood flow. Furthermore, PA blood flow increases in response to lower leg immersion in carbonated water likely due to a large increase in skin blood flow.

Title: Carbonate Ion-Enriched Hot Spring Water Promotes Skin Wound Healing in Nude Rats

Authors: Liang J, Kang D, Wang Y, Yu Y, Fan J, Takashi E.

Journal: PLoS One. 2015 Feb 11;10(2):e0117106. doi: 10.1371/journal.pone.0117106. PMID: 25671581; PMCID: PMC4324962.

Link to full text: Carbonate Ion-Enriched Hot Spring Water Promotes Skin Wound Healing in Nude Rats

Abstract: Hot spring or hot spa bathing (Onsen) is a traditional therapy for the treatment of certain ailments. There is a common belief that hot spring bathing has therapeutic effects for wound healing, yet the underlying molecular mechanisms remain unclear. To examine this hypothesis, we investigated the effects of Nagano hot spring water (rich in carbonate ion, 42°C) on the healing process of the skin using a nude rat skin wound model. We found that hot spring bathing led to an enhanced healing speed compared to both the unbathed and hot-water (42°C) control groups. Histologically, the hot spring water group showed increased vessel density and reduced inflammatory cells in the granulation tissue of the wound area. Real-time RT-PCR analysis along with zymography revealed that the wound area of the hot spring water group exhibited a higher expression of matrix metalloproteinases-2 and -9 compared to the two other control groups. Furthermore, we found that the enhanced wound healing process induced by the carbonate ion-enriched hot spring water was mediated by thermal insulation and moisture maintenance. Our results provide the evidence that carbonate ion-enriched hot spring water is beneficial for the treatment of skin wounds.

Title: A Novel Bathing Therapeutic Approach for Diabetic Foot Ulcers

Authors: Tappia PS, Hiebert B, Sanjanwala R, Komenda P, Sathianathan C, Arneja AS, Ramjiawan B.

Journal: Applied Sciences. 2021; 11(18):8402. https://doi.org/10.3390/app11188402

Link to full text: A Novel Bathing Therapeutic Approach for Diabetic Foot Ulcers

Abstract: Ineffective healing and treatment of foot ulcers can lead to an infection and gangrene of the wound area that ultimately results in the loss of the limb. The incidence of foot ulcers is higher in patients with diabetes, peripheral vascular disease and kidney disease. Accordingly, this study was undertaken to assess the ability of foot bathing in CO2-enriched water to heal foot ulcers. The design was a double-blinded, randomized, placebo-controlled study. Patients with at least one foot ulcer were randomized to receive either a treatment with bath therapy at 37 ± 0.5 °C containing either 1000–1200 ppm CO2-enriched tap water (the intervention) or non-carbonated tap water at 37 ± 0.5 °C (the control group). Treatment was conducted three times/week for 15 min per session for up to 16 weeks for a total of 48 treatment sessions. Before and at the end of every treatment month, wound size, wound area oxygenation and the ankle brachial index were measured. In addition, the McGill pain questionnaire was conducted. Blood was also collected at these time points (for a total of five collections) for the measurement of different biomarkers. While no significant differences (p < 0.05) in the group/time interaction effect were observed, a clear separation within the wound area reduction/wound area/oxygenated Hb outcomes was seen between placebo (control) and treatment (CO2) group. This pilot study is suggestive that bathing in CO2-enriched water may accelerate the healing of foot ulcers.

Title: Effect of Carbon Dioxide Therapy on Diabetic Foot Ulcer

Authors: Shalan, N. , Al-Bazzaz, A. , Al-Ani, I. , Najem, F. and Al-Masri, M.

Journal: Journal of Diabetes Mellitus, 5, 284-289. doi: 10.4236/jdm.2015.54035.

Link to full text: Effect of Carbon Dioxide Therapy on Diabetic Foot Ulcer

Abstract: Ulceration of diabetic foot represents one of the most concerning complications associated with uncontrolled blood sugar in diabetes mellitus. The aim of this study is to evaluate the condition of twenty-two diabetic patients with different degrees of ulceration in their feet after daily secessions of carbon dioxide therapy. Blood flow to the affected foot was measured by Doppler; also the size, color, degree of ulceration and sensation of the ulcerative area were all evaluated. Results showed improvement of blood flow to the affected foot as well as improvement in the sensation and color of the ulcerative area. It was concluded that carbon dioxide therapy of diabetic foot was promising and needed thorough investigation to be brought widely into application.

Title: Cognitive factors in carbon dioxide therapy

Authors: van den Hout MA, Griez E.

Journal: J Psychosom Res. 1982;26(2):209-14. doi: 10.1016/0022-3999(82)90038-1. PMID: 6804625.

Link to PubMed: Cognitive factors in carbon dioxide therapy

Abstract: The effects of CO2 inhalation on subjective state, heart rate and blood pressure were studied in two differently instructed groups, with use of air for placebo. Participants who were told that inhalation would produce a state of pleasant relaxation reported a statistically significant, confirming change when administered CO2, whereas air was found not to alter the subjective state appreciably. Those who had expected unpleasant feelings of tension did show a non-significant change in this respect after CO2 inhalation and no such reaction at all to the placebo. Heart rate and diastolic blood pressure dropped significantly after CO2 intake. Surprisingly, participants were subject to a significant decrease in heart rate upon inhalation of air when relaxation was expected. Finally, a number of theoretical inferences are made and the role of cognitive processes in the clinical use of CO2 therapy is underscored.

Title: Cardiovascular and subjective responses to inhalation of carbon dioxide. A controlled test with anxious patients

Authors: van den Hout MA, Griez E.

Journal: Psychother Psychosom. 1982;37(2):75-82. doi: 10.1159/000287556. PMID: 6812154.

Link to PubMed: Cardiovascular and subjective responses to inhalation of carbon dioxide. A controlled test with anxious patients

Abstract: Cardiovascular and subjective responses to inhalation of a 35% CO2 mixture were studied in a sample of anxious neurotics. Inhalation of air was used as a placebo control treatment. In comparison with the inhalation of air, CO2 produced a significant drop in diastolic blood pressure. A drop in heart rate frequency was observed both after inhalation of CO2 and air. CO2 inhalation did not produce an average decline in subjective feelings of distress. Explanations of these data are offered and they are compared with earlier findings. Some speculations are made on the clinical usefulness of CO2 inhalation in the treatment of anxious neurotics.

Title: Effect of carbon dioxide-enriched water and fresh water on the cutaneous microcirculation and oxygen tension in the skin of the foot.

Authors: Hartmann BR, Bassenge E, Pittler M.

Journal: Angiology. 1997 Apr;48(4):337-43. doi: 10.1177/000331979704800406. PMID: 9112881.

Link to PubMed: Effect of carbon dioxide-enriched water and fresh water on the cutaneous microcirculation and oxygen tension in the skin of the foot.

Abstract: The effects of immersion of the lower leg and foot in fresh water and in carbon dioxide (CO2)-enriched water (1200 mg CO2 per kg water; succinate+sodium bicarbonate: Actibath, KAO Tokyo) on cutaneous circulation, vasomotion and oxygen tension (PO2) were measured by laser Doppler flowmetry and transcutaneous oximetry. On the first of two consecutive days patients were randomly assigned to have the lower extremities immersed in either fresh water or CO2-enriched water under standardized conditions (temperature, 34 degrees C; depth, 35 cm; immersion time, twenty minutes) with concurrent measurement. On the second day patients were switched to the other bath type. For both sets of measurements probes were attached symmetrically to the dorsum of each foot. Included in the study were 18 patients with mild, bilateral, peripheral, occlusive arterial disease (intermittent claudication, femoral or iliac type). During immersion in CO2-enriched water the Doppler laser signal and vasomotion amplitude rose by 300%, while PO2 increased by 10%. These increases were still apparent during the latter part of the measurement period, following withdrawal of the limbs from the bath, while patients were seated and supine. During immersion in fresh water and thereafter the Doppler laser signal was unchanged and the PO2 increase was considerably less marked. The authors were thus able to demonstrate vasodilation and increased oxygen utilization (Bohr effect) resulting from topical CO2 application, and hence, that the use of topical CO2 has an objective basis.

Title: Effects of serial percutaneous application of carbon dioxide in intermittent claudication: results of a controlled trial

Authors: Hartmann BR, Bassenge E, Hartmann M.

Journal: Angiology. 1997 Nov;48(11):957-63. doi: 10.1177/000331979704801104. PMID: 9373047.

Link to full text: Effects of serial percutaneous application of carbon dioxide in intermittent claudication: results of a controlled trial

Abstract: In a prospective, controlled clinical trial, serial application of carbon-dioxide-enriched water was compared with fresh water. Twenty-four patients with peripheral arterial occlusive disease (stable claudication) were randomly allocated to one of two serial intervention groups, lower extremities immersed in either fresh water or in CO2-enriched water (1000 mg CO2/kg) water under standardized conditions (temperature, 33 degrees C; depth, 40 cm; immersion time, 30 min; five times a week over 4 weeks). The serial application of carbon-dioxide-enriched water increased arterial peak flow (reactive hyperemia), transcutaneous oxygen tension (basal value and half-recovery-time), and pain-free walking distance. The serial fresh water application did not change these values. The authors conclude that serial carbon dioxide application can be clinically effective in patients with arterial obstructions in the lower extremities.

Title: Vasomotor effects of transcutaneous CO2 in stage II peripheral occlusive arterial disease

Authors: Savin E, Bailliart O, Bonnin P, Bedu M, Cheynel J, Coudert J, Martineaud JP.

Journal: Angiology. 1995 Sep;46(9):785-91. doi: 10.1177/000331979504600904. PMID: 7661381.

Link to full text: Vasomotor effects of transcutaneous CO2 in stage II peripheral occlusive arterial disease

Abstract: Vasomotor effects of skin exposure to carbon dioxide (CO2) have been described in normal subjects. It was of interest, therefore, to determine whether percutaneous CO2 is of therapeutic benefit. In a randomized, double-blind study, 10 patients with lower limb arteriopathy (stage II) were investigated before and after local exposure for twenty minutes to CO2-rich spa gas or to water-vapor-saturated air at the same temperature as that CO2-rich spa gas. Brachial and femoral blood flows, brachial and posterior tibial artery pressures, heart rate, and chest and foot transcutaneous oxygen tensions (tcPO2) were determined. Femoral blood flow, tibial pressure, and foot tcPO2 were significantly increased after exposure of the skin to CO2-rich spa gas. This effect was not accompanied with systemic hemodynamic modifications. Water-vapor-saturated air had no effect. These results suggest that transfer of CO2 across the skin can have beneficial local vasomotor effects in patients with lower limb stage II arteriopathy.

Title: The effect of carbonated mineral water and mofette treatment in Baile Tusnad after ischemic stroke – a case report

Authors: Dogaru, G., Marieta, M., Bulboacă, A.E., Ciumărnean, L., & Stanescu, I.C.

Journal:

Link to full text: The effect of carbonated mineral water and mofette treatment in Baile Tusnad after ischemic stroke – a case report

Abstract: Carbon dioxide baths might represent an effective therapeutic method in the rehabilitation of coronary heart disease, myocardial infarction and stroke, as well as in the treatment of chronic venous insufficiency, inflammatory diseases and functional disorders. According to the World Health Organization, 5.5 million deaths from stroke were recorded in 2001, and about 15 million people survive stroke every year. Mortality from stroke is 11% for women and 8.4% for men. According to the European Association for Cardiovascular Prevention Rehabilitation, phase II and III cardiovascular rehabilitation is performed in Romania only in a proportion of 10%. The therapeutic effects of carbonated mineral waters are due to the action of carbon dioxide. This induces cutaneous vasodilation, with a decrease in blood pressure values. It also causes an increase of cardiac output, while reducing blood pressure and heart rate. Mofettes are natural emanations along the Harghita volcanic massif, which contain CO2 in concentrations of 90-98% with cutaneous vasodilator effects, increasing cerebral and muscle blood flow. The natural therapeutic factors in Baile Tusnad, consisting of carbonated mineral water baths, mofettes, climate therapy, along with medical physical culture, indicated in the rehabilitation treatment of post-stroke patients had a beneficial effect on clinical and functional symptomatology, improving the quality of gait and balance, functionality and exercise capacity in a patient who suffered stroke five years before and was followed up for three years, while she attended an annual medical rehabilitation program in Baile Tusnad. Continuing medical rehabilitation programs, in the absence of contraindications, in Romanian spa resorts for cardiovascular treatment, as well as conducting randomized clinical studies on the efficiency of these treatments is important.

Title: Effects of inhaled carbon dioxide and oxygen on cheyne-stokes respiration in patients with heart failure

Authors: Lorenzi-Filho G, Rankin F, Bies I, Douglas Bradley T.

Journal: Am J Respir Crit Care Med. 1999 May;159(5 Pt 1):1490-8. doi: 10.1164/ajrccm.159.5.9810040. PMID: 10228116.

Link to full text: Effects of inhaled carbon dioxide and oxygen on cheyne-stokes respiration in patients with heart failure

Abstract: We hypothesized that in patients with congestive heart failure (CHF), reductions in PaCO2 sensed at the peripheral chemoreceptors trigger central apneas during Cheyne-Stokes respiration (CSR-CSA), and that raising PaCO2 by inhalation of a CO2 would eliminate these events. The effects of CO2 inhalation on the frequency of apneas and hypopneas during stage 2 (S2) sleep were studied in 10 CHF patients with CSR-CSA. The time from the breath with the minimal end tidal fraction of CO2 (FETCO2) during hyperpnea until the onset of apnea correlated strongly with the lung to ear circulation time (LECT) (r2 = 0.90, p < 0.0001), a measure of lung to carotid body circulatory delay. Among the six patients who also inhaled O2, CO2 inhalation increased transcutaneous PCO2 (PtcCO2) (36.4 +/- 4.6 mm Hg versus 38 +/- 4.4 mm Hg, p < 0.002), abolished central apneas and hypopneas (43.0 +/- 8.4 per hour on air versus 1.6 +/- 2.6 per hour on CO2, p < 0.0001), and increased SaO2. In contrast, O2 inhalation causing a similar rise in SaO2 had no significant impact on either PtcCO2 or the frequency of central events. We conclude that central apneas in patients with CHF are triggered by a low PaCO2 most likely sensed at the peripheral chemoreceptors, and that inhalation of CO2 reverses central apneas by raising PaCO2.

Title: Acute effects on cardiovascular oscillations during controlled slow yogic breathing

Authors: Bhagat OL, Kharya C, Jaryal A, Deepak KK.

Journal: Indian J Med Res. 2017 Apr;145(4):503-512. doi: 10.4103/ijmr.IJMR_830_15. PMID: 28862183; PMCID: PMC5663165.

Link to full text: Acute effects on cardiovascular oscillations during controlled slow yogic breathing

Abstract: Background & objectives: Breathing exercises are believed to modulate the cardiovascular oscillations in the body. To assess the validity of the assumption and understand the underlying mechanism, the key autonomic regulatory parameters such as heart rate variability (HRV), blood pressure variability (BPV) and baroreflex sensitivity (BRS) were recorded during controlled slow yogic breathing. Alternate nostril breathing (ANB) was selected as the yogic manoeuvre. Methods: Twelve healthy volunteers (age 30±3.8 yr) participated in the study. ANB was performed at a breathing frequency of 5 breaths per minute (bpm). In each participant, the electrocardiogram, respiratory movements, beat-to-beat BP and end-tidal carbon dioxide were recorded for five minutes each: before, during and after ANB. The records were analyzed for HRV, BPV and BRS. Results: During ANB, HRV analysis showed significant increase in the standard deviation of all NN intervals, low-frequency (LF) component, LF/HF (low frequency/high frequency) ratio and significant decrease in the HF component. BPV analysis showed a significant increase in total power in systolic BPV (SBPV), diastolic BPV (DBPV) and mean BPV. BRS analysis showed a significant increase in the total number of sequences in SBPV and DBPV and significant augmentation of α-LF and reduction in α-HF. The power spectrum showed a dominant peak in HRV at 0.08 Hz (LF component) similar to the respiratory frequency. The acute short-term change in circulatory control system declined immediately after the cessation of slow yogic breathing (ANB) and remained elevated in post-ANB stage as compared to the pre-ANB. Interpretation & conclusions: Significant increase in cardiovascular oscillations and baroreflex recruitments during-ANB suggested a dynamic interaction between respiratory and cardiovascular system. Enhanced phasic relationship with some delay indicated the complexity of the system. It indicated that respiratory and cardiovascular oscillations were coupled through multiple regulatory mechanisms, such as mechanical coupling, baroreflex and central cardiovascular control.

Title: Effect of slow abdominal breathing combined with biofeedback on blood pressure and heart rate variability in prehypertension

Authors: Wang SZ, Li S, Xu XY, Lin GP, Shao L, Zhao Y, Wang TH.

Journal: J Altern Complement Med. 2010 Oct;16(10):1039-45. doi: 10.1089/acm.2009.0577. PMID: 20954960.

Link to PubMed: Effect of slow abdominal breathing combined with biofeedback on blood pressure and heart rate variability in prehypertension

Abstract: Objective: Prehypertension is a new category designated by the Seventh Report of the Joint National Committee on Prevention, Detection, Evaluation, and Treatment of High Blood Pressure (JNC7) in 2003. Managing prehypertension with nonpharmacological intervention is possibly beneficial to the prevention of hypertension. In this study, we observed the effect of slow abdominal breathing combined with electromyographic (EMG) biofeedback training on blood pressure (BP) in prehypertensives and assessed the changes of heart rate variability (HRV) in order to find an optional intervention to prevent hypertension and acquire some experimental data to clarify the underlying neural mechanism. Methods: Twenty-two (22) postmenopausal women with prehypertension were randomly assigned to either the experiment group or the control group. The experiment group performed 10 sessions of slow abdominal breathing (six cycles/min) combined with frontal electromyographic (EMG) biofeedback training and daily home practice, while the control group only performed slow abdominal breathing and daily home practice. BP and HRV (including R-R interval and standard deviation of the normal-normal intervals [SDNN]) were measured. Results: Participants with prehypertension could lower their systolic blood pressure (SBP) 8.4 mm Hg (p < 0.001) and diastolic blood pressure (DBP) 3.9 mm Hg (p < 0.05) using slow abdominal breathing combined with EMG biofeedback. The slow abdominal breathing also significantly decreased the SBP 4.3 mm Hg (p < 0.05), while it had no effect on the DBP (p > 0.05). Repeated-measures analyses showed that the biofeedback group + abdominal respiratory group (AB+BF) training was more effective in lowering the BP than the slow breathing (p < 0.05). Compared with the control group, the R-R interval increased significantly during the training in the AB+BF group (p < 0.05). The SDNN increased remarkably in both groups during the training (p < 0.05). Conclusions: Slow abdominal breathing combined with EMG biofeedback is an effective intervention to manage prehypertension. The possible mechanism is that slow abdominal breathing combined with EMG biofeedback could reduce sympathetic activity and meanwhile could enhance vagal activity.

Title: Hyperventilation-induced changes of blood cell counts depend on hypocapnia

Authors: Stäubli M, Vogel F, Bärtsch P, Flückiger G, Ziegler WH.

Journal: Eur J Appl Physiol Occup Physiol. 1994;69(5):402-7. doi: 10.1007/BF00865403. PMID: 7875136.

Link to PubMed: Hyperventilation-induced changes of blood cell counts depend on hypocapnia

Abstract: Voluntary hyperventilation for 20 min causes haemoconcentration and an increase of white blood cell and thrombocyte numbers. In this study, we investigated whether these changes depend on the changes of blood gases or on the muscle work of breathing. A group of 12 healthy medical students breathed 36 l.min-1 of air, or air with 5% CO2 for a period of 20 min. The partial pressure of CO2 decreased by 21.4 mmHg (2.85 kPa; P < 0.001) with air and by 4.1 mmHg (0.55 kPa; P < 0.005) with CO2 enriched air. This was accompanied by haemoconcentration of 8.9% with air (P < 0.01) and of 1.6% with CO2 enriched air (P < 0.05), an increase in the lymphocyte count of 42% with air (P < 0.001) and no change with CO2 enriched air, and an increase of the platelet number of 8.4% with air (P < 0.01) and no change with CO2 enriched air. The number of neutrophil granulocytes did not change during the experiments, but 75 min after deep breathing of air, band-formed neutrophils had increased by 82% (P < 0.025), whereas they were unchanged 75 min after the experiment with CO2 enriched air. Adrenaline and noradrenaline increased by 360% and 151% during the experiment with air, but remained unchanged with CO2 enriched air. It was concluded that the changes in the white blood cell and platelet counts and of the plasma catecholamine concentrations during and after voluntary hyperventilation for 20 min were consequences of marked hypocapnic alkalosis.

Title: Reversal of Hypoxia in Murine Atherosclerosis Prevents Necrotic Core Expansion by Enhancing Efferocytosis

Authors: Marsch E, Theelen TL, Demandt JA, Jeurissen M, van Gink M, Verjans R, Janssen A, Cleutjens JP, Meex SJ, Donners MM, Haenen GR, Schalkwijk CG, Dubois LJ, Lambin P, Mallat Z, Gijbels MJ, Heemskerk JW, Fisher EA, Biessen EA, Janssen BJ, Daemen MJ, Sluimer JC.

Journal: Arterioscler Thromb Vasc Biol. 2014 Dec;34(12):2545-53. doi: 10.1161/ATVBAHA.114.304023. Epub 2014 Sep 25. PMID: 25256233.

Link to full text: Reversal of Hypoxia in Murine Atherosclerosis Prevents Necrotic Core Expansion by Enhancing Efferocytosis

Abstract: Objective: Advanced murine and human plaques are hypoxic, but it remains unclear whether plaque hypoxia is causally related to atherogenesis. Here, we test the hypothesis that reversal of hypoxia in atherosclerotic plaques by breathing hyperoxic carbogen gas will prevent atherosclerosis. Approach and results: Low-density lipoprotein receptor-deficient mice (LDLR(-/-)) were fed a Western-type diet, exposed to carbogen (95% O2, 5% CO2) or air, and the effect on plaque hypoxia, size, and phenotype was studied. First, the hypoxic marker pimonidazole was detected in murine LDLR(-/-)) plaque macrophages from plaque initiation onwards. Second, the efficacy of breathing carbogen (90 minutes, single exposure) was studied. Compared with air, carbogen increased arterial blood pO2 5-fold in LDLR(-/-)) mice and reduced plaque hypoxia in advanced plaques of the aortic root (-32%) and arch (-84%). Finally, the effect of repeated carbogen exposure on progression of atherosclerosis was studied in LDLR(-/-)) mice fed a Western-type diet for an initial 4 weeks, followed by 4 weeks of diet and carbogen or air (both 90 min/d). Carbogen reduced plaque hypoxia (-40%), necrotic core size (-37%), and TUNEL(+) (terminal uridine nick-end labeling positive) apoptotic cell content (-50%) and increased efferocytosis of apoptotic cells by cluster of differentiation 107b(+) (CD107b, MAC3) macrophages (+36%) in advanced plaques of the aortic root. Plaque size, plasma cholesterol, hematopoiesis, and systemic inflammation were unchanged. In vitro, hypoxia hampered efferocytosis by bone marrow-derived macrophages, which was dependent on the receptor Mer tyrosine kinase. Conclusions: Carbogen restored murine plaque oxygenation and prevented necrotic core expansion by enhancing efferocytosis, likely via Mer tyrosine kinase. Thus, plaque hypoxia is causally related to necrotic core expansion. Keywords: atherosclerosis; carbogen; hypoxia; macrophages.

Title: The Effect of Dry Carbon Dioxide Bathing on Peripheral Blood Circulation Measured by Thermal Imaging among Patients with Risk Factors of PAD

Authors: Zbroja H, Kowalski M, Lubkowska A.

Journal: Int J Environ Res Public Health. 2021 Feb 4;18(4):1490. doi: 10.3390/ijerph18041490. PMID: 33557430; PMCID: PMC7916006.

Link to full text: The Effect of Dry Carbon Dioxide Bathing on Peripheral Blood Circulation Measured by Thermal Imaging among Patients with Risk Factors of PAD

Abstract: Peripheral artery disease (PAD) is becoming a serious health problem of present times. It appears crucial to explore therapies that might help to restore blood flow or increase tissue oxygenation. The most effective methods of detecting early-stage changes in blood circulation in the extremities need to be identified. The aim of this study was to identify the effect of carbon dioxide (CO2) bathing on peripheral blood circulation measured by thermal imaging among patients with risk factors of PAD and ankle-brachial index (ABI) in the normal range or ABI indicating some or moderate arterial disease (ABI > 0.5). The correlation between surface temperature change and PAD-relevant characteristics was also examined. Forty-six patients who were over 65 years old who had a minimum of two additional PAD risk factors were recruited. A series of ten dry CO2 baths was performed. Thermal images were taken before and after the intervention. The CO2 therapy caused a significant change in the body surface temperature of many body areas. Numerous moderate correlations between temperature change and health-related characteristics were identified. Therefore, patients with PAD risk factors could benefit from CO2 therapy. Improvements in blood flow change the body surface temperature, and these changes could be successfully detected by thermal imaging. Keywords: carbon dioxide; cardiovascular disease; peripheral artery disease; therapy; thermography.

Title: Carboxytherapy - an innovative trend in resort medicine

Authors: Bunyatyan ND, Drogovoz SM, Kononenko AV, Prokofiev AB.

Journal: Vopr Kurortol Fizioter Lech Fiz Kult. 2018;95(5):72-76. Russian. doi: 10.17116/kurort20189505172. PMID: 30412151.

Link to PubMed: Carboxytherapy - an innovative trend in resort medicine

Abstract: Carboxytherapy (the treatment based on carbon dioxide injections) is a multipurpose and widely used medical technology. The use of CO2 injections (intracutaneous, subcutaneous, and pneumopuncture) have substantially supplemented and increased the practical relevance of carboxytherapy as a method for the treatment of many diseases. Thanks to it physiological properties, CO2 has antihypoxic, antioxidant, vasodilatory, anti-inflammatory, analgesic, and spasmolytic activities; moreover, it improves blood viscosity, stimulates neoangiogenesis, and regenerative processes. Carbon dioxide is a sort of biochemical 'peacemaker' in tissue oxygenation: when blood cells are exposed to high CO2 concentrations (Bohr effect), the rate of gas exchange (CO2 and O2) increases. The human organism interprets carboxytherapy (local hypercapnia) as oxygen deficiency and responses to it by boosting not only the blood flow, but also the vascular endothelial growth factor which stimulates neoangiogenesis and in the long run improves blood supply and tissue trophism. The multiple mechanisms of action, polymodal efficacy, a tool kit with a wide range of detectors and various modes of treatment make carboxytherapy a popular medical technology all over the world, namely in cosmetology, dermatology, aesthetic medicine, angiology, orthopaedics, cardiology, neurology, pulmonology, gynaecology, urology, proctology, plastic and general surgery, and other areas. Carboxytherapy provides a perfect example of the off-label usage in medicine that made it one of the most extensively applied medical technology for the treatment of various diseases despite the lack of the preclinical data and scarce relevant information available in textbooks, reference books and booklets. Keywords: carboxytherapy; carbon dioxide; injectable; pharmacological effect; indication.

Title: Effect of Artificial Carbon Dioxide-Rich Water Immersion on Peripheral Blood Flow in Healthy Volunteers: Preliminary Study about Artificial Carbon Dioxide-Rich Water

Authors: Hasyar, A.R.A., Rasyid, H., Idris, I. and Yusuf, I.

Journal: Open Access Macedonian Journal of Medical Sciences 9.A (2021): 527-531.

Link to full text: Effect of Artificial Carbon Dioxide-Rich Water Immersion on Peripheral Blood Flow in Healthy Volunteers: Preliminary Study about Artificial Carbon Dioxide-Rich Water

Abstract: BACKGROUND: Peripheral blood circulation disorder is one of the global health problems. Balneotherapy that uses CO2 springs may be one of the complementary treatment options. The device to produce artificial CO2-rich water is needed to achieve an improvement effect, at least almost like the improvement effect of natural balneotherapy.AIM: This study aims to investigate the effect of artificial CO2-rich water immersion on peripheral blood flow using Bicarbonated JesC CREA BC-2000.METHODS AND MATERIALS: Thirty-nine healthy volunteers participated in this study. Each subject immersed both of their legs in a mixed solution from water and CO2 at temperature 38°C. This solution was mixed using a device, namely, "Bicarbonated JesC CREA BC-2000". Peripheral blood flow was measured for 5 min before immersion (in this study, we denoted it as the mean basal blood flow), 10 min during immersion, and 5 min after immersion using pocket JMS laser Doppler flowmetry MBF-IIA. Repeated analysis of variance was used for statistical analysis.RESULTS: There is the difference in peripheral blood flow among before, during, and after immersing the legs into artificial CO2-rich water using Bicarbonated JesC CREA BC-2000 (p < 0.001).CONCLUSION: Bicarbonated JesC CREA BC-2000 may be used as the device to produce an artificial CO2-rich water bath that may affect peripheral blood flow in healthy volunteers.

Title: Neonatal encephalopathy therapy optimization for better neuroprotection with inhalation of CO2: the HENRIC feasibility and safety trial

Authors: Szakmar E, Kovacs K, Meder U, Bokodi G, Andorka C, Lakatos A, Szabo AJ, Belteki G, Szabo M, Jermendy A.

Journal: Vopr Kurortol Fizioter Lech Fiz Kult. 2018;95(5):72-76. Russian. doi: 10.17116/kurort20189505172. PMID: 30412151.

Link to full text: Neonatal encephalopathy therapy optimization for better neuroprotection with inhalation of CO2: the HENRIC feasibility and safety trial

Abstract: Background: There is an association between hypocapnia and adverse neurodevelopmental outcome in infants with neonatal encephalopathy (NE). Our aim was to test the safety and feasibility of 5% CO2 and 95% air inhalation to correct hypocapnia in mechanically ventilated infants with NE undergoing therapeutic hypothermia. Methods: Ten infants were assigned to this open-label, single-center trial. The gas mixture of 5% CO2 and 95% air was administered through patient circuits if the temperature-corrected PCO2 ≤40 mm Hg. The CO2 inhalation was continued for 12 h or was stopped earlier if the base deficit (BD) level decreased <5 mmol/L. Follow-up was performed using Bayley Scales of Infant Development II. Results: The patients spent a median 95.1% (range 44.6-98.5%) of time in the desired PCO2 range (40-60 mm Hg) during the inhalation. All PCO2 values were >40 mm Hg, the lower value of the target range. Regression modeling revealed that BD and lactate had a tendency to decrease during the intervention (by 0.61 and 0.55 mmol/L/h, respectively), whereas pH remained stable. The rate of moderate disabilities and normal outcome was 50%. Conclusions: Our results suggest that inhaled 5% CO2 administration is a feasible and safe intervention for correcting hypocapnia.

Title: Carboxytherapy as an alternative off lable method for diabetes mellitus treatment: a review

Authors: Drogovoz, S. M., K. O. Kalko, I. A. Hailat, L. B. Ivantsyk, and I. V. Kireyev.

Journal: (2021).

Link to full text: Carboxytherapy as an alternative off lable method for diabetes mellitus treatment: a review

Abstract: Diabetes mellitus (DM) is one of the global medical and social problems of the XXI century. It was found that patients with diabetes mellitus showed an increased severity of coronavirus disease (COVID19) caused by infection with coronavirus 2 (SARS-CoV-2). At the same time, the development of new modern technologies allows the use of innovative methods for the treatment of diabetes. The purpose of this literature review was to analyze the possibilities of using carboxytherapy in the treatment of diabetes mellitus. Materials and research methods. Scientific publications in foreign and domestic journals on relevant topics over the past 5 years, Internet resources. Research results and their discussion. In the course of the literature review, the mechanisms of realization of the pharmacological effects of carboxytherapy have been summarized with the substantiation of the effect on individual links of pathogenesis in the body of patients with diabetes mellitus or with its complications. The scientific understanding of the role of carboxytherapy as an effective alternative method for treating diabetes mellitus and its complications has been expanded. Conclusions. Thus, the analyzed data indicate the rationality of the use of carboxytherapy as an alternative method for the treatment of diabetes mellitus and its complications. Keywords: diabetes mellitus (DM).

Title: CO2-induced acral blood flow and the oxygen partial pressure in arterial occlusive disease

Authors: Hartmann B, Drews B, Bassenge E.

Journal: Dtsch Med Wochenschr. 1991 Oct 25;116(43):1617-21. German. doi: 10.1055/s-2008-1063797. PMID: 1935632.

Link to PubMed: CO2-induced acral blood flow and the oxygen partial pressure in arterial occlusive disease

Abstract: The effect of CO2-containing water on foot skin blood flow (laser-Doppler flux) and transcutaneous oxygen partial pressure (tcpO2) was measured in 16 patients (15 men and 1 woman; mean age 58 [49-71] years) with peripheral arterial occlusive disease (stage II of Fontaine: bilateral occlusions of the superficial femoral arteries). After 30 minutes lying down and 10 minutes sitting up, one lower leg was immersed in fresh water, the other in CO2-containing water (1400 mg CO2/kg H2O), both at 34 degrees C for 20 minutes. On the next day the measurements were repeated changing sides. Skin blood flow of the dorsum of the foot and rhythmic flow oscillations (flux motion) increased threefold in the CO2 water-bath, after a latency period averaging 2 minutes, within 3 minutes from 1.1 to 3.9 relative units (P less than 0.005). Average tcpO2 increased during the bath in CO2 water by 9 mm Hg (P less than 0.005). There was no change in fresh water. The increase in cutaneous blood flow and oxygen partial pressure, which persisted throughout the period in CO2 water, is interpreted as an increase in microcirculation and can explain part of the effectiveness of this largely empirical treatment.

Title: Carbon dioxide balneotherapy and cardiovascular disease

Authors: Pagourelias ED, Zorou PG, Tsaligopoulos M, Athyros VG, Karagiannis A, Efthimiadis GK.

Journal: Int J Biometeorol. 2011 Sep;55(5):657-63. doi: 10.1007/s00484-010-0380-7. Epub 2010 Oct 22. PMID: 20967468.

Link to PubMed: Carbon dioxide balneotherapy and cardiovascular disease

Abstract: Carbon dioxide (CO(2)) balneotherapy is a kind of remedy with a wide spectrum of applications which have been used since the Middle Ages. However, its potential use as an adjuvant therapeutic option in patients with cardiovascular disease is not yet fully clarified. We performed a thorough review of MEDLINE Database, EMBASE, ISI WEB of Knowledge, COCHRANE database and sites funded by balneotherapy centers across Europe in order to recognize relevant studies and aggregate evidence supporting the use of CO(2) baths in various cardiovascular diseases. The three main effects of CO(2) hydrotherapy during whole body or partial immersion, including decline in core temperature, an increase in cutaneous blood flow, and an elevation of the score on thermal sensation, are analyzed on a pathophysiology basis. Additionally, the indications and contra-indications of the method are presented in an evidence-based way, while the need for new methodologically sufficient studies examining the use of CO(2) baths in other cardiovascular substrates is discussed.

Title: Influence of solutes in plasma on the total CO2 content determination: implications for clinical disorders

Authors: Pichette C, Chen CB, Goldstein M, Stinebaugh B, Halperin M.

Journal: Clin Biochem. 1983 Apr;16(2):91-3. doi: 10.1016/s0009-9120(83)90552-0. PMID: 6411383.

Link to PubMed: Influence of solutes in plasma on the total CO2 content determination: implications for clinical disorders

Abstract: The purpose of these experiments was to determine the effect of non-aqueous solids on the total CO2 content of plasma. Two major groups of compounds were explored; those which reduced the total CO2 content, such as saline, and those which did not, such as albumin. To assess the former, sufficient sodium chloride was added to a bicarbonate solution gassed with 5% CO2 to increase the total volume by 10%. The total CO2 content fell the predicted 10% from 27.2 to 24.5 mmol/L when sodium chloride was added. In contrast, when the aqueous volume of a bicarbonate solution gassed with 5% CO2 was decreased by isohydric albumin, the total CO2 content was not reduced. We hypothesize that carbamino-compounds were formed with albumin and this raised the volume of CO2 released by an excess of acid. Therefore, the calculated pK' for the bicarbonate buffer system, which is derived from solutions lacking proteins, is not solely determined by the concentrations of bicarbonate, dissolved CO2, carbonate and carbonic acid when albumin is present. The clinical implications of these results will be discussed.

Title: Effect of immersion in CO2-enriched water on free radical release and total antioxidant status in peripheral arterial occlusive disease

Authors: Dogliotti G, Galliera E, Iorio E, De Bernardi Di Valserra M, Solimene U, Corsi MM.

Journal: Int Angiol. 2011 Feb;30(1):12-7. PMID: 21248668.

Link to PubMed: Effect of immersion in CO2-enriched water on free radical release and total antioxidant status in peripheral arterial occlusive disease

Abstract: Aim: The aim of this paper was to investigate the release of oxygen free radicals in patients with peripheral occlusive arterial disease and the effects of immersion of the legs and feet in carbon dioxide (CO(2))-enriched water.

Methods: Twenty-five patients with peripheral occlusive arterial disease (Fontaine stage II) and 15 healthy controls were treated by immersing the lower legs in either CO(2)-enriched or normal spa water. Blood samples were collected in heparinized tubes and total antioxidant status (TAS) and reactive oxygen metabolites (ROMs) were measured after five treatments a week for two weeks.

Results: d-ROM plasma levels decreased in patients with peripheral occlusive disease after immersion in CO(2)-enriched water (P<0.001), and in healthy controls (P<0.01), in line with a significant increase in TAS (P<0.001).

Conclusion: CO(2)-enriched water immersion had a positive effect, reducing free radical plasma levels and raising the levels of antioxidants, suggesting an improvement in the microcirculation.

Title: CO2 Balneotherapy for Arterial Occlusion Diseases: Physiology and Clinical Practice

Authors: Hartmann, B., Pittler, M., & Drews, B.

Journal: White paper. Institute of Applied Physiology and Balneology, University of Freidburg (2009).

Link to full text: CO2 Balneotherapy for Arterial Occlusion Diseases: Physiology and Clinical Practice

Abstract: Carbon dioxide, along with being the final product of metabolism and a basic ingredient of photosynthesis, is also a remedy which when applied to the skin has effects that are empirically held in high regard. Already in the Middle Ages, acidic water and gases vented from the ground (carbon dioxide fumaroles) were recognized to have strong curative powers and to be effective against "St. Anthony's fire" caused by ergot poisoning which often occurred at the time. There was no other effective method of treatment. In 1624, the medical scholar van Helmont (1577-1644) confirmed that these gases contained carbon dioxide. The anti-infective properties of carbon dioxide were discovered and analyzed by Boyle (1627-1691) and Lavoisier (1743- 1794). The first systematic medical research of CO2 use was conducted by Lalouette (1777), who showed that chronic and inveterate skin damage is cured by the serial application of CO2.

Title: Improved resuscitation outcome in emergency medical systems with increased usage of sodium bicarbonate during cardiopulmonary resuscitation

Authors: Bar-Joseph G, Abramson NS, Kelsey SF, Mashiach T, Craig MT, Safar P; Brain Resuscitation Clinical Trial III (BRCT III) Study Group.

Journal: Acta Anaesthesiol Scand. 2005 Jan;49(1):6-15. doi: 10.1111/j.1399-6576.2005.00572.x. PMID: 15675975.

Link to PubMed: Improved resuscitation outcome in emergency medical systems with increased usage of sodium bicarbonate during cardiopulmonary resuscitation

Abstract: Background: The use of sodium bicarbonate (SB) in cardiopulmonary resuscitation (CPR) is controversial. This study analyzes the effects of SB use on CPR outcome in the Brain Resuscitation Clinical Trial III (BRCT III), which was a multicenter randomized trial comparing high-dose to standard-dose epinephrine during CPR. Sodium bicarbonate use in BRCT III was optional.

Methods: The entire BRCT III database was reviewed. Analysis included only patients who arrested out of the hospital and whose time from collapse to initiation of ACLS was no longer than 30 min (total n = 2122 patients). Sodium bicarbonate use by the 16 participating study sites was analyzed. The study sites were divided according to their SB usage profile: 'low SB user' sites administered SB in less than 50% of CPRs and their first epinephrine to SB time exceeded 10 min; and 'high SB user' sites used SB in over 50% of CPRs and their first epinephrine to SB time was <10 min.

Results: Sites' SB usage rates ranged between 3.1% and 98.2% of CPRs. Sodium bicarbonate usage rates correlated inversely with the sites' intervals from collapse (r = - 0.579 P = 0.018) from initiation of ACLS (r = - 0.685 P = 0.003) and from first epinephrine (r = - 0.611 P = 0.012) to SB administration. Mean ROSC rate in the 'high SB user' sites was 33.5% (CI = 30.0-37.0) compared to 25.7% (CI = 23.1-28.4) in the 'low SB user' sites. In the 'high SB user' sites, hospital discharge rate was 5.3% (CI = 3.6-7.0) compared to 3% (CI = 2.0-4.0) in the 'low SB user' sites, and 5.3% (CI = 3.6-7.0) had a favorable neurological outcome compared to 2.1% (CI = 1.2-3.0) in the 'low SB user' sites. Collapse to ACLS interval was 8.5 min (CI = 8.1-9.0) in the 'high SB user' sites compared to 10.2 min (CI = 9.8-10.6) in the 'low SB user' sites, and their ACLS to first epinephrine interval was 7.0 min (CI = 6.5-7.5) compared to 9.7 min (CI = 9.3-10.2). Multivariate regression analysis found that belonging to 'high SB user' sites independently increased the chances for ROSC (OR 1.36, CI 1.08-1.7) and for achieving a good neurological outcome (OR 2.18, CI 1.23-3.86).

Conclusions: Earlier and more frequent use of SB was associated with higher early resuscitability rates and with better long-term outcome. Sodium bicarbonate may be beneficial during CPR, and it should be subjected to a randomized clinical trial.

Title: Decrease in heart rates by artificial CO2 hot spring bathing is inhibited by beta1-adrenoceptor blockade in anesthetized rats

Authors: Hashimoto M, Yamamoto N.

Journal: J Appl Physiol (1985). 2004 Jan;96(1):226-32. doi: 10.1152/japplphysiol.00812.2003. Epub 2003 Aug 29. PMID: 12949009.

Link to full text: Decrease in heart rates by artificial CO2 hot spring bathing is inhibited by beta1-adrenoceptor blockade in anesthetized rats

Abstract: To investigate the effects of carbon dioxide (CO2) hot spring baths on physiological functions, head-out immersion of urethane-anesthetized, fursheared male Wistar rats was performed. Animals were immersed in water (30 or 35 degrees C) with high-CO2 content ( approximately 1,000 parts/million; CO2-water). CO2-water for bathing was made by using an artificial spa maker with normal tap water and high-pressure CO2 from a gas cylinder. When a human foot was immersed for 10 min in the CO2-water at 35 degrees C, the immersed skin reddened, whereas skin color did not change in normal tap water at the same temperature. Arterial blood pressure, heart rate (HR), underwater skin tissue blood flow, and temperatures of the colon and immersed skin were continuously measured while animals were immersed in a bathtub of water for approximately 30 min at room temperature (26 degrees C). Immersed skin vascular resistance, computed from blood pressure and tissue blood flow, was significantly lower in the CO2-water bath than in tap water at 30 degrees C, but no differences were apparent at 35 degrees C. HR of rats in CO2-water was significantly slower than in tap water at 35 degrees C. Decreased HR in CO2-water was inhibited by infusion of atenolol (beta1-adrenoceptor blocker), but it was unaffected by atropine (muscarinic cholinoceptor blocker). Theses results suggest that bradycardia in CO2 hot spring bathing is caused by inhibition of the cardiac sympathetic innervation. This CO2-water maker should prove a useful device for acquiring physiological evidence of balneotherapy.

Title: The effect of transcutaneous application of carbon dioxide (CO2) on skeletal muscle

Authors: Oe K, Ueha T, Sakai Y, Niikura T, Lee SY, Koh A, Hasegawa T, Tanaka M, Miwa M, Kurosaka M.

Journal: Biochem Biophys Res Commun. 2011 Apr 1;407(1):148-52. doi: 10.1016/j.bbrc.2011.02.128. Epub 2011 Mar 1. PMID: 21371433.

Link to PubMed: The effect of transcutaneous application of carbon dioxide (CO2) on skeletal muscle

Abstract: In Europe, carbon dioxide therapy has been used for cardiac disease and skin problems for a long time. However there have been few reports investigating the effects of carbon dioxide therapy on skeletal muscle. Peroxisome proliferators-activated receptor (PPAR)-gamma coactivator-1 (PGC-1α) is up-regulated as a result of exercise and mediates known responses to exercise, such as mitochondrial biogenesis and muscle fiber-type switching, and neovascularization via up-regulation of vascular endothelial growth factor (VEGF). It is also known that silent mating type information regulation 2 homologs 1 (SIRT1) enhances PGC-1α-mediated muscle fiber-type switching. Previously, we demonstrated transcutaneous application of CO2 increased blood flow and a partial increase of O2 pressure in the local tissue known as the Bohr effect. In this study, we transcutaneously applied CO2 to the lower limbs of rats, and investigated the effect on the fast muscle, tibialis anterior (TA) muscle. The transcutaneous CO2 application caused: (1) the gene expression of PGC-1α, silent mating type information regulation 2 homologs 1 (SIRT1) and VEGF, and increased the number of mitochondria, as proven by real-time PCR and immunohistochemistry, (2) muscle fiber switching in the TA muscle, as proven by isolation of myosin heavy chain and ATPase staining. Our results suggest the transcutaneous application of CO2 may have therapeutic potential for muscular strength recovery resulting from disuse atrophy in post-operative patients and the elderly population.

Title: Improved exercise ventilatory efficiency with nasal compared to oral breathing in cardiac patients

Authors: Eser P, Calamai P, Kalberer A, Stuetz L, Huber S, Kaesermann D, Guler S, Wilhelm M.

Journal: Front Physiol. 2024 Aug 6;15:1380562. doi: 10.3389/fphys.2024.1380562.PMID: 39165283; PMCID: PMC11334221.

Link to full text: Improved exercise ventilatory efficiency with nasal compared to oral breathing in cardiac patients

Abstract: Objectives: To assess whether nasal breathing improves exercise ventilatory efficiency in patients with heart failure (HF) or chronic coronary syndromes (CCS). Background: Exercise inefficient ventilation predicts disease progression and mortality in patients with cardiovascular diseases. In healthy people, improved ventilatory efficiency with nasal compared to oral breathing was found. Methods: Four study groups were recruited: Patients with HF, patients with CCS, old (age≥45 years) and young (age 20-40 years) healthy control subjects. After a 3-min warm-up, measurements of 5 min with once nasal and once oral breathing were performed in randomized order at 50% peak power on cycle ergometer. Ventilation and gas exchange parameters measured with spiroergometry were analysed by Wilcoxon paired-sample tests and linear mixed models adjusted for sex, height, weight and test order. Results: Groups comprised 15 HF, CCS, and young control and 12 old control. Ventilation/carbon dioxide production (V E/V CO2), ventilation (V E), breathing frequency (fR), and end-tidal oxygen partial pressure (PETO2) were significantly lower and tidal volume and end-tidal carbon dioxide partial pressure (PETCO2) significantly higher during nasal compared to oral breathing in all groups, with large effect sizes for most parameters. For patients with HF, median V E/V CO2 was 35% lower, fR 26% lower, and PETCO2 10% higher with nasal compared to oral breathing, respectively. Exercise oscillatory ventilation (EOV) was present in 6 patients and markedly reduced with nasal breathing. Conclusion: Nasal breathing during submaximal exercise significantly improved ventilatory efficiency and abnormal breathing patterns (rapid shallow breathing and EOV) in 80% of our patients with HF and CCS.

Title: Increase in carbon dioxide accelerates the performance of endurance exercise in rats

Authors: Ueha T, Oe K, Miwa M, Hasegawa T, Koh A, Nishimoto H, Lee SY, Niikura T, Kurosaka M, Kuroda R, Sakai Y.

Journal: J Physiol Sci. 2018 Jul;68(4):463-470. doi: 10.1007/s12576-017-0548-6. Epub 2017 Jun 10. PMID: 28601950; PMCID: PMC10717130.

Link to full text: Increase in carbon dioxide accelerates the performance of endurance exercise in rats

Abstract: Endurance exercise generates CO2 via aerobic metabolism; however, its role remains unclear. Exogenous CO2 by transcutaneous delivery promotes muscle fibre-type switching to increase endurance power in skeletal muscles. Here we determined the performance of rats running in activity wheels with/without transcutaneous CO2 exposure to clarify its effect on endurance exercise and recovery from muscle fatigue. Rats were randomised to control, training and CO2 groups. Endurance exercise included activity-wheel running with/without transcutaneous CO2 delivery. Running performance was measured after exercise initiation. We also analysed changes in muscle weight and muscle fibres in the tibialis anterior muscle. Running performance improved over the treatment period in the CO2 group, with a concomitant switch in muscle fibres to slow-type. The mitochondrial DNA content and capillary density in the CO2 group increased. CO2 was beneficial for performance and muscle development during endurance exercise: it may enhance recovery from fatigue and support anabolic metabolism in skeletal muscles.

Title: Effects of an artificially carbonated bath on athletic warm-up

Authors: Akamine T, Taguchi N

Journal: J Hum Ergol (Tokyo). 1998 Dec;27(1-2):22-9. PMID: 11579696.

Link to full text: Effects of an artificially carbonated bath on athletic warm-up

Abstract: The effects of an artificially carbonated bath (36 degrees C, CO2 300 ppm, 20 minutes) on the warm-up of swimmers was compared with those of a freshwater bath (36 degrees C, 20 minutes). Carbon dioxide is reported to have a vasodilatory effect on peripheral blood vessels of cutaneous and muscular tissue and to promote blood flow. We observed that the warm-up effects of a carbonated bath before swimming on the hematocrit, white blood cell, total plasma protein, and total cholesterol levels in venous blood were significantly increased more than those of a freshwater bath before swimming in recovery period (p < 0.05). Thus the carbonated bath tended to be more effective for increasing the concentrations of blood components. In the recovery period, the carbonated bath before swimming also resulted in significantly smaller changes in blood lactic acid and heart rate than those of a freshwater bath before swimming (p < 0.05). The decrease in electromyography of the M. rectus femoris during swimming suggested more efficient muscle activity after a carbonated bath. Therefore after a carbonated bath, swimmers should have a higher reserve left in the cardiovascular system, resulting in better performance during swimming and less accumulation of fatigue-related metabolites after swimming.

Title: Enhanced Athletic Recovery Using Cold-Water Immersion with Dissolved CO2 and H2

Authors: Yoshimura M, Fukuoka Y, Sawada Y, Ichikawa H, Nakamura M

Journal: International Journal of Physical Medicine & Rehabilitation

Link to full text: Enhanced Athletic Recovery Using Cold-Water Immersion with Dissolved CO2 and H2

Abstract: A Cold therapy is widely recognized as a prescription for acute-phase recovery in athletes. However, it has been suggested that vasoconstriction caused by cooling may, in fact, delay the recovery process after exercise. Therefore, we are researching a new cooling method that involves supplementing ice baths with carbon dioxide gas, which has vasodilatory effects, and hydrogen gas, which has anti-inflammatory properties. This new approach aims to improve muscle blood flow and performance while maintaining the benefits of cooling, such as pain alleviation. This mini review provides an overview of the acute recovery effects of this innovative method and its potential applications.

Title: Repeated-sprint training in hypoxia induced by voluntary hypoventilation improves running repeated-sprint ability in rugby players.

Authors: Fornasier-Santos C, Millet GP, Woorons X.

Journal: Eur J Sport Sci. 2018 May;18(4):504-512. doi: 10.1080/17461391.2018.1431312. Epub 2018 Feb 5. PMID: 29400616.

Link to PubMed: Repeated-sprint training in hypoxia induced by voluntary hypoventilation improves running repeated-sprint ability in rugby players.

Abstract: Purpose: The goal of this study was to determine the effects of repeated-sprint training in hypoxia induced by voluntary hypoventilation at low lung volume (VHL) on running repeated-sprint ability (RSA) in team-sport players. Methods: Twenty-one highly trained rugby players performed, over a 4-week period, seven sessions of repeated 40-m sprints either with VHL (RSH-VHL, n = 11) or with normal breathing (RSN, n = 10). Before (Pre-) and after training (Post-), performance was assessed with an RSA test (40-m all-out sprints with a departure every 30 s) until task failure (85% of the reference velocity assessed in an isolated sprint). Results: The number of sprints completed during the RSA test was significantly increased after the training period in RSH-VHL (9.1 ± 2.8 vs. 14.9 ± 5.3; +64%; p < .01) but not in RSN (9.8 ± 2.8 vs. 10.4 ± 4.7; +6%; p = .74). Maximal velocity was not different between Pre- and Post- in both groups whereas the mean velocity decreased in RSN and remained unchanged in RSH-VHL. The mean SpO2 recorded over an entire training session was lower in RSH-VHL than in RSN (90.1 ± 1.4 vs. 95.5 ± 0.5%, p < .01). Conclusion: RSH-VHL appears to be an effective strategy to produce a hypoxic stress and to improve running RSA in team-sport players.

Title: Application of carbon dioxide to the skin and muscle oxygenation of human lower-limb muscle sites during cold water immersion

Authors: Yoshimura M, Hojo T, Yamamoto H, Tachibana M, Nakamura M, Tsutsumi H, Fukuoka Y.

Journal: PeerJ. 2020 Aug 21;8:e9785. doi: 10.7717/peerj.9785. PMID: 32884861; PMCID: PMC7444506.

Link to full text: Application of carbon dioxide to the skin and muscle oxygenation of human lower-limb muscle sites during cold water immersion

Abstract: Background: Cold therapy has the disadvantage of inducing vasoconstriction in arterial and venous capillaries. The effects of carbon dioxide (CO2) hot water depend mainly on not only cutaneous vasodilation but also muscle vasodilation. We examined the effects of artificial CO2 cold water immersion (CCWI) on skin oxygenation and muscle oxygenation and the immersed skin temperature. Subjects and methods: Fifteen healthy young males participated. CO2-rich water containing CO2 >1,150 ppm was prepared using a micro-bubble device. Each subject's single leg was immersed up to the knee in the CO2-rich water (20 °C) for 15 min, followed by a 20-min recovery period. As a control study, a leg of the subject was immersed in cold tap-water at 20 °C (CWI). The skin temperature at the lower leg under water immersion (Tsk-WI) and the subject's thermal sensation at the immersed and non-immersed lower legs were measured throughout the experiment. We simultaneously measured the relative changes of local muscle oxygenation/deoxygenation compared to the basal values (Δoxy[Hb+Mb], Δdeoxy[Hb+Mb], and Δtotal[Hb+Mb]) at rest, which reflected the blood flow in the muscle, and we measured the tissue O2 saturation (StO2) by near-infrared spectroscopy on two regions of the tibialis anterior (TA) and gastrocnemius (GAS) muscles. Results: Compared to the CWI results, the Δoxy[Hb+Mb] and Δtotal[Hb+Mb] in the TA muscle at CCWI were increased and continued at a steady state during the recovery period. In GAS muscle, the Δtotal[Hb+Mb] and Δdeoxy[Hb+Mb] were increased during CCWI compared to CWI. Notably, StO2values in both TA and GAS muscles were significantly increased during CCWI compared to CWI. In addition, compared to the CWI, a significant decrease in Tsk at the immersed leg after the CCWI was maintained until the end of the 20-min recovery, and the significant reduction continued. Discussion: The combination of CO2 and cold water can induce both more increased blood inflow into muscles and volume-related (total heme concentration) changes in deoxy[Hb+Mb] during the recovery period. The Tsk-WI stayed lower with the CCWI compared to the CWI, as it is associated with vasodilation by CO2.

Title: Anaerobic performance after 3-day consecutive CO2-rich cold-water immersion in physically active males

Authors: FFujita M, Yoshimura M, Nakamura M, Hojo T, Fukuoka Y.

Journal: J Exerc Sci Fit. 2022 Apr;20(2):148-154. doi: 10.1016/j.jesf.2022.02.004. Epub 2022 Mar 4. PMID: 35356104; PMCID: PMC8921317.

Link to full text: Anaerobic performance after 3-day consecutive CO2-rich cold-water immersion in physically active males

Abstract:
Background objective: We investigated the effects of a 3-day consecutive CO2-rich cold (20 °C) water immersion (CCWI) following a high-intensity intermittent test (HIIT) on subjects' sublingual temperature (Tsub), blood lactate ([La]b), and heart rate (HR) compared to cold (20 °C) tap-water immersion (CWI) or passive recovery (PAS).

Methods: Thirty-two subjects were randomly allocated into three groups (CCWI, CWI, and PAS), each of which completed 4 consecutive days of cycling experiments. HR, Tsub, and [La]b were recorded on each day of exercise testing (immersion from Day 1 to Day 3 and Day 4). HIIT consisted of 8 sets of 20-sec maximum exercise at an intensity of 120% of VO2max with 10-sec passive rest. The mean and peak power, and peak pedal repetitions (PPR) within HIIT were averaged and the decline in PPR (ΔPPR) from Day 1 to Day 4 was measured.

Results: In CCWI and CWI, HR declined significantly following each immersion, with CCWI showing the larger reduction (p < 0.001). At Day 2, CCWI showed a significantly lower [La]b compared to PAS (p < 0.01). The changes in mean and peak power from Day 1 to Day 4 did not differ among the groups (p = 0.302). ΔPPR of HIIT was significantly correlated with the HR and [La]b values after immersions (ΔPPR-HR: r2 = 0.938, p < 0.001, ΔPPR-[La]b: r2 = 0.999, p < 0.001).

Conclusions: These findings indicate that CCWI is a promising intervention for maintaining peak performance in high-intensity intermittent exercise, which is associated with a reduction in [La]b and HR.

Title: The field study about the effects of artificial CO2-rich cool-water immersion after outdoor sports activity in a hot environment

Authors: Yoshimura M, Nakamura M, Hojo T, Arai A, Fukuoka Y.

Journal: J Exerc Sci Fit. 2023 Jul;21(3):268-274. doi: 10.1016/j.jesf.2023.05.001. Epub 2023 May 10. PMID: 37250065; PMCID: PMC10209124.

Link to full text: The field study about the effects of artificial CO2-rich cool-water immersion after outdoor sports activity in a hot environment

Abstract:
Background/objective: In our previous laboratory experiment (room temperature of 25 °C), CO2-rich cool-water immersion (CCWI) suppressed subjects' core body temperature even during repeated exercise. It is unclear whether the suppression of body temperature elevation would also continue after CCWI in a hot outdoor environment. Herein we investigated the thermal effects of CCWI after regular exercise training in heat on subjects' core temperature (Tcore), three skin temperatures (Tskin), heart rate (HR), and the rate of perceived ice (RPI).

Methods: Thirty-six subjects (25 males, 11 females) were randomly allocated into three groups (CCWI, CWI, and control). After training at their competitive clubs, each subject was immersed up to the chest in CCWI or CWI at 20 °C for 20 min, followed by a 60-min recovery period. Tcore, Tskin, HR, and RPI were measured at the initial rest, the end of immersion, and every 10 min during the recovery period.

Results: Compared to the control, the CCWI subjects' Tcore was significantly lower at 50-60 min after the end of immersion (p < 0.05). Tskin at abdominal and lower-leg regions during the recovery period was maintained at significantly lower values in the CWI and CCWI groups versus control (p < 0.05). The CCWI subjects maintained lower Tskin for a longer time than the CWI subjects.

Conclusions: These findings indicate that CCWI suppresses the rise in body temperatures more than CWI, even in a hot environment, suggesting that CCWI may be a more effective countermeasure against increasing body temperature in a hot outdoor environment.

Title: Increase in carbon dioxide accelerates the performance of endurance exercise in rats

Authors: Ueha T, Oe K, Miwa M, Hasegawa T, Koh A, Nishimoto H, Lee SY, Niikura T, Kurosaka M, Kuroda R, Sakai Y.

Journal: J Physiol Sci. 2018 Jul;68(4):463-470. doi: 10.1007/s12576-017-0548-6. Epub 2017 Jun 10. PMID: 28601950; PMCID: PMC10717130.

Link to full text: Increase in carbon dioxide accelerates the performance of endurance exercise in rats

Abstract: Endurance exercise generates CO2 via aerobic metabolism; however, its role remains unclear. Exogenous CO2 by transcutaneous delivery promotes muscle fibre-type switching to increase endurance power in skeletal muscles. Here we determined the performance of rats running in activity wheels with/without transcutaneous CO2 exposure to clarify its effect on endurance exercise and recovery from muscle fatigue. Rats were randomised to control, training and CO2 groups. Endurance exercise included activity-wheel running with/without transcutaneous CO2 delivery. Running performance was measured after exercise initiation. We also analysed changes in muscle weight and muscle fibres in the tibialis anterior muscle. Running performance improved over the treatment period in the CO2 group, with a concomitant switch in muscle fibres to slow-type. The mitochondrial DNA content and capillary density in the CO2 group increased. CO2 was beneficial for performance and muscle development during endurance exercise: it may enhance recovery from fatigue and support anabolic metabolism in skeletal muscles.

Keywords: Activity wheel; Carbon dioxide; Endurance exercise; Running performance.

Title: Inspired carbon dioxide during hypoxia: effects on task performance and cerebral oxygen saturation

Authors: Van Dorp E, Los M, Dirven P, Sarton E, Valk P, Teppema L, Stienstra R, Dahan A.

Journal: Aviat Space Environ Med. 2007 Jul;78(7):666-72. PMID: 17679563.

Link to PubMed: Inspired carbon dioxide during hypoxia: effects on task performance and cerebral oxygen saturation

Abstract: Introduction: Exposure to a hypoxic environment has a deleterious effect on physiological and mental functions. We studied the effect of added inspired CO2 during artificially induced hypoxic normobaric hypoxia (oxygen saturation approximately 80%) on complex task performance.

Methods: In random order, 22 healthy volunteers were exposed to 3 gas mixtures for 50 min each: sham hypoxia (SH, PetO2 103 mmHg without inspired CO2); isocapnic hypoxia (IH, PetO2 approximately 40 mmHg, PetcO2 clamped to 0.4 mmHg above resting values); and poikilocapnic hypoxia (PH, PetO2 approximately 40 mmHg; no inspired CO2). Brain oxygenation was measured using near infrared spectroscopy. During minutes 25-45 of hypoxia, subjects performed vigilance and task performance tests used in aviation research: the Vigilance and Tracking test and the Multi-Attribute Task battery (MAT-bat). The tests varied in difficulty with the tracking tests considered most difficult.

Results: PetCO2 levels differed significantly among groups: IH 42.8 +/- 0.7, SH 39.0 +/- 0.7, and PH 36.8 +/- 0.7 mmHg. Brain oxygenation levels were significantly higher during IH than PH (62.2 +/- 1.0 vs. 59.1 +/- 1.3%). The results of the performance tests indicated a negative effect of PH vs. SH on most function tests. For the Tracking test of the MAT-bat, performance was worst during PH, but returned to baseline during IH.

Discussion: We demonstrate the ability of added inspired CO2 to improve performance during hypoxia by preventing PH-associated hypocapnia-induced vasoconstriction of brain blood vessels. Our results are relevant to aerospace medicine and other circumstances in which complex tasks are performed in a hypoxic environment such as mountain climbing and working in confined spaces.

Title: Breathing Practices for Stress and Anxiety Reduction: Conceptual Framework of Implementation Guidelines Based on a Systematic Review of the Published Literature

Authors: Bentley TGK, D’Andrea-Penna G, Rakic M, Arce N, LaFaille M, Berman R, Cooley K, Sprimont P.

Journal: Brain Sciences. 2023; 13(12):1612. https://doi.org/10.3390/brainsci13121612

Link to full text: Breathing Practices for Stress and Anxiety Reduction: Conceptual Framework of Implementation Guidelines Based on a Systematic Review of the Published Literature

Abstract: Anxiety and stress plague populations worldwide. Voluntary regulated breathing practices offer a tool to address this epidemic. We reviewed the peer-reviewed published literature to evaluate practice components associated with stress reduction effectiveness. We identified 58 clinical studies, comprising 41 RCTs and 17 single-arm, pre-post, non-randomized trials; 5407 total participants were included, with a mean age of 35.9 years and 43.92% female. Among the 72 included interventions, breathing practices significantly reduced stress/anxiety in 54 interventions. Findings were used to develop a conceptual framework of breath practice effectiveness.

While commonly emphasized factors such as specific breath pace, population, and group versus individual practice were not found to be associated with effectiveness, five core components were: avoiding fast-only and <5 min practices; and incorporating human-guided training during initial sessions, multiple sessions, and long-term practice. Following this simple, evidence-based framework can help maximize the stress reduction benefits of breathing practices among broad populations.

Title: Slow-Breathing Curriculum for Stress Reduction in High School Students: Lessons Learned From a Feasibility Pilot

Authors: Bentley TGK, Seeber C, Hightower E, Mackenzie B, Wilson R, Velazquez A, Cheng A, Arce NN, Lorenz KA.

Journal: Front Rehabil Sci. 2022 Jul 1;3:864079. doi: 10.3389/fresc.2022.864079. PMID: 36189008; PMCID: PMC9397716.

Link to full text: Slow-Breathing Curriculum for Stress Reduction in High School Students: Lessons Learned From a Feasibility Pilot

Abstract: Purpose: Nearly one in three US adolescents meet the criteria for anxiety, an issue that has worsened with the COVID-19 pandemic. We developed a video-based slow diaphragmatic breathing stress-reduction curriculum for high school students and evaluated its feasibility, tolerability, and preliminary effectiveness.

Methods: This cluster-randomized feasibility pilot compared 5-min slow diaphragmatic breathing for 5 weeks with treatment-as-usual control among four 12th-grade public high school classes. Students individually participated after school during COVID-19-related hybrid teaching, with slow diaphragmatic breathing three times/week and breath science education once/week. Feasibility was based on completion of breathing exercises, breath science education, and preliminary effectiveness assessments, and ease/tolerability was based on qualitative assessments. Results: Forty-three students consented to participate. Breath practice compliance ranged from 29 to 83% across classes and weeks, and decreased on average over the 5 weeks. Compliance with the breath science videos ranged from 43 to 86%, and that with the weekly STAI-State and CO2TT measures varied from 36 to 86%. Compliance with ease/tolerability assessments ranged from 0 to 60%. Preliminary effectiveness assessments' compliance varied across classes from 83 to 89% during baseline, and 29 to 72% at follow-up. The curriculum was rated as somewhat-to-definitely useful/beneficial, and definitely-to-very easy/tolerable. Students reported enjoying the diaphragmatic breathing, CO2TT, and breath science education; some found the extended exhales challenging and the curriculum and assessments time-consuming. Preliminary effectiveness analyses indicated no significant changes in STAI or CO2TT from baseline to followup or from before to after breathing exercises (p > 0.05 for all).

Conclusions: Implementation of this 5-week slow breathing curriculum was feasible and tolerable to this cohort. Compliance, tolerability, and effectiveness may be improved with in-class participation. Future research on simple and accessible slow-breathing exercises is warranted to address today's adolescent stress-management crisis.

Title: Effects of conscious connected breathing on cortical brain activity, mood and state of consciousness in healthy adults

Authors: Bahi, C., Irrmischer, M., Franken, K. et al.

Journal: Curr Psychol 43, 10578–10589 (2024). https://doi.org/10.1007/s12144-023-05119-6

Link to full text: Effects of conscious connected breathing on cortical brain activity, mood and state of consciousness in healthy adults

Abstract: Breathwork as a means of inducing non-ordinary states of consciousness is gaining traction as a potential therapeutic modality. We examined the effects of breathwork (in the form of connected breathing) on electroencephalography (EEG) and mood in 20 healthy participants (aged between 23 and 39 years (female = 11, Mage = 29). In addition, to compare with other means of inducing non-ordinary states of consciousness, we assessed the subjective effects of breathwork using the 11 Dimension Altered State of Consciousness questionnaire. EEG spectral power analysis of eyes closed rest recordings before and after the breathwork session showed a decrease in delta (1–4 Hz) and theta (4–8 Hz) frequencies in frontotemporal and parietal regions, respectively no changes were seen in Alpha (9–12 Hz) and Beta (12–30 Hz) bands. However, after decomposing the beta waves in Beta 1 (12–15 Hz), Beta 2 (15–20 Hz), Beta 3 (20–30 Hz), decreases in power were observed across Beta1 and Beta 2 in parietotemporal regions. Notably, the spectral power in gamma increased in experienced practitioners. Scores on the Profile of Mood States questionnaire showed a reduction in negative affect (anger, tension, confusion, and depression) and an increase in esteem. Scores on the 11D-ASC scale indicated that subjective experiences during breathwork were similar to those after medium to high doses of psilocybin, suggesting the occurrence of experiences of mystical quality. Present results indicate that breathwork changes brain activity and mood, and induces mystical experiences. These results are promising and suggest that such techniques could be useful to improve mental well-being.

Title: Breathwork interventions for adults with clinically diagnosed anxiety disorders: A scoping review

Authors: Banushi B, Brendle M, Ragnhildstveit A, et al.

Journal: Brain Sci. 2023 Feb 2;13(2):256. doi: 10.3390/brainsci13020256. PMID: 36831799; PMCID: PMC9954474.

Link to full text: Breathwork interventions for adults with clinically diagnosed anxiety disorders: A scoping review

Abstract: Anxiety disorders are the most common group of mental disorders, but they are often underrecognized and undertreated in primary care. Dysfunctional breathing is a hallmark of anxiety disorders; however, mainstays of treatments do not tackle breathing in patients suffering anxiety. This scoping review aims to identify the nature and extent of the available research literature on the efficacy of breathwork interventions for adults with clinically diagnosed anxiety disorders using the DSM-5 classification system. Using the PRISMA extension for scoping reviews, a search of PubMed, Embase, and Scopus was conducted using terms related to anxiety disorders and breathwork interventions. Only clinical studies using breathwork (without the combination of other interventions) and performed on adult patients diagnosed with an anxiety disorder using the DSM-5 classification system were included. From 1081 articles identified across three databases, sixteen were included for the review. A range of breathwork interventions yielded significant improvements in anxiety symptoms in patients clinically diagnosed with anxiety disorders. The results around the role of hyperventilation in treatment of anxiety were contradictory in few of the examined studies. This evidence-based review supports the clinical utility of breathwork interventions and discusses effective treatment options and protocols that are feasible and accessible to patients suffering anxiety. Current gaps in knowledge for future research directions have also been identified.

Title: Transcutaneous Carbon Dioxide Improves Contractures After Spinal Cord Injury in Rats

Authors: Inoue S, Moriyama H, Yakuwa T, Mizuno E, Suzuki R, Nomura M, Sakai Y, Akisue T.

Journal: Clin Orthop Relat Res. 2019 Aug;477(8):1934-1946. doi: 10.1097/CORR.0000000000000808. PMID: 31135536; PMCID: PMC7000005.

Link to full text: Transcutaneous Carbon Dioxide Improves Contractures After Spinal Cord Injury in Rats

Abstract: Background: Joint contractures are a major complication in patients with spinal cord injuries. Positioning, stretching, and physical therapy are advocated to prevent and treat contractures; however, many patients still develop them. Joint motion (exercise) is crucial to correct contractures. Transcutaneous carbon dioxide (CO2) therapy was developed recently, and its effect is similar to that of exercise. This therapy may be an alternative or complementary approach to exercise.

Question/purposes: Using an established model of spinal cord injury in rats with knee flexion contractures, we sought to clarify whether transcutaneous CO2 altered (1) contracture, as measured by ROM; (2) muscular and articular factors contributing to the loss of ROM; (3) fibrosis and fibrosis-related gene expression in muscle; and (4) the morphology of and fibrosis-related protein expression in the joint capsule.

Methods: Thirty-six Wistar rats were divided into three equal groups: caged control, those untreated after spinal cord injury, and those treated with CO2 after spinal cord injury. The rats were treated with CO2 from either the first day (prevention) or 15th day (treatment) after spinal cord injury for 2 or 4 weeks. The hindlimbs of rats in the treated group were exposed to CO2 gas for 20 minutes once daily. Knee extension ROM was measured with a goniometer and was measured again after myotomy. We calculated the muscular and articular factors responsible for contractures by subtracting the post-myotomy ROM from that before myotomy. We also quantified histologic muscle fibrosis and evaluated fibrosis-related genes (collagen Type 1, α1 and transforming growth factor beta) in the biceps femoris muscle with real-time polymerase chain reaction. The synovial intima's length was measured, and the distribution of fibrosis-related proteins (Type I collagen and transforming growth factor beta) in the joint capsule was observed with immunohistochemistry. Knee flexion contractures developed in rats after spinal cord injuries at all timepoints.

Results: CO2 therapy improved limited-extension ROM in the prevention group at 2 weeks (22° ± 2°) and 4 weeks (29° ± 1°) and in the treatment group at 2 weeks (31° ± 1°) compared with untreated rats after spinal cord injuries (35° ± 2°, mean difference, 13°; 39° ± 1°, mean difference, 9°; and 38° ± 1°, mean difference, 7°, respectively) (95% CI, 10.50-14.86, 8.10-10.19, and 4.73-9.01, respectively; all p < 0.001). Muscular factors decreased in treated rats in the prevention group at 2 weeks (8° ± 2°) and 4 weeks (14°± 1°) and in the treatment group at 2 weeks (14 ± 1°) compared with untreated rats (15° ± 1°, 4.85-9.42; 16° ± 1°, 1.24-3.86; and 17° ± 2°, 1.16-5.34, respectively; all p < 0.05). The therapy improved articular factors in the prevention group at 2 weeks (4° ± 1°) and 4 weeks (6° ± 1°) and in the treatment group at 2 weeks (8° ± 1°) compared with untreated rats (10° ± 1°, 4.05-7.05; 12° ± 1°, 5.18-8.02; and 11° ± 2°, 1.73-5.50, respectively; all p < 0.05). CO2 therapy decreased muscle fibrosis in the prevention group at 2 weeks (p < 0.001). The expression of collagen Type 1, α1 mRNA in the biceps femoris decreased in treated rats in the prevention group at 2 and 4 weeks compared with untreated rat (p = 0.002 and p = 0.008, respectively), although there was little difference in the expression of transforming growth factor beta (p > 0.05). CO2 therapy did not improve shortening of the synovial intima at all timepoints (all p > 0.05). CO2 therapy decreased transforming growth factor beta immunolabeling in joint capsules in the rats in the prevention group at 2 weeks. The staining intensity and Type I collagen pattern showed no differences among all groups at all timepoints.

Conclusion: CO2 therapy may be useful for preventing and treating contractures after spinal cord injuries. CO2 therapy particularly appears to be more effective as a prevention and treatment strategy in early-stage contractures before irreversible degeneration occurs, as shown in a rat model.

Title: Role of the medullary raphe nuclei in the respiratory response to CO2

Authors: Dreshaj IA, Haxhiu MA, Martin RJ.

Journal: Respir Physiol. 1998 Jan;111(1):15-23. doi: 10.1016/s0034-5687(97)00110-2. PMID: 9496468.

Link to PubMed: Role of the medullary raphe nuclei in the respiratory response to CO2

Abstract: We characterized the role of neurons within the midline of the medulla oblongata on phrenic and hypoglossal nerve responses to hypercapnia during early-development. Studies were performed on decorticate or anesthetized; vagotomized and mechanically ventilated 14-20 day old piglets. Reversible withdrawal of midline neuronal activity was induced by microinjections of lidocaine (2%, 300 nl; n = 10) and lesioning was caused by microinjections of the neurotoxic agent, ibotenic acid (n = 12), at the same sites. At any given end-tidal CO2, peak phrenic and hypoglossal activities after lidocaine were significantly lower than in the control period (P < 0.01). Similarly, 1-2 h after injections of ibotenic acid, both phrenic and hypoglossal nerve responses to CO2 were significantly lower than in the control period (P < 0.01). The results indicate for the first time that the medullary midline neurons are required for full expression of ventilatory responses to hypercapnia and raise the possibility that dysfunction of these nuclei may contribute to respiratory instability during early postnatal life.

Title: Smaller amygdala is associated with anxiety in patients with panic disorder

Authors: Hayano F, Nakamura M, Asami T, Uehara K, Yoshida T, Roppongi T, Otsuka T, Inoue T, Hirayasu Y.

Journal: Psychiatry Clin Neurosci. 2009 Jun;63(3):266-76. doi: 10.1111/j.1440-1819.2009.01960.x. PMID: 19566756.

Link to full text: Smaller amygdala is associated with anxiety in patients with panic disorder

Abstract: Aims: Anxiety a core feature of panic disorder, is linked to function of the amygdala. Volume alterations in the brain of patients with panic disorder have previously been reported, but there has been no report of amygdala volume association with anxiety.

Methods: Volumes of hippocampus and amygdala were manually measured using magnetic resonance imaging obtained from 27 patients with panic disorder and 30 healthy comparison subjects. In addition the amygdala was focused on, applying small volume correction to optimized voxel-based morphometry (VBM). State-Trait Anxiety Inventory and the NEO Personality Inventory Revised were also used to evaluate anxiety.

Results: Amygdala volumes in both hemispheres were significantly smaller in patients with panic disorder compared with control subjects (left: t = -2.248, d.f. = 55, P = 0.029; right: t = -2.892, d.f. = 55, P = 0.005). VBM showed that structural alteration in the panic disorder group occurred on the corticomedial nuclear group within the right amygdala (coordinates [x,y,z (mm)]: [26,-6,-16], Z score = 3.92, family-wise error-corrected P = 0.002). The state anxiety was negatively correlated with the left amygdala volume in patients with panic disorder (r = -0.545, P = 0.016).

Conclusions: These findings suggested that the smaller volume of the amygdala may be associated with anxiety in panic disorder. Of note, the smaller subregion in the amygdala estimated on VBM could correspond to the corticomedial nuclear group including the central nucleus, which may play a crucial role in panic attack.

Title: Induction of c-Fos in 'panic/defence'-related brain circuits following brief hypercarbic gas exposure

Authors: Johnson PL, Fitz SD, Hollis JH, Moratalla R, Lightman SL, Shekhar A, Lowry CA.

Journal: J Psychopharmacol. 2011 Jan;25(1):26-36. doi: 10.1177/0269881109353464. Epub 2010 Jan 15. PMID: 20080924.

Link to PubMed: Induction of c-Fos in 'panic/defence'-related brain circuits following brief hypercarbic gas exposure

Abstract: Inspiration of air containing high concentrations of carbon dioxide (CO(2); hypercarbic gas exposure) mobilizes respiratory, sympathetic and hypothalamic-pituitary-adrenal axis responses and increases anxiety-like behaviour in rats and humans. Meanwhile the same stimulus induces panic attacks in the majority of panic disorder patients. However, little is known about the neural circuits that regulate these acute effects. In order to determine the effects of acute hypercarbic gas exposure on forebrain and brainstem circuits, conscious adult male rats were placed in flow cages and exposed to either atmospheric air or increasing environmental CO(2) concentrations (from baseline concentrations up to 20% CO(2)) during a 5 min period. The presence of immunoreactivity for the protein product of the immediate-early gene c-fos was used as a measure of functional cellular responses. Exposing rats to hypercarbic gas increased anxiety-related behaviour and increased numbers of c-Fos-immunoreactive cells in subcortical regions of the brain involved in: (1) the initiation of fear- or anxiety-associated behavioural responses (i.e. the dorsomedial hypothalamus, perifornical nucleus and dorsolateral and ventrolateral periaqueductal gray); (2) mobilization of the hypothalamic-pituitary-adrenal axis (i.e. the dorsomedial hypothalamus, perifornical nucleus and paraventricular hypothalamic nucleus); and (3) initiation of stress-related sympathetic responses (i.e. the dorsomedial hypothalamus, dorsolateral periaqueductal grey and rostroventrolateral medulla). These findings have implications for understanding how the brain senses changes in environmental CO(2) concentrations and the neural mechanisms underlying the subsequent adaptive changes in stress-related physiology and behaviour.

Title: Amygdalar atrophy in panic disorder patients detected by volumetric magnetic resonance imaging

Authors: Massana G, Serra-Grabulosa JM, Salgado-Pineda P, Gastó C, Junqué C, Massana J, Mercader JM, Gómez B, Tobeña A, Salamero M.

Journal: Neuroimage. 2003 May;19(1):80-90. doi: 10.1016/s1053-8119(03)00036-3. PMID: 12781728.

Link to PubMed: Amygdalar atrophy in panic disorder patients detected by volumetric magnetic resonance imaging

Abstract: It has been suggested that the pathophysiology of panic disorder (PD) may involve abnormalities in several brain structures, including the amygdala. To date, however, no study has used quantitative structural neuroimaging techniques to examine amygdalar anatomy in this disorder. Volumetric magnetic resonance imaging (MRI) studies of the amygdalas, hippocampi, and temporal lobes were conducted in 12 drug-free, symptomatic PD patients (six females and six males), and 12 case-matched healthy comparison subjects. Volumetric MRI data were normalized for brain size. PD patients were found to have smaller left-sided and right-sided amygdalar volumes than controls. No differences were found in either hippocampi or temporal lobes. These findings provide new evidence of changes in amygdalar structure in PD and warrant further anatomical and MRI brain studies of patients with this disorder.

Title: Neural Correlates of Competing Fear Behaviors Evoked by an Innately Aversive Stimulus

Authors: Mongeau R, Miller GA, Chiang E, Anderson DJ.

Journal: J Neurosci. 2003 May 1;23(9):3855-68. doi: 10.1523/JNEUROSCI.23-09-03855.2003. PMID: 12736356; PMCID: PMC6742203.

Link to full text: Neural Correlates of Competing Fear Behaviors Evoked by an Innately Aversive Stimulus

Abstract: Environment and experience influence defensive behaviors, but the neural circuits mediating such effects are not well understood. We describe a new experimental model in which either flight or freezing reactions can be elicited from mice by innately aversive ultrasound. Flight and freezing are negatively correlated, suggesting a competition between fear motor systems. An unfamiliar environment or a previous aversive event, moreover, can alter the balance between these behaviors. To identify potential circuits controlling this competition, global activity patterns in the whole brain were surveyed in an unbiased manner by c-fos in situ hybridization, using novel experimental and analytical methods. Mice predominantly displaying freezing behavior had preferential neural activity in the lateral septum ventral and several medial and periventricular hypothalamic nuclei, whereas mice predominantly displaying flight had more activity in cortical, amygdalar, and striatal motor areas, the dorsolateral posterior zone of the hypothalamus, and the vertical limb of the diagonal band. These complementary patterns of c-fos induction, taken together with known connections between these structures, suggest ways in which the brain may mediate the balance between these opponent defensive behaviors.

Title: Panic and the Brainstem: Clues from Neuroimaging Studies

Authors: Perna G, Guerriero G, Brambilla P, Caldirola D.

Journal: CNS Neurol Disord Drug Targets. 2014;13(6):1049-56. doi: 10.2174/1871527313666140612112923. PMID: 24923341.

Link to PubMed: Panic and the Brainstem: Clues from Neuroimaging Studies

Abstract: One of the most influential theories has conceived unexpected panic attack (PA) as a primal defensive reaction to threat within the internal milieu of the body. This theory is based on findings suggesting the involvement of dysfunctional respiratory regulation and/or abnormally sensitive central neural network of carbon dioxide (CO2)/hydrogen ion (H+) chemoreception in PA. Thus, unexpected PA may be related to phylogenetically older brain structures, including the brainstem areas, which process basic functions related to the organism's internal milieu. The brainstem represents a crucial area for homeostatic regulation, including chemoreception and cardio-respiratory control. In addition, the midbrain dorsal periaqueductal gray may be involved in the unconditioned defense reactions to proximal threats, including internal physical stimuli. Our aim was to specifically consider the potential involvement of the brainstem in panic disorder (PD) by a comprehensive review of the available neuroimaging studies. Available data are limited and potentially affected by several limitations. However, preliminary evidence of a role of the brainstem in PD can be found and, secondly, the brainstem serotonergic system seems to be involved in panic modulation with indications of both altered serotonergic receptors and 5-HT transporter bindings. In conclusion, our review suggests that the brainstem may be involved in psychopathology of PD and supports the relevant role of subcortical serotonergic system in panic pathogenesis.

Title: fMRI of fearful facial affect recognition in panic disorder: the cingulate gyrus-amygdala connection

Authors: Pillay SS, Gruber SA, Rogowska J, Simpson N, Yurgelun-Todd DA.

Journal: J Affect Disord. 2006 Aug;94(1-3):173-81. doi: 10.1016/j.jad.2006.04.007. Epub 2006 Jun 16. PMID: 16782207.

Link to PubMed: fMRI of fearful facial affect recognition in panic disorder: the cingulate gyrus-amygdala connection

Abstract: Background: This study investigated cingulate cortex (CC) and amygdala response to fearful facial affect recognition in patients with panic disorder (PD) as measured by BOLD fMRI during the presentation of static facial images.

Methods: Eight patients with PD and eight controls were studied. Scanning was performed on a GE Signa 1.5-T scanner. Echo planar and high-resolution MR images were acquired.

Results: Controls produced greater CC activation compared to patients with PD in response to fearful faces. Furthermore, patients with PD produced less amygdala activation than controls in response to fearful faces. During the neutral face condition, overall activation for the CC was significantly greater in PD patients although anterior cingulate cortex (ACC) activation was not as markedly different between both groups. There were no between group differences in amygdala activation on exposure to the neutral face. Only left CC activation was significantly correlated negatively with HAM-A in PD patients in the fearful facial affect condition.

Limitations: Although comparable to similar studies, the sample size is small enough to warrant further investigation. Also, the effects of medication need to be considered when interpreting these results.

Conclusions: Patients with PD activate the ACC and amygdala significantly less than controls when asked to identify fearful facial affect during fMRI. The higher the anxiety, the lower the left CC activation. Thus, chronic hyperarousal in PD may diminish attentional resources and emotional response reflected in reduced ACC and amygdala activation. Even if these are medication effects, the differences from controls are clinically relevant.

Title: Neuroanatomical Correlates of a Lactate-Induced Anxiety Attack

Authors: Reiman EM, Raichle ME, Robins E, Mintun MA, Fusselman MJ, Fox PT, Price JL, Hackman KA.

Journal: Arch Gen Psychiatry. 1989 Jun;46(6):493-500. doi: 10.1001/archpsyc.1989.01810060013003. PMID: 2786401.

Link to PubMed: Neuroanatomical Correlates of a Lactate-Induced Anxiety Attack

Abstract: Positron emission tomographic measurements of regional blood flow were used to assess local neuronal activity in patients with panic disorder and in normal control subjects before and during the infusion of sodium lactate. A new technique for the analysis of positron emission tomographic data was employed to identify significant changes in regional blood flow associated with lactate infusion in the panicking patients, nonpanicking patients, and controls. Lactate-induced panic was associated with significant blood flow increases bilaterally in the temporal poles; bilaterally in insular cortex, claustrum, or lateral putamen; bilaterally in or near the superior colliculus; and in or near the left anterior cerebellar vermis. Lactate infusion was not associated with significant changes in regional blood flow in the nonpanicking patients or control subjects. Thus, the identified regions seemed to be involved in an anxiety attack.

Title: Amygdala circuitry mediating reversible and bidirectional control of anxiety

Authors: Tye KM, Prakash R, Kim SY, Fenno LE, Grosenick L, Zarabi H, Thompson KR, Gradinaru V, Ramakrishnan C, Deisseroth K.

Journal: Nature. 2011 Mar 17;471(7338):358-62. doi: 10.1038/nature09820. Epub 2011 Mar 9. PMID: 21389985; PMCID: PMC3154022.

Link to full text: Amygdala circuitry mediating reversible and bidirectional control of anxiety

Abstract: Anxiety--a sustained state of heightened apprehension in the absence of immediate threat--becomes severely debilitating in disease states. Anxiety disorders represent the most common of psychiatric diseases (28% lifetime prevalence) and contribute to the aetiology of major depression and substance abuse. Although it has been proposed that the amygdala, a brain region important for emotional processing, has a role in anxiety, the neural mechanisms that control anxiety remain unclear. Here we explore the neural circuits underlying anxiety-related behaviours by using optogenetics with two-photon microscopy, anxiety assays in freely moving mice, and electrophysiology. With the capability of optogenetics to control not only cell types but also specific connections between cells, we observed that temporally precise optogenetic stimulation of basolateral amygdala (BLA) terminals in the central nucleus of the amygdala (CeA)--achieved by viral transduction of the BLA with a codon-optimized channelrhodopsin followed by restricted illumination in the downstream CeA--exerted an acute, reversible anxiolytic effect. Conversely, selective optogenetic inhibition of the same projection with a third-generation halorhodopsin (eNpHR3.0) increased anxiety-related behaviours. Importantly, these effects were not observed with direct optogenetic control of BLA somata, possibly owing to recruitment of antagonistic downstream structures. Together, these results implicate specific BLA-CeA projections as critical circuit elements for acute anxiety control in the mammalian brain, and demonstrate the importance of optogenetically targeting defined projections, beyond simply targeting cell types, in the study of circuit function relevant to neuropsychiatric disease.

Title: Neurobiology of panic and pH chemosensation in the brain

Authors: Wemmie JA.

Journal: Dialogues Clin Neurosci. 2011;13(4):475-83. doi: 10.31887/DCNS.2011.13.4/jwemmie. PMID: 22275852; PMCID: PMC3263394.

Link to full text: Neurobiology of panic and pH chemosensation in the brain

Abstract: Panic disorder is a common and disabling illness for which treatments are too frequently ineffective. Greater knowledge of the underlying biology could aid the discovery of better therapies. Although panic attacks occur unpredictably, the ability to provoke them in the laboratory with challenge protocols provides an opportunity for crucial insight into the neurobiology of panic. Two of the most well-studied panic provocation challenges are CO(2) inhalation and lactate infusion. Although it remains unclear how these challenges provoke panic animal models of CO(2) and lactate action are beginning to emerge, and offer unprecedented opportunities to probe the molecules and circuits underlying panic attacks. Both CO(2) and lactate alter pH balance and may generate acidosis that can influence neuron function through a growing list of pH-sensitive receptors. These observations suggest that a key to better understanding of panic disorder may He in more knowledge of brain pH regulation and pH-sensitive receptors.

Keywords: CO2; chemo-sensation; lactate, brain pH; panic disorder; provocation challenge.

Title: Behavioral and cardiovascular effects of 7.5% CO2 in human volunteers

Authors: Bailey JE, Argyropoulos SV, Kendrick AH, Nutt DJ.

Journal: Depress Anxiety. 2005;21(1):18-25. doi: 10.1002/da.20048. PMID: 15782425.

Link to PubMed: Behavioral and cardiovascular effects of 7.5% CO2 in human volunteers

Abstract: The study of carbon dioxide (CO2) inhalation in psychiatry has a long and varied history, with recent interest in using inhaled CO2 as an experimental tool to explore the neurobiology and treatment of panic disorder. As a consequence, many studies have examined the panic-like response to the gas either using the single or double breath 35% CO2 inhalation or 5-7% CO2 inhaled for 15-20 min, or rebreathing 5% CO2 for a shorter time. However, this lower dose regime produces little physiological or psychological effects in normal volunteers. For this reason we have studied the effects of a higher concentration of CO2, 7.5%, given over 20 min. Twenty healthy volunteers were recruited to a double blind, placebo-controlled study where air and 7.5% CO2 were inhaled for 20 min. Cardiovascular measures and subjective ratings were obtained. When compared to air, inhaling 7.5% CO2 for 20 min increases systolic blood pressure and heart rate, indicating increased autonomic arousal. It also increases ratings of anxiety and fear and other subjective symptoms associated with an anxiety state. The inhalation of 7.5% CO2 for 20 min is safe for use in healthy volunteers and produces robust subjective and objective effects. It seems promising as an anxiety provocation test that could be beneficial in the study of the effects of anxiety on sustained performance, the discovery of novel anxiolytic agents, and the study of brain circuits and mechanisms of anxiety.

Title: The 35% CO2 challenge in panic disorder: optimization by receiver operating characteristic (ROC) analysis

Authors: Battaglia M, Perna G.

Journal: J Psychiatr Res. 1995 Mar-Apr;29(2):111-9. doi: 10.1016/0022-3956(94)00045-s. PMID: 7666379.

Link to PubMed: The 35% CO2 challenge in panic disorder: optimization by receiver operating characteristic (ROC) analysis

Abstract: To test the power of the 35% CO2 test as a challenge for panic disorder (PD), and to set an ideal threshold of discrimination between patients and controls, we analysed by receiver operating characteristic (ROC) analysis the responses of 91 out-patients with PD and 46 controls who inhaled a 35% CO2/65% O2 gas mixture. ROC analysis confirmed that the CO2 challenge discriminates well between PD patients and controls, with 86% probability to classify them correctly on the basis of subjective anxiety after the test. A relatively modest increment in subjective anxiety (i.e. an absolute increment of 20 units, or a relative increment of 26% of subjective anxiety) proved to be the ideal threshold to separate the two groups of our sample.

Title: On the psychotropic effects of carbon dioxide

Authors: Colasanti A, Esquivel G, Schruers KJ, Griez EJ.

Journal: Curr Pharm Des. 2012;18(35):5627-37. doi: 10.2174/138161212803530745. PMID: 22632467.

Link to PubMed: On the psychotropic effects of carbon dioxide

Abstract: It has been well established that the inhalation of Carbon Dioxide (CO2) can induce in humans an emotion closely replicating spontaneous panic attacks, as defined by current psychiatry nosology. The purpose of this review is to provide a critical summary of the data regarding CO2's psychopharmacological properties and underlying mechanisms. The authors review the literature on the human and animal response for the exposure of exogenous CO2 focusing on five points of interest: 1) the early history of the use of CO2 as an anesthetic and therapeutic agent, 2) the subjective effects of breathing CO2 at different concentrations in humans, 3) the use of CO2 in experimental psychiatric research as an experimental model of panic, 4) the pharmacological modulation of CO2-induced responses, and 5) the putative neurobiological mechanisms underlying the affective state induced by CO2. The authors conclude with an evolutionary-inspired notion that CO2 might act as an agent of a primal emotion serving a homeostatic function, in the control of respiration and acid-base balance.

Title: Body fluids after CO2 inhalation: insight into panic mechanisms?

Authors: Griez E, van den Hout MA, Verstappen F.

Journal: Eur Arch Psychiatry Neurol Sci. 1987;236(6):369-71. doi: 10.1007/BF00377427. PMID: 3119345.

Link to PubMed: Body fluids after CO2 inhalation: insight into panic mechanisms?

Abstract: Blood gases and electrolyte fluctuations were studied in arterial blood, after a single 35% CO2-65% O2 inhalation, which is known to trigger panic attacks in patients with panic disorder. The immediate effect of this maneuver was a brief hypercapnic acidosis followed by a slight alkalotic rebound, with shifts in Ca2+ and K+. The possible effect of these changes on neuronal membrane excitability is discussed, referring to recent experimental findings in panic provocation.

Title: A Long-Term Prospective Evaluation of First-Degree Relatives of Panic Patients Who Underwent the 35% CO2 Challenge

Authors: Perna G, Cocchi S, Allevi L, Bussi R, Bellodi L.

Journal: Biol Psychiatry. 1999 Feb 1;45(3):365-7. doi: 10.1016/s0006-3223(98)00030-4. PMID: 10023515.

Link to PubMed: A Long-Term Prospective Evaluation of First-Degree Relatives of Panic Patients Who Underwent the 35% CO2 Challenge

Abstract: This follow-up study investigated the potential priming effect of the 35% CO2 challenge on the development of anxiety disorders and/or panic attacks in healthy first-degree relatives of panic patients across a period of 3-4 years subsequent to the challenge. Thirty-one relatives who underwent the 35% CO2 challenge 3-4 years before and 14 relatives, free from psychiatric diagnoses in the same period, were directly reevaluated for the presence of anxiety disorders and panic attacks. None developed anxiety disorders and only 1, among relatives previously tested with the 35% CO2 challenge, reported sporadic panic attacks. The 35% CO2 challenge is a safe research paradigm in the investigation of healthy subjects with a familial vulnerability to panic, and CO2 hypersensitivity might be considered a trait marker of an underlying familial vulnerability to panic disorder.

Title: Examining the latent class structure of CO2 hypersensitivity using time course trajectories of panic response systems

Authors: Roberson-Nay R, Beadel JR, Gorlin EI, Latendresse SJ, Teachman BA.

Journal: J Behav Ther Exp Psychiatry. 2015 Jun;47:68-76. doi: 10.1016/j.jbtep.2014.10.013. Epub 2014 Nov 15. PMID: 25496936; PMCID: PMC4324118.

Link to full text: Examining the latent class structure of CO2 hypersensitivity using time course trajectories of panic response systems

Abstract: Carbon dioxide (CO2) hypersensitivity is hypothesized to be a robust endophenotypic marker of panic spectrum vulnerability. The goal of the current study was to explore the latent class trajectories of three primary response systems theoretically associated with CO2 hypersensitivity: subjective anxiety, panic symptoms, and respiratory rate (fR). Participants (n = 376; 56% female) underwent a maintained 7.5% CO2 breathing task that included three phases: baseline, CO2 air breathing, and recovery. Growth mixture modeling was used to compare response classes (1…n) to identify the best-fit model for each marker. Panic correlates also were examined to determine class differences in panic vulnerability. For subjective anxiety ratings, a three-class model was selected, with individuals in one class reporting an acute increase in anxiety during 7.5% CO2 breathing and a return to pre-CO2 levels during recovery. A second, smaller latent class was distinguished by elevated anxiety across all three phases. The third class reported low anxiety reported during room air, a mild increase in anxiety during 7.5% CO2 breathing, and a return to baseline during recovery. Latent class trajectories for fR yielded one class whereas panic symptom response yielded two classes. This study examined CO2 hypersensitivity in one of the largest samples to date, but did not ascertain a general population sample thereby limiting generalizability. Moreover, a true resting baseline measure of fR was not measured. Two classes potentially representing different risk pathways were observed. Implications of results will be discussed in the context of panic risk research.

Title: Five percent carbon dioxide challenge: valid analogue and marker of panic disorder?

Authors: Sanderson WC, Wetzler S.

Journal: Biol Psychiatry. 1990 Apr 1;27(7):689-701. doi: 10.1016/0006-3223(90)90584-o. PMID: 2109638.

Link to PubMed: Five percent carbon dioxide challenge: valid analogue and marker of panic disorder?

Abstract: The administration of 5% carbon dioxide (CO2) to patients with panic disorder (PD) induces a behavioral response similar to a naturally occurring panic attack. This article reviews the literature on the nature and incidence of this response. We conclude that the 5% CO2 challenge test is a valid and useful laboratory analogue of naturally occurring panic attacks, and shows promise as a marker to identify a subset of PD patients. Though further research on reliability, validity, and dose-response effects must be conducted, the CO2 challenge test provides important information regarding the phenomenology of panic states.

Title: Fear Reactivity to Bodily Sensations Among Heavy Smokers and Nonsmokers

Authors: Abrams K, Zvolensky MJ, Dorflinger L, Galatis A, Blank M, Eissenberg T.

Journal: Exp Clin Psychopharmacol. 2008 Jun;16(3):230-9. doi: 10.1037/1064-1297.16.3.230. PMID: 18540783.

Link to PubMed: Fear Reactivity to Bodily Sensations Among Heavy Smokers and Nonsmokers

Abstract: Individuals who smoke are more likely to experience panic attacks and develop panic disorder than those in the general population. One possible explanation is that smokers may experience a heightened fear response to somatic disturbances. To date, few laboratory studies have tested this hypothesis directly. The present study examined 24 adult heavy smokers (10 females) in 12-hr nicotine withdrawal and 24 adult nonsmokers (12 females) on subjective and physiological reactivity to a 4-min carbon dioxide rebreathing challenge. Results indicate that, despite an attenuated acceleration in respiration during the challenge, smokers experienced a significantly greater increase in self-reported panic symptoms than nonsmokers. In addition, smokers reported significantly greater trait levels of suffocation fear prior to the challenge. Findings are discussed with respect to the role of smoking in panic vulnerability.

Title: Interoceptive Fear Conditioning and Panic Disorder: The Role of Conditioned Stimulus–Unconditioned Stimulus Predictability

Authors: Acheson DT, Forsyth JP, Moses E.

Journal: Behav Ther. 2012 Mar;43(1):174-89. doi: 10.1016/j.beth.2011.06.001. Epub 2011 Jun 15. PMID: 22304889.

Link to PubMed: Interoceptive Fear Conditioning and Panic Disorder: The Role of Conditioned Stimulus–Unconditioned Stimulus Predictability

Abstract: Interoceptive fear conditioning is at the core of contemporary behavioral accounts of panic disorder. Yet, to date only one study has attempted to evaluate interoceptive fear conditioning in humans (see Acheson, Forsyth, Prenoveau, & Bouton, 2007). That study used brief (physiologically inert) and longer-duration (panicogenic) inhalations of 20% CO(2)-enriched air as an interoceptive conditioned (CS) and unconditioned (US) stimulus and evaluated fear learning in three conditions: CS only, CS-US paired, and CS-US unpaired. Results showed fear conditioning in the paired condition, and fearful responding and resistance to extinction in an unpaired condition. The authors speculated that such effects may be due to difficulty discriminating between the CS and the US. The aims of the present study are to (a) replicate and expand this line of work using an improved methodology, and (b) clarify the role of CS-US discrimination difficulties in either potentiating or depotentiating fear learning. Healthy participants (N=104) were randomly assigned to one of four conditions: (a) CS only, (b) contingent CS-US pairings, (c) unpaired CS and US presentations, or (d) an unpaired "discrimination" contingency, which included an exteroceptive discrimination cue concurrently with CS onset. Electrodermal and self-report ratings served as indices of conditioned responding. Consistent with expectation, the paired contingency and unpaired contingencies yielded elevated fearful responding to the CS alone. Moreover, adding a discrimination cue to the unpaired contingency effectively attenuated fearful responding. Overall, findings are consistent with modern learning theory accounts of panic and highlight the role of interoceptive conditioning and unpredictability in the etiology of panic disorder.

Title: A genetic study of the acute anxious response to carbon dioxide stimulation in man

Authors: Battaglia M, Ogliari A, Harris J, Spatola CA, Pesenti-Gritti P, Reichborn-Kjennerud T, Torgersen S, Kringlen E, Tambs K.

Journal: J Psychiatr Res. 2007 Dec;41(11):906-17. doi: 10.1016/j.jpsychires.2006.12.002. Epub 2007 Jan 24. PMID: 17254605.

Link to PubMed: A genetic study of the acute anxious response to carbon dioxide stimulation in man

Abstract: People with panic disorder-agoraphobia and their relatives often react anxiously to CO(2)-enriched gas mixtures. Available data are not suited to disentangle genetic from common environmental causes of familial aggregation of CO(2) reactivity, nor provide quantitative estimations of the sources of trait variation. Three-hundred-forty-six twin pairs belonging to the general population-based Norwegian NIPH Mental Health Study underwent self-assessments of anxiety and of DSM-IV panic symptoms after inhalation of a 35%CO(2)-65%O(2) mixture. Two thresholds were employed - at sample's 75th and 90th percentiles of responses - to define provoked panic attacks and to calculate polychoric correlations. Variance components were estimated by structural equation modelling (SEM). For definitions of responses based on the sum of all 13 panic symptoms, SEM could not discriminate between shared environmental versus genetic causes of familial resemblance for provoked attacks. For definitions of responses based on global anxiety, or on the sums of those symptoms (dyspnea, dizziness, palpitations) with highest variance post-CO(2), the best-fitting models indicated additive genetic factors as the sole causes for within-family resemblance. Best-fit heritability estimates ranged from 0.42 to 0.57. Genetic and idiosyncratic environmental factors explain most of individual differences in reactivity to hypercapnia. Within-family similarities for this trait are largely explained by genetic determinants.

Title: A Genetically Informed Study of the Association Between Childhood Separation Anxiety, Sensitivity to CO2, Panic Disorder, and the Effect of Childhood Parental Loss

Authors: Battaglia M, Pesenti-Gritti P, Medland SE, Ogliari A, Tambs K, Spatola CA.

Journal: Arch Gen Psychiatry. 2009 Jan;66(1):64-71. doi: 10.1001/archgenpsychiatry.2008.513. PMID: 19124689.

Link to PubMed: A Genetically Informed Study of the Association Between Childhood Separation Anxiety, Sensitivity to CO2, Panic Disorder, and the Effect of Childhood Parental Loss

Abstract: Context: Childhood separation anxiety disorder can predate panic disorder, which usually begins in early adulthood. Both disorders are associated with heightened sensitivity to inhaled CO(2) and can be influenced by childhood parental loss. Objectives: To find the sources of covariation between childhood separation anxiety disorder, hypersensitivity to CO(2), and panic disorder in adulthood and to measure the effect of childhood parental loss on such covariation. Design: Multivariate twin study. Participants: Seven hundred twelve young adults from the Norwegian Institute of Public Health Twin Panel, a general population cohort. Main outcome measures: Personal direct assessment of lifetime panic disorder through structured psychiatric interviews, history of childhood parental loss, and separation anxiety disorder symptoms. Subjective anxiety response to a 35% CO(2)/65% O(2) inhaled mixture compared with compressed air (placebo). Results: Our best-fitting solution yielded a common pathway model, implying that covariation between separation anxiety in childhood, hypersensitivity to CO(2), and panic disorder in adulthood can be explained by a single latent intervening variable influencing all phenotypes. The latent variable governing the 3 phenotypes' covariation was in turn largely (89%) influenced by genetic factors and childhood parental loss (treated as an identified element of risk acting at a family-wide level), which accounted for the remaining 11% of covariance. Residual variance was explained by 1 specific genetic variance component for separation anxiety disorder and variable-specific unique environmental variance components. Conclusions: Shared genetic determinants appear to be the major underlying cause of the developmental continuity of childhood separation anxiety disorder into adult panic disorder and the association of both disorders with heightened sensitivity to CO(2). Inasmuch as childhood parental loss is a truly environmental risk factor, it can account for a significant additional proportion of the covariation of these 3 developmentally related phenotypes.

Title: CO2-Induced Panic Attacks: A Twin Study

Authors: Bellodi L, Perna G, Caldirola D, Arancio C, Bertani A, Di Bella D.

Journal: Am J Psychiatry. 1998 Sep;155(9):1184-8. doi: 10.1176/ajp.155.9.1184. PMID: 9734540.

Link to PubMed: CO2-Induced Panic Attacks: A Twin Study

Abstract: Objective: The authors investigated the role of genetic factors in 35% CO2-induced panic attacks. Method: Ninety twins recruited from the general population were challenged with one-vital-capacity inhalations of 35% CO2-65% O2. Probandwise concordance rates were calculated and rates compared for monozygotic and for dizygotic twins. Results: A significantly higher concordance was found for 35% CO2-induced panic attacks among monozygotic than dizygotic twins (55.6% versus 12.5%). Conclusions: These results suggest a relevant role of genetic factors in 35% CO2-induced panic attacks.

Title: Panic disorder subtypes: differential responses to CO2 challenge

Authors: Biber B, Alkin T.

Journal: Am J Psychiatry. 1999 May;156(5):739-44. doi: 10.1176/ajp.156.5.739. PMID: 10327907.

Link to PubMed: Panic disorder subtypes: differential responses to CO2 challenge

Abstract: Objective: The purpose of this study was to investigate the possibility of a differential sensitivity to CO2 in patients diagnosed with panic disorder subtypes that were defined by the presence of prominent respiratory symptoms. Method: The authors used a 35% CO2 and 65% O2 mixture as a challenge agent. Fifty-one unmedicated subjects with DSM-III-R panic disorder, who were divided into respiratory (N = 28) and nonrespiratory (N = 23) subtypes by their symptom profiles, underwent a CO2 challenge procedure. Patients in the two groups were compared with regard to physiological and psychological measures, pulmonary function tests, panic rates, and smoking habits. Results: The patients in the respiratory group were significantly more sensitive to CO2 than were the patients in the nonrespiratory group. The respiratory group also had higher scores on the Panic and Agoraphobia Scale and had a longer duration of illness; both of these factors can be indicators of illness severity. In addition, the respiratory group's higher cigarette consumption (mean = 12.46 package-years, SD = 2.49) may have been a contributory factor not only for illness severity but also for the pathogenesis of panic disorder. Conclusions: The CO2 challenge procedure appears to be a good dissection tool in the understanding of different subtypes of panic disorder. Moreover, there may be a more specific association with prominent respiratory symptom subtype and CO2 hypersensitivity.

Title: Experiential, autonomic, and respiratory correlates of CO2 reactivity in individuals with high and low anxiety sensitivity

Authors: Blechert J, Wilhelm FH, Meuret AE, Wilhelm EM, Roth WT.

Journal: Psychiatry Res. 2013 Oct 30;209(3):566-73. doi: 10.1016/j.psychres.2013.02.010. Epub 2013 Mar 13. PMID: 23489596.

Link to PubMed: Experiential, autonomic, and respiratory correlates of CO2 reactivity in individuals with high and low anxiety sensitivity

Abstract: Psychometric studies indicate that anxiety sensitivity (AS) is a risk factor for anxiety disorders such as panic disorder (PD). To better understand the psychophysiological basis of AS and its relation to clinical anxiety, we examined whether high-AS individuals show similarly elevated reactivity to inhalations of carbon dioxide (CO2) as previously reported for PD and social phobia in this task. Healthy individuals with high and low AS were exposed to eight standardized inhalations of 20% CO2-enriched air, preceded and followed by inhalations of room air. Anxiety and dyspnea, in addition to autonomic and respiratory responses were measured every 15 s. Throughout the task, high AS participants showed a respiratory pattern of faster, shallower breathing and reduced inhalation of CO2 indicative of anticipatory or contextual anxiety. In addition, they showed elevated dyspnea responses to the second set of air inhalations accompanied by elevated heart rate, which could be due to sensitization or conditioning. Respiratory abnormalities seem to be common to high AS individuals and PD patients when considering previous findings with this task. Similarly, sensitization or conditioning of anxious and dyspneic symptoms might be common to high AS and clinical anxiety. Respiratory conditionability deserves greater attention in anxiety disorder research.

Title: Respiratory, autonomic, and experiential responses to repeated inhalations of 20% CO₂ enriched air in panic disorder, social phobia, and healthy controls

Authors: Blechert J, Wilhelm FH, Meuret AE, Wilhelm EM, Roth WT.

Journal: Biol Psychol. 2010 Apr;84(1):104-11. doi: 10.1016/j.biopsycho.2010.01.002. Epub 2010 Jan 12. PMID: 20064582; PMCID: PMC2891596.

Link to full text: Respiratory, autonomic, and experiential responses to repeated inhalations of 20% CO₂ enriched air in panic disorder, social phobia, and healthy controls

Abstract: Inhalation of carbon dioxide (CO₂) enriched air triggers anxiety in panic disorder (PD) patients, which is often interpreted as a sign of biological vulnerability. However, most studies have not measured respiration in these tasks. We compared patients with PD (n=20) and social phobia (SP, n=19) to healthy controls (n=18) during eight inhalations of 20% CO₂, preceded and followed by two inhalations of room air, while continuously measuring subjective anxiety and dyspnea as well as autonomic and respiratory variables. PD patients showed increased reactivity and delayed recovery during CO₂ inhalations for most measures. Unlike both other groups, the PD group's tidal volume responses did not habituate across CO₂ inhalations. However, PD patients did not differ from SP patients on most other measures, supporting a continuum model of CO₂ sensitivity across anxiety disorders. Both patient groups showed continued reactivity during the last air inhalations, which is unlikely to be due to a biological sensitivity.

Title: The 35% CO2 challenge test in patients with social phobia

Authors: Caldirola D, Perna G, Arancio C, Bertani A, Bellodi L.

Journal: Psychiatry Res. 1997 Jun 16;71(1):41-8. doi: 10.1016/s0165-1781(97)00038-3. PMID: 9247980.

Link to PubMed: The 35% CO2 challenge test in patients with social phobia

Abstract: Panic disorder (PD) and social phobia (SP) share many clinical, demographic and biological characteristics. To investigate the relationships between the two disorders, the responses to inhalation of a 35% carbon-dioxide (CO2) and 65% oxygen (O2) gas mixture were assessed. Sixteen patients with PD, 16 patients with SP, 13 patients with both SP and PD, seven patients with SP who experienced sporadic unexpected panic attacks and 16 healthy control subjects inhaled one vital capacity of 35% CO2 or compressed air. A double-blind, randomized, crossover design was used. PD patients and SP patients showed similar anxiogenic reactions to 35% CO2, both stronger than seen in control subjects. Patients with both disorders and SP patients with sporadic unexpected panic attacks reacted similarly to subjects with PD or SP alone. These results suggest that PD and SP share a common hypersensitivity to CO2 and thus might belong to the same spectrum of vulnerability.

Title: Carbon Dioxide-Induced Emotion and Respiratory Symptoms in Healthy Volunteers

Authors: Colasanti A, Salamon E, Schruers K, van Diest R, van Duinen M, Griez EJ.

Journal: Neuropsychopharmacology. 2008 Dec;33(13):3103-10. doi: 10.1038/npp.2008.31. Epub 2008 Mar 19. PMID: 18354390.

Link to full text: Carbon Dioxide-Induced Emotion and Respiratory Symptoms in Healthy Volunteers

Abstract: A number of evidences have established that panic and respiration are closely related. Clinical studies indicated that respiratory sensations constitute a discrete cluster of panic symptoms and play a major role in the pathophysiology of panic. The aim of the present study was to explore the phenomenology of an experimental model of panic in healthy volunteers based on the hypothesis that: (1) we can isolate discrete clusters of panic symptoms, (2) respiratory symptoms represent a distinct cluster of panic symptoms, and (3) respiratory symptoms are the best predictor of the subjective feeling of panic, as defined in the DSM IV criteria. Sixty-four healthy volunteers received a double inhalation of four mixtures containing 0, 9, 17.5 and 35% CO(2,) respectively, in a double-blind, cross-over, random design. An electronic visual analog scale and the Panic Symptom List (PSL) were used to assess subjective 'fear/discomfort' and panic symptoms, respectively. Statistical analyses consisted of Spearman's correlations, a principal component factor analysis of the 13 PSL symptoms, and linear regressions analyses. The factor analysis extracted three clusters of panic symptoms: respiratory, cognitive, and neurovegetative (r(2)=0.65). Respiratory symptoms were highly related to subjective feeling of fear/discomfort specifically in the CO(2)-enriched condition. Moreover, the respiratory component was the most important predictor of the subjective feeling of 'fear/discomfort' (beta=0.54). The discrete clusters of symptoms observed in this study were similar to those elicited in panic attacks naturally occurring in patients affected by panic disorder. Consistent with the idea that respiration plays a crucial role in the pathophysiology of panic, we found that respiratory symptoms were the best predictors the subjective state defined in the DSM IV criteria for panic.

Title: Hypersensitivity to carbon dioxide as a disease-specific trait marker

Authors: Coryell W.

Journal: Biol Psychiatry. 1997 Feb 1;41(3):259-63. doi: 10.1016/s0006-3223(97)87457-4. PMID: 9024948.

Link to PubMed: Hypersensitivity to carbon dioxide as a disease-specific trait marker

Abstract: There is now substantial evidence that an abnormal threshold for suffocation alarm underlies panic disorder. Because this disorder is highly familial, evidence of an abnormal suffocation threshold may be apparent in high-risk individuals before they develop clinical illness. To explore this possibility, we used a single inhalation of 35% CO2 vs. air to evaluate 11 subjects who had at least one first-degree relative with DSM-III-R panic disorder, 13 who had at least two relatives treated for mania or for depression (HR-AD), and 15 low-risk controls who had no family history of panic disorder, affective disorder, or alcoholism (LR-C). All were aged 18-34 and had no history of panics or of any Research Diagnostic Criteria disorder. Five (45.5%) of the subjects at high risk for panic disorder, but none of the LR-C subjects (p = .007), nor any of the HR-AD subjects (p = .011), developed a panic attack following inhalation of the CO2 mixture.

Title: Anxiety responses to CO2 inhalation in subjects at high-risk for panic disorder

Authors: Coryell W, Pine D, Fyer A, Klein D.

Journal: J Affect Disord. 2006 May;92(1):63-70. doi: 10.1016/j.jad.2005.12.045. Epub 2006 Mar 9. PMID: 16527360.

Link to PubMed: Anxiety responses to CO2 inhalation in subjects at high-risk for panic disorder

Abstract: Background: A number of reports have shown that patients with panic disorder have greater anxiety responses to the inhalation of enhanced carbon dioxide mixtures than do well controls or patients with other psychiatric illnesses. Three earlier studies have shown that well individuals who have first-degree relatives with panic disorder also experience more anxiety following CO(2) than do controls without such a family history. The following was undertaken to confirm and extend these findings. Methods: Well subjects at high risk for panic disorder (

Title: The 35% CO2 Inhalation Procedure: Test–Retest Reliability

Authors: Coryell W, Arndt S.

Journal: Biol Psychiatry. 1999 Apr 1;45(7):923-7. doi: 10.1016/s0006-3223(98)00241-8. PMID: 10202581.

Link to PubMed: The 35% CO2 Inhalation Procedure: Test–Retest Reliability

Abstract: The subjective response to a single-breath, 35% carbon dioxide challenge test shows promise as a tool for the study of panic disorder and may comprise a trait marker for that disorder. Little has been done to measure the reliability of test results, however. Subjects took a single breath at 35% CO2 and completed a self-rating of anxiety symptoms immediately thereafter. This procedure was repeated after a mean interval of 29 days. One group, considered at high risk for panic disorder, consisted of well, first-degree relatives of individuals treated for panic disorder. The control group included well subjects at high risk for affective disorder and subjects who had family histories negative for both affective disorder and panic disorder. On both testing occasions, subjects at high risk for panic disorder had symptom scores that were significantly higher than those of control subjects. Group differences in the portions who experienced a panic attack were dependent on the symptom threshold used to define an attack. A lower threshold was optimal with the second testing and a single, positive test result appeared to be more meaningful than a single negative result. The majority of individual symptom ratings were highly correlated across tests. Ratings for "smothering sensations," in particular, correlated highly across tests and consistently discriminated high-risk from control subjects. The sources of test result variability are unclear and warrant more investigation before the tests can be clinically useful. Research efforts should seek optimal thresholds to define positive test results within given data sets.

Title: Restoring Acid-Sensing Ion Channel-1a in the Amygdala of Knock-Out Mice Rescues Fear Memory But Not Unconditioned Fear Responses

Authors: Coryell MW, Wunsch AM, Haenfler JM, Allen JE, McBride JL, Davidson BL, Wemmie JA.

Journal: J Neurosci. 2008 Dec 17;28(51):13738-41. doi: 10.1523/JNEUROSCI.3907-08.2008. PMID: 19091964; PMCID: PMC2651157.

Link to full text: Restoring Acid-Sensing Ion Channel-1a in the Amygdala of Knock-Out Mice Rescues Fear Memory But Not Unconditioned Fear Responses

Abstract: Acid-sensing ion channel-1a (ASIC1a) contributes to multiple fear behaviors, however the site of ASIC1a action in behavior is not known. To explore a specific location of ASIC1a action, we expressed ASIC1a in the basolateral amygdala of ASIC1a-/- mice using viral vector-mediated gene transfer. This rescued context-dependent fear memory, but not the freezing deficit during training or the unconditioned fear response to predator odor. These data pinpoint the basolateral amygdala as the site where ASIC1a contributes to fear memory. They also discriminate fear memory from fear expressed during training and from unconditioned fear. Furthermore, this work illustrates a strategy for identifying discrete brain regions where specific genes contribute to complex behaviors.

Title: The effects of acceptance versus control contexts on avoidance of panic-related symptoms

Authors: Eifert GH, Heffner M.

Journal: J Behav Ther Exp Psychiatry. 2003 Sep-Dec;34(3-4):293-312. doi: 10.1016/j.jbtep.2003.11.001. PMID: 14972675.

Link to full text: The effects of acceptance versus control contexts on avoidance of panic-related symptoms

Abstract: The present study compared the effects of creating an acceptance versus a control treatment context on the avoidance of aversive interoceptive stimulation. Sixty high anxiety sensitive females were exposed to two 10-min periods of 10% carbon dioxide enriched air, an anxiogenic stimulus. Before each inhalation period, participants underwent a training procedure aimed at encouraging them either to mindfully observe (acceptance context) or to control symptoms via diaphragmatic breathing (control context). A third group was given no particular training or instructions. We hypothesized that an acceptance rather than control context would be more useful in the reduction of anxious avoidance. Compared to control context and no-instruction participants, acceptance context participants were less avoidant behaviorally and reported less intense fear and cognitive symptoms and fewer catastrophic thoughts during the CO(2) inhalations. We discuss the implications of our findings for an acceptance-focused vs. control-focused context when conducting clinical interventions for panic and other anxiety disorders.

Title: The effects of acute exercise and high lactate levels on 35% CO2 challenge in healthy volunteers

Authors: Esquivel G, Schruers K, Kuipers H, Griez E.

Journal: Acta Psychiatr Scand. 2002 Nov;106(5):394-7. doi: 10.1034/j.1600-0447.2002.01333.x. PMID: 12366475.

Link to PubMed: The effects of acute exercise and high lactate levels on 35% CO2 challenge in healthy volunteers

Abstract: Objective: To test the possible antipanic effects of acute exercise in healthy volunteers exposed to an inhalation of 35% CO2 challenge. Method: Twenty healthy subjects in a randomized separate group design, performed exercise in a bicycle ergometer reaching >6 mm of blood lactate and a control condition of minimal activity in the same fashion with no lactate elevation. Immediately afterwards an inhalation of a vital capacity using a mixture of 35% CO2/65% O2 through a mask was given on both conditions. Results: Subjects under the exercise condition reported less panic symptoms than controls after a CO2 challenge on the diagnostic statistical manual-IV (DSM-IV) Panic Symptom List but no difference on the Visual Analogue Anxiety Scale. Conclusion: Subjects under the exertion condition had lactate levels comparable with those of lactate infusions but an inhibitory rather than accumulative effect was seen when combined with a CO2 challenge.

Title: To inhale or not to inhale: Conditioned avoidance in breathing behavior in an odor—20% CO2 paradigm

Authors: Fannes S, Van Diest I, Meulders A, De Peuter S, Vansteenwegen D, Van den Bergh O.

Journal: Biol Psychol. 2008 Apr;78(1):87-92. doi: 10.1016/j.biopsycho.2008.01.003. Epub 2008 Jan 31. PMID: 18316154.

Link to PubMed: To inhale or not to inhale: Conditioned avoidance in breathing behavior in an odor—20% CO2 paradigm

Abstract: This study investigated breathing behavior in an odor-CO2-inhalation fear conditioning paradigm. A differential conditioning paradigm was applied in 55 participants. Both acquisition and extinction consisted of three CS+ and three CS- trials. Diluted ammonia and butyric acid served as conditional odor cues (CSs); inhalation of 20% CO2-enriched as US. The US was presented 10s after CS+ onset and both stimuli co-terminated 30s later. Subjective anxiety and US-expectancy were measured online upon presentation of the CSs. Respiratory behavior showed a biphasic pattern during CS+ acquisition trials. Participants paradoxically lowered their ventilation first; an increased ventilation was observed only towards the end of the trial. Extinction of this breathing inhibition was found. Participants avoiding the CO2 during acquisition did not show a reduction in fear from acquisition to extinction, whereas Non-avoiders did. We conclude that paradoxical decreases in ventilation constitute a relevant behavioral index of fear in CO2-inhalation paradigms.

Title: Emotional avoidance: an experimental test of individual differences and response suppression using biological challenge

Authors: Feldner MT, Zvolensky MJ, Eifert GH, Spira AP.

Journal: Behav Res Ther. 2003 Apr;41(4):403-11. doi: 10.1016/s0005-7967(02)00020-7. PMID: 12643964.

Link to PubMed: Emotional avoidance: an experimental test of individual differences and response suppression using biological challenge

Abstract: The present study examined the affective consequences of response inhibition during a state of anxiety-related physical stress. Forty-eight non-clinical participants were selected on the basis of pre-experimental differences in emotional avoidance (high versus low) and subjected to four inhalations of 20% carbon dioxide-enriched air. Half of the participants were instructed to inhibit the challenge-induced aversive emotional state, whereas the other half was instructed to simply observe their emotional response. Participants high in emotional avoidance compared to those low in emotional avoidance responded with greater levels of anxiety and affective distress but not physiological arousal. Individuals high in emotional avoidance also reported greater levels of anxiety relative to the low emotional avoidance group when suppressing compared to observing bodily sensations. These findings are discussed in terms of the significance of emotional avoidance processes during physical stress, with implications for better understanding the nature of panic disorder.

Title: Anxiogenic effects of repeated administrations of 20% CO2-enriched air: stability within sessions and habituation across time

Authors: Forsyth JP, Lejuez CW, Finlay C.

Journal: J Behav Ther Exp Psychiatry. 2000 Jun;31(2):103-21. doi: 10.1016/s0005-7916(00)00014-8. PMID: 11132115.

Link to PubMed: Anxiogenic effects of repeated administrations of 20% CO2-enriched air: stability within sessions and habituation across time

Abstract: Increasingly carbon dioxide-enriched air is being used as an aversive unconditioned stimulus in laboratory examinations of anxiety. Yet, little is known about the stability of the autonomic and subjective effects of this stimulus across repeated inhalations and sessions. We examined whether repeated administrations of high concentrations of CO2-enriched air produced either habituation, stability, or sensitization across several autonomic and self-report indices within one session (Experiment 1) and then several sessions (Experiment 2) of exposure. Results suggest that non-clinical participants do not habituate to CO2 within sessions, but do show habituation on cardiac and subjective report of anxiety across sessions. Individual difference factors such as anxiety sensitivity and suffocation fear seem to moderate some of these effects, including self-reported distress and anxiety in response to the challenge. These results support the use of CO2 as a panicogenic aversive stimulus in laboratory models of fear onset and in clinical settings for interoceptive exposure treatments of panic.

Title: The absence of relation between anxiety sensitivity and fear conditioning using 20% versus 13% CO2-enriched air as unconditioned stimuli

Authors: Forsyth JP, Palav A, Duff K.

Journal: Behav Res Ther. 1999 Feb;37(2):143-53. doi: 10.1016/s0005-7967(98)00113-2. PMID: 9990745.

Link to PubMed: The absence of relation between anxiety sensitivity and fear conditioning using 20% versus 13% CO2-enriched air as unconditioned stimuli

Abstract: Anxiety sensitivity has been implicated as a risk factor in the development and maintenance of anxiety and fear-related disorders. Indeed, persons who score high on the anxiety sensitivity index (ASI) are generally more responsive to biological challenge procedures such as CO2-inhalation that directly evoke the feared bodily events. One would expect, therefore, that persons high on anxiety sensitivity should be more conditionable and hence more likely to acquire fears, than persons low on anxiety sensitivity when CO2-enriched air is used as an unconditioned stimulus (UCS). Undergraduates (N = 96), scoring high, medium and low on the ASI received 8 repeated 20-s inhalations of either 20 or 13% CO2-enriched air (UCSs) paired with one of three CSs differing in fear-relevance (snake, heart and flowers). Several autonomic and self-report measures were assessed. Contrary to expectation, electrodermal and cardiac conditioned responses failed to discriminate between ASI groups. Yet, SUDS and severity and frequency of DSM-IV panic symptoms varied reliably as a function of anxiety sensitivity. Overall, the findings suggest that anxiety sensitivity is related to subjective fear-related complaints, but not autonomic responding and conditionability. We discuss clinical and theoretical implications for understanding the place fo anxiety sensitivity in fear onset.

Title: High-Dose Carbon Dioxide Challenge Test in Anxiety Disorder Patients

Authors: Gorman JM, Papp LA, Martinez J, Goetz RR, Hollander E, Liebowitz MR, Jordan F.

Journal: Biol Psychiatry. 1990 Nov 1;28(9):743-57. doi: 10.1016/0006-3223(90)90510-9. PMID: 2124151.

Link to full text: High-Dose Carbon Dioxide Challenge Test in Anxiety Disorder Patients

Abstract: Many investigators have shown that panic disorder patients and possibly social phobics are hypersensitive to the anxiogenic effects of inhaled carbon dioxide (CO2). In this study we administered double-breath inhalation of 35% CO2 and 65% oxygen (O2) to panic disorder patients, social phobics, and normal controls. At baseline, panic disorder patients were characterized by higher pulse, anxiety score, and evidence of hyperventilation. Panic patients and social phobics panicked more often to 35% CO2 than to room air; normal controls did not have a higher rate of panic to CO2 than to room air. However, we did not find significant group differences in anxiety level, physiological measures, or biochemical measures in response to CO2 breathing compared with room air breathing. These results confirm earlier reports of baseline hyperventilation in panic disorder patients. However, 35% CO2 may be too high a dose to differentiate respiratory responses of patients compared with normals.

Title: Anxiogenic effects of CO2 and hyperventilation in patients with panic disorder

Authors: Gorman JM, Papp LA, Coplan JD, Martinez JM, Lennon S, Goetz RR, Ross D, Klein DF.

Journal: Am J Psychiatry. 1994 Apr;151(4):547-53. doi: 10.1176/ajp.151.4.547. PMID: 8147452.

Link to PubMed: Anxiogenic effects of CO2 and hyperventilation in patients with panic disorder

Abstract: Objective: Previous studies have indicated that patients with panic disorder are more likely than normal subjects to have acute panic attacks during inhalation of CO2, but methodological objections have been raised. In this study the authors attempted to address three of these methodological problems by ensuring that raters who assessed whether panic attacks occurred were blind to subjects' diagnoses, by randomizing the order of administration of 5% CO2 and hyperventilation, and by challenging a greater number of subjects with 7% CO2. Method: Patients with panic disorder and normal subjects underwent 20-minute inhalations of 5% CO2 and 7% CO2 and 15 minutes of room-air hyperventilation. Ratings of panic/no panic during each condition were made separately by an assessor blind to diagnosis and by the subject. Scores on four panic rating scales were also recorded before and after each intervention. Results: Room-air hyperventilation caused panic attacks in a small number of patients; the difference in panic rate between patients and comparison subjects was statistically significant by the subjects' but not by the raters' assessment. Panic rates during 5% CO2 and 7% CO2 were significantly greater among the patients by both assessments; the panic rate was greatest during 7% CO2. Order of administration did not significantly affect panic rates for hyperventilation and 5% CO2. Conclusions: Panic patients were clearly more sensitive to the anxiogenic effects of CO2 than comparison subjects, and CO2 was a more potent anxiogenic stimulus than room-air hyperventilation. Seven percent CO2 discriminated best between patients and comparison subjects and should be the focus of further research.

Title: Response to hyperventilation in a group of patients with panic disorder

Authors: Gorman JM, Askanazi J, Liebowitz MR, Fyer AJ, Stein J, Kinney JM, Klein DF.

Journal: Am J Psychiatry. 1984 Jul;141(7):857-61. doi: 10.1176/ajp.141.7.857. PMID: 6428243.

Link to PubMed: Response to hyperventilation in a group of patients with panic disorder

Abstract: Twelve patients with DSM-III diagnoses of panic disorder and four normal volunteers were studied during hyperventilation of room air and increased ventilation stimulated by 5% CO2. Patients also underwent sodium lactate infusion. Eight of the patients had a panic attack during sodium lactate infusion, seven during CO2 inhalation, and three during room-air hyperventilation. Since CO2 increases the firing of the locus ceruleus, and since sodium lactate infusion is known to increase cerebral CO2, these findings are consistent with other evidence implicating the locus ceruleus in panic anxiety.

Title: The Effect of Successful Treatment on the Emotional and Physiological Response to Carbon Dioxide Inhalation in Patients with Panic Disorder

Authors: Gorman JM, Martinez J, Coplan JD, Kent J, Kleber M.

Journal: Biol Psychiatry. 2004 Dec 1;56(11):862-7. doi: 10.1016/j.biopsych.2004.08.016. PMID: 15576063.

Link to PubMed: The Effect of Successful Treatment on the Emotional and Physiological Response to Carbon Dioxide Inhalation in Patients with Panic Disorder

Abstract: Background: A number of studies have shown that patients with panic disorder are more likely to have panic attacks during carbon dioxide inhalation than are normal comparison subjects. Some studies have shown that antipanic medications can reduce the anxiogenic response to carbon dioxide, but none have shown if this is the case for cognitive behavioral therapy or if successful treatment reduces the respiratory physiologic response to carbon dioxide. Methods: Twenty-five patients with panic disorder and 13 normal comparison subjects underwent baseline testing with 5% and 7% carbon dioxide inhalation. The patients were then retested after at least 12 weeks of treatment with either antipanic medication or cognitive behavioral therapy. Comparison subjects were retested after a similar interval. Results: Successful treatment resulted in lower panic rates, and reduced anxiogenic response. Treatment had no effect, however, on the respiratory physiologic response. Conclusions: There is dissociation in treatment response between the subjective and objective responses to carbon dioxide inhalation in panic disorder patients, with the former but not the latter showing positive change. We hypothesize that the strengthening of higher cortical control over subcortical fear-related structures, whether via medication or cognitive behavioral therapy treatment, results in less anxiety and fear in response to provoked symptoms reminiscent of naturally occurring panic.

Title: Ventilatory physiology of patients with panic disorder

Authors: Gorman JM, Fyer MR, Goetz R, Askanazi J, Liebowitz MR, Fyer AJ, Kinney J, Klein DF.

Journal: Arch Gen Psychiatry. 1988 Jan;45(1):31-9. doi: 10.1001/archpsyc.1988.01800250035006. Erratum in: Arch Gen Psychiatry 1991 Feb;48(2):181. PMID: 2827599.

Link to PubMed: Ventilatory physiology of patients with panic disorder

Abstract: Thirty-one patients with DSM-III panic disorder or agoraphobia with panic attacks, 13 normal controls, and 12 patients with other anxiety disorders were studied during ventilatory challenge with room air hyperventilation and 5% carbon dioxide inhalation. Patients also underwent sodium lactate infusion. Among the patients with panic disorder, 58% panicked with sodium lactate, 39% with 5% CO2, and 23% with room air hyperventilation. Of the other patients, four panicked with sodium lactate, none with 5% CO2, and one with room air hyperventilation. One normal control panicked with both sodium lactate and 5% CO2. Panic with CO2 was associated with an exaggerated ventilatory response and increases in plasma norepinephrine level and diastolic blood pressure. Patients with panic disorder may have hypersensitive CO2 receptors that, when triggered, evoke a subjective panic associated with an exaggerated ventilatory response and consequent hypocapnic alkalosis.

Title: CO2 Vulnerability in Panic Disorder

Authors: Griez EJ, Lousberg H, van den Hout MA, van der Molen GM.

Journal: Psychiatry Res. 1987 Feb;20(2):87-95. doi: 10.1016/0165-1781(87)90001-1. PMID: 3107009.

Link to PubMed: CO2 Vulnerability in Panic Disorder

Abstract: The immediate effects of a single inhalation of a 35% CO2 mixture in oxygen were examined in 12 patients with panic disorders and 11 normal control subjects. Compared to a placebo air inhalation, the CO2 inhalation provoked short-lived autonomic panic symptoms in both patients and normals; it also elicited high subjective anxiety in patients with panic disorders. The latter rated the overall CO2-induced state as very similar to a real-life panic attack.

Title: Carbon Dioxide Inhalation Induces Dose-Dependent and Age-Related Negative Affectivity

Authors: Griez EJ, Colasanti A, van Diest R, Salamon E, Schruers K.

Journal: PLoS One. 2007 Oct 3;2(10):e987. doi: 10.1371/journal.pone.0000987. PMID: 17912364; PMCID: PMC1991589.

Link to full text: Carbon Dioxide Inhalation Induces Dose-Dependent and Age-Related Negative Affectivity

Abstract: Background: Carbon dioxide inhalation is known to induce an emotion similar to spontaneous panic in Panic Disorder patients. The affective response to carbon dioxide in healthy subjects was not clearly characterized yet. Methodology/principal findings: Sixty-four healthy subjects underwent a double inhalation of four mixtures containing respectively 0, 9, 17.5 and 35% CO(2) in compressed air, following a double blind, cross-over, randomized design. Affective responses were assessed according to DSM IV criteria for panic, using an Electronic Visual Analogue Scale and the Panic Symptom List. It was demonstrated that carbon dioxide challenges induced a dose dependent negative affect (p<0.0001). This affect was semantically identical to the DSM IV definition of panic. Older individuals were subjectively less sensitive to Carbon Dioxide (p<0.05). Conclusions/significance: CO(2) induced affectivity may lay on a continuum with pathological panic attacks. Consistent with earlier suggestions that panic is a false biological alarm, the affective response to CO(2) may be part of a protective system triggered by suffocation and acute metabolic distress.

Title: Induction of c-Fos in 'panic/defence'-related brain circuits following brief hypercarbic gas exposure

Authors: Johnson PL, Fitz SD, Hollis JH, Moratalla R, Lightman SL, Shekhar A, Lowry CA.

Journal: J Psychopharmacol. 2011 Jan;25(1):26-36. doi: 10.1177/0269881109353464. Epub 2010 Jan 15. PMID: 20080924.

Link to PubMed: Induction of c-Fos in 'panic/defence'-related brain circuits following brief hypercarbic gas exposure

Abstract: Inspiration of air containing high concentrations of carbon dioxide (CO(2); hypercarbic gas exposure) mobilizes respiratory, sympathetic and hypothalamic-pituitary-adrenal axis responses and increases anxiety-like behaviour in rats and humans. Meanwhile the same stimulus induces panic attacks in the majority of panic disorder patients. However, little is known about the neural circuits that regulate these acute effects. In order to determine the effects of acute hypercarbic gas exposure on forebrain and brainstem circuits, conscious adult male rats were placed in flow cages and exposed to either atmospheric air or increasing environmental CO(2) concentrations (from baseline concentrations up to 20% CO(2)) during a 5 min period. The presence of immunoreactivity for the protein product of the immediate-early gene c-fos was used as a measure of functional cellular responses. Exposing rats to hypercarbic gas increased anxiety-related behaviour and increased numbers of c-Fos-immunoreactive cells in subcortical regions of the brain involved in: (1) the initiation of fear- or anxiety-associated behavioural responses (i.e. the dorsomedial hypothalamus, perifornical nucleus and dorsolateral and ventrolateral periaqueductal gray); (2) mobilization of the hypothalamic-pituitary-adrenal axis (i.e. the dorsomedial hypothalamus, perifornical nucleus and paraventricular hypothalamic nucleus); and (3) initiation of stress-related sympathetic responses (i.e. the dorsomedial hypothalamus, dorsolateral periaqueductal grey and rostroventrolateral medulla). These findings have implications for understanding how the brain senses changes in environmental CO(2) concentrations and the neural mechanisms underlying the subsequent adaptive changes in stress-related physiology and behaviour.

Title: Neural Pathways Underlying Lactate-Induced Panic

Authors: Johnson PL, Truitt WA, Fitz SD, Lowry CA, Shekhar A.

Journal: Neuropsychopharmacology. 2008 Aug;33(9):2093-107. doi: 10.1038/sj.npp.1301621. Epub 2007 Dec 5. PMID: 18059441; PMCID: PMC3065200.

Link to full text: Neural Pathways Underlying Lactate-Induced Panic

Abstract: Panic disorder is a severe anxiety disorder characterized by susceptibility to induction of panic attacks by subthreshold interoceptive stimuli such as 0.5 M sodium lactate infusions. Although studied for four decades, the mechanism of lactate sensitivity in panic disorder has not been understood. The dorsomedial hypothalamus/perifornical region (DMH/PeF) coordinates rapid mobilization of behavioral, autonomic, respiratory and endocrine responses to stress, and rats with disrupted GABA inhibition in the DMH/PeF exhibit panic-like responses to lactate, similar to panic disorder patients. Utilizing a variety of anatomical and pharmacological methods, we provide evidence that lactate, via osmosensitive periventricular pathways, activates neurons in the compromised DMH/PeF, which relays this signal to forebrain limbic structures such as the bed nucleus of the stria terminalis to mediate anxiety responses, and specific brainstem sympathetic and parasympathetic pathways to mediate the respiratory and cardiovascular components of the panic-like response. Acutely restoring local GABAergic tone in the DMH/PeF blocked lactate-induced panic-like responses. Autonomic panic-like responses appear to be a result of DMH/PeF-mediated mobilization of sympathetic responses (verified with atenolol) and resetting of the parasympathetically mediated baroreflex. Based on our findings, DMH/PeF efferent targets such as the C1 adrenergic neurons, paraventricular hypothalamus, and the central amygdala are implicated in sympathetic mobilization; the nucleus of the solitary tract is implicated in baroreflex resetting; and the parabrachial nucleus is implicated in respiratory responses. These results elucidate neural circuits underlying lactate-induced panic-like responses and the involvement of both sympathetic and parasympathetic systems.

Title: Sex differences in response to a panicogenic challenge procedure: an experimental evaluation of panic vulnerability in a non-clinical sample

Authors: Kelly MM, Forsyth JP, Karekla M.

Journal: Behav Res Ther. 2006 Oct;44(10):1421-30. doi: 10.1016/j.brat.2005.10.012. Epub 2005 Dec 20. PMID: 16364237.

Link to PubMed: Sex differences in response to a panicogenic challenge procedure: an experimental evaluation of panic vulnerability in a non-clinical sample

Abstract: Epidemiological studies show that women are twice as likely as men to develop panic disorder (PD) during their lifetimes. Data from retrospective studies also suggest that women are more distressed by panic and other negative emotional states than men, and that this tendency may precede the development of PD. The present prospective study sought to expand this work by evaluating sex differences in the predisposition to panic in individuals without PD or other forms of psychopathology. Participants (N=96; 52 women) were exposed to 12 panicogenic inhalations of 20% CO2-enriched air while physiological (e.g., heart rate, electrodermal response, Frontalis EMG) and self-report (e.g., subject distress, fear, frequency and severity of DSM-IV panic symptoms) response domains were monitored. As expected, magnitude of autonomic responding failed to reliably discriminate between women and men. Yet, women reported more fear and panic immediately following the challenge procedure relative to men, and this sex difference persisted when assessed again 30 min post-challenge. Implications of these results are discussed broadly in the context of biopsychosocial accounts of sex differences in the clinical presentation of PD.

Title: Carbon dioxide inhalation as a human experimental model of panic: The relationship between emotions and cardiovascular physiology

Authors: Leibold NK, Viechtbauer W, Goossens L, De Cort K, Griez EJ, Myin-Germeys I, Steinbusch HW, van den Hove DL, Schruers KR.

Journal: Biol Psychol. 2013 Oct;94(2):331-40. doi: 10.1016/j.biopsycho.2013.06.004. Epub 2013 Jun 28. PMID: 23816952.

Link to PubMed: Carbon dioxide inhalation as a human experimental model of panic: The relationship between emotions and cardiovascular physiology

Abstract: Inhaling carbon dioxide (CO2)-enriched air induces fear and panic symptoms resembling real-life panic attacks, the hallmark of panic disorder. The present study aimed to describe the emotional and cardiovascular effects evoked by inhaling CO2, taking shortcomings of previous studies into account. Healthy volunteers underwent a double inhalation of 0, 9, 17.5, and 35% CO2, according to a randomized, cross-over design. In addition to fear, discomfort, and panic symptom ratings, blood pressure and heart rate were continuously monitored. Results showed a dose-dependent increase in all self-reports. Systolic and diastolic blood pressure rose with increasing CO2 concentration, whereas heart rate results were less consistent. Diastolic blood pressure and heart rate variation correlated with fear and discomfort. Based on this relationship and the observation that the diastolic blood pressure most accurately mimicked the degree of self-reported emotions, it might serve as a putative biomarker to assess the CO2-reactivity in the future.

Title: Avoidance of 20% carbon dioxide-enriched air with humans

Authors: Lejuez CW, O'Donnell J, Wirth O, Zvolensky MJ, Eifert GH.

Journal: J Exp Anal Behav. 1998 Jul;70(1):79-86. doi: 10.1901/jeab.1998.70-79. PMID: 9684345; PMCID: PMC1284669.

Link to full text: Avoidance of 20% carbon dioxide-enriched air with humans

Abstract: Four college students were exposed to a Sidman avoidance procedure to determine if an avoidance contingency involving 20% carbon dioxide-enriched air (CO2) would produce and maintain responding. In Phase 1, two conditions (contingent and noncontingent) were conducted each day. These conditions were distinguished by the presence or absence of a blue or green box on a computer screen. In the contingent condition, CO2 presentation were delivered every 3 s unless a subject pulled a plunger. Each plunger pull postponed CO2 presentations for 10 s. In the noncontingent condition, CO2 presentations occurred on the average of every 5 min independent of responding. Following stable responding in Phase 1, condition-correlated stimuli were reversed. In both conditions, plunger response rate was high during the contingent condition and low or zero during the noncontingent condition. Furthermore, subjects avoided most CO2 presentations. However, CO2 presentations did not increase verbal reports of fear. Overall, the results from the present study suggest that CO2 can be used effectively in basic studies of aversive control and in laboratory analogues of response patterns commonly referred to as anxiety.

Title: Examination of mask disturbance behavior during a carbon dioxide-enriched air challenge

Authors: Marshall-Berenz EC, Gonzalez A, Leyro TM, Zvolensky MJ.

Journal: J Behav Ther Exp Psychiatry. 2011 Sep;42(3):253-7. doi: 10.1016/j.jbtep.2011.01.004. Epub 2011 Jan 12. PMID: 21349246; PMCID: PMC3086949.

Link to full text: Examination of mask disturbance behavior during a carbon dioxide-enriched air challenge

Abstract: Background and objectives: The present investigation examined mask disturbance behavior in the context of a biological challenge. Method: Participants included 128 adults (63.3% women; M(age) = 23.2, SD = 8.9) who underwent a 10% carbon dioxide-enriched air challenge. Results: Mask disturbance behavior, including complete or partial mask removal, was common during the challenge. Moreover, mask removal behavior during the challenge was significantly related to a greater increase in anxiety pre- to post-challenge, as well as greater levels of self-reported avoidance of a future challenge administration. However, mask touching and lifting behaviors were not related to other challenge criterion variables. Limitations: The current study consisted primarily of young adults and did not include individuals with a panic disorder diagnosis. Replication and extension of the current findings is recommended. Conclusions: Results suggest that mask removal behavior during the challenge may represent a possible behavioral marker of panic-relevant risk, although further work is necessary to determine its usefulness as a challenge measure.

Title: Shared Variance Among Self-Report and Behavioral Measures of Distress Intolerance

Authors: McHugh RK, Daughters SB, Lejuez CW, Murray HW, Hearon BA, Gorka SM, Otto MW.

Journal: Cognit Ther Res. 2011 Jun;35(3):266-275. doi: 10.1007/s10608-010-9295-1. PMID: 23894216; PMCID: PMC3721199.

Link to full text: Shared Variance Among Self-Report and Behavioral Measures of Distress Intolerance

Abstract: Distress intolerance may be an important individual difference variable in understanding maladaptive coping responses across diagnostic categories. However, the measurement of distress intolerance remains inconsistent across studies and little evidence for convergent validity among existing measures is available. This study evaluated the overlap among self-report and behavioral measures of distress intolerance in four samples, including an unselected sample, a sample of patients with drug dependence, and two samples of cigarette smokers. Results suggested that the self-report measures were highly correlated, as were the behavioral measures; however, behavioral and self-report measures did not exhibit significant associations with each other. There was some evidence of domain specificity, with anxiety sensitivity demonstrating strong associations with somatic distress intolerance, and a lack of association between behavioral measures that elicit affective distress and those that elicit somatic distress. These findings highlight a potential divergence in the literature relative to the conceptualization of distress intolerance as either sensitivity to distress or as the inability to persist at a task when distressed. Further research is needed to elucidate the conceptualization and measurement of distress intolerance to facilitate future clinical and research applications of this construct.

Title: History of suffocation, state-trait anxiety, and anxiety sensitivity in predicting 35% carbon dioxide-induced panic

Authors: Monkul ES, Onur E, Tural U, Hatch JP, Alkın T, Yücel B, Fidaner H.

Journal: Psychiatry Res. 2010 Sep 30;179(2):194-7. doi: 10.1016/j.psychres.2009.06.015. Epub 2010 May 16. PMID: 20478634.

Link to PubMed: History of suffocation, state-trait anxiety, and anxiety sensitivity in predicting 35% carbon dioxide-induced panic

Abstract: The aim of this study was to examine the effects of history of suffocation, state-trait anxiety, and anxiety sensitivity on response to a 35% carbon dioxide (CO₂) challenge in panic disorder patients, their healthy first-degree relatives and healthy comparisons. Thirty-two patients with panic disorder, 32 first-degree relatives, and 34 healthy volunteers underwent the 35% CO₂ challenge. We assessed baseline anxiety with the Anxiety Sensitivity Index (ASI) and State-Trait Anxiety Inventory (STAI1), and panic symptoms with the Panic Symptom List (PSL III-R). A history of suffocation was associated with greater risk of CO₂ reactivity in the combined sample. Patients had more anxiety sensitivity and state and trait anxiety than relatives and healthy comparisons; the difference between relatives and healthy comparisons was not significant. In female patients, trait anxiety predicted CO₂-induced panic. Having a CO₂-sensitive panic disorder patient as a first-degree relative did not predict CO₂-induced panic in a healthy relative. History of suffocation may be an important predictor of CO₂-induced panic. Trait anxiety may have a gender-specific relation to CO₂ reactivity.

Title: Acute panicogenic, anxiogenic and dissociative effects of carbon dioxide inhalation in patients with post-traumatic stress disorder (PTSD)

Authors: Muhtz C, Yassouridis A, Daneshi J, Braun M, Kellner M.

Journal: J Psychiatr Res. 2011 Jul;45(7):989-93. doi: 10.1016/j.jpsychires.2011.01.009. Epub 2011 Feb 15. PMID: 21324483.

Link to PubMed: Acute panicogenic, anxiogenic and dissociative effects of carbon dioxide inhalation in patients with post-traumatic stress disorder (PTSD)

Abstract: Background: Increased anxiety and panic to inhalation of carbon dioxide (CO(2)) has been described in patients with anxiety disorders, especially panic disorder, compared to healthy subjects. Post-traumatic stress disorder (PTSD) has been hypothesised to resemble panic disorder and is currently classified as an anxiety disorder in DSM-IV. However, there are only very few data available about the sensitivity of patients with PTSD to CO(2). Methods: In 10 patients with PTSD, 10 sex- and age-matched healthy subjects and 8 patients with panic disorder we assessed anxiety, panic, dissociative and PTSD symptoms before and after a single vital capacity inhalation of 35% CO(2). Results: Patients with PTSD showed an increased anxiety, panic and dissociative reaction to the inhalation of 35% CO(2) compared to healthy participants. PTSD subjects' responses were indistinguishable from those of panic patients. Additionally, PTSD-typical symptoms like post-traumatic flashbacks were provoked in patients with PTSD after the inhalation of CO(2). Conclusions: In our sample, PTSD was associated with an increased CO(2) reactivity, pointing to an increased susceptibility of PTSD patients to CO(2) challenge.

Title: Panicogens in patients with Post-Traumatic Stress Disorder (PTSD)

Authors: Muhtz C, Wiedemann K, Kellner M.

Journal: Curr Pharm Des. 2012;18(35):5608-18. doi: 10.2174/138161212803530817. PMID: 22632476.

Link to PubMed: Panicogens in patients with Post-Traumatic Stress Disorder (PTSD)

Abstract: Symptom provocation has proved its worth for understanding the pathophysiology of diseases and in general for the development of new therapeutic approaches in the medical field. In the research of anxiety disorders, investigations using experimentally induced panic attacks by various agents, such as sodium lactate, carbon dioxide, cholezystokinine-tetrapetid etc., have a long tradition and allow the exploration of usually naturally occuring spontaneous psychopathological phenomena under controlled conditions. Post-Traumatic Stress Disorder (PTSD) is a prevalent disorder that can develop following exposure to an extreme traumatic event. In DSM-IV it is currently classified as an anxiety disorder and shares phenomenological similarities with panic disorder. The use of panicogenic challenge tests is also an interesting neurobiological approach to learn more about the nature of PTSD and may be a possibility to develop new therapeutic strategies for the treatment of PTSD symptoms. Not only panic anxiety, but also flashbacks and other dissociative symptoms can be provoked by several panicogens in PTSD. The purpose of this review is to evaluate studies using panicogens in PTSD. Methodological short-comings of current studies and needed directions of further research are discussed.

Title: 35% Carbon dioxide and breath-holding challenge tests in panic disorder: a comparison with spontaneous panic attacks

Authors: Nardi AE, Valença AM, Mezzasalma MA, Lopes FL, Nascimento I, Veras AB, Freire RC, de-Melo-Neto VL, Zin WA.

Journal: Depress Anxiety. 2006;23(4):236-44. doi: 10.1002/da.20165. PMID: 16528718.

Link to PubMed: 35% Carbon dioxide and breath-holding challenge tests in panic disorder: a comparison with spontaneous panic attacks

Abstract: Respiration and its control mechanisms may represent an important system involved in abnormal anxiety. Our aim was to compare the demographic and clinical features of patients with panic disorder (PD) with agoraphobia (DSM-IV) who had a panic attack after both the 35% carbon dioxide (CO(2)) test and the breath-holding test (CPA group), and compare them with PD patients who did not have a panic attack after both tests (NPA group). We examined 76 patients with PD who were administered a 35% CO(2)test and a breath-holding test within a 1-week interval. Anxiety scales were applied before and after each test. A panic attack was induced in 50 (65.8%) patients during the CO(2)test (chi(2) = 28.44, df = 1, P<.001) and in 40 (52.6%) patients during the breath-holding test (chi(2) = 15.35, df = 1, P = .036). All patients who had a panic attack during the breath-holding test also had a panic attack during the CO(2)test (n = 40; CPA group). Twenty-six (34.2%) patients with PD did not have a panic attack after both respiratory tests (NPA group). The CPA group had more (chi(2) = 21.67, df = 1, P = .011) respiratory PD subtype. In the CPA group, the disorder started earlier (Mann-Whitney, P<.001), had a higher familial prevalence of PD (chi(2) = 18.34, df = 1, P = .028), and had more previous depressive episodes (chi(2) = 23.59, df = 1, P<.001). Our data suggest that there is an association between respiratory PD subtype and the response to respiratory challenge tests: CO(2)and breath-holding. The CPA may be confirmed as a subgroup of respiratory PD subtype.

Title: Sex differences in panic-relevant responding to a 10% carbon dioxide-enriched air biological challenge

Authors: Nillni YI, Berenz EC, Rohan KJ, Zvolensky MJ.

Journal: J Anxiety Disord. 2012 Jan;26(1):165-72. doi: 10.1016/j.janxdis.2011.10.008. Epub 2011 Nov 6. PMID: 22115836; PMCID: PMC3254702.

Link to full text: Sex differences in panic-relevant responding to a 10% carbon dioxide-enriched air biological challenge

Abstract: The current study examined sex differences in psychological (i.e., self-reported anxiety, panic symptoms, and avoidance) and physiological (i.e., heart rate and skin conductance level) response to, and recovery from, a laboratory biological challenge. Participants were a community-recruited sample of 128 adults (63.3% women; M(age)=23.2 years, SD=8.9) who underwent a 4-min 10% CO(2)-enriched air biological challenge. As predicted, women reported more severe physical panic symptoms and avoidance (i.e., less willingness to participate in another challenge) and demonstrated increased heart rate as compared to men above and beyond the variance accounted for by other theoretically relevant variables (recent panic attack history, neuroticism, and anxiety sensitivity). Additionally, women demonstrated a faster rate of recovery with respect to heart rate compared to men. These results are in line with literature documenting sex-specific differences in panic psychopathology, and results are discussed in the context of possible mechanisms underlying sex differences in panic vulnerability.

Title: The Relationships between Adverse Events, Early Antecedents, and Carbon Dioxide Reactivity as an Intermediate Phenotype of Panic Disorder

Authors: Ogliari A, Tambs K, Harris JR, Scaini S, Maffei C, Reichborn-Kjennerud T, Battaglia M.

Journal: Psychother Psychosom. 2010;79(1):48-55. doi: 10.1159/000259417. Epub 2009 Nov 18. PMID: 19923875.

Link to PubMed: The Relationships between Adverse Events, Early Antecedents, and Carbon Dioxide Reactivity as an Intermediate Phenotype of Panic Disorder

Abstract: Background: Although adverse events have been consistently described to precede and potentially precipitate the onset of panic disorder, there is no information about their ability to alter the individual reactivity to inhaled carbon dioxide, a putative intermediate phenotype of susceptibility to panic disorder. Method: Seven-hundred twelve subjects belonging to the general population-based Norwegian Institute of Public Health Twin Panel underwent a 35% CO(2)/65% O(2) inhalation challenge test and interview-based lifetime assessments of DSM-IV panic disorder, separation anxiety disorder, childhood parental separation/loss, major life events, adverse events of suffocative nature and common stressful life events. Regression models were applied to predict global subjective anxiety and DSM-IV panic symptoms after 35% CO(2)/65% O(2) inhalation. Results: The responses to the challenge measured as semicontinuous variables were predicted by symptoms of childhood separation anxiety, childhood parental loss, common stressful events, major life events, suffocative events and the female gender. The role of most of these predictors was confirmed and held true after the exclusion of subjects with lifetime panic attacks/disorder from the analyses. Conclusions: Several factors which have been reported by previous clinical studies to influence the individual susceptibility to develop panic disorder seem to affect the individual reactivity to inhaled carbon dioxide in people from the general population. Some elements of risk may impact simultaneously upon the individual liability to panic and exaggerated sensitivity to hypercapnia.

Title: Respiratory Psychophysiology of Panic Disorder: Three Respiratory Challenges in 98 Subjects

Authors: Papp LA, Martinez JM, Klein DF, Coplan JD, Norman RG, Cole R, de Jesus MJ, Ross D, Goetz R, Gorman JM.

Journal: Am J Psychiatry. 1997 Nov;154(11):1557-65. doi: 10.1176/ajp.154.11.1557. PMID: 9356564.

Link to PubMed: Respiratory Psychophysiology of Panic Disorder: Three Respiratory Challenges in 98 Subjects

Abstract: Objective: Respiratory abnormalities may play a central role in the pathophysiology of panic disorder. The current study was undertaken to examine the respiratory response in the largest series of subjects to date during three respiratory challenges that used improved methodology. Method: Fifty-nine patients with DSM-III-R panic disorder and 39 normal volunteers were challenged with 5% and 7% CO2 inhalation and room air hyperventilation separated by room air breathing with continuous spirometry. Results: Patients with panic disorder were more sensitive to the anxiogenic effects of CO2 than were normal subjects, and CO2 was a more potent stimulus to panic than hyperventilation. Patients increased their respiratory rate more quickly during CO2 inhalation than did comparison subjects, and this increase preceded the panic attacks. Patients who panicked in response to 5% CO2 demonstrated continued rise in end-tidal CO2, while the end-tidal CO2 of the comparison groups stabilized. Low end-tidal CO2 and high variance in minute ventilation at baseline predicted panic attacks during CO2 inhalation. Following CO2 or hyperventilation challenges, respiratory rate dropped sharply, while tidal volume remained elevated longer in patients than in comparison subjects. Conclusions: The findings confirm the greater behavioral and physiological sensitivity of patients with panic disorder to CO2 inhalation and identify a series of respiratory abnormalities. Panic attacks in panic disorder may be explained by inefficient compensatory mechanisms, primarily of respiratory rate.

Title: Diagnostic and substance specificity of carbon-dioxide-induced panic

Authors: Papp LA, Klein DF, Martinez J, Schneier F, Cole R, Liebowitz MR, Hollander E, Fyer AJ, Jordan F, Gorman JM.

Journal: Am J Psychiatry. 1993 Feb;150(2):250-7. doi: 10.1176/ajp.150.2.250. PMID: 8123056.

Link to PubMed: Diagnostic and substance specificity of carbon-dioxide-induced panic

Abstract: Objective: The authors assessed the substance and diagnostic specificity of carbon-dioxide-induced panic since, in addition to the specific biochemical effects of inhaled carbon dioxide (CO2), simple physiologic distress is also frequently implicated as a panicogenic factor during respiratory challenge studies with CO2 in patients with anxiety disorders. Method: Eighteen patients with panic disorder, 20 with social phobia, and 23 psychiatrically normal subjects inhaled a mixture of 35% CO2 and 65% O2 for 30 seconds through a face mask. They also breathed for 30 seconds through a valve reducing the diameter of the airway. A double-blind, counterbalanced, randomized design was used. Results: In spite of important similarities between the two interventions, including the induction of equal amounts of subjective respiratory distress, carbon dioxide inhalation was significantly more potent than increased airway resistance in provoking panic in the anxiety disorder patients. The patients with panic disorder were significantly more sensitive to CO2 than were the patients with social phobia or the normal subjects. Conclusions: Carbon dioxide inhalation appears to have a specific panicogenic effect in panic patients that goes beyond simple breathlessness.

Title: Psychophysiological responses to CO2 inhalation

Authors: Pappens M, De Peuter S, Vansteenwegen D, Van den Bergh O, Van Diest I.

Journal: Int J Psychophysiol. 2012 Apr;84(1):45-50. doi: 10.1016/j.ijpsycho.2012.01.008. Epub 2012 Jan 20. PMID: 22265938.

Link to PubMed: Psychophysiological responses to CO2 inhalation

Abstract: Inhalation of CO(2)-enriched air has been used as a laboratory model for a number of anxiety disorders, such as general anxiety disorder and panic disorder. Because studies describing psychophysiological responses to this challenge are scarce, the present studies investigated skin conductance level, eyeblink startle, self-reported anxiety and fractional end-tidal carbon dioxide during inhalation of CO(2)-enriched air. In study 1, thirty-five healthy volunteers inhaled 7.5% CO(2) for 2min. In study 2, twenty healthy volunteers inhaled 20% CO(2) for 30s. Control groups (N=20 in each study) inhaled room air during the same time periods. Compared to room air breathing, both CO(2)-mixtures were associated with increases in skin conductance levels, self-reported anxiety and fractional end-tidal CO(2.) Eyeblink startles were inhibited during CO(2) compared to room air breathing in both experiments. Our findings suggest that inhalation of CO(2)-enriched air is associated with a circa-strike defensive response pattern, corroborating its application as an interoceptive, panic-relevant stimulus in fear research.

Title: Carbon dioxide/oxygen challenge test in panic disorder

Authors: Perna G, Battaglia M, Garberi A, Arancio C, Bertani A, Bellodi L.

Journal: Psychiatry Res. 1994 May;52(2):159-71. doi: 10.1016/0165-1781(94)90085-x. PMID: 7972572.

Link to PubMed: Carbon dioxide/oxygen challenge test in panic disorder

Abstract: The effects of a single inhalation of a 35% CO2/65% O2 gas mixture were examined in 71 patients with panic disorder with or without agoraphobia and 44 normal control subjects. Compared with the placebo condition, inhalation of air, the CO2/O2 mixture elicited a clear anxiety reaction only in panic disorder patients, who experienced a sudden rise of subjective anxiety as well as of several panic symptoms. Respiratory symptoms and the fear of dying best distinguished the patients from the control subjects. Baseline anxiety was not the key factor in explaining this differential reaction. The clinical features of panic disorder (namely, frequency of panic attacks, agoraphobia, anticipatory anxiety, and duration of illness) were not significantly related to the response to the challenge test, suggesting that CO2 reactivity might be a trait marker of panic disorder.

Title: 35% CO2 challenge in panic and mood disorders

Authors: Perna G, Barbini B, Cocchi S, Bertani A, Gasperini M.

Journal: J Affect Disord. 1995 Mar 14;33(3):189-94. doi: 10.1016/0165-0327(94)00088-q. PMID: 7790671.

Link to PubMed: 35% CO2 challenge in panic and mood disorders

Abstract: 20 patients with Panic Disorder (PD), 19 patients with Mood Disorder (MD) and 20 healthy controls inhaled one vital capacity of 35% CO2-65% O2 gas mixture and of compressed air in a double-blind, random, cross-over design. Only PD patients showed a strong reaction to 35% CO2 while MD patients and controls did not react significantly. These results support the specificity of the 35% CO2 challenge in PD patients and suggest that PD and MD are separate disorders.

Title: Antipanic drug modulation of 35% CO2 hyperreactivity and short-term treatment outcome

Authors: Perna G, Bertani A, Caldirola D, Gabriele A, Cocchi S, Bellodi L.

Journal: J Clin Psychopharmacol. 2002 Jun;22(3):300-8. doi: 10.1097/00004714-200206000-00011. PMID: 12006901.

Link to PubMed: Antipanic drug modulation of 35% CO2 hyperreactivity and short-term treatment outcome

Abstract: Carbon dioxide (CO2) inhalation induces acute anxiety and panic attacks in patients with Panic Disorder (PD). Anti-panic drugs decrease CO2 reactivity after the first days of treatment; however, the clinical meaning of this finding has not yet been established. This study investigated the effects of treatment with tricyclic antidepressants and selective serotonin re-uptake inhibitors (SSRIs) on CO2 reactivity and compared the relationships between 35% CO2 hyperreactivity modulation and short-term clinical outcome. One hundred twenty-three patients with PD with or without agoraphobia who were hyperreactive to CO2 were randomly assigned to treatment groups with imipramine, clomipramine, paroxetine, sertraline, or fluvoxamine. A double-blind, randomized design was applied. Each patient received the 35% CO2 challenge on days 0, 7, and 30. The severity of clinical symptomatology was measured on days 0 and 30. Decreased hyperreactivity to 35% CO2 in all five treatment groups was already evident after the first week. The decrease in CO2 reactivity at the end of treatment was proportional to the degree of clinical improvement. Multiple regression analyses showed that the decrease in CO2 reactivity after the first week was a significant predictor for good clinical outcome after one month. The results of this study confirm evidence that psychoactive drugs effective in the treatment of PD decrease CO2 hyperreactivity. They also suggest that precocious modulation of CO2 reactivity might fairly reliably predict short-term clinical outcome in patients with "respiratory" PD.

Title: Sensitivity to 35% CO2 in healthy first-degree relatives of patients with panic disorder

Authors: Perna G, Cocchi S, Bertani A, Arancio C, Bellodi L.

Journal: Am J Psychiatry. 1995 Apr;152(4):623-5. doi: 10.1176/ajp.152.4.623. PMID: 7694916.

Link to PubMed: Sensitivity to 35% CO2 in healthy first-degree relatives of patients with panic disorder

Abstract: Objective: The authors tested the hypothesis that hyperreactivity to CO2 in healthy subjects represents an underlying familial vulnerability to panic disorder. Method: One vital-capacity inhalation of 35% CO2 and 65% O2 was administered to each of 84 patients with panic disorder, 23 healthy first-degree relatives of probands with panic disorder, and 44 healthy subjects with no family history of panic disorder. Results: The first-degree relatives of the probands with panic disorder reacted significantly more than the healthy subjects and significantly less than the probands. Conclusions: These findings suggest an association between family history of panic disorder and hyperreactivity to 35% CO2 in healthy subjects.

Title: Differential carbon dioxide sensitivity in childhood anxiety disorders and nonill comparison group

Authors: Pine DS, Klein RG, Coplan JD, Papp LA, Hoven CW, Martinez J, Kovalenko P, Mandell DJ, Moreau D, Klein DF, Gorman JM.

Journal: Arch Gen Psychiatry. 2000 Oct;57(10):960-7. doi: 10.1001/archpsyc.57.10.960. PMID: 11015814.

Link to PubMed: Differential carbon dioxide sensitivity in childhood anxiety disorders and nonill comparison group

Abstract: Background: To examine the relationship between respiratory regulation and childhood anxiety disorders, this study considered the relationship between anxiety disorders and symptoms during carbon dioxide (CO(2)) exposure, CO(2) sensitivity in specific childhood anxiety disorders, and the relationship between symptomatic and physiological responses to CO(2). Methods: Following procedures established in adults, 104 children (aged 9-17 years), including 25 from a previous study, underwent 5% CO(2) inhalation. The sample included 57 probands with an anxiety disorder (social phobia, generalized anxiety disorder, separation anxiety disorder, and panic disorder) and 47 nonill comparison subjects. Symptoms of anxiety were assessed before, during, and after CO(2) inhalation. Results: All children tolerated the procedure well, experiencing transient or no increases in anxiety symptoms. Children with an anxiety disorder, particularly separation anxiety disorder, exhibited greater changes in somatic symptoms during inhalation of CO(2)-enriched air, relative to the comparison group. During CO(2) inhalation, symptom ratings were positively correlated with respiratory rate increases, as well as with levels of tidal volume, minute ventilation, end-tidal CO(2), and irregularity in respiratory rate during room-air breathing. Conclusions: Childhood anxiety disorders, particularly separation anxiety disorder, are associated with CO(2) hypersensitivity, as defined by symptom reports. Carbon dioxide hypersensitivity is associated with physiological changes similar to those found in panic disorder. These and other data suggest that certain childhood anxiety disorders may share pathophysiological features with adult panic disorder.

Title: Response to 5% carbon dioxide in children and adolescents: relationship to panic disorder in parents and anxiety disorders in subjects

Authors: Pine DS, Klein RG, Roberson-Nay R, Mannuzza S, Moulton JL 3rd, Woldehawariat G, Guardino M.

Journal: Arch Gen Psychiatry. 2005 Jan;62(1):73-80. doi: 10.1001/archpsyc.62.1.73. PMID: 15630075.

Link to PubMed: Response to 5% carbon dioxide in children and adolescents: relationship to panic disorder in parents and anxiety disorders in subjects

Abstract: Background: Carbon dioxide (CO(2)) sensitivity is postulated to be a familial risk marker of panic disorder (PD). Exaggerated responses to CO(2) inhalation have been reported in adults with PD and their unaffected adult relatives, as well as in clinic-referred children with anxiety disorders.

Objective: To test in a family-based design whether CO(2) hypersensitivity is a familial risk marker for PD and associated with current anxiety disorders in children and adolescents.

Setting and participants: One hundred forty-two offspring (aged 9-19 years) of parents with PD, major depressive disorder, or no disorder. Forty-five (32%) had a current anxiety disorder, excluding specific phobia.

Design and main outcome measures: Parents and offspring received diagnostic assessments. Offspring underwent 5% CO(2) inhalation at home. Panic symptoms and panic attacks were rated with the Acute Panic Inventory at baseline, while anticipating CO(2) delivery ("threat"), and during CO(2) inhalation. Respiratory rate and volume were measured with spirometry.

Results: No group differences were found in Acute Panic Inventory ratings at baseline or in respiratory measures during threat. Risk for PD was not associated with CO(2) sensitivity (panic symptoms and respiratory physiologic response). During CO(2) inhalation, offspring with anxiety disorders, relative to offspring without anxiety disorders, experienced significantly more panic symptoms and panic attacks, as well as elevated respiratory rates. During threat, panic symptoms were significantly and independently associated with both parental PD and offspring anxiety disorders.

Conclusions: No support was obtained for CO(2) hypersensitivity as a familial risk marker for PD in children and adolescents. Links between childhood anxiety disorders and CO(2) sensitivity were replicated. Familial risk for PD in children and adolescents may be associated with vulnerability to anticipatory anxiety.

Title: Cognitive mediation in the affective component of spontaneous panic attacks

Authors: Rapee R, Mattick R, Murrell E.

Journal: J Behav Ther Exp Psychiatry. 1986 Dec;17(4):245-53. doi: 10.1016/0005-7916(86)90059-5. PMID: 3100580.

Link to PubMed: Cognitive mediation in the affective component of spontaneous panic attacks

Abstract: Single inhalations of a 50% carbon dioxide/50% oxygen gas mixture were administered to 16 subjects with spontaneous panic attacks and to 16 social phobics who did not experience such attacks. Half of each diagnostic category was randomly allocated to either a no explanation condition in which minimal instructions on expected outcome were provided, or an explanation condition in which all possible sensations were described and attributed to the effects of the gas. Subjects with panic attacks who were given no explanation reported a greater proportion of catastrophic cognitions, greater panic, and a greater similarity of the overall experience to a naturally-occurring panic attack than those with panic attacks who received a full explanation. In contrast, both groups of social phobics reported similar effects to each other, regardless of the explanation given. The results provide support for cognitive mediation in the "panic" component of spontaneous panic attacks.

Title: Response to hyperventilation and inhalation of 5.5% carbon dioxide-enriched air across the DSM-III-R anxiety disorders

Authors: Rapee RM, Brown TA, Antony MM, Barlow DH.

Journal: J Abnorm Psychol. 1992 Aug;101(3):538-52. doi: 10.1037//0021-843x.101.3.538. PMID: 1500611.

Link to PubMed: Response to hyperventilation and inhalation of 5.5% carbon dioxide-enriched air across the DSM-III-R anxiety disorders

Abstract: Anxiety disorder patients (n = 198; under criteria of the Diagnostic and Statistical Manual of Mental Disorders; rev. 3rd ed.; American Psychiatric Association, 1987) and nonanxious control subjects (n = 25) underwent challenges of 90 s of voluntary hyperventilation and 15 min of 5.5% carbon dioxide in air. Panic disorder subjects showed a greater subjective response to both challenges than did subjects with other anxiety disorders, who in turn responded more than did control subjects. Furthermore, subjects with panic disorder as an additional diagnosis tended to report more subjective response than did anxiety disorder subjects without panic disorder. The best prechallenge predictor of response to each procedure was a measure of fear of physical symptoms. The findings support previous results that have pointed to a greater fear or anxiety-inducing effect of these challenge procedures in panic disorder patients, as compared with other subjects.

Title: Evidence for distinct genetic effects associated with response to 35% CO₂

Authors: Roberson-Nay R, Moruzzi S, Ogliari A, Pezzica E, Tambs K, Kendler KS, Battaglia M.

Journal: Depress Anxiety. 2013 Mar;30(3):259-66. doi: 10.1002/da.22038. Epub 2013 Jan 24. PMID: 23349098; PMCID: PMC4096694.

Link to full text: Evidence for distinct genetic effects associated with response to 35% CO₂

Abstract: Background: Carbon dioxide (CO2 ) hypersensitivity represents an individual difference response to breathing CO2 enriched air. People with a history of panic attacks or panic disorder are particularly prone to anxious response, suggesting that CO2 hypersensitivity is a robust risk marker of panic spectrum vulnerability.

Methods: Twin pairs (n = 346) from the general population-based Norwegian NIPH Mental Health Study completed a measure of anxiety before and after vital capacity inhalation of 35% CO2 air and before and after inhalation of regular air. Three hypotheses regarding genetic factors for CO2 hypersensitivity were examined: (1) a single set of genetic risk factors impacts anxiety before exposure to CO2 and these same genes constitute the only genetic influences on anxiety in response to CO2 , (2) the genetic effects on pre-CO2 anxiety are entirely different from the genetic effects on anxiety in response to exposure to CO2 (i.e., new genetic effects), and (3) pre-CO2 anxiety influences anxiety in response to CO2 as well as unique genetic factors that become activated by respiratory stimulation.

Results: Our results support the latter hypothesis for response to 35% CO2 , with additive genetic and unique environmental factors best fitting the data. Evidence of new genetic effects was observed, accounting for 20% unique variance in post 35% CO2 anxiety response. New genetic effects were not observed for anxiety ratings made post regular air where only preregular air anxiety ratings explained significant variance in this outcome.

Conclusions: These data suggest that there are distinct genetic factors associated with responsivity to respiratory stimulation via 35% CO2 .

Title: Examining the latent class structure of CO2 hypersensitivity using time course trajectories of panic response systems

Authors: Roberson-Nay R, Beadel JR, Gorlin EI, Latendresse SJ, Teachman BA.

Journal: J Behav Ther Exp Psychiatry. 2015 Jun;47:68-76. doi: 10.1016/j.jbtep.2014.10.013. Epub 2014 Nov 15. PMID: 25496936; PMCID: PMC4324118.

Link to full text: Examining the latent class structure of CO2 hypersensitivity using time course trajectories of panic response systems

Abstract: Background and objectives: Carbon dioxide (CO2) hypersensitivity is hypothesized to be a robust endophenotypic marker of panic spectrum vulnerability. The goal of the current study was to explore the latent class trajectories of three primary response systems theoretically associated with CO2 hypersensitivity: subjective anxiety, panic symptoms, and respiratory rate (fR).

Methods: Participants (n = 376; 56% female) underwent a maintained 7.5% CO2 breathing task that included three phases: baseline, CO2 air breathing, and recovery. Growth mixture modeling was used to compare response classes (1…n) to identify the best-fit model for each marker. Panic correlates also were examined to determine class differences in panic vulnerability.

Results: For subjective anxiety ratings, a three-class model was selected, with individuals in one class reporting an acute increase in anxiety during 7.5% CO2 breathing and a return to pre-CO2 levels during recovery. A second, smaller latent class was distinguished by elevated anxiety across all three phases. The third class reported low anxiety reported during room air, a mild increase in anxiety during 7.5% CO2 breathing, and a return to baseline during recovery. Latent class trajectories for fR yielded one class whereas panic symptom response yielded two classes.

Limitations: This study examined CO2 hypersensitivity in one of the largest samples to date, but did not ascertain a general population sample thereby limiting generalizability. Moreover, a true resting baseline measure of fR was not measured.

Conclusions: Two classes potentially representing different risk pathways were observed. Implications of results will be discussed in the context of panic risk research.

Keywords: Anxiety; Carbon dioxide hypersensitivity; Latent class; Panic; Respiratory; Risk.

Title: The influence of an illusion of control on panic attacks induced via inhalation of 5.5% carbon dioxide-enriched air

Authors: Sanderson WC, Rapee RM, Barlow DH.

Journal: Arch Gen Psychiatry. 1989 Feb;46(2):157-62. doi: 10.1001/archpsyc.1989.01810020059010. PMID: 2492423.

Link to PubMed: The influence of an illusion of control on panic attacks induced via inhalation of 5.5% carbon dioxide-enriched air

Abstract: The current study tested the notion that a sense of control can mitigate anxiety and panic attacks caused by the inhalation of 5.5% carbon dioxide (CO2)-enriched air. Twenty patients with panic disorder inhaled a mixture of 5.5% CO2-enriched air for 15 minutes. All patients were instructed that illumination of a light directly in front of them would signal that they could decrease the amount of CO2 that they were receiving, if desired, by turning a dial attached to their chair. For ten patients, the light was illuminated during the entire administration of CO2. For the remaining ten patients, the light was never illuminated. In fact, all patients experienced the full CO2 mixture, and the dial was ineffective. When compared with patients who believed they had control, patients who believed they could not control the CO2 administration (1) reported a greater number of DSM-III-revised panic attack symptoms, (2) rated the symptoms as more intense, (3) reported greater subjective anxiety, (4) reported a greater number of catastrophic cognitions, (5) reported a greater resemblance of the overall inhalation experience to a naturally occurring panic attack, and (6) were significantly more likely to report panic attacks. These data illustrate the contribution of psychologic factors to laboratory induction of panic attacks through inhalation of 5.5% CO2-enriched air.

Title: Extinction of panicogenic effects of a 35% CO2 challenge in patients with panic disorder

Authors: Schmidt NB, Trakowski JH, Staab JP.

Journal: J Abnorm Psychol. 1997 Nov;106(4):630-8. doi: 10.1037//0021-843x.106.4.630. PMID: 9358693.

Link to PubMed: Extinction of panicogenic effects of a 35% CO2 challenge in patients with panic disorder

Abstract: Inhalations of high concentrations of carbon dioxide (CO2) reliably produce panic attacks in patients with panic disorder. The present study evaluated whether cognitive-behavioral treatment (CBT) for panic disorder would extinguish CO2-induced panic and whether changes in panic and arousal-related cognitions were associated with the induction of panic. Patients with panic disorder (N = 54) were assigned to 1 of 3 experimental conditions: CBT with respiratory training (CBT-R), CBT without respiratory training (CBT), or delayed treatment. Participants received 5 repeated vital-capacity inhalations of 35% CO2/65% O2 prior to and following either 12 treatment sessions or a 12-week waiting period. During pretreatment assessments, 74% of patients experienced a panic attack during at least 1 inhalation. At posttreatment, only 20% of treated participants (CBT-R = 19%, CBT = 22%), compared with 64% of untreated participants, panicked. Forty-four percent of treated participants, compared with 0% of untreated participants, reported no anxiety during all posttreatment inhalations. Anxiety sensitivity as well as panic appraisals regarding the likelihood of panic and self-efficacy with coping with panic were significantly related to fearful responding to the CO2 challenge.

Title: Biological Challenge Manipulation of PCO2 Levels: A Test of Klein's (1993) Suffocation Alarm Theory of Panic

Authors: Schmidt NB, Telch MJ, Jaimez TL.

Journal: J Abnorm Psychol. 1996 Aug;105(3):446-54. doi: 10.1037//0021-843x.105.3.446. PMID: 8772015.

Link to PubMed: Biological Challenge Manipulation of PCO2 Levels: A Test of Klein's (1993) Suffocation Alarm Theory of Panic

Abstract: D.F. Klein (1993) proposed that patients with panic disorder (PD) have a hypersensitive suffocation monitor that predisposes them to experience panic attacks under certain conditions. The suffocation alarm theory predicts differential emotional responding to biological challenges that affect arterial partial pressure of carbon dioxide (PCO2). These PD patients should exhibit (a) lower fear and less likelihood of panic in response to biological challenges that lower PCO2 levels (e.g., hyperventilation), and (b) increased fear and greater likelihood of panic in response to biological challenges that raise PCO2 levels (e.g., inhalation of 35% CO2 gas). The following indicators of the suffocation monitor were assessed: (a) severity of dyspnea symptoms, (b) frequency of dyspnea symptoms, (c) heightened respiration rate, and (d) lowered PCO2 levels. Ratings of physiological and subjective responding, as well as panic, were obtained during both a hyperventilation and a 35% CO2 challenge. None of the classification methods predicted differential emotional responding to hyperventilation versus 35% CO2 challenge.

Title: Genetic moderation of CO2-induced fear by 5-HTTLPR genotype

Authors: Schruers K, Esquivel G, van Duinen M, Wichers M, Kenis G, Colasanti A, Knuts I, Goossens L, Jacobs N, van Rozendaal J, Smeets H, van Os J, Griez E.

Journal: J Psychopharmacol. 2011 Jan;25(1):37-42. doi: 10.1177/0269881110372543. Epub 2010 Jun 28. PMID: 20584994.

Link to PubMed: Genetic moderation of CO2-induced fear by 5-HTTLPR genotype

Abstract: Inhalation of an increased concentration of carbon dioxide (CO(2)) has been shown to induce a state of negative affect in healthy subjects that is closely related to the clinical phenomenon of panic. It has been suggested that the vulnerability to CO(2) is moderated by differences in serotonin (5-HT) activity, caused by a functional polymorphism in the promoter region of the 5-HT transporter (5-HTTLPR) gene. Our aim was to examine the relationship between bi- and tri-allelic 5-HTTLPR genotype and the affective response to different dosages of inhaled CO(2) in healthy volunteers. Ninety-six subjects performed a double inhalation of four mixtures containing, respectively, 0%, 9%, 17.5% and 35% CO(2), following a double-blind, cross-over, randomized design. Affective responses were measured with a visual analogue scale for fear and the Panic Symptom List. 5-HTTLPR genotype was expressed as LL, SL and SS. Subjects with the SL and SS genotype reported less fear than LL subjects. A significant interaction effect was found between genotype and CO(2) dosage: the SS genotype showed lower fear scores than the LL genotype, particularly in the 17.5% CO(2) dose condition. The present study suggests that the dose-dependent fear reaction to CO(2) is moderated by a polymorphism in the 5-HT transporter gene, particularly at intermediate CO(2) dosages. It also underscores the usefulness of the introduction of an intermediate phenotype related to panic to reveal an underlying genetic vulnerability otherwise staying elusive. These results are in line with current theories on the role of 5-HT in both panic and respiration.

Title: Symptom profiles of natural and laboratory panic attacks

Authors: Schruers KR, van de Mortel H, Overbeek T, Griez E.

Journal: Acta Neuropsychiatr. 2004 Apr;16(2):101-6. doi: 10.1111/j.0924-2708.2004.0084.x. PMID: 26984003.

Link to PubMed: Symptom profiles of natural and laboratory panic attacks

Abstract: Background: Little accurate information is available about the symptomatology of real-life panic attacks and about how well they are reproduced by an experimental model such as the 35% CO2 challenge.

Method: Real-life panic symptoms were assessed in a group of 67 panic disorder patients, using daily life monitoring. Panic symptoms elicited by a 35% CO2 challenge were assessed in 61 panic disorder patients, and their frequency was compared with the real-life symptoms.

Results: The most frequent real-life symptoms were palpitations, dizziness and trembling. The 35% CO2 challenge reproduced well the majority of real-life symptoms.

Conclusion: The findings suggest that the 35% CO2 challenge is a marker for spontaneous panic attacks, which are considered the core of panic disorder.

Title: Effects of 7.5% CO2 challenge in generalized anxiety disorder

Authors: Seddon K, Morris K, Bailey J, Potokar J, Rich A, Wilson S, Bettica P, Nutt DJ.

Journal: J Psychopharmacol. 2011 Jan;25(1):43-51. doi: 10.1177/0269881110364270. Epub 2010 Mar 16. PMID: 20233897.

Link to PubMed: Effects of 7.5% CO2 challenge in generalized anxiety disorder

Abstract: We have previously developed a putative model of generalized anxiety disorder in healthy volunteers using a 20-minute 7.5% carbon dioxide (CO(2)) inhalation challenge. The aim of this study was to validate the 7.5% CO(2) paradigm by assessing its effects in patients with generalized anxiety disorder in a test-retest design. Twelve medication-free generalized anxiety disorder patients attended our lab for two study days. On each study day placebo (compressed air) and 7.5% CO(2) mixture were randomly administered over 20 min, at least 30 min apart, in a single blind, randomized, placebo-controlled cross-over design. Subjective ratings, cardiovascular measures and cortisol levels were collected throughout. CO(2) challenge significantly increased ratings for anxiety and other subjective symptoms associated with generalized anxiety disorder, compared with air. It also significantly increased systolic blood pressure on day 2, indicating increased autonomic arousal. There was no change between the two test days in mean anxiety rating scores, and there also appeared to be a correlation for individual scores on a number of the subjective measures. In conclusion, 20 min of 7.5% CO(2) gas inhalation increases anxiety responses in patients with generalized anxiety disorder, and this is reliable over time.

Title: Respiratory sensations in subjects who lack a ventilatory response to CO2

Authors: Shea SA, Andres LP, Shannon DC, Guz A, Banzett RB.

Journal: Respir Physiol. 1993 Aug;93(2):203-19. doi: 10.1016/0034-5687(93)90006-v. PMID: 8210759.

Link to PubMed: Respiratory sensations in subjects who lack a ventilatory response to CO2

Abstract: An urge to breath is perceived during breath hold and hypercapnia (termed 'air hunger') and during heavy exercise (often termed 'shortness of breath'). To better understand the neural mechanisms responsible for these sensations we studied five patients (8-17 years old) with congenital central hypoventilation syndrome (CCHS) who lack ventilatory response to CO2. CCHS patients reported no respiratory discomfort during CO2 inhalation or during maximal breath hold which was of much longer duration than age-matched controls. However, all 3 CCHS patients who exercised heavily reported some sensations akin to shortness of breath (they increased breathing nearly as much as controls). Our results are consistent with two possibilities. First, the air hunger of hypercapnia and breath hold is caused by projection to the forebrain of respiratory chemoreceptor afferents which bypass the respiratory centers, while exercise shortness of breath is caused by direct projections of limb afferents or locomotory center activity. Second, air hunger and shortness of breath share the same origin--projection of increased brain stem respiratory center motor activity (corollary discharge) to the forebrain.

Title: The interplay between physical activity and anxiety sensitivity in fearful responding to carbon dioxide challenge

Authors: Smits JA, Tart CD, Rosenfield D, Zvolensky MJ.

Journal: Psychosom Med. 2011 Jul-Aug;73(6):498-503. doi: 10.1097/PSY.0b013e3182223b28. Epub 2011 Jun 23. PMID: 21700713; PMCID: PMC3131468.

Link to full text: The interplay between physical activity and anxiety sensitivity in fearful responding to carbon dioxide challenge

Abstract: Objective: Physical activity may confer protective effects in the development of anxiety and its disorders. These effects may be particularly strong among individuals who have elevated levels of anxiety sensitivity (AS; i.e., the fear of somatic arousal), an established cognitive-based risk factor for anxiety and its disorders. The present study performed a laboratory test of the interplay between physical activity and AS.

Methods: The participants were adults free of Axis I psychopathology (n = 145) who completed measures of physical activity and AS before undergoing a recurrent 20% carbon dioxide-enriched air (CO(2)) challenge.

Results: Consistent with the hypothesis, physical activity was significantly related to CO(2) challenge reactivity among persons with elevated levels of AS, at high levels of physical activity (p < .001) but not at low levels of physical activity (p = .90). Also consistent with hypothesis, irrespective of the level of physical activity, physical activity did not relate significantly to CO(2) challenge reactivity among persons with normative levels of AS (p = .28).

Conclusions: These findings provide novel empirical insight into the role that physical activity may play in terms of resiliency for the development of anxiety disorders. Specifically, the protective effects of physical activity may only be evident at higher doses and among persons who are at increased risk of developing anxiety disorders because they have elevated AS.

Title: Unexpected arousal, anxiety sensitivity, and their interaction on CO₂-induced panic: further evidence for the context-sensitivity vulnerability model

Authors: Telch MJ, Harrington PJ, Smits JA, Powers MB.

Journal: J Anxiety Disord. 2011 Jun;25(5):645-53. doi: 10.1016/j.janxdis.2011.02.005. Epub 2011 Mar 3. PMID: 21474277.

Link to PubMed: Unexpected arousal, anxiety sensitivity, and their interaction on CO₂-induced panic: further evidence for the context-sensitivity vulnerability model

Abstract: The present experiment tested several predictions derived from the context-sensitivity vulnerability model of panic. Participants (N=79) scoring either high or low in anxiety sensitivity (AS) and with no history of unexpected panic were randomly assigned to one of two instructional sets: expected arousal (EA) or expected relaxation (ER). All participants were administered inhalation of room air and 35% CO(2) in a counterbalanced order. Consistent with theoretical predictions, High-AS participants who received ER instructions showed greater emotional responding compared to High-AS participants who received EA instructions, while instructional set did not affect responding among Low-AS participants. Panic attacks were observed in 52% of the High-AS-ER group compared to 17%, 5%, and 5% in the High-AS-EA, Low-AS-ER, and Low-AS-EA groups respectively. These findings are consistent with the theory's assertion that dispositional tendencies, such as anxiety sensitivity potentiate the panicogenic effects of threat-relevant context variables.

Title: Emotional reactivity to a single inhalation of 35% carbon dioxide and its association with later symptoms of posttraumatic stress disorder and anxiety in soldiers deployed to Iraq

Authors: Telch MJ, Rosenfield D, Lee HJ, Pai A.

Journal: Arch Gen Psychiatry. 2012 Nov;69(11):1161-8. doi: 10.1001/archgenpsychiatry.2012.8. PMID: 23117637.

Link to PubMed: Emotional reactivity to a single inhalation of 35% carbon dioxide and its association with later symptoms of posttraumatic stress disorder and anxiety in soldiers deployed to Iraq

Abstract: Context: The identification of modifiable predeployment vulnerability factors that increase the risk of combat stress reactions among soldiers once deployed to a war zone offers significant potential for the prevention of posttraumatic stress disorder (PTSD) and other combat-related stress disorders. Adults with anxiety disorders display heightened emotional reactivity to a single inhalation of 35% carbon dioxide (CO(2)); however, data investigating prospective linkages between emotional reactivity to CO(2) and susceptibility to war-zone stress reactions are lacking.

Objective: To investigate the association of soldiers' predeployment emotional reactivity to 35% CO(2) challenge with several indices of subsequent war-zone stress symptoms assessed monthly while deployed in Iraq.

Design, setting, and participants: Prospective cohort study of 158 soldiers with no history of deployment to a war zone were recruited from the Texas Combat Stress Risk Study between April 2, 2007, and August 28, 2009.

Main outcome measures: Multilevel regression models were used to investigate the association between emotional reactivity to 35% CO(2) challenge (assessed before deployment) and soldiers' reported symptoms of general anxiety/stress, PTSD, and depression while deployed to Iraq.

Results: Growth curves of PTSD, depression, and general anxiety/stress symptoms showed a significant curvilinear relationship during the 16-month deployment period. War-zone stressors reported in theater were associated with symptoms of general anxiety/stress, PTSD, and depression. Consistent with the prediction, soldiers' emotional reactivity to a single inhalation of 35% CO(2)-enriched air before deployment significantly potentiated the effects of war-zone stressors on the subsequent development of PTSD symptoms and general anxiety/stress symptoms but not on the development of depression, even after accounting for the effects of trait anxiety and the presence of past or current Axis I mental disorders.

Conclusion: Soldiers' emotional reactivity to a 35% CO(2) challenge may serve as a vulnerability factor for increasing soldiers' risk for PTSD and general anxiety/stress symptoms in response to war-zone stressors.

Title: Objective and subjective measures in recovery from a 35% carbon dioxide challenge

Authors: Niccolai V, van Duinen MA, Griez EJ.

Journal: Can J Psychiatry. 2008 Nov;53(11):737-44. doi: 10.1177/070674370805301105. PMID: 19087467.

Link to PubMed: Objective and subjective measures in recovery from a 35% carbon dioxide challenge

Abstract: Objectives: Because hyperventilation, dyspnea, and a feeling of choking are often core features of a panic attack, respiration has been one of the most widely studied physiological parameters in panic disorder (PD) patients. A respiratory subgroup of PD, with distinct etiological pathways, has also been suggested. Investigation of the recovery phase following a respiratory challenge may be a reliable way to establish respiratory impairment in PD patients. The objective of the present study was to investigate the recovery phase from a 35% carbon dioxide challenge in PD patients and in healthy controls, and to test the hypothesis of a different respiratory pattern in patients, compared to control subjects.

Methods: Eleven nonmedicated PD patients with or without agoraphobia, 11 medicated PD patients, and 11 control subjects took part in a 35% carbon dioxide and 65% oxygen inhalation challenge. Respiratory rate, partial pressure of carbon dioxide, heart rate, and blood pressure were recorded during the baseline phase (10 minutes) and the recovery phase (10 minutes). Visual Analogue Scale of Anxiety and Panic Symptom List scores were collected pre- and post-challenge.

Results: Nonmedicated patients had increased variability in respiratory rate and partial pressure of carbon dioxide during recovery, compared with control subjects and medicated PD patients. Also, PD patients tended to have higher heart rates and to need more time to recover from the challenge than control subjects.

Conclusions: Results suggest that PD patients have less effective homeostatic control after their physiological equilibrium has been disrupted by a respiratory stressor.

Title: Reactivity to a 35% CO2 challenge in healthy first-degree relatives of patients with panic disorder

Authors: Van Beek N, Griez E.

Journal: Biol Psychiatry. 2000 May 1;47(9):830-5. doi: 10.1016/s0006-3223(99)00265-6. PMID: 10812042.

Link to PubMed: Reactivity to a 35% CO2 challenge in healthy first-degree relatives of patients with panic disorder

Abstract: Background: The effects of a 35% CO2 challenge were examined in healthy first-degree relatives of panic disorder patients and in healthy control subjects matched for age and gender.

Methods: One single inhalation of a 35% CO2/65% O2 challenge was administered to 50 first-degree relatives of panic disorder patients and 50 control subjects.

Results: The first-degree relatives were more reactive to the 35% CO2 challenge than the control subjects.

Conclusions: These findings indicate that being a member of a family with a panic disorder patient is, in itself an important factor in CO2 hypersensitivity among subjects who have never experienced a panic attack. Both panic disorder patients and their first-degree relatives have a tendency to be more reactive to the CO2 challenge.

Title: Reliability of the 35% carbon dioxide panic provocation challenge

Authors: Verburg K, Pols H, de Leeuw M, Griez E.

Journal: Psychiatry Res. 1998 May 8;78(3):207-14. doi: 10.1016/s0165-1781(98)00009-2. PMID: 9657425.

Link to PubMed: Reliability of the 35% carbon dioxide panic provocation challenge

Abstract: The objective of this study was to determine the test-retest reliability of the 35% carbon dioxide (CO2) panic provocation challenge. Thirty patients with panic disorder were included in this study. Twenty-four patients were challenged twice, with 1 week between the two challenges. Six patients dropped out after the first test. The 35% CO2 challenge appeared to have a good test-retest reliability; both on induced subjective anxiety, measured on a Visual Analogue Scale for Anxiety (VAS-A), and induced panic symptoms, measured with a Panic Symptom List. Assessing the state of anxiety immediately after the challenge gave the most reliable results. Calculating increase in anxiety from the pre- and post-scores on the VAS-A rendered less reliable scores. This study completes a series of studies in which the criteria for an ideal model of panic are tested for the 35% CO2 challenge. Apart from an absolute specificity for panic disorder, the challenge meets these criteria.

Title: The amygdala is a chemosensor that detects carbon dioxide and acidosis to elicit fear behavior

Authors: Ziemann AE, Allen JE, Dahdaleh NS, Drebot II, Coryell MW, Wunsch AM, Lynch CM, Faraci FM, Howard MA 3rd, Welsh MJ, Wemmie JA.

Journal: Cell. 2009 Nov 25;139(5):1012-21. doi: 10.1016/j.cell.2009.10.029. PMID: 19945383; PMCID: PMC2808123.

Link to full text: The amygdala is a chemosensor that detects carbon dioxide and acidosis to elicit fear behavior

Abstract: The amygdala processes and directs inputs and outputs that are key to fear behavior. However, whether it directly senses fear-evoking stimuli is unknown. Because the amygdala expresses acid-sensing ion channel-1a (ASIC1a), and ASIC1a is required for normal fear responses, we hypothesized that the amygdala might detect a reduced pH. We found that inhaled CO(2) reduced brain pH and evoked fear behavior in mice. Eliminating or inhibiting ASIC1a markedly impaired this activity, and localized ASIC1a expression in the amygdala rescued the CO(2)-induced fear deficit of ASIC1a null animals. Buffering pH attenuated fear behavior, whereas directly reducing pH with amygdala microinjections reproduced the effect of CO(2). These data identify the amygdala as an important chemosensor that detects hypercarbia and acidosis and initiates behavioral responses. They also give a molecular explanation for how rising CO(2) concentrations elicit intense fear and provide a foundation for dissecting the bases of anxiety and panic disorders.

Title: The effects of offset control over 20% carbon-dioxide-enriched air on anxious responding

Authors: Zvolensky MJ, Eifert GH, Lejuez CW, McNeil DW.

Journal: J Abnorm Psychol. 1999 Nov;108(4):624-32. doi: 10.1037//0021-843x.108.4.624. PMID: 10609427.

Link to PubMed: The effects of offset control over 20% carbon-dioxide-enriched air on anxious responding

Abstract: Control over the offset of repeated administrations of 20% carbon-dioxide-enriched air was assessed in nonclinical participants (n = 30) reporting elevated levels of anxiety sensitivity--a population at an increased risk for experiencing panic attacks and possibly developing panic disorder. In Phase I, participants were randomly assigned to 1 of 2 conditions: one that permitted offset control over gas inhalation and one that did not. These conditions were reversed in Phase II. Across phases, a lack of offset control resulted in greater self-reported anxiety compared with having control, although no significant differences were observed for heart rate. Whereas all participants demonstrated a Stroop interference effect for general (e.g., coffin) compared with specific (e.g., dizzy) physical threat word types prior to the first experimental phase, this effect persisted only for participants who had offset control in Phase I. We discuss these results in relation to the differential effects of offset control, with implications for better understanding anxious responding during elevated bodily arousal.

Title: Offset control during recurrent 20% carbon dioxide-enriched air induction: relation to individual difference variables

Authors: Zvolensky MJ, Eifert GH, Lejuez CW.

Journal: Emotion. 2001 Jun;1(2):148-65. doi: 10.1037/1528-3542.1.2.148. PMID: 12899194.

Link to PubMed: Offset control during recurrent 20% carbon dioxide-enriched air induction: relation to individual difference variables

Abstract: Although control over aversive events maintains a central role in contemporary models of anxiety pathology, particularly panic disorder, there is little understanding about the emotional consequences of specific types of control processes. In the present study, offset control over 8 20% carbon dioxide-enriched air administrations was experimentally manipulated in a large nonclinical population (n = 96) varying in anxiety sensitivity (high or low) and gender. Dependent measures included self-reported anxiety, affective reports of valence, arousal, emotional control, and physiological indices of heart rate and skin conductance. High anxiety-sensitive participants who lacked offset control reported significantly greater elevations in self-reported anxiety, emotional displeasure, arousal, and dyscontrol relative to their yoked counterparts with offset control. In contrast, low anxiety-sensitive individuals responded with similar levels of cognitive and affective distress regardless of the offset control manipulation. Although the provocation procedure reliably produced bodily arousal relative to baseline, at a physiological level of analysis, no significant differences emerged across conditions. These findings are discussed in relation to offset control during recurrent interoceptive arousal, with implications for better understanding anxiety about abrupt bodily sensations.

Title: Cognitive therapy versus interoceptive exposure as treatment of panic disorder without agoraphobia

Authors: Arntz A.

Journal: Behav Res Ther. 2002 Mar;40(3):325-41. doi: 10.1016/s0005-7967(01)00014-6. PMID: 11863242.

Link to PubMed: Cognitive therapy versus interoceptive exposure as treatment of panic disorder without agoraphobia

Abstract: Cognitive therapy (CT) and interoceptive exposure (IE) as treatments of panic disorder without agoraphobia were compared in a sample of 69 patients, randomly allocated to condition. There were no significant differences between treatments as to reductions in panic frequency, daily anxiety levels and a composite questionnaire score, at posttest after the 12-session treatment, and at both follow-ups (4 weeks, 6 months). In both conditions, high percentages of patients were panic free at post and follow-up tests (range 75-92%). Although the reduction in idiosyncratic beliefs about the catastrophic nature of bodily sensations was equally strong in both conditions, post-treatment beliefs correlated strongly with symptoms at post and follow-up tests in the CT condition, but not in the IE condition. Reduction of beliefs may be essential in CT, but not in IE. This suggests that the two treatments utilize different change mechanisms.

Title: Anxiety sensitivity and interoceptive exposure: a transdiagnostic construct and change strategy

Authors: Boswell JF, Farchione TJ, Sauer-Zavala S, Murray HW, Fortune MR, Barlow DH.

Journal: Behav Ther. 2013 Sep;44(3):417-31. doi: 10.1016/j.beth.2013.03.006. Epub 2013 Apr 2. PMID: 23768669; PMCID: PMC3727659.

Link to full text: Anxiety sensitivity and interoceptive exposure: a transdiagnostic construct and change strategy

Abstract: Recent findings support the relevance of anxiety sensitivity (AS) and interoceptive exposure (IE) across emotional disorders. This study (a) evaluated levels of AS across different anxiety disorders, (b) examined change in AS over the course of transdiagnostic psychological intervention, and its relationship with outcome, and (c) described the implementation of IE to address AS with patients with different anxiety disorders. Participants (N=54) were patients who received treatment with the Unified Protocol for Transdiagnostic Treatment of Emotional Disorders (UP) in two consecutive treatment trials. Participants completed a measure of AS at pre- and posttreatment, and multiple occasions during treatment. Symptom severity was assessed at pre- and posttreatment, and clinical information related to physical symptoms and IE were collected as part of routine clinical practice. Elevated AS was observed at pretreatment across diagnoses and decreases in AS were observed from pre- to posttreatment. Similar changes occurred across the diagnostic categories, notably coinciding with the introduction of IE. Change in AS was correlated with reduced symptom levels at posttreatment and 6-month follow-up. Patients with different anxiety disorders endorsed similar physical symptoms and practiced similar IE exercises with similar effects. Results provide preliminary support for the usefulness of IE as a treatment strategy across the spectrum of anxiety disorders, and additional support for the transdiagnostic relevance of AS.

Title: Efficacy of interoceptive exposure therapy combined with trauma-related exposure therapy for posttraumatic stress disorder: a pilot study

Authors: Wald J, Taylor S.

Journal: J Anxiety Disord. 2007;21(8):1050-60. doi: 10.1016/j.janxdis.2006.10.010. Epub 2007 Jan 10. PMID: 17270392.

Link to PubMed: Efficacy of interoceptive exposure therapy combined with trauma-related exposure therapy for posttraumatic stress disorder: a pilot study

Abstract: The aim of this case series was to examine efficacy of interoceptive exposure (IE) combined with trauma-related exposure therapy (TRE) for posttraumatic stress disorder (PTSD). Seven participants completed treatment consisting of four weekly sessions of IE followed by eight weekly sessions of TRE (four sessions of imaginal exposure and four sessions of in vivo exposure). Assessments were conducted at pretreatment, posttreatment, 1- and 3-month follow-up. Outcome measures included PTSD symptoms, anxiety sensitivity, posttraumatic cognitions, anxiety, and depression. Five of the seven participants showed pre- to posttreatment improvements on these measures, and two participants showed less symptom reduction. Results at the 1-month follow-up showed that treatment gains were generally maintained in five (of the seven) participants and four of these individuals no longer met PTSD diagnostic criteria. Four individuals completed the 3-month follow-up and their symptoms and diagnostic status remained unchanged. These preliminary findings are promising. The next step in this line of research is to conduct a randomized, controlled trial to further to examine the efficacy, tolerability, and mechanisms of using IE in the treatment of PTSD.

Title: Responses to interoceptive exposure in people with posttraumatic stress disorder (PTSD): a preliminary analysis of induced anxiety reactions and trauma memories and their relationship to anxiety sensitivity and PTSD symptom severity

Authors: Wald J, Taylor S.

Journal: Cogn Behav Ther. 2008;37(2):90-100. doi: 10.1080/16506070801969054. PMID: 18470740.

Link to PubMed: Responses to interoceptive exposure in people with posttraumatic stress disorder (PTSD): a preliminary analysis of induced anxiety reactions and trauma memories and their relationship to anxiety sensitivity and PTSD symptom severity

Abstract: A growing body of evidence suggests that anxiety sensitivity (AS; fear of arousal-related sensations) plays a role in posttraumatic stress disorder (PTSD). Consistent with this, evidence indicates that interoceptive exposure (IE), which is a method for reducing AS, reduces PTSD symptoms. Clinical observations from our treatment studies indicate that IE triggers both anxiety and trauma memories in people with PTSD. The primary aim of this study was to describe the anxiety responses to a series of IE exercises and to examine whether or not trauma memories were activated. A secondary aim was to explore the relationships among AS, PTSD symptom severity, and IE responses. Data were collected from 23 people with PTSD who completed measures of PTSD symptoms and AS and a standardized battery of 10 IE exercises. Elevated anxiety and strong arousal responses were frequently elicited by the exercises, and trauma memories were also frequently triggered. AS and IE-triggered trauma memories significantly predicted IE-induced peak anxiety. The implications of the findings are discussed in terms of how IE might exert its therapeutic effects in the treatment of PTSD.

Title: Response to repeated CO2 in individuals with elevated anxiety sensitivity: replication with 20% CO2

Authors: Beck JG, Wolf MS.

Journal: J Behav Ther Exp Psychiatry. 2001 Mar;32(1):1-16. doi: 10.1016/s0005-7916(01)00018-0. PMID: 11729942.

Link to PubMed: Response to repeated CO2 in individuals with elevated anxiety sensitivity: replication with 20% CO2

Abstract: The present report replicates and extends previous work examining response patterns to repeated presentation of CO2. In previous studies, two distinct response patterns to repeated presentation of 35% CO2 were noted, representing habituation and nonhabituation of anxiety. In this report, 21 participants who had never experienced a panic attack but who reported high levels of anxiety sensitivity were presented with 12 trials of 20% CO2, followed by a trial involving inhalation of room air (to examine dishabituation) and two more trials of 20% CO2. Results indicated that 67% of the sample reported habituation of anxiety. Reductions in anxiety across inhalations were paralleled by changes in tidal volume, perceived panic symptom severity, and feelings of panic. Notable dishabituation was observed in the nonhabituation sample. Results are discussed in the light of basic learning processes underlying the treatment of Panic Disorder (PD).

Title: How does interoceptive exposure for panic disorder work? An uncontrolled case study

Authors: Beck JG, Shipherd JC, Zebb BJ.

Journal: Beck JG, Shipherd JC, Zebb BJ.

Link to PubMed: How does interoceptive exposure for panic disorder work? An uncontrolled case study

Abstract: To examine the influence of interoceptive exposure (IE) when used alone in the treatment of Panic Disorder (PD), 17 PD patients were presented with six IE sessions, using 35% CO2 as the exposure medium. The data indicate that IE alone is effective in reducing panic, panic-related fears, and general anxiety. However, the positive effects of IE do not appear to extend to agoraphobia, related fears, or depressed mood. Two distinct within-session patterns of fear response to IE were noted, one indicating habituation and the other indicating a lack of fear reduction. Although both patterns were associated with reductions in panic and anxiety following IE, the Habituators appeared to have a more positive outcome, which occurred more rapidly. These data suggest that IE may operate via two different pathways. Implications for understanding fear reduction are discussed, along with directions for future study.

Title: Repeated exposure to interoceptive cues: does habituation of fear occur in panic disorder patients? A preliminary report

Authors: Beck JG, Shipherd JC.

Journal: Behav Res Ther. 1997 Jun;35(6):551-7. doi: 10.1016/s0005-7967(97)00010-7. PMID: 9159979.

Link to PubMed: Repeated exposure to interoceptive cues: does habituation of fear occur in panic disorder patients? A preliminary report

Abstract: In order to explore the mechanism of action of interoceptive exposure, 12 Panic Disorder (PD) patients were presented with two sessions of repeated CO2 inhalation. Two distinct patterns of responding were noted. The first pattern was described as habituation of fear (n = 6). These patients showed decrements in pre- and post-inhalation anxiety during both sessions (with more rapid decline during session 2), as well as spontaneous recovery of fear at the onset of session 2. The second pattern indicated fear sensitization. These patients showed relatively low levels of anticipatory anxiety preceding CO2 inhalation during both sessions but reported robust increases in fear following gas inhalation. The extent of this increase was slightly less during session 2 relative to session I and did not appear to be mediated by cardiovascular arousal, as both groups showed rapid HR habituation during both sessions. Results are discussed in light of current theories of PD and its treatment.

Title: Fearful responding to repeated CO2 inhalation: a preliminary investigation

Authors: Beck JG, Shipherd JC, Zebb BJ.

Journal: Behav Res Ther. 1996 Aug;34(8):609-20. doi: 10.1016/0005-7967(96)00039-3. PMID: 8870287.

Link to PubMed: Fearful responding to repeated CO2 inhalation: a preliminary investigation

Abstract: In an effort to explore factors which maintain fear of physical sensations, repeated administration of 35% CO2 was used with college students scoring high and low on the Anxiety Sensitivity Index. Half of each group was administered 12 CO2 trials, while the other half received 9 CO2 trials, followed by a dishabituation trial (Trial 10) and 2 more CO2 administrations (Trials 11 and 12). Measures included subjective anxiety, heart rate, skin conductance, and number of panic symptoms reported. Results indicated a nonsignificant trend for the High ASI group to show increased pre-inhalation anxiety across trials, while the Low ASI group showed a rapid reduction in pre-inhalation anxiety. Post-inhalation skin conductance mirrored this pattern, although rapid reduction in post-inhalation heart rate was observed. Overall, the High ASI participants showed a notable lack of fear reduction across trials. Results are discussed in light of sensitization as a factor contributing to anticipatory anxiety, with implications for understanding the etiology and maintenance of Panic Disorder.

Title: Response patterns to repeated CO2 inhalation in individuals with high anxiety sensitivity

Authors: Beck JG, Shipherd JC, Read J.

Journal: Behav Res Ther. 1999 Nov;37(11):1073-89. doi: 10.1016/s0005-7967(98)00196-x. PMID: 10500321.

Link to PubMed: Response patterns to repeated CO2 inhalation in individuals with high anxiety sensitivity

Abstract: The present report extends previous work which has documented two distinct response patterns to repeated presentation of interoceptive cues (using CO2 inhalation) in PD patients [Beck, J. G. & Shipherd, J. C. (1997). Repeated exposure to interoceptive cues: does habituation of fear occur in panic disorder patients? Behaviour Research and Therapy, 35, 551-557]. We were interested in determining if these two patterns of fear habituation and sensitization would be noted in panic-naive individuals who reported high levels of Anxiety Sensitivity. A second aspect of this report examined whether attention to bodily sensations versus to neutral material would impact fear habituation and sensitization. Participants included 43 panic-naive individuals who scored at least 1 standard deviation above norms on the Anxiety Sensitivity Index. Results indicated that 37% of the sample reported habituation of fear, 47% reported fear sensitization and 16% demonstrated relatively stable fear levels across 12 inhalations of CO2 during session 1. The attentional manipulation did not exert a pronounced influence on anxiety, panic symptom severity, skin conductance, or heart rate in either Habituators or Sensitizers during session 2. These results are discussed in light of their relevance in understanding fundamental psychopathological processes underlying Panic Disorder.

Title: Respiratory, autonomic, and experiential responses to repeated inhalations of 20% CO₂ enriched air in panic disorder, social phobia, and healthy controls

Authors: Blechert J, Wilhelm FH, Meuret AE, Wilhelm EM, Roth WT.

Journal: Biol Psychol. 2010 Apr;84(1):104-11. doi: 10.1016/j.biopsycho.2010.01.002. Epub 2010 Jan 12. PMID: 20064582; PMCID: PMC2891596.

Link to full text: Respiratory, autonomic, and experiential responses to repeated inhalations of 20% CO₂ enriched air in panic disorder, social phobia, and healthy controls

Abstract: Inhalation of carbon dioxide (CO₂) enriched air triggers anxiety in panic disorder (PD) patients, which is often interpreted as a sign of biological vulnerability. However, most studies have not measured respiration in these tasks. We compared patients with PD (n=20) and social phobia (SP, n=19) to healthy controls (n=18) during eight inhalations of 20% CO₂, preceded and followed by two inhalations of room air, while continuously measuring subjective anxiety and dyspnea as well as autonomic and respiratory variables. PD patients showed increased reactivity and delayed recovery during CO₂ inhalations for most measures. Unlike both other groups, the PD group's tidal volume responses did not habituate across CO₂ inhalations. However, PD patients did not differ from SP patients on most other measures, supporting a continuum model of CO₂ sensitivity across anxiety disorders. Both patient groups showed continued reactivity during the last air inhalations, which is unlikely to be due to a biological sensitivity.

Keywords: anxiety disorder; biological challenge; carbon dioxide; habituation; psychophysiology; sensitization.

Title: Anxiogenic effects of repeated administrations of 20% CO2-enriched air: stability within sessions and habituation across time

Authors: Forsyth JP, Lejuez CW, Finlay C.

Journal: J Behav Ther Exp Psychiatry. 2000 Jun;31(2):103-21. doi: 10.1016/s0005-7916(00)00014-8. PMID: 11132115.

Link to PubMed: Anxiogenic effects of repeated administrations of 20% CO2-enriched air: stability within sessions and habituation across time

Abstract: Increasingly carbon dioxide-enriched air is being used as an aversive unconditioned stimulus in laboratory examinations of anxiety. Yet, little is known about the stability of the autonomic and subjective effects of this stimulus across repeated inhalations and sessions. We examined whether repeated administrations of high concentrations of CO2-enriched air produced either habituation, stability, or sensitization across several autonomic and self-report indices within one session (Experiment 1) and then several sessions (Experiment 2) of exposure. Results suggest that non-clinical participants do not habituate to CO2 within sessions, but do show habituation on cardiac and subjective report of anxiety across sessions. Individual difference factors such as anxiety sensitivity and suffocation fear seem to moderate some of these effects, including self-reported distress and anxiety in response to the challenge. These results support the use of CO2 as a panicogenic aversive stimulus in laboratory models of fear onset and in clinical settings for interoceptive exposure treatments of panic.

Title: The effect of successful treatment on the emotional and physiological response to carbon dioxide inhalation in patients with panic disorder

Authors: Gorman JM, Martinez J, Coplan JD, Kent J, Kleber M.

Journal: Biol Psychiatry. 2004 Dec 1;56(11):862-7. doi: 10.1016/j.biopsych.2004.08.016. PMID: 15576063.

Link to PubMed: The effect of successful treatment on the emotional and physiological response to carbon dioxide inhalation in patients with panic disorder

Abstract: Background: A number of studies have shown that patients with panic disorder are more likely to have panic attacks during carbon dioxide inhalation than are normal comparison subjects. Some studies have shown that antipanic medications can reduce the anxiogenic response to carbon dioxide, but none have shown if this is the case for cognitive behavioral therapy or if successful treatment reduces the respiratory physiologic response to carbon dioxide.

Methods: Twenty-five patients with panic disorder and 13 normal comparison subjects underwent baseline testing with 5% and 7% carbon dioxide inhalation. The patients were then retested after at least 12 weeks of treatment with either antipanic medication or cognitive behavioral therapy. Comparison subjects were retested after a similar interval.

Results: Successful treatment resulted in lower panic rates, and reduced anxiogenic response. Treatment had no effect, however, on the respiratory physiologic response.

Conclusions: There is dissociation in treatment response between the subjective and objective responses to carbon dioxide inhalation in panic disorder patients, with the former but not the latter showing positive change. We hypothesize that the strengthening of higher cortical control over subcortical fear-related structures, whether via medication or cognitive behavioral therapy treatment, results in less anxiety and fear in response to provoked symptoms reminiscent of naturally occurring panic.

Title: Effect of Antipanic Treatment on Response to Carbon Dioxide

Authors: Gorman JM, Browne ST, Papp LA, Martinez J, Welkowitz L, Coplan JD, Goetz RR, Kent J, Klein DF.

Journal: Biol Psychiatry. 1997 Dec 1;42(11):982-91. doi: 10.1016/s0006-3223(97)00160-1. PMID: 9386849.

Link to full text: Effect of Antipanic Treatment on Response to Carbon Dioxide

Abstract: Background: Disordered breathing among patients with panic disorder, including hyperventilation during attacks and increased anxiogenic response to carbon dioxide (CO2) inhalation, is well established. We wished to assess whether there is a change in the physiological response to CO2 after patients have undergone antipanic therapy with either tricyclic antidepressants or cognitive behavioral therapy (CBT).

Methods: Twenty-nine patients with panic disorder underwent baseline CO2 sensitivity testing using the traditional Read rebreathing method and then received either antidepressant treatment (n = 21) or CBT (n = 8). After completing treatment, CO2 testing was repeated. A comparison sample of 14 normal volunteers also had two CO2 sensitivity tests, separated by an average of 21.6 (SD = 8.8) weeks.

Results: Using a liberal standard, in which all CO2 sensitivity tests whose correlations between minute ventilation and end-tidal CO2 were at least .75 were used, patients, but not controls, demonstrated a significant reduction in CO2 sensitivity between the first and second test. Using a more conservative .90 correlation standard reduced the sample size available and resulted in trend reduction in patients but no significant change in controls. There was a suggestion that the change was most pronounced in treatment responders, although the number of patient nonresponders is extremely small in this sample.

Conclusions: These data indicate that treatment reduces CO2 sensitivity in patients with panic disorder. We speculate that manipulation of the serotonergic and noradrenergic neurotransmission systems, both known to play a role in the control of respiration, may have a specific effect in reducing respiratory hyperactivity in panic disorder.

Title: CO2 inhalation in the treatment of panic attacks

Authors: Griez E, van den Hout MA.

Journal: Behav Res Ther. 1986;24(2):145-50. doi: 10.1016/0005-7967(86)90085-9. PMID: 3083806.

Link to PubMed: CO2 inhalation in the treatment of panic attacks

Abstract: If panic disorder is conceptualized as a fear of interoceptive sensations, it can be treated by exposure to the autonomic symptoms that are induced by CO2 inhalation. In a cross-over study of 14 patients, propranolol was used as a control treatment. CO2 produced a significant reduction of fear of autonomic panic sensations, state and trait anxiety, agoraphobic anxiety and avoidance, and of the frequency of panicking. Propranolol resulted in improvements of state and of agoraphobic anxiety. Except for Trait Anxiety, CO2 was superior on all measures, most clearly on the Fear of Autonomic Sensations. At the 6-month follow-up, 3 patients required further treatment. Except for State Anxiety, the improvements were, on average, maintained. The clinical and theoretical implications are discussed.

Title: The relationship between the effect of lactate infusion on anxiety states, and their amelioration by carbon dioxide inhalation

Authors: Haslam MT.

Journal: Br J Psychiatry. 1974 Jul;125(0):88-90. doi: 10.1192/bjp.125.1.88. PMID: 4855036.

Link to PubMed: The relationship between the effect of lactate infusion on anxiety states, and their amelioration by carbon dioxide inhalation

Abstract: Anxiety is perhaps one of the commonest and most disabling symptoms in medicine, yet its satisfactory alleviation has been elusive. Work by Kluver and Bucy (1939) suggested that the amygdala generated anxiety, the function of the hippocampus being to decrease it. The hypothalamus appears to organize the level of arousal through the amygdala.

Title: Augmented respiratory relief. A new use of CO2 therapy in the treatment of phobic conditions: a preliminary report on two cases

Authors: Orwin A.

Journal: Br J Psychiatry. 1973 Feb;122(567):171-3. doi: 10.1192/bjp.122.2.171. PMID: 4714831.

Link to PubMed: Augmented respiratory relief. A new use of CO2 therapy in the treatment of phobic conditions: a preliminary report on two cases

Abstract: Respiratory relief (R.R.) appeared to be a simple and rapid treatment for specific phobias and for the phobic components of more complex neurotic conditions (Orwin, 1971). The basic hypothesis was that the R.R., satisfying an intense need to breathe, if simultaneous with the presentation of a phobic stimulus, would inhibit the degree of anxiety previously provoked and that this inhibition would persist. The need to breathe was induced by voluntary restriction up to maximum voluntary respiratory arrest (M.V.R.A.). As in conventional desensitization (Wolpe, 1958), a hierarchy was drawn up and the patients were desensitized using R.R. instead of relaxation. Carbon dioxide-oxygen mixtures were introduced because CO2 is known to stimulate respiration, and it was expected that by intensifying the need to breathe the speed and effectiveness of treatment would be increased by the augmented respiratory relief (A.R.R.).

Title: Antipanic drug modulation of 35% CO2 hyperreactivity and short-term treatment outcome

Authors: Perna G, Bertani A, Caldirola D, Gabriele A, Cocchi S, Bellodi L.

Journal: J Clin Psychopharmacol. 2002 Jun;22(3):300-8. doi: 10.1097/00004714-200206000-00011. PMID: 12006901.

Link to PubMed: Antipanic drug modulation of 35% CO2 hyperreactivity and short-term treatment outcome

Abstract: Carbon dioxide (CO2) inhalation induces acute anxiety and panic attacks in patients with Panic Disorder (PD). Anti-panic drugs decrease CO2 reactivity after the first days of treatment; however, the clinical meaning of this finding has not yet been established. This study investigated the effects of treatment with tricyclic antidepressants and selective serotonin re-uptake inhibitors (SSRIs) on CO2 reactivity and compared the relationships between 35% CO2 hyperreactivity modulation and short-term clinical outcome. One hundred twenty-three patients with PD with or without agoraphobia who were hyperreactive to CO2 were randomly assigned to treatment groups with imipramine, clomipramine, paroxetine, sertraline, or fluvoxamine. A double-blind, randomized design was applied. Each patient received the 35% CO2 challenge on days 0, 7, and 30. The severity of clinical symptomatology was measured on days 0 and 30. Decreased hyperreactivity to 35% CO2 in all five treatment groups was already evident after the first week. The decrease in CO2 reactivity at the end of treatment was proportional to the degree of clinical improvement. Multiple regression analyses showed that the decrease in CO2 reactivity after the first week was a significant predictor for good clinical outcome after one month. The results of this study confirm evidence that psychoactive drugs effective in the treatment of PD decrease CO2 hyperreactivity. They also suggest that precocious modulation of CO2 reactivity might fairly reliably predict short-term clinical outcome in patients with "respiratory" PD.

Title: Extinction of panicogenic effects of a 35% CO2 challenge in patients with panic disorder

Authors: Schmidt NB, Trakowski JH, Staab JP.

Journal: J Abnorm Psychol. 1997 Nov;106(4):630-8. doi: 10.1037//0021-843x.106.4.630. PMID: 9358693.

Link to PubMed: Extinction of panicogenic effects of a 35% CO2 challenge in patients with panic disorder

Abstract: Inhalations of high concentrations of carbon dioxide (CO2) reliably produce panic attacks in patients with panic disorder. The present study evaluated whether cognitive-behavioral treatment (CBT) for panic disorder would extinguish CO2-induced panic and whether changes in panic and arousal-related cognitions were associated with the induction of panic. Patients with panic disorder (N = 54) were assigned to 1 of 3 experimental conditions: CBT with respiratory training (CBT-R), CBT without respiratory training (CBT), or delayed treatment. Participants received 5 repeated vital-capacity inhalations of 35% CO2/65% O2 prior to and following either 12 treatment sessions or a 12-week waiting period. During pretreatment assessments, 74% of patients experienced a panic attack during at least 1 inhalation. At posttreatment, only 20% of treated participants (CBT-R = 19%, CBT = 22%), compared with 64% of untreated participants, panicked. Forty-four percent of treated participants, compared with 0% of untreated participants, reported no anxiety during all posttreatment inhalations. Anxiety sensitivity as well as panic appraisals regarding the likelihood of panic and self-efficacy with coping with panic were significantly related to fearful responding to the CO2 challenge.

Title: The effects of inhaling a 35 per cent CO2-65 per cent O2 mixture upon anxiety level in neurotic patients

Authors: Slater SL, Leavy A.

Journal: Behav Res Ther. 1966 Nov;4(4):309-16. doi: 10.1016/0005-7967(66)90027-1. PMID: 5978686.

Link to PubMed: The effects of inhaling a 35 per cent CO2-65 per cent O2 mixture upon anxiety level in neurotic patients

Abstract: Wolpe has reported the use of a 35 per cent CO2-65 per cent O2 mixture to relax patients who cannot otherwise be relaxed sufficiently to allow desensitization. In this investigation, the anxiety-reducing effects of single inhalations of a 35 per cent CO2-65 per cent O2 were studied on twelve patients, all of whom had been diagnosed by their doctors as having a high level of anxiety. On the basis of the subjects' ratings, this mixture was more effective than was an equal volume of air which was breathed through the same apparatus as was the experimental mixture. Evidence is offered which suggests that the decrement in anxiety which followed administration of the 35 per cent CO2-65 per cent O2 mixture was due to some effect of the gas other than the hyperventilation which it induces.

Title: Mechanism of change in cognitive-behavioral treatment of panic disorder: evidence for the fear of fear mediational hypothesis

Authors: Smits JA, Powers MB, Cho Y, Telch MJ.

Journal: J Consult Clin Psychol. 2004 Aug;72(4):646-52. doi: 10.1037/0022-006X.72.4.646. PMID: 15301649.

Link to PubMed: Mechanism of change in cognitive-behavioral treatment of panic disorder: evidence for the fear of fear mediational hypothesis

Abstract: Numerous clinical trials have demonstrated the efficacy of cognitive-behavioral treatment (CBT) for panic disorder. However, studies investigating the mechanisms responsible for improvement with CBT are lacking. The authors used regression analyses outlined by R. M. Baron and D. A. Kenny (1986) to test whether a reduction in fear of fear (FOF) underlies improvement resulting from CBT. Pre- and posttreatment measures were collected from 90 CBT-treated patients and 40 wait-list control participants. Overall, treatment accounted for 31% of the variance in symptom reduction. The potency of FOF as a mediator varied as a function of symptom facet, as full mediation was observed for the change in global disability, whereas the effects of CBT on agoraphobia, anxiety, and panic frequency were partially accounted for by reductions in FOF. Clinical implications and future research directions are discussed.

Title: Reduction of CO2-induced anxiety in patients with panic attacks after repeated CO2 exposure

Authors: van den Hout MA, van der Molen GM, Griez E, Lousberg H, Nansen A.

Journal: Am J Psychiatry. 1987 Jun;144(6):788-91. doi: 10.1176/ajp.144.6.788. PMID: 3109265.

Link to PubMed: Reduction of CO2-induced anxiety in patients with panic attacks after repeated CO2 exposure

Abstract: The authors compared the subjective reaction of 13 panic patients and eight control subjects to a 35% CO2 challenge, a treatment known to produce physical symptoms comparable to those of natural or lactate-induced panic, and to placebo treatment (inhalation of air). They found that patients had higher placebo scores than control subjects, patients tended to get highly anxious on CO2 and control subjects did not, and CO2-induced subjective anxiety in patients decreased as the number of CO2-induced exposures to interoceptive anxiety symptoms increased. The data support a behavioral account of the effects of anxiogenics.

Title: Carbon dioxide inhalation treatments of neurotic anxiety. An overview

Authors: Wolpe J.

Journal: J Nerv Ment Dis. 1987 Mar;175(3):129-33. doi: 10.1097/00005053-198703000-00001. PMID: 3102688.

Link to PubMed: Carbon dioxide inhalation treatments of neurotic anxiety. An overview

Abstract: A lucky chance more than 30 years ago revealed the remarkable efficacy of single inhalations of high concentrations of carbon dioxide in eliminating or markedly reducing free-floating anxiety. The reduction of anxiety lasts for days, weeks, or longer--well beyond the persistence of carbon dioxide in the body. The effects are explicable on the hypothesis that free-floating anxiety is anxiety conditioned to continuously present sources of stimulation, such as background noise or the awareness of space or time, and that the anxiety response habit is weakened when the anxiety is inhibited by the competition of responses that carbon dioxide induces. More recently, it has become apparent that inhalations of carbon dioxide, applied in a different manner, are effective in overcoming maladaptive anxiety responses to specific stimuli, e.g., social stimuli. The substance is also proving to be a valuable resource in the treatment of the common variety of panic attacks.

Title: Comparison of the panicogenic effect of cholecystokinin 30-33 and carbon dioxide in panic disorder

Authors: Bradwejn J, Koszycki D.

Journal: Prog Neuropsychopharmacol Biol Psychiatry. 1991;15(2):237-9. doi: 10.1016/0278-5846(91)90086-g. PMID: 1908105.

Link to PubMed: Comparison of the panicogenic effect of cholecystokinin 30-33 and carbon dioxide in panic disorder

Abstract: 1. Twenty-two patients who met DSM-III-R criteria for panic disorder received either cholecystokinin 30-33 (25 micrograms i.v.) or 33% carbon dioxide. 2. The principal outcome measures of the study included the number and sum intensity of panic symptoms and the incidence of panic attacks. 3. The incidence of panic attacks tended (P = .07) to be higher with cholecystokinin 30-33 than with carbon dioxide. Nevertheless, patients who panicked within each group did not differ significantly with regard to the number and sum intensity of symptoms or symptom profile. 4. That cholecystokinin 30-33 and 35% carbon dioxide induced panic attacks which were qualitatively and quantitatively similar suggest that these agents might act on distinct systems that have a final common target or a final common mechanism of action.

Title: Restoring Acid-sensing ion channel-1a in the amygdala of knock-out mice rescues fear memory but not unconditioned fear responses

Authors: Coryell MW, Wunsch AM, Haenfler JM, Allen JE, McBride JL, Davidson BL, Wemmie JA.

Journal: J Neurosci. 2008 Dec 17;28(51):13738-41. doi: 10.1523/JNEUROSCI.3907-08.2008. PMID: 19091964; PMCID: PMC2651157.

Link to full text: Restoring Acid-sensing ion channel-1a in the amygdala of knock-out mice rescues fear memory but not unconditioned fear responses

Abstract: Acid-sensing ion channel-1a (ASIC1a) contributes to multiple fear behaviors, however the site of ASIC1a action in behavior is not known. To explore a specific location of ASIC1a action, we expressed ASIC1a in the basolateral amygdala of ASIC1a-/- mice using viral vector-mediated gene transfer. This rescued context-dependent fear memory, but not the freezing deficit during training or the unconditioned fear response to predator odor. These data pinpoint the basolateral amygdala as the site where ASIC1a contributes to fear memory. They also discriminate fear memory from fear expressed during training and from unconditioned fear. Furthermore, this work illustrates a strategy for identifying discrete brain regions where specific genes contribute to complex behaviors.

Title: Baseline respiratory parameters in panic disorder: a meta-analysis

Authors: Grassi M, Caldirola D, Vanni G, Guerriero G, Piccinni M, Valchera A, Perna G.

Journal: J Affect Disord. 2013 Apr 5;146(2):158-73. doi: 10.1016/j.jad.2012.08.034. Epub 2012 Oct 27. PMID: 23107756.

Link to PubMed: Baseline respiratory parameters in panic disorder: a meta-analysis

Abstract: Background: The presence of abnormalities in baseline respiratory function of subjects with panic disorder (PD) is expected according to PD respiratory theories. We aimed to meta-analyze results from studies comparing baseline respiratory and hematic parameters related to respiration between subjects with PD and controls.

Methods: A literature research in bibliographic databases was performed. Fixed-effects models were applied for all parameters while random-effects models only when suitable (at least 10 independent studies). Several moderator analyses and publication bias diagnostics were performed.

Results: We found significantly higher mean minute ventilation and lower et-pCO(2) in subjects with PD than controls. Moreover we also found evidences of reduced HCO(3)(-) and PO(4)(-) hematic concentrations, higher indexes of respiratory variability/irregularity and higher rate of sighs and apneas. Evidence of heterogeneity was partly explained by moderator analyses. No relevant publication bias was found.

Limitations: Several shortcomings affected the included studies, such as over-inclusive recruitment criteria, samples unbalanced for socio-demographic characteristics, lack of statistical details and small number of studies available for several parameters.

Discussion: Our results support the idea of abnormalities in respiratory function of subjects with PD. Compared to controls, they showed baseline hyperventilation; the results from hematic parameters suggest that hyperventilation may be chronic and not simply caused by their high anxiety levels during respiratory assessment. Evidences of higher variability and irregularity in respiratory patterns of subjects with PD were also found. It is unclear to what extent the higher rate of sighs and apneas may explain the other baseline respiratory abnormalities found in PD.

Title: Induction of c-Fos in 'panic/defence'-related brain circuits following brief hypercarbic gas exposure

Authors: Johnson PL, Fitz SD, Hollis JH, Moratalla R, Lightman SL, Shekhar A, Lowry CA.

Journal: J Psychopharmacol. 2011 Jan;25(1):26-36. doi: 10.1177/0269881109353464. Epub 2010 Jan 15. PMID: 20080924.

Link to PubMed: Induction of c-Fos in 'panic/defence'-related brain circuits following brief hypercarbic gas exposure

Abstract: Inspiration of air containing high concentrations of carbon dioxide (CO(2); hypercarbic gas exposure) mobilizes respiratory, sympathetic and hypothalamic-pituitary-adrenal axis responses and increases anxiety-like behaviour in rats and humans. Meanwhile the same stimulus induces panic attacks in the majority of panic disorder patients. However, little is known about the neural circuits that regulate these acute effects. In order to determine the effects of acute hypercarbic gas exposure on forebrain and brainstem circuits, conscious adult male rats were placed in flow cages and exposed to either atmospheric air or increasing environmental CO(2) concentrations (from baseline concentrations up to 20% CO(2)) during a 5 min period. The presence of immunoreactivity for the protein product of the immediate-early gene c-fos was used as a measure of functional cellular responses. Exposing rats to hypercarbic gas increased anxiety-related behaviour and increased numbers of c-Fos-immunoreactive cells in subcortical regions of the brain involved in: (1) the initiation of fear- or anxiety-associated behavioural responses (i.e. the dorsomedial hypothalamus, perifornical nucleus and dorsolateral and ventrolateral periaqueductal gray); (2) mobilization of the hypothalamic-pituitary-adrenal axis (i.e. the dorsomedial hypothalamus, perifornical nucleus and paraventricular hypothalamic nucleus); and (3) initiation of stress-related sympathetic responses (i.e. the dorsomedial hypothalamus, dorsolateral periaqueductal grey and rostroventrolateral medulla). These findings have implications for understanding how the brain senses changes in environmental CO(2) concentrations and the neural mechanisms underlying the subsequent adaptive changes in stress-related physiology and behaviour.

Title: Neural pathways underlying lactate-induced panic

Authors: Johnson PL, Truitt WA, Fitz SD, Lowry CA, Shekhar A.

Journal: Neuropsychopharmacology. 2008 Aug;33(9):2093-107. doi: 10.1038/sj.npp.1301621. Epub 2007 Dec 5. PMID: 18059441; PMCID: PMC3065200.

Link to full text: Neural pathways underlying lactate-induced panic

Abstract: Panic disorder is a severe anxiety disorder characterized by susceptibility to induction of panic attacks by subthreshold interoceptive stimuli such as 0.5 M sodium lactate infusions. Although studied for four decades, the mechanism of lactate sensitivity in panic disorder has not been understood. The dorsomedial hypothalamus/perifornical region (DMH/PeF) coordinates rapid mobilization of behavioral, autonomic, respiratory and endocrine responses to stress, and rats with disrupted GABA inhibition in the DMH/PeF exhibit panic-like responses to lactate, similar to panic disorder patients. Utilizing a variety of anatomical and pharmacological methods, we provide evidence that lactate, via osmosensitive periventricular pathways, activates neurons in the compromised DMH/PeF, which relays this signal to forebrain limbic structures such as the bed nucleus of the stria terminalis to mediate anxiety responses, and specific brainstem sympathetic and parasympathetic pathways to mediate the respiratory and cardiovascular components of the panic-like response. Acutely restoring local GABAergic tone in the DMH/PeF blocked lactate-induced panic-like responses. Autonomic panic-like responses appear to be a result of DMH/PeF-mediated mobilization of sympathetic responses (verified with atenolol) and resetting of the parasympathetically mediated baroreflex. Based on our findings, DMH/PeF efferent targets such as the C1 adrenergic neurons, paraventricular hypothalamus, and the central amygdala are implicated in sympathetic mobilization; the nucleus of the solitary tract is implicated in baroreflex resetting; and the parabrachial nucleus is implicated in respiratory responses. These results elucidate neural circuits underlying lactate-induced panic-like responses and the involvement of both sympathetic and parasympathetic systems.

Title: A key role for orexin in panic anxiety

Authors: Johnson PL, Truitt W, Fitz SD, Minick PE, Dietrich A, Sanghani S, Träskman-Bendz L, Goddard AW, Brundin L, Shekhar A.

Journal: Nat Med. 2010 Jan;16(1):111-5. doi: 10.1038/nm.2075. Epub 2009 Dec 27. PMID: 20037593; PMCID: PMC2832844.

Link to full text: A key role for orexin in panic anxiety

Abstract: Panic disorder is a severe anxiety disorder with recurrent, debilitating panic attacks. In individuals with panic disorder there is evidence of decreased central gamma-aminobutyric acid (GABA) activity as well as marked increases in autonomic and respiratory responses after intravenous infusions of hypertonic sodium lactate. In a rat model of panic disorder, chronic inhibition of GABA synthesis in the dorsomedial-perifornical hypothalamus of rats produces anxiety-like states and a similar vulnerability to sodium lactate-induced cardioexcitatory responses. The dorsomedial-perifornical hypothalamus is enriched in neurons containing orexin (ORX, also known as hypocretin), which have a crucial role in arousal, vigilance and central autonomic mobilization, all of which are key components of panic. Here we show that activation of ORX-synthesizing neurons is necessary for developing a panic-prone state in the rat panic model, and either silencing of the hypothalamic gene encoding ORX (Hcrt) with RNAi or systemic ORX-1 receptor antagonists blocks the panic responses. Moreover, we show that human subjects with panic anxiety have elevated levels of ORX in the cerebrospinal fluid compared to subjects without panic anxiety. Taken together, our results suggest that the ORX system may be involved in the pathophysiology of panic anxiety and that ORX antagonists constitute a potential new treatment strategy for panic disorder.

Title: Respiratory Feedback for Treating Panic Disorder

Authors: Meuret AE, Wilhelm FH, Roth WT.

Journal: J Clin Psychol. 2004 Feb;60(2):197-207. doi: 10.1002/jclp.10245. PMID: 14724927.

Link to PubMed: Respiratory Feedback for Treating Panic Disorder

Abstract: Panic disorder patients often complain of shortness of breath or other respiratory complaints, which has been used as evidence for both hyperventilation and false suffocation alarm theories of panic. Training patients to change their breathing patterns is a common intervention, but breathing rarely has been measured objectively in assessing the patient or monitoring therapy results. We report a new breathing training method that makes use of respiratory biofeedback to teach individuals to modify four respiratory characteristics: increased ventilation (Respiratory Rate x Tidal Volume), breath-to-breath irregularity in rate and depth, and chest breathing. As illustrated by a composite case, feedback of respiratory rate and end-tidal pCO2 can facilitate voluntary control of respiration and reduce symptoms. Respiratory monitoring may provide relevant diagnostic, prognostic, and outcome information.

Title: Changes in respiration mediate changes in fear of bodily sensations in panic disorder

Authors: Meuret AE, Rosenfield D, Hofmann SG, Suvak MK, Roth WT.

Journal: J Psychiatr Res. 2009 Mar;43(6):634-41. doi: 10.1016/j.jpsychires.2008.08.003. Epub 2008 Oct 5. PMID: 18835608; PMCID: PMC3327292.

Link to full text: Changes in respiration mediate changes in fear of bodily sensations in panic disorder

Abstract: The purpose of the study was to examine whether changes in pCO(2) mediate changes in fear of bodily sensation (as indexed by anxiety sensitivity) in a bio-behavioral treatment for panic disorder that targets changes in end-tidal pCO(2). Thirty-five panic patients underwent 4 weeks of capnometry-assisted breathing training targeting respiratory dysregulation. Longitudinal mediation analyses of the changes in fear of bodily symptoms over time demonstrated that pCO(2), but not respiration rate, was a partial mediator of the changes in anxiety sensitivity. Results were supported by cross lag panel analyses, which indicated that earlier pCO(2) levels predicted later levels of anxiety sensitivity, but not vice versa. PCO(2) changes also led to changes in respiration rate, questioning the importance of respiration rate in breathing training. The results provide little support for changes in fear of bodily sensations leading to changes in respiration, but rather suggest that breathing training targeting pCO(2) reduced fear of bodily sensations in panic disorder.

Title: Feedback of end-tidal pCO2 as a therapeutic approach for panic disorder

Authors: Meuret AE, Wilhelm FH, Ritz T, Roth WT.

Journal: J Psychiatr Res. 2008 Jun;42(7):560-8. doi: 10.1016/j.jpsychires.2007.06.005. Epub 2007 Aug 3. PMID: 17681544; PMCID: PMC2890048.

Link to full text: Feedback of end-tidal pCO2 as a therapeutic approach for panic disorder

Abstract: Background: Given growing evidence that respiratory dysregulation is a central feature of panic disorder (PD) interventions for panic that specifically target respiratory functions could prove clinically useful and scientifically informative. We tested the effectiveness of a new, brief, capnometry-assisted breathing therapy (BRT) on clinical and respiratory measures in PD.

Methods: Thirty-seven participants with PD with or without agoraphobia were randomly assigned to BRT or to a delayed-treatment control group. Clinical status, respiration rate, and end-tidal pCO(2) were assessed at baseline, post-treatment, 2-month and 12-month follow-up. Respiratory measures were also assessed during homework exercises using a portable capnometer as a feedback device.

Results: Significant improvements (in PD severity, agoraphobic avoidance, anxiety sensitivity, disability, and respiratory measures) were seen in treated, but not untreated patients, with moderate to large effect sizes. Improvements were maintained at follow-up. Treatment compliance was high for session attendance and homework exercises; dropouts were few.

Conclusions: The data provide preliminary evidence that raising end-tidal pCO(2) by means of capnometry feedback is therapeutically beneficial for panic patients. Replication and extension will be needed to verify this new treatment's efficacy and determine its mechanisms.

Title: Does fear reactivity during exposure predict panic symptom reduction?

Authors: Meuret AE, Seidel A, Rosenfield B, Hofmann SG, Rosenfield D.

Journal: J Consult Clin Psychol. 2012 Oct;80(5):773-85. doi: 10.1037/a0028032. Epub 2012 Apr 9. PMID: 22486408; PMCID: PMC3404244.

Link to full text: Does fear reactivity during exposure predict panic symptom reduction?

Abstract: Objective: Fear reactivity during exposure is a commonly used indicator of learning and overall therapy outcome. The objective of this study was to assess the predictive value of fear reactivity during exposure using multimodal indicators and an advanced analytical design. We also investigated the degree to which treatment condition (cognitive training vs. respiratory skill training) moderated fear reactivity and therapeutic outcome.

Method: Thirty-four patients with panic disorder and agoraphobia completed a total of 123 in-vivo exposure sessions, comprising 3 weekly sessions and a 4th session 2 months following therapy completion. Sessions varied in length and phobic stimuli. Cardiorespiratory physiology (heart rate, carbon dioxide partial pressure [PCO2], respiration rate) and experiential symptoms (panic symptoms and anxiety) were assessed repeatedly throughout exposure sessions, in addition to weekly assessments of panic cognitions, avoidance, and functioning.

Results: Panic symptomatology decreased substantially in both treatment conditions during therapy and follow-up. Significant cardiorespiratory and experiential reactivity was observed during all exposures, characterized by activation followed by reduction. Greater within-session activation of anxiety and panic symptoms was inversely related to improvement in panic symptoms severity, but neither physiological activation nor within- or between-session reduction of either physiological or experiential variables was predictive of outcome. No moderating effects of treatment condition were found.

Conclusions: Fear activation and reduction during exposure are weak predictors of corrective learning and fear extinction. Clinical implications for exposure therapy and directions for future research are discussed.

Title: Do unexpected panic attacks occur spontaneously?

Authors: Meuret AE, Rosenfield D, Wilhelm FH, Zhou E, Conrad A, Ritz T, Roth WT.

Journal: Biol Psychiatry. 2011 Nov 15;70(10):985-91. doi: 10.1016/j.biopsych.2011.05.027. Epub 2011 Jul 23. PMID: 21783179; PMCID: PMC3327298.

Link to full text: Do unexpected panic attacks occur spontaneously?

Abstract: Background: Spontaneous or unexpected panic attacks, per definition, occur "out of the blue," in the absence of cues or triggers. Accordingly, physiological arousal or instability should occur at the onset of, or during, the attack, but not preceding it. To test this hypothesis, we examined if points of significant autonomic changes preceded the onset of spontaneous panic attacks.

Methods: Forty-three panic disorder patients underwent repeated 24-hour ambulatory monitoring. Thirteen natural panic attacks were recorded during 1960 hours of monitoring. Minute-by-minute epochs beginning 60 minutes before and continuing to 10 minutes after the onset of individual attacks were examined for respiration, heart rate, and skin conductance level. Measures were controlled for physical activity and vocalization and compared with time matched control periods within the same person.

Results: Significant patterns of instability across a number of autonomic and respiratory variables were detected as early as 47 minutes before panic onset. The final minutes before onset were dominated by respiratory changes, with significant decreases in tidal volume followed by abrupt carbon dioxide partial pressure increases. Panic attack onset was characterized by heart rate and tidal volume increases and a drop in carbon dioxide partial pressure. Symptom report was consistent with these changes. Skin conductance levels were generally elevated in the hour before, and during, the attacks. Changes in the matched control periods were largely absent.

Conclusions: Significant autonomic irregularities preceded the onset of attacks that were reported as abrupt and unexpected. The findings invite reconsideration of the current diagnostic distinction between uncued and cued panic attacks.

Title: Brain glucose and lactate levels during ventilator-induced hypo- and hypercapnia

Authors: van Hulst RA, Lameris TW, Haitsma JJ, Klein J, Lachmann B.

Journal: Clin Physiol Funct Imaging. 2004 Jul;24(4):243-8. doi: 10.1111/j.1475-097X.2004.00557.x. PMID: 15233840.

Link to PubMed: Brain glucose and lactate levels during ventilator-induced hypo- and hypercapnia

Abstract: Objective: Levels of glucose and lactate were measured in the brain by means of microdialysis in order to evaluate the effects of ventilator-induced hypocapnia and hypercapnia on brain metabolism in healthy non-brain-traumatized animals.

Design and setting: Prospective animal study in a university laboratory.

Subjects: Eight adult Landrace/Yorkshire pigs.

Interventions: The microdialysis probe was inserted in the brain along with a multiparameter sensor and intracranial pressure (ICP) probe. The animals were ventilated in a pressure-controlled mode according to the open lung concept with an inspired oxygen fraction of 0.4/1.0. Starting at normoventilation (PaCO(2) +/-40 mmHg) two steps of both hypercapnia (PCO(2) +/- 70 and 100 mmHg) and hypocapnia (PaCO(2) +/- 20 and 30 mmHg) were performed. Under these conditions, brain glucose and lactate levels as well as brain oxygen (PbrO(2)), brain carbon dioxide (PbrCO(2)), brain pH (brpH), brain temperature and ICP were measured.

Results: At hypercapnia (PaCO(2) = 102.7 mmHg) there were no significant changes in brain glucose and lactate but there was a significant increase in PbrCO(2), PbrO(2) and ICP. In contrast, at hypocapnia (PCO(2) = 19.8 mmHg) there was a significant increase in brain lactate and a significant decrease in both brain glucose and PbrCO(2).

Conclusions: Hypocapnia decreases brain glucose and increases brain lactate concentration, indicating anaerobic metabolism, whereas hypercapnia has no influence on levels of brain glucose and brain lactate.

Title: Voluntary hyperventilation in the treatment of panic disorder--functions of hyperventilation, their implications for breathing training, and recommendations for standardization

Authors: Meuret AE, Ritz T, Wilhelm FH, Roth WT.

Journal: Clin Psychol Rev. 2005 May;25(3):285-306. doi: 10.1016/j.cpr.2005.01.002. PMID: 15792851.

Link to PubMed: Voluntary hyperventilation in the treatment of panic disorder--functions of hyperventilation, their implications for breathing training, and recommendations for standardization

Abstract: Hyperventilation has numerous theoretical and empirical links to anxiety and panic. Voluntary hyperventilation (VH) tests have been applied experimentally to understand psychological and physiological mechanisms that produce and maintain anxiety, and therapeutically in the treatment of anxiety disorders. From the theoretical perspective of hyperventilation theories of anxiety, VH is useful diagnostically to the clinician and educationally to the patient. From the theoretical perspective of cognitive-behavior therapy, VH is a way to expose patients with panic disorder to sensations associated with panic and to activate catastrophic cognitions that need restructuring. Here we review panic disorder treatment studies using breathing training that have included VH. We differentiate the roles of VH in diagnosis, education about symptoms, training of breathing strategies, interoceptive exposure, and outcome measurement--discussing methodological issues specific to these roles and VH test reliability and validity. We propose how VH procedures might be standardized in future studies.

Title: Carbon dioxide hypersensitivity, hyperventilation, and panic disorder

Authors: Papp LA, Klein DF, Gorman JM.

Journal: Am J Psychiatry. 1993 Aug;150(8):1149-57. doi: 10.1176/ajp.150.8.1149. PMID: 8392296.

Link to PubMed: Carbon dioxide hypersensitivity, hyperventilation, and panic disorder

Abstract: Objective: The purpose of this article is to offer a comprehensive, data-based explanation of the relationship between hyperventilation and panic disorder linking CO2 hypersensitivity, cognitive/behavioral factors, and the respiratory effects of antipanic pharmacologic and psychological treatments.

Method: The authors conducted a computerized search of MEDLINE for relevant articles.

Results: Some panic patients have a chronic, subtle respiratory disturbance. Acute hyperventilation is neither necessary nor sufficient for panic to occur. Respiratory abnormalities in panic patients may adaptively aim at coping with a hypersensitive CO2 chemoreceptor system. Pharmacologic panicogens also stimulate the respiratory system, causing hyperventilation. Triggering this hypersensitive respiratory control mechanism may incite panic. Antipanic medications may reset the receptor threshold. Misattribution and catastrophic interpretation of somatic symptoms or the sense of loss of control may contribute to panic symptoms. Behavioral interventions such as desensitization or breathing retraining may block the full-blown attack. Cognitive strategies through cognitive control of respiration may supplement and accentuate these interventions.

Conclusions: Panic disorder may be due to an inherently unstable autonomic nervous system, coupled with cognitive distress.

Title: Respiratory muscle tension as symptom generator in individuals with high anxiety sensitivity

Authors: Ritz T, Meuret AE, Bhaskara L, Petersen S.

Journal: Psychosom Med. 2013 Feb;75(2):187-95. doi: 10.1097/PSY.0b013e31827d1072. Epub 2013 Jan 16. PMID: 23324873.

Link to PubMed: Respiratory muscle tension as symptom generator in individuals with high anxiety sensitivity

Abstract: Objective: Anxiety and panic are associated with the experience of a range of bodily symptoms, in particular unpleasant breathing sensations (dyspnea). Respiratory theories of panic disorder have focused on disturbances in blood gas regulation, but respiratory muscle tension as a source of dyspnea has not been considered. We therefore examined the potential of intercostal muscle tension to elicit dyspnea in individuals with high anxiety sensitivity, a risk factor for developing panic disorder.

Methods: Individuals high and low in anxiety sensitivity (total N=62) completed four tasks: electromyogram biofeedback for tensing intercostal muscle, electromyogram biofeedback for tensing leg muscles, paced breathing at three different speeds, and a fine motor task. Global dyspnea, individual respiratory sensations, nonrespiratory sensations, and discomfort were assessed after each task, whereas respiratory pattern (respiratory inductance plethysmography) and end-tidal carbon dioxide (capnography) were measured continuously.

Results: In individuals with high compared to low anxiety sensitivity, intercostal muscle tension elicited a particularly strong report of obstruction (M=5.1, SD=3.6 versus M=2.5, SD=3.0), air hunger (M=1.9, SD=2.1 versus M=0.4, SD=0.8), hyperventilation symptoms (M=0.6, SD=0.6 versus M=0.1, SD=0.1), and discomfort (M=5.1, SD=3.2 versus M=2.2, SD=2.1) (all p values<.05). This effect was not explained by site-unspecific muscle tension, voluntary manipulation of respiration, or sustained task-related attention. Nonrespiratory control sensations were not significantly affected by tasks (F<1), and respiratory variables did not reflect any specific responding of high-Anxiety Sensitivity Index participants to intercostal muscle tension.

Conclusions: Respiratory muscle tension may contribute to the respiratory sensations experienced by panic-prone individuals. Theories and treatments for panic disorder should consider this potential source of symptoms.

Title: Autonomic function in panic disorder: cardiorespiratory and plasma catecholamine responsivity to multiple challenges of the autonomic nervous system

Authors: Stein MB, Asmundson GJ.

Journal: Biol Psychiatry. 1994 Oct 15;36(8):548-58. doi: 10.1016/0006-3223(94)90619-x. PMID: 7827218.

Link to PubMed: Autonomic function in panic disorder: cardiorespiratory and plasma catecholamine responsivity to multiple challenges of the autonomic nervous system

Abstract: Panic disorder has been widely hypothesized to be associated with dysfunction of the autonomic nervous system. In this study, 24 patients with panic disorder and 26 healthy control subjects took part in a broad battery of autonomic function tests, each designed to stress the autonomic nervous system in a particular fashion. Testing consisted of postural challenge, isometric exercise, cold pressor, and Valsalva maneuver. Dependent measures included heart rate, vagal tone, blood pressure, respiratory frequency, end-tidal CO2 levels, and plasma norepinephrine and epinephrine levels. The testing procedures reliably produced changes in autonomic output in the expected directions, but patients with panic disorder were not found to differ from healthy controls in their cardiorespiratory or plasma catecholaminergic responses. This pattern of normal autonomic responsivity in the patients with panic disorder was evident across multiple test conditions with varying autonomic demand characteristics, thereby supporting the integrity of autonomic regulatory systems in this illness. These data run counter to a simple notion of autonomic dysfunction in panic disorder.

Title: The bed nucleus of the stria terminalis is critical for anxiety-related behavior evoked by CO2 and acidosis

Authors: Taugher RJ, Lu Y, Wang Y, Kreple CJ, Ghobbeh A, Fan R, Sowers LP, Wemmie JA.

Journal: J Neurosci. 2014 Jul 30;34(31):10247-55. doi: 10.1523/JNEUROSCI.1680-14.2014. PMID: 25080586; PMCID: PMC4115136.

Link to full text: The bed nucleus of the stria terminalis is critical for anxiety-related behavior evoked by CO2 and acidosis

Abstract: Carbon dioxide (CO2) inhalation lowers brain pH and induces anxiety, fear, and panic responses in humans. In mice, CO2 produces freezing and avoidance behavior that has been suggested to depend on the amygdala. However, a recent study in humans with bilateral amygdala lesions revealed that CO2 can trigger fear and panic even in the absence of amygdalae, suggesting the importance of extra-amygdalar brain structures. Because the bed nucleus of the stria terminalis (BNST) contributes to fear- and anxiety-related behaviors and expresses acid-sensing ion channel-1A (ASIC1A), we hypothesized that the BNST plays an important role in CO2-evoked fear-related behaviors in mice. We found that BNST lesions decreased both CO2-evoked freezing and CO2-conditioned place avoidance. In addition, we found that CO2 inhalation caused BNST acidosis and that acidosis was sufficient to depolarize BNST neurons and induce freezing behavior; both responses depended on ASIC1A. Finally, disrupting Asic1a specifically in the BNST reduced CO2-evoked freezing, whereas virus-vector-mediated expression of ASIC1A in the BNST of Asic1a(-/-) and Asic1a(+/+) mice increased CO2-evoked freezing. Together, these findings identify the BNST as an extra-amygdalar fear circuit structure important in CO2-evoked fear-related behavior.

Keywords: ASIC1A; CO2; anxiety; bed nucleus of the stria terminalis; pH; panic.

Title: Acid-base dysregulation and chemosensory mechanisms in panic disorder: a translational update

Authors: Vollmer LL, Strawn JR, Sah R.

Journal: Transl Psychiatry. 2015 May 26;5(5):e572. doi: 10.1038/tp.2015.67. PMID: 26080089; PMCID: PMC4471296.

Link to full text: Acid-base dysregulation and chemosensory mechanisms in panic disorder: a translational update

Abstract: Panic disorder (PD), a complex anxiety disorder characterized by recurrent panic attacks, represents a poorly understood psychiatric condition which is associated with significant morbidity and an increased risk of suicide attempts and completed suicide. Recently however, neuroimaging and panic provocation challenge studies have provided insights into the pathoetiology of panic phenomena and have begun to elucidate potential neural mechanisms that may underlie panic attacks. In this regard, accumulating evidence suggests that acidosis may be a contributing factor in induction of panic. Challenge studies in patients with PD reveal that panic attacks may be reliably provoked by agents that lead to acid-base dysbalance such as CO2 inhalation and sodium lactate infusion. Chemosensory mechanisms that translate pH into panic-relevant fear, autonomic, and respiratory responses are therefore of high relevance to the understanding of panic pathophysiology. Herein, we provide a current update on clinical and preclinical studies supporting how acid-base imbalance and diverse chemosensory mechanisms may be associated with PD and discuss future implications of these findings.

Title: The amygdala is a chemosensor that detects carbon dioxide and acidosis to elicit fear behavior

Authors: Ziemann AE, Allen JE, Dahdaleh NS, Drebot II, Coryell MW, Wunsch AM, Lynch CM, Faraci FM, Howard MA 3rd, Welsh MJ, Wemmie JA.

Journal: Cell. 2009 Nov 25;139(5):1012-21. doi: 10.1016/j.cell.2009.10.029. PMID: 19945383; PMCID: PMC2808123.

Link to full text: The amygdala is a chemosensor that detects carbon dioxide and acidosis to elicit fear behavior

Abstract: The amygdala processes and directs inputs and outputs that are key to fear behavior. However, whether it directly senses fear-evoking stimuli is unknown. Because the amygdala expresses acid-sensing ion channel-1a (ASIC1a), and ASIC1a is required for normal fear responses, we hypothesized that the amygdala might detect a reduced pH. We found that inhaled CO(2) reduced brain pH and evoked fear behavior in mice. Eliminating or inhibiting ASIC1a markedly impaired this activity, and localized ASIC1a expression in the amygdala rescued the CO(2)-induced fear deficit of ASIC1a null animals. Buffering pH attenuated fear behavior, whereas directly reducing pH with amygdala microinjections reproduced the effect of CO(2). These data identify the amygdala as an important chemosensor that detects hypercarbia and acidosis and initiates behavioral responses. They also give a molecular explanation for how rising CO(2) concentrations elicit intense fear and provide a foundation for dissecting the bases of anxiety and panic disorders.

Title: Carbon dioxide for neurogenic orthostatic hypotension in adults: a novel therapy

Authors: Baker JR, Ranada SI, Incognito AV, Sheldon RS, Morillo CA, Wilson RJA, Phillips AA, Raj SR.

Journal: Eur Heart J. 2024 Oct 7:ehae653. doi: 10.1093/eurheartj/ehae653. Epub ahead of print. PMID: 39374345.

Link to PubMed: Carbon dioxide for neurogenic orthostatic hypotension in adults: a novel therapy

Abstract: Neurogenic orthostatic hypotension (nOH) is a hallmark feature of autonomic nervous system failure. On standing, people with nOH experience large reductions in blood pressure (≥20/10 mmHg) and debilitating symptoms (e.g. light-headedness, blurred vision, and syncope). Current blood pressure therapies have limited efficacy and potentially serious side effects, including supine hypertension, creating a strong clinical need for new therapeutic approaches. Previous studies examining the effects of hypocapnia [i.e. low arterial carbon dioxide (CO2)] on blood pressure have shown that hyperventilation-induced hypocapnia decreases supine blood pressure in nOH patients and can be readily prevented with exogenous CO2. Whether exogenous CO2 can prevent OH in these patients is unknown. In this proof-of-concept study, we tested the hypothesis that increased inspired CO2 increases standing blood pressure in patients with nOH.

Title: Hyperventilation syndrome after general anesthesia: Our experience

Authors: Bansal T, Hooda S.

Journal: J Anaesthesiol Clin Pharmacol. 2016 Oct-Dec;32(4):536-537. doi: 10.4103/0970-9185.168192. PMID: 28096595; PMCID: PMC5187629.

Link to full text: Hyperventilation syndrome after general anesthesia: Our experience

Abstract: We read with interest the article published in J Anaesthesiol Clin Pharmacol 2015;31:284-5. Tomioka et al. have reported two cases of hyperventilation syndrome after general anesthesia. These authors have described that hyperventilation syndrome reflects the enhancement of sympathoadrenal tone. The first case was managed with midazolam. The second case was managed with beta-adrenergic blocker, propranolol hydrochloride. Hyperventilation syndrome often occurs under stressful conditions and has been reported before, during or after anesthesia and surgery. Hyperventilation is associated with multiple somatic symptoms due to hypocalcemia induced by respiratory alkalosis. We wish to share our experience of a 30 years old full term parturient presenting in labor in a highly anxious state with symptoms and signs of hypocalcemia in the form of circumoral numbness and severe carpopedal spasm. The patient was managed successfully with reassurance and infusion of 10 ml of 10% calcium gluconate over 10 min. The blood sample taken for ionised serum calcium before administering calcium gluconate showed a low level of 0.7 mmol/L (normal range 1–1.5 mmol/L). We did not use anxiolytic or beta blocker for fear of any untoward effect on the fetus as the patient was in labor.

Title: Carbon dioxide influence on nitric oxide production in endothelial cells and astrocytes: cellular mechanisms

Authors: Fathi AR, Yang C, Bakhtian KD, Qi M, Lonser RR, Pluta RM.

Journal: Brain Res. 2011 Apr 22;1386:50-7. doi: 10.1016/j.brainres.2011.02.066. Epub 2011 Feb 26. PMID: 21362408; PMCID: PMC3073030.

Link to full text: Carbon dioxide influence on nitric oxide production in endothelial cells and astrocytes: cellular mechanisms

Abstract: Cerebral vessels may regulate cerebral blood flow by responding to changes in carbon dioxide (CO(2)) through nitric oxide (NO) production. To better determine the role of NO production by human adult cerebral microvascular endothelial cells and human fetal astrocytes under different CO(2) conditions, we studied endothelial cell and astrocyte production of NO under hypo-, normo- and hypercapnic conditions. Human cerebral endothelial cell and fetal astrocyte cultures were exposed to hypocapnic (pCO(2) 21.7±6.7mmHg), normocapnic (pCO(2) 40.1±0.9mmHg) and hypercapnic (pCO(2) 56.3±8.7mmHg) conditions. NO production was recorded continuously over 24hours with stable pH. N-nitro-l-arginine [NLA; a nitric oxide synthase (NOS) inhibitor] and l-arginine (substrate for NO production via NOS) were used to further define the role of NOS in chemoregulation. NO levels in endothelial cells increased during hypercapnia by 36% in 8hours and remained 25% above baseline. NO increase in astrocytes was 30% after 1hour but returned to baseline at 8hours. NLA blocked NO increase in endothelial cells under hypercapnia. During hypocapnia, NO levels in the endothelial cells decreased by 30% at 8hours but were unchanged in astrocytes. l-arginine prevented NO decrease in endothelial cells under hypocapnia. NO changes in the endothelial cells correlated with changes in pCO(2) (R=0.99) and were independent of pH. This study suggests that cerebral endothelial cells and astrocytes release NO under normocapnic conditions and NO production is increased during hypercapnia and decreased during hypocapnia independent of pH. Further, this demonstrates that endothelial cells may play a pivotal role in chemoregulation by modulating NOS activity.

Title: The influence of carbon dioxide on brain activity and metabolism in conscious humans

Authors: Xu F, Uh J, Brier MR, Hart J Jr, Yezhuvath US, Gu H, Yang Y, Lu H.

Journal: J Cereb Blood Flow Metab. 2011 Jan;31(1):58-67. doi: 10.1038/jcbfm.2010.153. Epub 2010 Sep 15. PMID: 20842164; PMCID: PMC3049465.

Link to full text: The influence of carbon dioxide on brain activity and metabolism in conscious humans

Abstract: A better understanding of carbon dioxide (CO(2)) effect on brain activity may have a profound impact on clinical studies using CO(2) manipulation to assess cerebrovascular reserve and on the use of hypercapnia as a means to calibrate functional magnetic resonance imaging (fMRI) signal. This study investigates how an increase in blood CO(2), via inhalation of 5% CO(2), may alter brain activity in humans. Dynamic measurement of brain metabolism revealed that mild hypercapnia resulted in a suppression of cerebral metabolic rate of oxygen (CMRO(2)) by 13.4% ± 2.3% (N=14) and, furthermore, the CMRO(2) change was proportional to the subject's end-tidal CO(2) (Et-CO(2)) change. When using functional connectivity MRI (fcMRI) to assess the changes in resting-state neural activity, it was found that hypercapnia resulted in a reduction in all fcMRI indices assessed including cluster volume, cross-correlation coefficient, and amplitude of the fcMRI signal in the default-mode network (DMN). The extent of the reduction was more pronounced than similar indices obtained in visual-evoked fMRI, suggesting a selective suppression effect on resting-state neural activity. Scalp electroencephalogram (EEG) studies comparing hypercapnia with normocapnia conditions showed a relative increase in low frequency power in the EEG spectra, suggesting that the brain is entering a low arousal state on CO(2) inhalation.

Title: High Behavioral Sensitivity to Carbon Dioxide Associates with Enhanced Fear Memory and Altered Forebrain Neuronal Activation

Authors: McMurray KMJ, Gray A, Horn P, Sah R.

Journal: Neuroscience. 2020 Mar 1;429:92-105. doi: 10.1016/j.neuroscience.2019.12.009. Epub 2020 Jan 10. PMID: 31930959; PMCID: PMC8418542.

Link to full text: High Behavioral Sensitivity to Carbon Dioxide Associates with Enhanced Fear Memory and Altered Forebrain Neuronal Activation

Abstract: There is considerable interest in pre-trauma individual differences that may contribute to increased risk for developing post-traumatic stress disorder (PTSD). Identification of underlying vulnerability factors that predict differential responses to traumatic experiences is important. Recently, the relevance of homeostatic perturbations in shaping long-term behavior has been recognized. Sensitivity to CO2 inhalation, a homeostatic threat to survival, was shown to associate with the later development of PTSD symptoms in veterans. Here, we investigated whether behavioral sensitivity to CO2 associates with PTSD-relevant behaviors and alters forebrain fear circuitry in mice. Mice were exposed to 5% CO2 or air inhalation and tested one week later on acoustic startle and footshock contextual fear conditioning, extinction and reinstatement. CO2 inhalation evoked heterogenous freezing behaviors (high freezing CO2-H and low freezing CO2-L) that significantly associated with fear conditioning and extinction behaviors. CO2-H mice elicited potentiated conditioned fear and delayed extinction while behavioral responses in CO2-L mice were similar to the air group. Persistent neuronal activation marker ΔFosB immunostaining revealed altered regional neuronal activation within the hippocampus, amygdala and medial pre-frontal cortex that correlated with conditioned fear and extinction. Inter-regional co-activation mapping revealed disruptions in the coordinated activity of hippocampal dentate-amygdala-infralimbic regions and infralimbic-prelimbic associations in CO2-H mice that may explain their enhanced fear phenotype. In conclusion, our data support an association of behavioral sensitivity to interoceptive threats such as CO2 with altered fear responding to exteroceptive threats and suggest that "CO2-sensitive" individuals may be susceptible to developing PTSD.

Title: Respiratory relief: a new and rapid method for the treatment of phobic states

Authors: Orwin A.

Journal: Br J Psychiatry. 1971 Dec;119(553):635-7. doi: 10.1192/bjp.119.553.635. PMID: 5143663.

Link to PubMed: Respiratory relief: a new and rapid method for the treatment of phobic states

Abstract: The treatment of phobic states by desensitization is tedious and not always completely successful (Marks, 1969). Recently newer methods claiming improvement in speed and efficacy have been introduced, such as flooding or implosion (for example, Boulougouris et al., 1971) and prolonged exposure (Watson et al., 1971). An approach based on respira tory control which seems to be as effective as other methods and even more rapid in action is described in this short paper. Since anxiety caused by maximum voluntary respiratory arrest (M.V.R.A.) is abolished on breathing, it was decided to determine whether voluntary manipulation of respiratory function could lead to the use of this respiratory relief (R.R.) as an antagonist to the anxiety induced by specific phobias. It was hypothesized that the temporal pairing of R.R. with the anxiety induced by presentation of the feared object, the timing of which could be accurate to within less than a second, should result in the extinc tion of the anxiety response. The methods described are simple, but more complex variants have been used (Orwin, 1971).

Title: Metabolism and physiological effects of carbon dioxide. Implications in anaesthetic management

Authors: Victoria R.

Journal: The Moldovan Medical Journal 60.3 (2017): 42-45.

Link to full text: Metabolism and physiological effects of carbon dioxide. Implications in anaesthetic management

Abstract: Background: Carbon dioxide is a normal by-product of aerobic metabolism that maintains the equilibrium of respiratory act, being eliminated from the lungs. Despite of an increasing number of researches concerning carbon dioxide metabolism and its effects on human homeostasis, there are still discussions about carbon dioxide metabolism, physiology and its implication in anaesthetic management, ICU, critically ill patient. The use of mild to moderate hypercapnia during general anaesthesia and in mechanically ventilated patients is growing, based on scientifical researches of last years. Material and methods: There has been scientifically evaluated data from PubMed, 2002-2017. Key words used in search are: "carbon dioxide", "ventilation", "metabolism", "gas change". There were selected articles, taking in consideration their title, and chosen abstracts. The article contains a qualitative analysis and synthesis of the recommendation, concerning anaesthetic management and metabolism of carbon dioxide. Conclusions: Carbon dioxide metabolism and its anaesthetic management, represents a challenge that will be actual for many years in future. The amount of controversial studies about effects of carbon dioxide on patients under general anaesthesia or mechanical ventilation, determined us to perform a review of literature, and evaluate it. There are known facts about carbon dioxide metabolism, such as normal values, how it is produced in human body, how it is evacuated, effects on cardiovascular, nervous systems, and still there are many controversial studies on that topic, that determine to study it more and find new research results. Key words: carbon dioxide, ventilation, metabolism, gas change.

Title: The Effects of altered arterial tensions of carbon dioxide and oxygen on cerebral blood flow and cerebral oxygen consumption of normal young men

Authors: Kety SS, Schmidt CF.

Journal: J Clin Invest. 1948 Jul;27(4):484-92. doi: 10.1172/JCI101995. PMID: 16695569; PMCID: PMC439519.

Link to full text: The Effects of altered arterial tensions of carbon dioxide and oxygen on cerebral blood flow and cerebral oxygen consumption of normal young men

Abstract: A method for measuring quantitatively the volume of cerebral blood flow in man by inhalation of nitrous oxide (1) found its first application in a study of the cerebral circulatory effects of low CO2 tension achieved by hyperventilation; of high CO2 tension, and of high and low 02 tensions obtained by inhalation of appropriate gas mixtures (2). Only the first part of this study, the effects of active and passive hyperventilation, has been published in detail (3). The purpose of the present paper is to present the remainder of these findings and to derive from them, together with those of the hyperventilation experiments, evidence bearing on the intrinsic control of the human cerebral circulation as revealed by quantitative measurements.

Title: Five percent CO₂ is a potent, fast-acting inhalation anticonvulsant

Authors: Tolner EA, Hochman DW, Hassinen P, Otáhal J, Gaily E, Haglund MM, Kubová H, Schuchmann S, Vanhatalo S, Kaila K.

Journal: 2011 Jan;52(1):104-14. doi: 10.1111/j.1528-1167.2010.02731.x. Epub 2010 Sep 30. PMID: 20887367; PMCID: PMC3017646.

Link to full text: Five percent CO₂ is a potent, fast-acting inhalation anticonvulsant

Abstract: Purpose: CO₂ has been long recognized for its anticonvulsant properties. We aimed to determine whether inhaling 5% CO₂ can be used to suppress seizures in epilepsy patients. The effect of CO₂ on cortical epileptic activity accompanying behavioral seizures was studied in rats and nonhuman primates, and based on these data, preliminary tests were carried out in humans.

Methods: In freely moving rats, cortical afterdischarges paralleled by myoclonic convulsions were evoked by sensorimotor cortex stimulation. Five percent CO₂ was applied for 5 min, 3 min before stimulation. In macaque monkeys, hypercarbia was induced by hypoventilation while seizure activity was electrically or chemically evoked in the sensorimotor cortex. Seven patients with drug-resistant partial epilepsy were examined with video-EEG (electroencephalography) and received 5% CO₂ in medical carbogen shortly after electrographic seizure onset.

Results: In rats, 5% CO₂ strongly suppressed cortical afterdischarges, by approximately 75%, whereas responses to single-pulse stimulation were reduced by about 15% only. In macaques, increasing pCO₂) from 37 to 44-45 mm Hg (corresponding to inhalation of 5% CO₂ or less) suppressed stimulation-induced cortical afterdischarges by about 70% and single, bicuculline-induced epileptiform spikes by approximately 25%. In a pilot trial carried out in seven patients, a rapid termination of electrographic seizures was seen despite the fact that the application of 5% CO₂ was started after seizure generalization.

Conclusions: Five percent CO₂ has a fast and potent anticonvulsant action. The present data suggest that medical carbogen with 5% CO₂ can be used for acute treatment to suppress seizures in epilepsy patients.

Title: Carbon dioxide influence on nitric oxide production in endothelial cells and astrocytes: Cellular mechanisms

Authors: Fathi AR, Yang C, Bakhtian KD, Qi M, Lonser RR, Pluta RM.

Journal: Brain Res. 2011 Apr 22;1386:50-7. doi: 10.1016/j.brainres.2011.02.066. Epub 2011 Feb 26. PMID: 21362408; PMCID: PMC3073030.

Link to full text: Carbon dioxide influence on nitric oxide production in endothelial cells and astrocytes: Cellular mechanisms

Abstract: Cerebral vessels may regulate cerebral blood flow by responding to changes in carbon dioxide (CO(2)) through nitric oxide (NO) production. To better determine the role of NO production by human adult cerebral microvascular endothelial cells and human fetal astrocytes under different CO(2) conditions, we studied endothelial cell and astrocyte production of NO under hypo-, normo- and hypercapnic conditions. Human cerebral endothelial cell and fetal astrocyte cultures were exposed to hypocapnic (pCO(2) 21.7±6.7mmHg), normocapnic (pCO(2) 40.1±0.9mmHg) and hypercapnic (pCO(2) 56.3±8.7mmHg) conditions. NO production was recorded continuously over 24hours with stable pH. N-nitro-l-arginine [NLA; a nitric oxide synthase (NOS) inhibitor] and l-arginine (substrate for NO production via NOS) were used to further define the role of NOS in chemoregulation. NO levels in endothelial cells increased during hypercapnia by 36% in 8hours and remained 25% above baseline. NO increase in astrocytes was 30% after 1hour but returned to baseline at 8hours. NLA blocked NO increase in endothelial cells under hypercapnia. During hypocapnia, NO levels in the endothelial cells decreased by 30% at 8hours but were unchanged in astrocytes. l-arginine prevented NO decrease in endothelial cells under hypocapnia. NO changes in the endothelial cells correlated with changes in pCO(2) (R=0.99) and were independent of pH. This study suggests that cerebral endothelial cells and astrocytes release NO under normocapnic conditions and NO production is increased during hypercapnia and decreased during hypocapnia independent of pH. Further, this demonstrates that endothelial cells may play a pivotal role in chemoregulation by modulating NOS activity.

Title: Out of thin air: Hyperventilation-triggered seizures

Authors: Salvati KA, Beenhakker MP.

Journal: Brain Res. 2019 Jan 15;1703:41-52. doi: 10.1016/j.brainres.2017.12.037. Epub 2017 Dec 27. PMID: 29288644; PMCID: PMC6546426.

Link to full text: Out of thin air: Hyperventilation-triggered seizures

Abstract: Voluntary hyperventilation triggers seizures in the vast majority of people with absence epilepsy. The mechanisms that underlie this phenomenon remain unknown. Herein, we review observations - many made long ago - that provide insight into the relationship between breathing and absence seizures.

Keywords: Circuit; Epilepsy; Respiration; Seizure; Thalamus; pH.

Title: Hyperventilation syndromes in medicine and psychiatry: a review.

Authors: Lum LC.

Journal: J R Soc Med. 1987 Apr;80(4):229-31. doi: 10.1177/014107688708000413. PMID: 3295237; PMCID: PMC1290766.

Link to full text: Hyperventilation syndromes in medicine and psychiatry: a review.

Abstract: In 1940 Lewis' described DaCosta's syndrome (a frequent presentation of hyperventilation) as 'one of the commonest afflictions of sedentary town dwellers'. In this issue ofthe JRSM(p. 216), the paper 'Role of the nurse counsellor in managing patients with the hyperventilation syndrome' prompts a review of recent advances in this field. The 1980s have seen a remarkable upsurge of interest in the role of hyperventilation in functional illness, anxiety disorder, panic and phobic states2-4. In a previous review5 the author traced the changing concepts over the last 50 years, from the early realization6 that overbreathing was the physiological mechanism whereby anxiety produces somatic disturbance, to the later view7 that anxiety disorders arose from the persistence of symptoms due to unrecognized hyperventilation. Yet, even in 1982, a Lancet editorial8 commented on how rarely this diagnosis was even considered. Snaith9, reviewing panic disorder, failed to mention it

Title: Hyperventilation syndrome: a diagnosis begging for recognition.

Authors: Magarian GJ, Middaugh DA, Linz DH.

Journal: West J Med. 1983 May;138(5):733-6. PMID: 6880192; PMCID: PMC1010816.

Link to full text: Hyperventilation syndrome: a diagnosis begging for recognition.

Abstract: Beginning with the American Civil War, military physicians seeing soldiers under the stress of combat have described a syndrome characterized by breathlessness, lightheadedness or dizziness, pronounced fatigue and exercise intolerance, numbness and paresthesias and chest pain. Rarely have organic diseases been found to account for the symptoms in such cases, yet despite reassurance, symptoms commonly persist for prolonged periods despite removal from the apparent stress setting. This syndrome has been given many names including irritable heart, soldier's heart, Da Costa's syndrome, effort syndrome, neurocirculatory asthenia and, more recently, hyperventilation syndrome. Since the original descriptions in soldiers, it is now recognized that hyperventilation occurs in many persons under stresses of daily living. It is manifest not only in those overtly stressed, anxious and depressed but also in those who appear outwardly calm as they "bottle up" their feelings, often because of undeveloped or lack of acceptable emotional outlets. Physicians and lay persons alike readily recognize acute hyperventilatory attacks occurring under acute stress. However, chronic or recurrent hyperventilation problems often are unrecognized probably for a variety of reasons, including the frequent lack of obvious overbreathing, a tendency to focus on one or two complaints that alone are not particularly suggestive of hyperventilation, minimal discussion of the topic in medical school and cursory coverage in medical textbooks.

Title: Carbon dioxide flooding of the pericardium-An old practice revisited

Authors: Bhaskar, J.

Journal: Indian Journal of Thoracic and Cardiovascular Surgery 24 (2008): 37-43.

Link to Springer: Carbon dioxide flooding of the pericardium-An old practice revisited

Abstract: Background
Cerebral air embolism is a known complication of open-heart surgery with devastating consequences. Neurological injury after cardio-pulmonary bypass can be broadly divided into 2 categories:
Type I: Focal injury
Type II: Intellectual deterioration/neurocognitive impairment.
Increasing evidence points to cerebral embolization as the principal culprit. Since the early days of modern cardiac surgery, surgeons have attempted to reduce embolization by various means. One of the early techniques propounded was carbon dioxide insufflation of the pericardial cavity. Carbon dioxide, was uniquely positioned for this, being denser than air, more soluble than air, and easily managed perioperatively if the blood levels became unphysiological.. The technique did not find favor, since there was no instrumentation, in the early days of cardiopulmonary bypass to measure and establish its impact on the actual outcomes.
Methods
A literature search was done using the key words carbon dioxide, open heart, cardiac across Pubmed, Google and Cochrane databases in an attempt to find out the role of carbon dioxide flooding in modern cardiac surgery. Our effort yielded over 244 articles, of which 48 were deemed to be relevant in answering our question.
Conclusions
Reviewing all the information we conclude that with proper technique, carbon dioxide flooding does significantly reduce embolic counts. However, there are no studies at this point in time, which show a direct correlation between the practice and improved neurocognitive outcome.

Title: Regulation of the Cerebral Circulation by Arterial Carbon Dioxide

Authors: Hoiland RL, Fisher JA, Ainslie PN.

Journal: Compr Physiol. 2019 Jun 12;9(3):1101-1154. doi: 10.1002/cphy.c180021. PMID: 31187899.

Link to PubMed: Regulation of the Cerebral Circulation by Arterial Carbon Dioxide

Abstract: Intact, coordinated, and precisely regulated cerebrovascular responses are required for the maintenance of cerebral metabolic homeostasis, adequate perfusion, oxygen delivery, and acid-base balance during deviations from homeostasis. Increases and decreases in the partial pressure of arterial carbon dioxide (PaCO2 ) lead to robust and rapid increases and decreases in cerebral blood flow (CBF). In awake and healthy humans, PaCO2 is the most potent regulator of CBF, and even small fluctuations can result in large changes in CBF. Alterations in the responsiveness of the cerebral vasculature can be detected with carefully controlled stimulus-response paradigms and hold relevance for cerebrovascular risk in steno-occlusive disease. As changes in PaCO2 do not typically occur in isolation, the integrative influence of physiological factors such as intracranial pressure, arterial oxygen content, cerebral perfusion pressure, and sympathetic nervous activity must be considered. Further, age and sex, as well as vascular pathologies are also important to consider. Following a brief summary of key historical events in the development of our understanding of cerebrovascular physiology and an overview of the measurement techniques to index CBF this review provides an in-depth description of CBF regulation in response to alterations in PaCO2 . Cerebrovascular reactivity and regional flow distribution are described, with further consideration of how differences in reactivity of parallel networks can lead to the "steal" phenomenon. Factors that influence cerebrovascular reactivity are discussed and the mechanisms and regulatory pathways mediating the exquisite sensitivity of the cerebral vasculature to changes in PaCO2 are outlined. Finally, topical avenues for future research are proposed. © 2019 American Physiological Society. Compr Physiol 9:1101-1154, 2019.

Title: Carbon Dioxide for the treatment of Febrile seizures: rationale, feasibility, and design of the CARDIF-study

Authors: Ohlraun S, Wollersheim T, Weiß C, Martus P, Weber-Carstens S, Schmitz D, Schuelke M.

Journal: J Transl Med. 2013 Jun 27;11:157. doi: 10.1186/1479-5876-11-157. PMID: 23806032; PMCID: PMC3700755.

Link to full text: Carbon Dioxide for the treatment of Febrile seizures: rationale, feasibility, and design of the CARDIF-study

Abstract: Background: 2-8% of all children aged between 6 months and 5 years have febrile seizures. Often these seizures cease spontaneously, however depending on different national guidelines, 20-40% of the patients would need therapeutic intervention. For seizures longer than 3-5 minutes application of rectal diazepam, buccal midazolam or sublingual lorazepam is recommended. Benzodiazepines may be ineffective in some patients or cause prolonged sedation and fatigue. Preclinical investigations in a rat model provided evidence that febrile seizures may be triggered by respiratory alkalosis, which was subsequently confirmed by a retrospective clinical observation. Further, individual therapeutic interventions demonstrated that a pCO2-elevation via re-breathing or inhalation of 5% CO2 instantly stopped the febrile seizures. Here, we present the protocol for an interventional clinical trial to test the hypothesis that the application of 5% CO2 is effective and safe to suppress febrile seizures in children.

Methods: The CARDIF (CARbon DIoxide against Febrile seizures) trial is a monocentric, prospective, double-blind, placebo-controlled, randomized study. A total of 288 patients with a life history of at least one febrile seizure will be randomized to receive either carbogen (5% CO2 plus 95% O2) or placebo (100% O2). As recurrences of febrile seizures mainly occur at home, the study medication will be administered by the parents through a low-pressure can fitted with a respiratory mask. The primary outcome measure is the efficacy of carbogen to interrupt febrile seizures. As secondary outcome parameters we assess safety, practicability to use the can, quality of life, contentedness, anxiousness and mobility of the parents.

Prospect: The CARDIF trial has the potential to develop a new therapy for the suppression of febrile seizures by redressing the normal physiological state. This would offer an alternative to the currently suggested treatment with benzodiazepines. This study is an example of academic translational research from the study of animal physiology to a new therapy.

Title: New breathing therapy reduces panic and anxiety by reversing hyperventilation

Authors: Rosenfield, D., Seidel, A., Bhaskara, L., & Hofmann, S.

Journal: J. Med. Sci 11, no. 8 (2011): 327-328.

Link to full text: New breathing therapy reduces panic and anxiety by reversing hyperventilation

Abstract: A new treatment that helps people with panic disorder to normalize their breathing works better to reduce panic symptoms and hyperventilation than traditional cognitive therapy, according to a new study. The new 'CART' treatment was found to be better than traditional cognitive therapy at altering hyperventilation and panic symptoms.

Title: Spinal cord transection inhibits HR reduction in anesthetized rats immersed in an artificial CO2-hot spring bath

Authors: Yamamoto N, Hashimoto M.

Journal: Int J Biometeorol. 2007 Jan;51(3):201-8. doi: 10.1007/s00484-006-0055-6. Epub 2006 Sep 7. PMID: 16957940.

Link to PubMed: Spinal cord transection inhibits HR reduction in anesthetized rats immersed in an artificial CO2-hot spring bath

Abstract: Like humans, the heart rate (HR) of anesthetized rats immersed in CO(2)-water is lower than that when immersed in tap water at the same temperature. To investigate the afferent signal pathway in the mechanism of HR reduction, Wistar rats were anesthetized with urethane and then the spinal cord was transected between T(4) and T(5). The animals were immersed up to the axilla in a bathtub of tap-water (CO(2) contents: 10-20 mg.l(-1)) or of CO(2)-water (965-1,400 mg.l(-1)) at 35 degrees C while recording HR, arterial blood pressure, and arterial blood gas parameters (PaCO(2), PaO(2), pH). Arterial blood gas parameters did not change during immersion, irrespective of CO(2) concentration of the bath water, whereas the HR was reduced in the CO(2)-water bath. The inhalation of CO(2)-mixed gas (5% CO(2), 20% O(2), 75% N(2)) resulted in increased levels of blood gases and an increased HR during immersion in all types of water tested. The HR reduction observed in sham transected control animals immersed in CO(2)-water disappeared after subsequent spinal cord transection. These results show that the dominant afferent signal pathway to the brain, which is involved in inducing the reduced HR during immersion in CO(2)-water, is located in the neuronal route and not in the bloodstream.

Title: Hypocapnia related changes in pain-induced brain activation as measured by functional MRI

Authors: Terekhin P, Forster C.

Journal: Neurosci Lett. 2006 May 29;400(1-2):110-4. doi: 10.1016/j.neulet.2006.02.040. Epub 2006 Mar 6. PMID: 16517071.

Link to PubMed: Hypocapnia related changes in pain-induced brain activation as measured by functional MRI

Abstract: Stress, acute pain and chronic pain may often result in hyperventilation (HV) which produces hypocapnia. The aim of this fMRI-study was to investigate the influence of hypocapnia on cortical activation during noxious stimulation in 14 healthy volunteers. The intensity of voluntary HV was controlled by capnometry Three tasks were performed in the fMRI sessions: (I) three 3-min HV periods with 7-min periods of recovery in between; (II) mechanically induced phasic pain stimulation--pain task (PT); (III) tapping--motor task (MT). The last two of these protocols were performed under normocapnic and hypocapnic conditions. HV decreased the fMRI signal by 3-7% in all regions of the cortex and subcortical nuclei. This decrease was most prominent in the opercular, frontal and temporal areas. When the PT was performed during hypocapnia a strong reduction in cluster sizes and lower t-values in S1 and insular cortex were found. In contrast MT was accompanied by an increase in cluster sizes and higher t-values. From this we conclude that hypocapnia significantly influences the BOLD signal in nociceptive and motor systems, indicating that either the coupling between the BOLD effect and neuronal processing changed or that the activity in the cortical network which represents the pain processing is decreased. These effects should be considered for functional brain imaging studies on the nociceptive system.

Title: Adenosine and ATP link PCO2 to cortical excitability via pH

Authors: Dulla CG, Dobelis P, Pearson T, Frenguelli BG, Staley KJ, Masino SA.

Journal: Neuron. 2005 Dec 22;48(6):1011-23. doi: 10.1016/j.neuron.2005.11.009. PMID: 16364904; PMCID: PMC1924599.

Link to full text: Adenosine and ATP link PCO2 to cortical excitability via pH

Abstract: In addition to affecting respiration and vascular tone, deviations from normal CO(2) alter pH, consciousness, and seizure propensity. Outside the brainstem, however, the mechanisms by which CO(2) levels modify neuronal function are unknown. In the hippocampal slice preparation, increasing CO(2), and thus decreasing pH, increased the extracellular concentration of the endogenous neuromodulator adenosine and inhibited excitatory synaptic transmission. These effects involve adenosine A(1) and ATP receptors and depend on decreased extracellular pH. In contrast, decreasing CO(2) levels reduced extracellular adenosine concentration and increased neuronal excitability via adenosine A(1) receptors, ATP receptors, and ecto-ATPase. Based on these studies, we propose that CO(2)-induced changes in neuronal function arise from a pH-dependent modulation of adenosine and ATP levels. These findings demonstrate a mechanism for the bidirectional effects of CO(2) on neuronal excitability in the forebrain.

Title: Application of carbon-based nanomaterials in Alzheimer's disease

Authors: Bai M, Shao X, Wang C, Wang J, Wang X, Guan P, Hu X.

Journal: Mater Horiz. 2025 Feb 3;12(3):673-693. doi: 10.1039/d4mh01256a. Erratum in: Mater Horiz. 2025 Feb 3;12(3):1018. doi: 10.1039/d4mh90125h. PMID: 39526325.

Link to PubMed: Application of carbon-based nanomaterials in Alzheimer's disease

Abstract: Alzheimer's disease (AD) is a chronic, progressive neurodegenerative disorder marked by permanent impairment of brain function across the whole brain. This condition results in a progressive deterioration of cognitive function in patients and is frequently associated with psychological symptoms such as agitation and anxiety, imposing a significant burden on both patients and their families. Nanomaterials possess numerous distinctive physical and chemical features that render them extensively utilized. In the biomedical domain, nanomaterials can be utilized for disease prevention and therapy, including medication delivery systems, biosensors, and tissue engineering. This article explores the etiology and potential molecular processes of AD, as well as the application of carbon-based nanomaterials in the diagnosis and treatment of AD. Some of such nanomaterials are carbon quantum dots, carbon nanotubes, and graphene, among others. These materials possess distinctive physicochemical features that render them highly promising for applications in biosensing, drug delivery, neuroprotection, and photothermal treatment. In addition, this review explored various therapeutic approaches for AD in terms of reducing inflammation, preventing oxidative damage, and inhibiting Aβ aggregation. The advent of carbon nanomaterials in nanotechnology has facilitated the development of novel treatment approaches for Alzheimer's disease. These strategies provide promising approaches for early diagnosis, effective intervention and neuroprotection of the disease.

Title: Association of Hypocapnia in Children with Febrile Seizures

Authors: Sachan D, Goyal S.

Journal: J Pediatr Neurosci. 2018 Oct-Dec;13(4):388-391. doi: 10.4103/JPN.JPN_73_18. PMID: 30937077; PMCID: PMC6413613.

Link to full text: Association of Hypocapnia in Children with Febrile Seizures

Abstract: Introduction: Febrile seizure is a benign condition in children. Susceptibility genes associated with febrile convulsions have been identified, but the precise pathophysiologic mechanism that triggers febrile seizure is unclear. Using animal models, it has been demonstrated that hyperthermia causes respiratory alkalosis with consequent brain alkalosis and seizures. This study was conducted to find out any association of febrile seizures with fever induced hypocapnia.

Methods: We conducted a cross sectional observational study and enrolled 45 children presenting with febrile seizures. Axillary temperature was measured and venous blood gas analysis was done soon after admission and within 24 hour of seizure onset. Mean pH and pCO2 from venous blood gas analysis was measured and compared with standard normal values. Data was analyzed using SPSS software version 17.0 software.

Results: The mean pCO2 (27.95 ± 5.31mmHg) was much below normal range, and 91% of children had hypocapnia (pCO2 <35) after the febrile seizures. However alkalosis (pH > 7.45) was demonstrated in only 20% of children. Also pCO2 levels in samples drawn before 2 hours were significantly less than those taken after 2 hours (23.24 ± 3.44 vs 29.29 ± 4.99 respectively; p = 0.001).

Conclusion: Our data indicates that febrile seizures may be associated with fever induced hyperventilation and ensuing hypocapnia may be one of the precipitating factor in inducing seizures. However, well-structured human trials are needed to demonstrate the same.

Keywords: Febrile seizures; hypocapnia; respiratory alkalosis; venous blood gas.

Title: Cerebrospinal fluid lipidomic fingerprint of obstructive sleep apnoea in Alzheimer's disease

Authors: Dakterzada F, Benítez ID, Targa A, Carnes A, Pujol M, Jové M, Mínguez O, Vaca R, Sánchez-de-la-Torre M, Barbé F, Pamplona R, Piñol-Ripoll G.

Journal: Alzheimers Res Ther. 2023 Aug 7;15(1):134. doi: 10.1186/s13195-023-01278-7. PMID: 37550750; PMCID: PMC10408111.

Link to full text: Cerebrospinal fluid lipidomic fingerprint of obstructive sleep apnoea in Alzheimer's disease

Abstract: Background: Obstructive sleep apnoea (OSA) has a high prevalence in patients with Alzheimer's disease (AD). Both conditions have been shown to be associated with lipid dysregulation. However, the relationship between OSA severity and alterations in lipid metabolism in the brains of patients with AD has yet to be fully elucidated. In this context, we examined the cerebrospinal fluid (CSF) lipidome of patients with suspected OSA to identify potential diagnostic biomarkers and to provide insights into the pathophysiological mechanisms underlying the effect of OSA on AD.

Methods: The study included 91 consecutive AD patients who underwent overnight polysomnography (PSG) to diagnose severe OSA (apnoea-hypopnea index ≥ 30/h). The next morning, CSF samples were collected and analysed by liquid chromatography coupled to mass spectrometry in an LC-ESI-QTOF-MS/MS platform.

Results: The CSF levels of 11 lipid species were significantly different between AD patients with (N = 38) and without (N = 58) severe OSA. Five lipids (including oxidized triglyceride OxTG(57:2) and four unknown lipids) were significantly correlated with specific PSG measures of OSA severity related to sleep fragmentation and hypoxemia. Our analyses revealed a 4-lipid signature (including oxidized ceramide OxCer(40:6) and three unknown lipids) that provided an accuracy of 0.80 (95% CI: 0.71-0.89) in the detection of severe OSA. These lipids increased the discriminative power of the STOP-Bang questionnaire in terms of the area under the curve (AUC) from 0.61 (0.50-0.74) to 0.85 (0.71-0.93).

Conclusions: Our results reveal a CSF lipidomic fingerprint that allows the identification of AD patients with severe OSA. Our findings suggest that an increase in central nervous system lipoxidation may be the principal mechanism underlying the association between OSA and AD.

Keywords: Alzheimer's disease; Biomarker; Cerebrospinal fluid; Diagnosis; Lipidomics; Obstructive sleep apnoea; STOP-Bang questionnaire.

Title: Cerebrovascular reactivity to carbon dioxide in Alzheimer's disease

Authors: Glodzik L, Randall C, Rusinek H, de Leon MJ.

Journal: J Alzheimers Dis. 2013;35(3):427-40. doi: 10.3233/JAD-122011. PMID: 23478306; PMCID: PMC3776495.

Link to full text: Cerebrovascular reactivity to carbon dioxide in Alzheimer's disease

Abstract: There is growing evidence that cerebrovascular reactivity to carbon dioxide (CVRCO2) is impaired in Alzheimer's disease (AD). Preclinical and animal studies suggest chronic hypercontractility in brain vessels in AD. We review (a) preclinical studies of mechanisms for impaired CVRCO2 in AD; (b) clinical studies of cerebrovascular function in subjects with AD dementia, mild cognitive impairment (MCI), and normal cognition. Although results of clinical studies are inconclusive, an increasing number of reports reveal an impairment of vascular reactivity to carbon dioxide in subjects with AD, and possibly also in MCI. Thus, CVRCO2 may be an attractive means to detect an early vascular dysfunction in subjects at risk.

Title: Carbon dioxide inhalation treatments of neurotic anxiety. An overview

Authors: Wolpe J.

Journal: J Nerv Ment Dis. 1987 Mar;175(3):129-33. doi: 10.1097/00005053-198703000-00001. PMID: 3102688.

Link to PubMed: Carbon dioxide inhalation treatments of neurotic anxiety. An overview

Abstract: A lucky chance more than 30 years ago revealed the remarkable efficacy of single inhalations of high concentrations of carbon dioxide in eliminating or markedly reducing free-floating anxiety. The reduction of anxiety lasts for days, weeks, or longer--well beyond the persistence of carbon dioxide in the body. The effects are explicable on the hypothesis that free-floating anxiety is anxiety conditioned to continuously present sources of stimulation, such as background noise or the awareness of space or time, and that the anxiety response habit is weakened when the anxiety is inhibited by the competition of responses that carbon dioxide induces. More recently, it has become apparent that inhalations of carbon dioxide, applied in a different manner, are effective in overcoming maladaptive anxiety responses to specific stimuli, e.g., social stimuli. The substance is also proving to be a valuable resource in the treatment of the common variety of panic attacks.

Title: Corticospinal excitability is associated with hypocapnia but not changes in cerebral blood flow

Authors: Hartley GL, Watson CL, Ainslie PN, Tokuno CD, Greenway MJ, Gabriel DA, O'Leary DD, Cheung SS.

Journal: J Physiol. 2016 Jun 15;594(12):3423-37. doi: 10.1113/JP271914. Epub 2016 Feb 24. PMID: 26836470; PMCID: PMC4908026.

Link to full text: Corticospinal excitability is associated with hypocapnia but not changes in cerebral blood flow

Abstract: Key points: Reductions in cerebral blood flow (CBF) may be implicated in the development of neuromuscular fatigue; however, the contribution from hypocapnic-induced reductions (i.e. P ETC O2) in CBF versus reductions in CBF per se has yet to be isolated. We assessed neuromuscular function while using indomethacin to selectively reduce CBF without changes in P ETC O2 and controlled hyperventilation-induced hypocapnia to reduce both CBF and P ETC O2. Increased corticospinal excitability appears to be exclusive to reductions in P ETC O2 but not reductions in CBF, whereas sub-optimal voluntary output from the motor cortex is moderately associated with decreased CBF independent of changes in P ETC O2. These findings suggest that changes in CBF and P ETC O2 have distinct roles in modulating neuromuscular function.

Abstract: Although reductions in cerebral blood flow (CBF) may be involved in central fatigue, the contribution from hypocapnia-induced reductions in CBF versus reductions in CBF per se has not been isolated. This study examined whether reduced arterial PCO2 (P aC O2), independent of concomitant reductions in CBF, impairs neuromuscular function. Neuromuscular function, as indicated by motor-evoked potentials (MEPs), maximal M-wave (Mmax ) and cortical voluntary activation (cVA) of the flexor carpi radialis muscle during isometric wrist flexion, was assessed in ten males (29 ± 10 years) during three separate conditions: (1) cyclooxygenase inhibition using indomethacin (Indomethacin, 1.2 mg kg(-1) ) to selectively reduce CBF by 28.8 ± 10.3% (estimated using transcranial Doppler ultrasound) without changes in end-tidal PCO2 (P ETC O2); (2) controlled iso-oxic hyperventilation-induced reductions in P aC O2 (Hypocapnia), P ETC O2 = 30.1 ± 4.5 mmHg with related reductions in CBF (21.7 ± 6.3%); and (3) isocapnic hyperventilation (Isocapnia) to examine the potential direct influence of hyperventilation-mediated activation of respiratory control centres on CBF and changes in neuromuscular function. Change in MEP amplitude (%Mmax ) from baseline was greater in Hypocapnia tha in Isocapnia (11.7 ± 9.8%, 95% confidence interval (CI) [2.6, 20.7], P = 0.01) and Indomethacin (13.3 ± 11.3%, 95% CI [2.8, 23.7], P = 0.01) with a large Cohen's effect size (d ≥ 1.17). Although not statistically significant, cVA was reduced with a moderate effect size in Indomethacin (d = 0.7) and Hypocapnia (d = 0.9) compared to Isocapnia. In summary, increased corticospinal excitability - as reflected by larger MEP amplitude - appears to be exclusive to reduced P aC O2, but not reductions in CBF per se. Sub-optimal voluntary output from the motor cortex is moderately associated with decreased CBF, independent of reduced P aC O2.

Title: Intermittent hypoxia can aggravate motor neuronal loss and cognitive dysfunction in ALS mice

Authors: Kim SM, Kim H, Lee JS, Park KS, Jeon GS, Shon J, Ahn SW, Kim SH, Lee KM, Sung JJ, Lee KW.

Journal: PLoS One. 2013 Nov 26;8(11):e81808. doi: 10.1371/journal.pone.0081808. PMID: 24303073; PMCID: PMC3841127.

Link to full text: Intermittent hypoxia can aggravate motor neuronal loss and cognitive dysfunction in ALS mice

Abstract: Background: Patients with ALS may be exposed to variable degrees of chronic intermittent hypoxia. However, all previous experimental studies on the effects of hypoxia in ALS have only used a sustained hypoxia model and it is possible that chronic intermittent hypoxia exerts effects via a different molecular mechanism from that of sustained hypoxia. No study has yet shown that hypoxia (either chronic intermittent or sustained) can affect the loss of motor neurons or cognitive function in an in vivo model of ALS.

Objective: To evaluate the effects of chronic intermittent hypoxia on motor and cognitive function in ALS mice.

Methods: Sixteen ALS mice and 16 wild-type mice were divided into 2 groups and subjected to either chronic intermittent hypoxia or normoxia for 2 weeks. The effects of chronic intermittent hypoxia on ALS mice were evaluated using the rotarod, Y-maze, and wire-hanging tests. In addition, numbers of motor neurons in the ventral horn of the spinal cord were counted and western blot analyses were performed for markers of oxidative stress and inflammatory pathway activation.

Results: Compared to ALS mice kept in normoxic conditions, ALS mice that experienced chronic intermittent hypoxia had poorer motor learning on the rotarod test, poorer spatial memory on the Y-maze test, shorter wire hanging time, and fewer motor neurons in the ventral spinal cord. Compared to ALS-normoxic and wild-type mice, ALS mice that experienced chronic intermittent hypoxia had higher levels of oxidative stress and inflammation.

Conclusions: Chronic intermittent hypoxia can aggravate motor neuronal death, neuromuscular weakness, and probably cognitive dysfunction in ALS mice. The generation of oxidative stress with activation of inflammatory pathways may be associated with this mechanism. Our study will provide insight into the association of hypoxia with disease progression, and in turn, the rationale for an early non-invasive ventilation treatment in patients with ALS.

Title: Involvement of Lipids in Alzheimer's Disease Pathology and Potential Therapies

Authors: Chew H, Solomon VA, Fonteh AN.

Journal: Front Physiol. 2020 Jun 9;11:598. doi: 10.3389/fphys.2020.00598. PMID: 32581851; PMCID: PMC7296164.

Link to full text: Involvement of Lipids in Alzheimer's Disease Pathology and Potential Therapies

Abstract: Lipids constitute the bulk of the dry mass of the brain and have been associated with healthy function as well as the most common pathological conditions of the brain. Demographic factors, genetics, and lifestyles are the major factors that influence lipid metabolism and are also the key components of lipid disruption in Alzheimer's disease (AD). Additionally, the most common genetic risk factor of AD, APOE ϵ4 genotype, is involved in lipid transport and metabolism. We propose that lipids are at the center of Alzheimer's disease pathology based on their involvement in the blood-brain barrier function, amyloid precursor protein (APP) processing, myelination, membrane remodeling, receptor signaling, inflammation, oxidation, and energy balance. Under healthy conditions, lipid homeostasis bestows a balanced cellular environment that enables the proper functioning of brain cells. However, under pathological conditions, dyshomeostasis of brain lipid composition can result in disturbed BBB, abnormal processing of APP, dysfunction in endocytosis/exocytosis/autophagocytosis, altered myelination, disturbed signaling, unbalanced energy metabolism, and enhanced inflammation. These lipid disturbances may contribute to abnormalities in brain function that are the hallmark of AD. The wide variance of lipid disturbances associated with brain function suggest that AD pathology may present as a complex interaction between several metabolic pathways that are augmented by risk factors such as age, genetics, and lifestyles. Herewith, we examine factors that influence brain lipid composition, review the association of lipids with all known facets of AD pathology, and offer pointers for potential therapies that target lipid pathways.

Keywords: amyloid precursor protein; apolipoproteins; blood-brain barrier; energy metabolism; inflammation; late-onset Alzheimer's disease; mitochondria; myelination.

Title: One Small Step for Mouse: High CO2 Inhalation as a New Therapeutic Strategy for Parkinson's Disease

Authors: Nadeev AD, Kritskaya KA, Fedotova EI, Berezhnov AV.

Journal: Biomedicines. 2022 Nov 6;10(11):2832. doi: 10.3390/biomedicines10112832. PMID: 36359351; PMCID: PMC9687253.

Link to full text: One Small Step for Mouse: High CO2 Inhalation as a New Therapeutic Strategy for Parkinson's Disease

Abstract: Parkinson's disease (PD) is a ubiquitous neurodegenerative disorder for which no effective treatment strategies are available. Existing pharmacotherapy is aimed only at correcting symptoms and slowing the progression of the disease, mainly by replenishing dopamine deficiency. It is assumed that mitochondrial dysfunction plays a key role in the pathogenesis of PD. It has been suggested that activation of specific degradation of damaged mitochondria (mitophagy) may prevent cell death. An almost exclusive way to initiate mitophagy is acidification of intracellular pH. We attempted to implement transient brain acidification using two experimental therapy strategies: forced moderate physical activity and high CO2 inhalation. The beneficial effects of CO2 supplementation on behavioral aspects were demonstrated in a rotenone-induced PD model. Mice treated with CO2 restored their exploratory behavior and total locomotor activity lost after rotenone administration. Additionally, this treatment enabled the removal of impaired coordination. We have illustrated this therapeutic strategy using histological studies of brain sections to confirm the survival of nigrostriatal areas. These findings suggest that high CO2 inhalation presumably initiates mitophagy via transient brain acidification, and can treat PD-like symptoms in a rodent rotenone model of PD.

Keywords: Parkinson's disease; experimental therapy; hypercapnia; neurodegeneration; neuroprotection.

Title: Oxidative stress and cellular pathologies in Parkinson's disease

Authors: Puspita L, Chung SY, Shim JW.

Journal: Mol Brain. 2017 Nov 28;10(1):53. doi: 10.1186/s13041-017-0340-9. PMID: 29183391; PMCID: PMC5706368.

Link to full text: Oxidative stress and cellular pathologies in Parkinson's disease

Abstract: Parkinson's disease (PD) is a chronic and progressive neurodegeneration of dopamine neurons in the substantia nigra. The reason for the death of these neurons is unclear; however, studies have demonstrated the potential involvement of mitochondria, endoplasmic reticulum, α-synuclein or dopamine levels in contributing to cellular oxidative stress as well as PD symptoms. Even though those papers had separately described the individual roles of each element leading to neurodegeneration, recent publications suggest that neurodegeneration is the product of various cellular interactions. This review discusses the role of oxidative stress in mediating separate pathological events that together, ultimately result in cell death in PD. Understanding the multi-faceted relationships between these events, with oxidative stress as a common denominator underlying these processes, is needed for developing better therapeutic strategies.

Keywords: Alpha-synuclein; Dopamine neurons; Mitochondria; Oxidative stress; Parkinson's disease; Reactive oxygen species; Unfolded protein response.

Title: An essay on the shaking palsy. 1817

Authors: Parkinson J.

Journal: 1817. J Neuropsychiatry Clin Neurosci. 2002 Spring;14(2):223-36; discussion 222. doi: 10.1176/jnp.14.2.223. PMID: 11983801.

Link to full text: An essay on the shaking palsy. 1817

Abstract: The advantages which have been derived from the caution with which hypothetical statements are admitted, are in no instance more obvious than in those sciences which more particularly belong to the healing art. It therefore is necessary, that some conciliatory explanation should be offered for the present publication: in which, it is acknowledged, that mere conjecture takes the place of experiment; and, that analogy is the substitute for anatomical examination, the only sure foundation for pathological knowledge.
When, however, the nature of the subject, and the circumstances under which it has been here taken up, are considered, it is hoped that the offering of the following pages to the attention of the medical public, will not be severely censured. The disease, respecting which the present inquiry is made, is of a nature highly afflictive. Notwithstanding which, it has not yet obtained a place in the classification of nosologists; some have regarded its characteristic symptoms as distinct and different diseases, and others have given its name to diseases differing essentially from it; whilst the unhappy sufferer has considered it as an evil, from the domination of which he had no prospect of escape.
The disease is of long duration: to connect, therefore, the symptoms which occur in its later stages with those which mark its commencement, requires a continuance of observation of the same case, or at least a correct history of its symptoms, even for several years. Of both these advantages the writer has had the opportunities of availing himself, and has hence been led particularly to observe several other cases in which the disease existed in different stages of its progress. By these repeated observations, he hoped that he had been led to a probable conjecture as to the nature of the malady, and that analogy had suggested such means as might be productive of relief, and perhaps even of cure, if employed before the disease had been too long established. He therefore considered it to be a duty to submit his opinions to the examination of others, even in their present state of immaturity and imperfection.
To delay their publication did not, indeed, appear to be warrantable. The disease had escaped particular notice; and the task of ascertaining its nature and cause by anatomical investigation, did not seem likely to be taken up by those who, from their abilities and opportunities, were most likely to accomplish it. That these friends to humanity and medical science, who have already unveiled to us many of the morbid processes by which health and life is abridged, might be excited to extend their researches to this malady, was much desired; and it was hoped, that this might be procured by the publication of these remarks.
Should the necessary information be thus obtained, the writer will repine at no censure which the precipitate publication of mere conjectural suggestions may incur: but shall think himself fully rewarded by having excited the attention of those, who may point out the most appropriate means of relieving a tedious and most distressing malady.

Title: Panic anxiety, dyspnea, and respiratory disease. Theoretical and clinical considerations

Authors: Smoller JW, Pollack MH, Otto MW, Rosenbaum JF, Kradin RL.

Journal: Am J Respir Crit Care Med. 1996 Jul;154(1):6-17. doi: 10.1164/ajrccm.154.1.8680700. PMID: 8680700.

Link to PubMed: Panic anxiety, dyspnea, and respiratory disease. Theoretical and clinical considerations

Abstract: There is intriguing evidence suggesting pathophysiologic relationships among dyspnea, hyperventilation, and panic anxiety. The symptoms of panic attacks and pulmonary disease overlap, so that panic anxiety can reflect underlying cardiopulmonary disease and dyspnea can reflect an underlying anxiety disorder. The pathogenesis of panic may be related to respiratory physiology by several mechanisms: the anxiogenic effects of hyperventilation, the catastrophic misinterpretation of respiratory symptoms, and/or a neurobiologic sensitivity to CO2, lactate, or other signals of suffocation. In a subset of patients with PD, incipient pulmonary dysfunction may also contribute to their anxiety symptoms. Patients with pulmonary disease, particularly those with obstructive lung disease, have a high rate of panic symptoms and PD. There is reason to believe that pulmonary disease constitutes a risk factor for the development of panic related to repeated experiences with dyspnea and life-threatening exacerbations of pulmonary dysfunction, repeated episodes of hypercapnia or hyperventilation, the use of anxiogenic medications, and the stress of coping with chronic disease. Panic in pulmonary patients may carry significant morbidity, including phobic avoidance of activity, overly aggressive treatment with anxiogenic medications, and more prolonged and frequent hospitalization. Successful treatment of panic in these patients can improve functional status and quality of life by relieving anxiety and dyspnea. Nonpharmacologic treatment of panic, including cognitive-behavioral approaches, can be useful in patients with concomitant respiratory disease. Sedating medications such as benzodiazepines should be used with caution in patients with pulmonary disease to avoid respiratory depression. Serotonergic antidepressants (SSRIs) and anxiolytics (buspirone) may be effective treatments for panic or generalized anxiety in pulmonary patients and have relatively little potential for significant adverse effects.

Title: The Effects of Hypocapnia on Brain Tissue Pulsations

Authors: Alharbi M, Turner P, Ince J, Oura M, Ebirim KU, Almudayni A, Lecchini-Visintini A, Minhas JS, Chung EML.

Journal: Brain Sci. 2020 Sep 6;10(9):614. doi: 10.3390/brainsci10090614. PMID: 32899967; PMCID: PMC7565182.

Link to full text: The Effects of Hypocapnia on Brain Tissue Pulsations

Abstract: Hypocapnia is known to affect patients with acute stroke and plays a key role in governing cerebral autoregulation. However, the impact of hypocapnia on brain tissue pulsations (BTPs) is relatively unexplored. As BTPs are hypothesised to result from cerebrovascular resistance to the inflow of pulsatile arterial blood, it has also been hypothesised that cerebral autoregulation changes mediated by hypocapnia will alter BTP amplitude. This healthy volunteer study reports measurements of BTPs obtained using transcranial tissue Doppler (TCTD). Thirty participants underwent hyperventilation to induce mild hypocapnia. BTP amplitude, EtCO2, blood pressure, and heart rate were then analysed to explore the impact of hypocapnia on BTP amplitude. Significant changes in BTP amplitude were noted during recovery from hypocapnia, but not during the hyperventilation manoeuvre itself. However, a significant increase in heart rate and pulse pressure and decrease in mean arterial pressure were also observed to accompany hypocapnia, which may have confounded our findings. Whilst further investigation is required, the results of this study provide a starting point for better understanding of the effects of carbon dioxide levels on BTPs. Further research in this area is needed to identify the major physiological drivers of BTPs and quantify their interactions with other aspects of cerebral haemodynamics.

Keywords: BTP; brain tissue pulsations; cerebral autoregulation; hypocapnia.

Title: Humans without a sense of smell breathe differently

Authors: Gorodisky, L., Honigstein, D., Weissbrod, A. et al.

Journal: Nat Commun 15, 8809 (2024). https://doi.org/10.1038/s41467-024-52650-6

Link to full text: Humans without a sense of smell breathe differently

Abstract: Olfaction may play a restricted role in human behavior, yet paradoxically, its absence in anosmia is associated with diverse deleterious outcomes, culminating in reduced life expectancy. The mammalian nose serves two purposes: olfaction and breathing. Because respiratory patterns are impacted by odors, we hypothesized that nasal respiratory airflow may be altered in anosmia. We apply a wearable device that precisely logs nasal airflow for 24-hour-long sessions in participants with isolated congenital anosmia and controls. We observe significantly altered patterns of respiratory nasal airflow in anosmia in wake and in sleep. These differences allow classification of anosmia at 83% accuracy using the respiratory trace alone. Patterns of respiratory airflow have pronounced impact on health, emotion and cognition. We therefore suggest that a portion of the deleterious outcomes associated with anosmia may be attributed to altered patterns of respiratory nasal airflow rather than a direct result of lost odor perception per se.

Title: Dismantling the Component-Specific Effects of Yogic Breathing: Feasibility of a Fully Remote Three-Arm RCT with Virtual Laboratory Visits and Wearable Physiology

Authors: Ma Y, Yang H, Vazquez M, Buraks O, Haack M, Mullington JM, Goldstein MR.

Journal: Int J Environ Res Public Health. 2023 Feb 11;20(4):3180. doi: 10.3390/ijerph20043180. PMID: 36833875; PMCID: PMC9958552.

Link to full text: Dismantling the Component-Specific Effects of Yogic Breathing: Feasibility of a Fully Remote Three-Arm RCT with Virtual Laboratory Visits and Wearable Physiology

Abstract: Despite the growing research base examining the benefits and physiological mechanisms of slow-paced breathing (SPB), mindfulness (M), and their combination (as yogic breathing, SPB + M), no studies have directly compared these in a ”dismantling” framework. To address this gap, we conducted a fully remote three-armed feasibility study with wearable devices and video-based laboratory visits. Eighteen healthy participants (age 18–30 years, 12 female) were randomized to one of three 8-week interventions: slow-paced breathing (SPB, N = 5), mindfulness (M, N = 6), or yogic breathing (SPB + M, N = 7). The participants began a 24-h heart rate recording with a chest-worn device prior to the first virtual laboratory visit, consisting of a 60-min intervention-specific training with guided practice and experimental stress induction using a Stroop test. The participants were then instructed to repeat their assigned intervention practice daily with a guided audio, while concurrently recording their heart rate data and completing a detailed practice log. The feasibility was determined using the rates of overall study completion (100%), daily practice adherence (73%), and the rate of fully analyzable data from virtual laboratory visits (92%). These results demonstrate feasibility for conducting larger trial studies with a similar fully remote framework, enhancing the ecological validity and sample size that could be possible with such research designs.

Title: Increased net water loss by oral compared to nasal expiration in healthy subjects

Authors: Svensson S, Olin AC, Hellgren J.

Journal: Rhinology. 2006 Mar;44(1):74-7. PMID: 16550955.

Link to full text: Increased net water loss by oral compared to nasal expiration in healthy subjects

Abstract:
Aim of the study: To compare the difference in respiratory water loss during expiration through the nose and through the mouth, in healthy subjects.

Methods: The study included 19 healthy, non-smoking volunteers without any present history of non-infectious rhinitis, presenting with symptoms of rhinitis, asthma or previous nasal surgery. Nasal and oral expiratory breath condensates were collected using a breath condenser during tidal respiration at indoor resting conditions. During the nasal breath condensate sampling, the subjects were breathing into a transparent face mask covering the nose and the mouth with the mouth closed. During the oral breath condensate sampling, the subjects inhaled through the nose and exhaled through a mouthpiece connected to the condenser. The airflow during the sampling was assessed with a dry-spirometer connected to the condenser. Sampling was stopped after 100 litres of expired air for each breathing mode. Nasal sampling was done before and after decongestion of the nasal mucosa with oxymetazoline, 0.5 mg/ml. The effect on the nasal mucosa was assessed with acoustic rhinometry.

Results: The mean loss of expired water was 42% less by nasal expiration before decongestion than by oral expiration (1.9 x 10(-3) g/L min compared to 2.7 x 10(-3) g/L min, p < 0.001). The mean expiratory minute ventilation was 9.0 L/min by nasal respiration and 9.8 L/min by oral respiration. Decongestion of the nasal mucosa showed a mean increase of the cross-sectional area at 4 cm from the nostril (1.44 to 1.67 cm2, p = 0.0024), but there was no effect on the net water loss (1.9 x 10(-3) g/Lmin vs 1.9 x 10(-3) g/Lmin).

Conclusion: This study showed that the net water loss increased by 42% when the breathing mode was switched from nasal to oral expiration during tidal breathing in healthy subjects. Increased water and energy loss by oral breathing could be a contributing factor to the symptoms seen in patients suffering from nasal obstruction.

Title: Levels of end-tidal carbon dioxide are low despite normal respiratory rate in individuals with long COVID

Authors: Wood J, Tabacof L, Tosto-Mancuso J, McCarthy D, Kontorovich A, Putrino D.

Journal: J Breath Res. 2021 Dec 8;16(1). doi: 10.1088/1752-7163/ac3c18. PMID: 34808607.

Link to full text: Levels of end-tidal carbon dioxide are low despite normal respiratory rate in individuals with long COVID

Abstract: ETCO2 and respiratory rate (RR) were measured in patients attending an outpatient clinic using an EMMA capnograph (Masimo Corporation, Irvine, CA). Approval for the collection and publication of this data was provided by the relevant Institutional Review Board (STUDY-20-001537). All patients were previously reviewed by a physician and diagnosed with confirmed (via PCR and/or antibody test) or probable (according to World Health Organization and Center for Disease Control and Prevention guidelines [6, 7]) past COVID-19 infection. The criteria for diagnosis of long COVID was the presence of persistent symptoms for at least 12 weeks after initial infection [2]. The capnograph was calibrated and a healthy control was used to check device readings at the commencement of each clinic. At the time of assessment, patients were seated and at rest, and had not performed other assessments or procedures immediately prior. Patients were instructed to breathe orally through the capnograph adapter at their resting rate and depth for a minimum of 15 s, and until the ETCO2 level was stable for a further period of at least 5 s. Data were analyzed using Stata (StataCorp LLC, TX), with descriptive statistics and Pearson correlation coefficient provided.

Title: Effects of decortication and carotid sinus nerve section on ventilation of the rat

Authors: Maskrey M, Megirian D, Nicol SC.

Journal: Respir Physiol. 1981 Mar;43(3):263-73. doi: 10.1016/0034-5687(81)90108-0. PMID: 6792671.

Link to PubMed: Effects of decortication and carotid sinus nerve section on ventilation of the rat

Abstract: The effect on ventilation of exposure to hypoxic, hypercapnic and hypoxic/hypercapnic gas mixtures was studied before and after functional decortication of intact rats and rats in which the carotid chemoreceptors had been disconnected. Unanaesthetized rats responded to both hypoxia and hypercapnia with an increase in minute ventilation (V) through increases in both frequency (f) and tidal volume (VT). Decortication led to a greater V response to CO2. This was through an effect on f, rather than VT. Carotid sinus nerve section (CSNS) caused a lessening in the V response to gas mixtures, f and VT being equally affected. Decortication, following CSNS, increased the V response but this time through increased VT rather than f. This effect on VT was not specific to any particular gas mixture. It is concluded that the carotid body chemoreceptors, together with the bulbopontine rate controller, influence the response to CO2. It is further suggested that this integration takes place in the reticular formation and is normally under some degree of inhibition from the cerebral cortex.

Title: CO2, brainstem chemoreceptors and breathing

Authors: Nattie E.

Journal: Prog Neurobiol. 1999 Nov;59(4):299-331. doi: 10.1016/s0301-0082(99)00008-8. PMID: 10501632.

Link to full text: CO2, brainstem chemoreceptors and breathing

Abstract: The regulation of breathing relies upon chemical feedback concerning the levels of CO2 and O2. The carotid bodies, which detect O2, provide tonic excitation to brainstem respiratory neurons under normal conditions and dramatic excitation if O2 levels fall. Feedback for CO2 involves the carotid body and receptors in the brainstem, central chemoreceptors. Small increases in CO2 produce large increases in breathing. Decreases in CO2 below normal can, in sleep and anesthesia, decrease breathing, even to apnea. Central chemoreceptors, once thought localized to the surface of the ventral medulla, are likely distributed more widely with sites presently identified in the: (1) ventrolateral medulla; (2) nucleus of the solitary tract; (3) ventral respiratory group; (4) locus ceruleus; (5) caudal medullary raphé; and (6) fastigial nucleus of the cerebellum. Why so many chemoreceptor sites? Hypotheses, some with supporting data, include the following. Geographical specificity; all regions of the brainstem with respiratory neurons contain chemoreceptors. Stimulus intensity; some sites operate in the physiological range of CO2 values, others only with more extreme changes. Stimulus specificity; CO2 or pH may be sensed by multiple mechanisms. Temporal specificity; some sites respond more quickly to changes on blood or brain CO2 or pH. Syncytium; chemosensitive neurons may be connected via low resistance, gap junctions. Arousal state: sites may vary in effectiveness and importance dependent on state of arousal. Overall, as judged by experiments of nature, and in the laboratory, central chemoreceptors are critical for adequate breathing in sleep, but other aspects of the control system can maintain breathing in wakefulness.

Title: Ventilatory response of decorticate and decerebrate cats to hypoxia and CO2

Authors: Tenney SM, Ou LC.

Journal: Respir Physiol. 1977 Feb;29(1):81-92. doi: 10.1016/0034-5687(77)90119-0. PMID: 847311.

Link to PubMed: Ventilatory response of decorticate and decerebrate cats to hypoxia and CO2

Abstract: The steady state ventilatory response of normal, fully awake cats was studied under graded hypoxia (at PAO2 = 110, 55, 45 torr) with PACO2 controlled throughout at the resting, normoxic level and at +5 torr. Subsequently, either a mid-collicular decerebration or a decortication was performed, and the ventilatory studies were repeated. Respiratory frequency, tidal volume, and ventilation in the decerebrate state responded to hypoxia and hypercapnia in a manner indistinguishable from the control. The decorticate cats, however, exhibited an exaggerated response to hypoxia, principally the result of increased frequency. The negative hypoxic, hypercapnic interaction, characteristic of awake cats, was demonstrable in both the decerebrate and decorticate animals. The findings are interpreted as revealing coupled descending influences on the medullary respiratory centers in hypoxia--one that is facilitatory and originates in the diencephalon, and the other, inhibitory, from the cerebrum. The significance of this suprapontine system in normal hypoxic ventilatory control is discussed.

Title: Air hunger' from increased Pco2 persists after complete neuromuscular block in humans

Authors: Banzett RB, Lansing RW, Brown R, Topulos GP, Yager D, Steele SM, Londoño B, Loring SH, Reid MB, Adams L, et al.

Journal: Respir Physiol. 1990 Jul;81(1):1-17. doi: 10.1016/0034-5687(90)90065-7. PMID: 2120757.

Link to PubMed: Air hunger' from increased Pco2 persists after complete neuromuscular block in humans

Abstract: The tolerance of totally curarized subjects for prolonged breath hold is viewed by many as evidence that respiratory muscle contraction is essential to generate the sensation of breathlessness. Although conflicting evidence exists, none of it was obtained during total neuromuscular block. We completely paralyzed four normal, unsedated subjects with vecuronium (a non-depolarizing neuromuscular blocker). Subjects were mechanically ventilated with hyperoxic gas mixtures at fixed rate and tidal volume. End-expiratory PCO2 (PETCO2) was varied surreptitiously by changing inspired PCO2. Subjects rated their respiratory discomfort or 'air hunger' every 45 sec. At low PETCO2 (median 35 Torr) they felt little or no air hunger. When PETCO2 was raised (median 44 Torr) all subjects reported severe air hunger. They had reported the same degree of air hunger at essentially the same PETCO2 before paralysis. When questioned afterwards all subjects said the sensation could be described by the terms 'air hunger', 'urge to breathe', and 'shortness of breath', and that is was like breath holding. They reported no fundamental difference in the sensation before and after paralysis. We conclude that respiratory muscle contraction is not important in the genesis of air hunger evoked by hypercapnia.

Title: Accuracy of respiratory symptom perception in different affective contexts

Authors: Bogaerts K, Notebaert K, Van Diest I, Devriese S, De Peuter S, Van den Bergh O.

Journal: J Psychosom Res. 2005 Jun;58(6):537-43. doi: 10.1016/j.jpsychores.2004.12.005. PMID: 16125521.

Link to PubMed: Accuracy of respiratory symptom perception in different affective contexts

Abstract: Objective: The accuracy of respiratory symptom perception was investigated in different affective contexts in participants (N=48) scoring high or low for negative affectivity (NA). Methods: Within-subject correlations were calculated between two subjective ratings and their respective physiological referent (the rating of "deeper breathing" and respiratory volume, and rating of "faster breathing" and breathing frequency) across nine consecutive breathing trials. Three different air mixtures were used: room air, air enriched with 7.5% CO(2), and with 10% CO(2). For half the participants, the trials were framed in a pleasant context, created by adding a pleasant odour to the air mixture in addition to information announcing pleasant feelings as a result of breathing the air mixtures. The other half received the trials in a distressing context: A foul smelling odour was added and the information announced unpleasant feelings. Results: High-NA persons were overall less accurate than were low-NA persons in the perception of respiratory volume. For breathing frequency, high-NA persons were significantly less accurate in the distressing condition than in the pleasant one, whereas for low-NA persons, the information frame did not matter. Conclusion: The study shows that the accuracy of respiratory symptom reports is reduced in high-NA persons, especially in a distressing context.

Title: A prospective examination of distress tolerance and early smoking lapse in adult self-quitters

Authors: Brown RA, Lejuez CW, Strong DR, Kahler CW, Zvolensky MJ, Carpenter LL, Niaura R, Price LH.

Journal: Nicotine Tob Res. 2009 May;11(5):493-502. doi: 10.1093/ntr/ntp041. Epub 2009 Apr 16. PMID: 19372572; PMCID: PMC2671465.

Link to full text: A prospective examination of distress tolerance and early smoking lapse in adult self-quitters

Abstract: Introduction: A significant percentage of smokers attempting cessation lapse to smoking within a matter of days, and current models of relapse devote insufficient attention to such early smoking lapse. Studies attempting to relate severity of nicotine withdrawal symptoms to short-term smoking cessation outcomes have yielded equivocal results. How one reacts to the discomfort of nicotine withdrawal and quitting smoking (i.e., distress tolerance) may be a more promising avenue of investigation with important treatment implications. Methods: The present investigation examined distress tolerance and early smoking lapse using a prospective design. Participants were 81 adult daily smokers recruited through newspaper advertisements targeted at smokers planning to quit smoking without assistance (i.e., no pharmacotherapy or psychosocial treatment; 42 males and 39 females; mean age = 42.6 years, SD = 12.20). Results: As hypothesized, both greater breath-holding duration and carbon dioxide-enriched air persistence were associated with a significantly lower risk of smoking lapse following an unaided quit attempt. These effects were above and beyond the risk associated with levels of nicotine dependence, education, and history of major depressive disorder, suggesting that distress tolerance and task persistence may operate independently of risk factors such as nicotine dependence and depressive history. In contrast to expectation, persistence on the Paced Auditory Serial Addition Test (a psychological challenge task) was not a significant predictor of earlier smoking lapse. Discussion: These results are discussed in relation to refining theoretical models of the role of distress tolerance in early smoking lapse and the utility of such models in the development of specialized treatment approaches for smoking cessation.

Title: Distress Tolerance and Duration of Past Smoking Cessation Attempts

Authors: Brown RA, Lejuez CW, Kahler CW, Strong DR.

Journal: J Abnorm Psychol. 2002 Feb;111(1):180-5. PMID: 11866171.

Link to PubMed: Distress Tolerance and Duration of Past Smoking Cessation Attempts

Abstract: The present study tested the hypothesis that limited ability to tolerate physical and psychological distress is associated with early relapse from smoking cessation. Specifically, the authors exposed 16 current smokers who had failed to sustain any previous quit attempt for more than 24 hr (immediate relapsers) and 16 smokers with at least 1 sustained quit attempt of 3 months or longer (delayed relapsers) to psychological (mental arithmetic) and physical (carbon dioxide inhalation-breath holding) stressors. Relative to delayed relapsers, immediate relapsers were characterized by higher baseline levels of affective vulnerability, by greater levels of dysphoria and urge to smoke after 12 hr of nicotine deprivation, and by less task persistence on the stressors, suggesting that these may be risk factors for early lapse in the context of quitting smoking.

Title: Rebreathing to cope with hyperventilation: experimental tests of the paper bag method

Authors: van den Hout MA, Boek C, van der Molen GM, Jansen A, Griez E.

Journal: J Behav Med. 1988 Jun;11(3):303-10. doi: 10.1007/BF00844435. PMID: 3139884.

Link to PubMed: Rebreathing to cope with hyperventilation: experimental tests of the paper bag method

Abstract: To explore if and how the common rebreathing (paper bag) approach to hyperventilation works, two experiments were carried out. In the first experiment, 12 normal volunteers, aware of the common physiological rebreathing rationale, were twice asked to overbreath intensely and then either to rebreath or to restart normal ventilation. Alveolar CO2 increased more quickly and physical symptoms disappeared earlier in the rebreathing condition. The second experiment had a similar design; however, this time the subjects were led to believe that, after both hyperventilation provocation tests, they were rebreathing in a semiclosed tube system. On one of the occasions, the tube system was, in fact, open. The CO2 restoration again differed in the two conditions. In this second experiment, the moment of symptom disappearance was not significantly earlier in the rebreathing condition. The combined results of the studies suggest that expectation and suggestion play a role in the effects of rebreathing approaches on hyperventilation.

Title: Ventilatory response to CO2. II. Studies in neurotic psychiatric patients and practitioners of transcendental meditation

Authors: Singh BS.

Journal: Psychosom Med. 1984 Jul-Aug;46(4):347-62. doi: 10.1097/00006842-198407000-00004. PMID: 6435149.

Link to PubMed: Ventilatory response to CO2. II. Studies in neurotic psychiatric patients and practitioners of transcendental meditation

Abstract: The prominence of respiratory symptoms in patients with neurotic psychiatric disorders is noted and the literature on the control of respiration is reviewed to attempt to explain this finding. A previous study demonstrated a positive correlation between the ventilatory response to CO2 (S) and neurotic personality traits in a group of normal subjects. This study attempted to follow up this finding and hypothesized that a group of neurotically disturbed patients would have a higher S value and a group of individuals who practiced a calming technique such as transcendental meditation (TM) would have a lower S value than normal subjects. The second hypothesis was confirmed, but not the first, in that the neurotically disturbed patients had the lowest mean values for S of the three groups, rather than the highest. Particular characteristics of the sample of psychiatric patients cast doubt, however, on the validity of this finding. Three additional findings of this study were that anxious, depressive, and hyperventilating subject groups were no different from one another in terms of S values; that very experienced TM practitioners (sidhas) could significantly lower their ventilatory response to CO2 in the meditating state as compared to the nonmeditating alert state; and that the S value did not increase in two male subjects with endogenous depression after successful treatment with electroconvulsive therapy.

Title: Pattern of breathing and ventilatory response to CO2 in subjects practicing hatha-yoga

Authors: Stănescu DC, Nemery B, Veriter C, Maréchal C.

Journal: J Appl Physiol Respir Environ Exerc Physiol. 1981 Dec;51(6):1625-9. doi: 10.1152/jappl.1981.51.6.1625. PMID: 6798002.

Link to PubMed: Pattern of breathing and ventilatory response to CO2 in subjects practicing hatha-yoga

Abstract: WE studied eight Belgian subjects well advanced in the practice of hatha-yoga and compared them with eight sex-, age-, and height-matched control subjects. Practice of yoga (range 4-12 yr) involves control of posture and manipulation of breathing, including slow near-vital capacity maneuvers accompanied by apnea at end inspiration and end expiration. Average values for the yoga and the control group (in parentheses) are as follows: ventilation (VE) 5.53 1 X min-1 (7.07); tidal volume (VT), 1.03 liters (0.56); rate of breathing, 5.5 min-1 (13.4); end-tidal PCO2, 39.0 Torr (35.3). All differences are significant (P less than 0.05). Ventilatory response to CO2 (rebreathing technique) was significantly lower in the yoga group (P less than 0.01). The regression relating VE to VT during rebreathing of CO2 was VE = 8.1 (VT - 0.23) for the yoga group and VE = 15.8 (VT - 0.16) for the control group (P less than 0.005). We attribute these changes to chronic manipulation of respiration.

Title: Effect of transcendental meditation on breathing and respiratory control

Authors: Wolkove N, Kreisman H, Darragh D, Cohen C, Frank H.

Journal: J Appl Physiol Respir Environ Exerc Physiol. 1984 Mar;56(3):607-12. doi: 10.1152/jappl.1984.56.3.607. PMID: 6423589.

Link to PubMed: Effect of transcendental meditation on breathing and respiratory control

Abstract: We studied the effect of transcendental meditation (TM) on breathing using 16 experienced meditators and 16 control subjects. In controls, there was no significant difference in minute ventilation (VE), respiratory pattern, or hypercapnic response, whether breathing with eyes open-awake (CA), or with eyes closed-relaxing (CR). In meditators, VE decreased significantly during quiet breathing from 14.0 +/- 0.7 1/min with eyes open-awake (MA) to 12.4 +/- 0.6 1/min during meditation (MM) (P less than 0.02). The change in VE during meditation was due to a decrease in tidal volume (VT) resulting from a shortened inspiratory time (TI). Meditation was associated with a decreased response to progressive hypercapnia from 3.7 +/- 0.4 to 2.5 +/- 0.21 X min-1 X Torr-1 during MA and MM trials, respectively (P less than 0.01). During meditation VT was smaller at a given alveolar PCO2 than during MA studies because of a decrease in mean inspiratory flow rate (VT/TI). These observations suggest that an alteration in wakefulness, more subtle than sleep or the unconscious state, can significantly affect the chemical and neural regulation of breathing.

Title: Does a change in end-tidal carbon dioxide level predict high altitude mountain sickness?

Authors: Thundiyil JG, Williams AT, Little I, Stutsman M, Ladde JG, Papa L.

Journal: Heliyon. 2023 May 5;9(5):e16000. doi: 10.1016/j.heliyon.2023.e16000. PMID: 37215892; PMCID: PMC10192757.

Link to full text: Does a change in end-tidal carbon dioxide level predict high altitude mountain sickness?

Abstract: Background It is postulated that lack of hypoxic ventilatory response is a predictor for AMS. End-tidal carbon dioxide (ETCO2) is an accurate, noninvasive surrogate measure of ventilation. Objectives We sought to determine if changes in baseline ETCO2 predicts the development of AMS. Methods This prospective cohort study took place in three separate high-altitude hiking treks. Subjects included a convenience sample of hikers. Predictor variable was change in ETCO2 levels and outcome variable was AMS. Measurements of ETCO2 levels were obtained at the base and repeated daily at various elevations and the summit of each hike. Concurrently, hikers were scored for AMS by a trained investigator. We utilized correlation coefficients and developed a linear regression model for analysis. Results 21 subjects in 3 separate hikes participated: 10 ascended to 19,341 ft over 7 days, 6 ascended to 8900 ft in 1 day, and 4 ascended to 11,006 ft in 1 day. Mean age was 40 years, 67% were males, mean daily elevation gain was 2150 ft, and 5 hikers developed AMS. The correlation coefficients for ETCO2 and development of AMS were −0.46 (95%CI -0.33 to −0.57), and −0.77 (95%CI -0.71 to −0.83) for ETCO2 and altitude. ETCO2 predicted the development of symptoms better than the elevation with AUCs of 0.90 (95%CI 0.81–0.99) versus 0.64 (95%CI 0.45–0.83). An ETCO2 measurement of ≤22 mmHg was 100% sensitive and 60% specific for predicting AMS. Conclusions ETCO2 was strongly correlated with altitude and moderately correlated with AMS and it was a better predictor than altitude.

Title: Effect of carbon dioxide in acute mountain sickness: a rediscovery

Authors: Harvey TC, Raichle ME, Winterborn MH, Jensen J, Lassen NA, Richardson NV, Bradwell AR.

Journal: Lancet. 1988 Sep 17;2(8612):639-41. doi: 10.1016/s0140-6736(88)90465-5. Erratum in: Lancet 1988 Oct 1;2(8614):808. PMID: 2901515.

Link to PubMed: Effect of carbon dioxide in acute mountain sickness: a rediscovery

Abstract: The effect of adding CO2 to inhaled air in six subjects with acute mountain sickness was investigated during a medical expedition to 5400 m.3% CO2 in ambient air increased ventilation and resulted in a rise in PaO2 of between 24% and 40%. There was a 9-28% increase in PaCO2 and a reduction of the respiratory alkalosis normally seen at high altitude. Symptoms of acute mountain sickness were rapidly relieved. In three subjects cerebral blood flow increased by 17-39%, so that oxygen delivery to the brain would have been considerably improved. This study confirms earlier suggestions of the beneficial effect of CO2 inhalation at high altitude.

Title: Supplemental CO2 improves oxygen saturation, oxygen tension, and cerebral oxygenation in acutely hypoxic healthy subjects

Authors: Stepanek J, Dunn RA, Pradhan GN, Cevette MJ.

Journal: Physiol Rep. 2020 Jul;8(14):e14513. doi: 10.14814/phy2.14513. PMID: 32725791; PMCID: PMC7387889.

Link to full text: Supplemental CO2 improves oxygen saturation, oxygen tension, and cerebral oxygenation in acutely hypoxic healthy subjects

Abstract: Oxygen is viewed in medicine as the sole determinant of tissue oxygenation, though carbon dioxide homeostasis is equally important and clinically often ignored. The aims of this study were as follows: (a) to examine the effects of different acute hypoxic conditions on partial pressure of arterial oxygen ( PaO2), arterial oxygen saturation of hemoglobin (SaO2), and regional cerebral saturation of hemoglobin (rSO2 ); and (b) to evaluate supplemental CO2 as a tool to improve oxygenation in acutely hypoxic individuals. We hypothesized that exposure to gas mixtures with added CO2 would improve oxygenation in hypoxic human subjects. Twenty healthy subjects were exposed to 5-min intervals of two gas mixtures: hypoxic gas mixture containing 8% oxygen, and a CO2-enriched mixture containing 8% oxygen plus either 3% or 5% CO2. Ten subjects received the 3% CO2 -enriched mixture, and the remaining 10 subjects received the 5% CO2 -enriched mixture. The order of exposure was randomized. Blood gases, pulse oximetry, end-tidal CO2, and cerebral oximetry were measured. Compared to the purely hypoxic gas group, PaO2 was increased in the 3% and 5% CO2-enriched groups by 14.9 and 9.5 mmHg, respectively. Compared to pure hypoxia, SaO2 was increased in the 3% and 5% CO2-enriched groups by 16.8% and 12.9%, respectively. Both CO2-enriched gas groups had significantly higher end-exposure rSO2 and recovered to baseline rSO2 within 1 min, compared to the pure hypoxic gas group, which returned to baseline in 5 min. These results suggest that in acutely hypoxic subjects, CO2 supplementation improves blood oxygen saturation and oxygen tension as well as cerebral oxygenation measures.

Title: Rapid elimination of CO through the lungs: coming full circle 100 years on

Authors: Fisher JA, Iscoe S, Fedorko L, Duffin J.

Journal: Exp Physiol. 2011 Dec;96(12):1262-9. doi: 10.1113/expphysiol.2011.059428. Epub 2011 Oct 3. PMID: 21967899; PMCID: PMC3274699.

Link to full text: Rapid elimination of CO through the lungs: coming full circle 100 years on

Abstract: At the start of the 20th century, CO poisoning was treated by administering a combination of CO2 and O2 (carbogen) to stimulate ventilation. This treatment was reported to be highly effective, even reversing the deep coma of severe CO poisoning before patients arrived at the hospital. The efficacy of carbogen in treating CO poisoning was initially attributed to the absorption of CO2; however, it was eventually realized that the increase in pulmonary ventilation was the predominant factor accelerating clearance of CO from the blood. The inhaled CO2 in the carbogen stimulated ventilation but prevented hypocapnia and the resulting reductions in cerebral blood flow. By then, however, carbogen treatment for CO poisoning had been abandoned in favour of hyperbaric O2. Now, a half-century later, there is accumulating evidence that hyperbaric O2 is not efficacious, most probably because of delays in initiating treatment. We now also know that increases in pulmonary ventilation with O2-enriched gas can clear CO from the blood as fast, or very nearly as fast, as hyperbaric O2. Compared with hyperbaric O2, the technology for accelerating pulmonary clearance of CO with hyperoxic gas is not only portable and inexpensive, but also may be far more effective because treatment can be initiated sooner. In addition, the technology can be distributed more widely, especially in developing countries where the prevalence of CO poisoning is highest. Finally, early pulmonary CO clearance does not delay or preclude any other treatment, including subsequent treatment with hyperbaric O2.

Title: A randomized controlled trial of theophylline versus CO2 inhalation for treating apnea of prematurity

Authors: Al-Saif S, Alvaro R, Manfreda J, Kwiatkowski K, Cates D, Qurashi M, Rigatto H.

Journal: J Pediatr. 2008 Oct;153(4):513-8. doi: 10.1016/j.jpeds.2008.04.025. Epub 2008 Jun 4. PMID: 18534618.

Link to PubMed: A randomized controlled trial of theophylline versus CO2 inhalation for treating apnea of prematurity

Abstract: Objective: To determine whether inhalation of 0.8% CO(2) in preterm infants decreases the duration and rate of apnea as effectively as or better than theophylline with fewer adverse side effects. Study design: A prospective, randomized, control study of 42 preterm infants of gestational age 27 to 32 weeks assigned to receive inhaled CO(2) (n = 21) or theophylline (n = 21). The study group had a mean (+/- standard error of the mean) birth weight of 1437 +/- 57 g, gestational age of 29.4 +/- 0.3 weeks, and postnatal age of 43 +/- 4 days. After a control period, 0.8% CO(2) or theophylline was given for 2 hours, followed by a recovery period. Results: In the CO(2) group, apneic time and rate decreased significantly, from 9.4 +/- 1.6 seconds/minute and 94 +/- 15 apneic episodes/hour to 3.0 +/- 0.5 seconds/minute and 34 +/- 5 apneic episodes/hour. In the theophylline group, apneic time and rate decreased significantly, from 8 +/- 1 seconds/minute and 80 +/- 8 apneic episodes/hour to 2.5 +/- 0.4 seconds/minute and 28 +/- 3 apneic episodes/hour. Cerebral blood flow velocity (CBFV) decreased only during theophylline administration. Conclusions: Our findings suggest that inhaled low (0.8%) CO(2) concentrations in preterm infants is at least as effective as theophylline in decreasing the duration and number of apneic episodes, has fewer side effects, and causes no changes in CBFV. We speculate that CO(2) may be a better treatment for apnea of prematurity than methylxanthines.

Title: A Modeling Study on Inspired CO2 Rebreathing Device for Sleep Apnea Treatment by Means of CFD Analysis and Experiment

Authors: Shokoueinejad, M., Pazouki, A., Levin, J. et al.

Journal: Journal of Medical and Biological Engineering. 37. 288-297. 10.1007/s40846-017-0223-7.

Link to Springer: A Modeling Study on Inspired CO2 Rebreathing Device for Sleep Apnea Treatment by Means of CFD Analysis and Experiment

Abstract: We present the device design, simulation, and measurement results of a therapy device that potentially prevents sleep apnea by slightly increasing inspired CO2 through added dead space (DS). The rationale for treatment of sleep apnea with CO2 manipulation is based on two recently reported premises: (i) preventing transient reductions in PaCO2 will prevent the patient from reaching their apneic threshold, thereby preventing “central” apnea and instabilities in respiratory motor output; and (ii) raising PaCO2 and end-tidal CO2, even by a minimal amount, provides a strong recruitment of upper airway dilator muscles, thereby preventing airway obstruction. We have also provided the simulation results, obtained from solving the Navier–Stokes (NS) equations within the device volume. Therein, the NS equations are coupled with a convection–diffusion equation that represents the transport of CO2 in the device, thus enabling the transient simulation of CO2 propagation. Using this procedure, a prototype of variable volume dead space reservoir device was designed. Volumetric factors influencing carbon dioxide increases in the added reservoir (open-ended DS) were investigated. The maximum/minimum amount of CO2 concentration were obtained for the maximum/minimum device volume; 3.4 and 2.4 mol/m3 for the DS volumes of 1.2 and 0.5 × 10−3 m3, respectively. In all case studies, the CO2 buildup reached a plateau after approximately 20 breathing cycles. The experimental measurement results are in agreement with the simulation and numerical results obtained using the proposed simplified modeling technique, with a maximum relative error of 3.5%.

Title: Oxidative Stress in the Lung - The Essential Paradox

Authors: Rogers LK, Cismowski MJ.

Journal: Curr Opin Toxicol. 2018 Feb;7:37-43. doi: 10.1016/j.cotox.2017.09.001. Epub 2017 Sep 19. PMID: 29308441; PMCID: PMC5754020.

Link to full text: Oxidative Stress in the Lung - The Essential Paradox

Abstract: As eukaryotic life evolved, so too did the need for a source of energy that meets the requirements of complex organisms. Oxygen provides this vast potential energy source, but the same chemical reactivity which provides this potential also can have detrimental effects. The lung evolved as an organ that can efficiently promote gas exchange for the entire organism but as such, the lung is highly susceptible to its external environment. Oxygen can be transformed through both enzymatic and non-enzymatic processes into reactive oxygen species (ROS) and reactive nitrogen species (RNS), which can lead to protein, lipid, and DNA damage. Under normal conditions ROS/RNS concentrations are minimized through the activity of antioxidants located both intracellularly and in the epithelial lining fluid of the lung. Oxidative stress in the lung results when the antioxidant capacity is overwhelmed or depleted through external exposures, such as altered oxygen tension or air pollution, or internally. Internal sources of oxidative stress include systemic disease and the activation of resident cells and inflammatory cells recruited in response to an exposure or systemic response. Pulmonary responses to oxidative stress include activation of oxidases, lipid peroxidation, increases in nitric oxide, and autophagy. These internal and external exposures with the subsequent pulmonary responses contribute to development of diseases directly linked to oxidative stress. These include asthma, COPD, and lung cancers. While the vulnerability of the lung to oxidative stress is acknowledged, few effective preventative strategies or therapeutics are currently available.

Title: Oxidative stress and free radicals in COPD--implications and relevance for treatment

Authors: Domej W, Oettl K, Renner W.

Journal: Int J Chron Obstruct Pulmon Dis. 2014 Oct 17;9:1207-24. doi: 10.2147/COPD.S51226. PMID: 25378921; PMCID: PMC4207545.

Link to full text: Oxidative stress and free radicals in COPD--implications and relevance for treatment

Abstract: Oxidative stress occurs when free radicals and other reactive species overwhelm the availability of antioxidants. Reactive oxygen species (ROS), reactive nitrogen species, and their counterpart antioxidant agents are essential for physiological signaling and host defense, as well as for the evolution and persistence of inflammation. When their normal steady state is disturbed, imbalances between oxidants and antioxidants may provoke pathological reactions causing a range of nonrespiratory and respiratory diseases, particularly chronic obstructive pulmonary disease (COPD). In the respiratory system, ROS may be either exogenous from more or less inhalative gaseous or particulate agents such as air pollutants, cigarette smoke, ambient high-altitude hypoxia, and some occupational dusts, or endogenously generated in the context of defense mechanisms against such infectious pathogens as bacteria, viruses, or fungi. ROS may also damage body tissues depending on the amount and duration of exposure and may further act as triggers for enzymatically generated ROS released from respiratory, immune, and inflammatory cells. This paper focuses on the general relevance of free radicals for the development and progression of both COPD and pulmonary emphysema as well as novel perspectives on therapeutic options. Unfortunately, current treatment options do not suffice to prevent chronic airway inflammation and are not yet able to substantially alter the course of COPD. Effective therapeutic antioxidant measures are urgently needed to control and mitigate local as well as systemic oxygen bursts in COPD and other respiratory diseases. In addition to current therapeutic prospects and aspects of genomic medicine, trending research topics in COPD are presented. Keywords: antioxidants; chronic obstructive pulmonary disease; reactive nitrogen species; reactive oxygen species.

Title: Reactive oxygen species and mitochondria: A nexus of cellular homeostasis

Authors: Dan Dunn J, Alvarez LA, Zhang X, Soldati T.

Journal: Redox Biol. 2015 Dec;6:472-485. doi: 10.1016/j.redox.2015.09.005. Epub 2015 Sep 10. PMID: 26432659; PMCID: PMC4596921.

Link to full text: Reactive oxygen species and mitochondria: A nexus of cellular homeostasis

Abstract: Reactive oxygen species (ROS) are integral components of multiple cellular pathways even though excessive or inappropriately localized ROS damage cells. ROS function as anti-microbial effector molecules and as signaling molecules that regulate such processes as NF-kB transcriptional activity, the production of DNA-based neutrophil extracellular traps (NETs), and autophagy. The main sources of cellular ROS are mitochondria and NADPH oxidases (NOXs). In contrast to NOX-generated ROS, ROS produced in the mitochondria (mtROS) were initially considered to be unwanted by-products of oxidative metabolism. Increasing evidence indicates that mtROS have been incorporated into signaling pathways including those regulating immune responses and autophagy. As metabolic hubs, mitochondria facilitate crosstalk between the metabolic state of the cell with these pathways. Mitochondria and ROS are thus a nexus of multiple pathways that determine the response of cells to disruptions in cellular homeostasis such as infection, sterile damage, and metabolic imbalance. In this review, we discuss the roles of mitochondria in the generation of ROS-derived anti-microbial effectors, the interplay of mitochondria and ROS with autophagy and the formation of DNA extracellular traps, and activation of the NLRP3 inflammasome by ROS and mitochondria. Keywords: Autophagy; Immunity; Inflammasome; Mitochondria; Neutrophil extracellular traps; Reactive oxygen species.

Title: Reactive Oxygen Species in Metabolic and Inflammatory Signaling

Authors: Forrester SJ, Kikuchi DS, Hernandes MS, Xu Q, Griendling KK.

Journal: Circ Res. 2018 Mar 16;122(6):877-902. doi: 10.1161/CIRCRESAHA.117.311401. PMID: 29700084; PMCID: PMC5926825.

Link to full text: Reactive Oxygen Species in Metabolic and Inflammatory Signaling

Abstract: Reactive oxygen species (ROS) are well known for their role in mediating both physiological and pathophysiological signal transduction. Enzymes and subcellular compartments that typically produce ROS are associated with metabolic regulation, and diseases associated with metabolic dysfunction may be influenced by changes in redox balance. In this review, we summarize the current literature surrounding ROS and their role in metabolic and inflammatory regulation, focusing on ROS signal transduction and its relationship to disease progression. In particular, we examine ROS production in compartments such as the cytoplasm, mitochondria, peroxisome, and endoplasmic reticulum and discuss how ROS influence metabolic processes such as proteasome function, autophagy, and general inflammatory signaling. We also summarize and highlight the role of ROS in the regulation metabolic/inflammatory diseases including atherosclerosis, diabetes mellitus, and stroke. In order to develop therapies that target oxidative signaling, it is vital to understand the balance ROS signaling plays in both physiology and pathophysiology, and how manipulation of this balance and the identity of the ROS may influence cellular and tissue homeostasis. An increased understanding of specific sources of ROS production and an appreciation for how ROS influence cellular metabolism may help guide us in the effort to treat cardiovascular diseases. Keywords: cardiovascular diseases; inflammation; metabolism; oxidative stress; signal transduction.

Title: The Effects of Hyperbaric Oxygenation on Oxidative Stress, Inflammation and Angiogenesis

Authors: De Wolde SD, Hulskes RH, Weenink RP, Hollmann MW, Van Hulst RA.

Journal: Biomolecules. 2021 Aug 14;11(8):1210. doi: 10.3390/biom11081210. PMID: 34439876; PMCID: PMC8394403.

Link to full text: The Effects of Hyperbaric Oxygenation on Oxidative Stress, Inflammation and Angiogenesis

Abstract: Hyperbaric oxygen therapy (HBOT) is commonly used as treatment in several diseases, such as non-healing chronic wounds, late radiation injuries and carbon monoxide poisoning. Ongoing research into HBOT has shown that preconditioning for surgery is a potential new treatment application, which may reduce complication rates and hospital stay. In this review, the effect of HBOT on oxidative stress, inflammation and angiogenesis is investigated to better understand the potential mechanisms underlying preconditioning for surgery using HBOT. A systematic search was conducted to retrieve studies measuring markers of oxidative stress, inflammation, or angiogenesis in humans. Analysis of the included studies showed that HBOT-induced oxidative stress reduces the concentrations of pro-inflammatory acute phase proteins, interleukins and cytokines and increases growth factors and other pro-angiogenesis cytokines. Several articles only noted this surge after the first HBOT session or for a short duration after each session. The anti-inflammatory status following HBOT may be mediated by hyperoxia interfering with NF-κB and IκBα. Further research into the effect of HBOT on inflammation and angiogenesis is needed to determine the implications of these findings for clinical practice. Keywords: angiogenesis; hyperbaric oxygen therapy; hyperbaric oxygenation; inflammation; neovascularization; oxidative stress.

Title: The Role of Reactive-Oxygen-Species in Microbial Persistence and Inflammation

Authors: Spooner R, Yilmaz O.

Journal: Int J Mol Sci. 2011 Jan 13;12(1):334-52. doi: 10.3390/ijms12010334. PMID: 21339989; PMCID: PMC3039955.

Link to full text: The Role of Reactive-Oxygen-Species in Microbial Persistence and Inflammation

Abstract: The mechanisms of chronic infections caused by opportunistic pathogens are of keen interest to both researchers and health professionals globally. Typically, chronic infectious disease can be characterized by an elevation in immune response, a process that can often lead to further destruction. Reactive-Oxygen-Species (ROS) have been strongly implicated in the aforementioned detrimental response by host that results in self-damage. Unlike excessive ROS production resulting in robust cellular death typically induced by acute infection or inflammation, lower levels of ROS produced by host cells are increasingly recognized to play a critical physiological role for regulating a variety of homeostatic cellular functions including growth, apoptosis, immune response, and microbial colonization. Sources of cellular ROS stimulation can include "danger-signal-molecules" such as extracellular ATP (eATP) released by stressed, infected, or dying cells. Particularly, eATP-P2X(7) receptor mediated ROS production has been lately found to be a key modulator for controlling chronic infection and inflammation. There is growing evidence that persistent microbes can alter host cell ROS production and modulate eATP-induced ROS for maintaining long-term carriage. Though these processes have yet to be fully understood, exploring potential positive traits of these "injurious" molecules could illuminate how opportunistic pathogens maintain persistence through physiological regulation of ROS signaling. Keywords: NADPH oxidase; NLRX1; P2X7 receptor; Porphyromonas gingivalis; epithelium; extracellular ATP; microbial persistence; opportunistic pathogens; reactive-oxygen-species.

Title: Humming, Nitric Oxide and Paranasal Sinus Ventilation

Authors: Maniscalco, M.

Journal: Karolinska Institutet (Sweden), 2006.

Link to full text: Humming, Nitric Oxide and Paranasal Sinus Ventilation

Abstract: The paranasal sinuses are air-filled bony cavities surrounding the nose. They communicate with the nose via the sinus ostia through which fluid and gases pass in both directions. A proper ventilation is crucial for sinus integrity and blockage of the ostia is a major risk factor for development of sinusitis. In this thesis we have explored an entirely new approach to monitor sinus ventilation- the nasal humming test. We show in human studies in vivo and in a sinus/nasal model that the oscillating airflow generated during humming produce a dramatic increase in sinus ventilation. Interestingly, this increased gas exchange can be readily monitored on-line by simultaneously measuring the levels of the gas nitric oxide (NO) in nasally exhaled air. The sinuses constitute a major natural reservoir of NO and when gas-exchange increases during humming NO escapes rapidly into the nasal cavity thereby creating a highly reproducible peak in exhaled NO. When exploring the different factors that determine the humming peak in NO we found that sinus ostium size was the most important but the humming frequency also influenced the sinus NO release. In patients with severe nasal polyposis and completely blocked sinus ostia the humming peak in NO was abolished. Moreover, in patients with allergic rhinitis, absence of a NO peak was associated with endoscopic signs suggestive of ostial obstruction. In the last study we went on to study if an oscillating airflow could be used not only to wash a gas out from the sinuses but also to enhance passage of substances into the sinuses. Indeed, we found evidence of an increased intra-sinus drug deposition by adding a sounding airflow to an aerosol. In conclusion, the ventilation of the paranasal sinuses increases greatly when a person is humming; a finding that could have both diagnostic and therapeutic implications. Measurements of nasal NO during humming may represent a test of sinus ostialfunction. In addition, aerosol in combination with a sounding airflow could possibly be useful to increase the delivery of drugs into the paranasal sinuses.

Title: Exhaled nasal nitric oxide during humming: potential clinical tool in sinonasal disease?

Authors: Maniscalco M, Pelaia G, Sofia M.

Journal: Biomark Med. 2013 Apr;7(2):261-6. doi: 10.2217/bmm.13.11. PMID: 23547821.

Link to PubMed: Exhaled nasal nitric oxide during humming: potential clinical tool in sinonasal disease?

Abstract: The use of nasal nitric oxide (nNO) in sinonasal disease has recently been advocated as a potential tool to explore upper inflammatory airway disease. However, it is currently hampered by some factors including the wide range of measurement methods, the presence of various confounding factors and the heterogeneity of the study population. The contribution of nasal airway and paranasal sinuses communicating with the nose through the ostia represents the main confounding factor. There is accumulating evidence that nasal humming (which is the production of a tone without opening the lips or forming words) during nNO measurement increases nNO levels due to a rapid gas exchange in the paranasal sinuses. The aim of this review is to discuss the basic concepts and clinical applications of nNO assessment during humming, which represents a simple and noninvasive method to approach sinonasal disease.

Title: Strong humming for one hour daily to terminate chronic rhinosinusitis in four days: a case report and hypothesis for action by stimulation of endogenous nasal nitric oxide production

Authors: Eby GA.

Journal: Med Hypotheses. 2006;66(4):851-4. doi: 10.1016/j.mehy.2005.11.035. Epub 2006 Jan 10. PMID: 16406689.

Link to PubMed: Strong humming for one hour daily to terminate chronic rhinosinusitis in four days: a case report and hypothesis for action by stimulation of endogenous nasal nitric oxide production

Abstract: Rhinosinusitis is an inflammation or infection of the nose and air pockets (sinuses) above, below and between the eyes which connect with the back of the nose through tiny openings (ostia). Rhinosinusitis can be caused by bacteria, viruses, fungi (molds) and possibly by allergies. Chronic rhinosinusitis (CRS) is an immune disorder caused by fungi. The immune response produced by eosinophils causes the fungi to be attacked, which leads to damage of the sinus membranes, resulting in full-blown rhinosinusitis symptoms. Gaseous nitric oxide (NO) is naturally released in the human respiratory tract. The major part of NO found in exhaled air originates in the nasal airways, although significant production of NO also takes place in the paranasal sinuses. Proper ventilation is essential for maintenance of sinus integrity, and blockage of the ostium is a central event in pathogenesis of sinusitis. Concentrations of NO in the healthy sinuses are high. Nasal NO is known to be increased 15- to 20-fold by humming compared with quiet exhalation. NO is known to be broadly antifungal, antiviral and antibacterial. This case report shows that a subject hummed strongly at a low pitch ( approximately 130 Hz) for 1h (18 hums per minute) at bedtime the first night, and hummed 60-120 times 4 times a day for the following 4 days as treatment for severe CRS. The humming technique was described as being one that maximally increased intranasal vibrations, but less than that required to produce dizziness. The morning after the first 1-h humming session, the subject awoke with a clear nose and found himself breathing easily through his nose for the first time in over 1 month. During the following 4 days, CRS symptoms slightly reoccurred, but with much less intensity each day. By humming 60-120 times four times per day (with a session at bedtime), CRS symptoms were essentially eliminated in 4 days. Coincidentally, the subject's cardiac arrhythmias (PACs) were greatly lessened. It is hypothesized that strong, prolonged humming increased endogenous nasal NO production, thus eliminating CRS by antifungal means.

Title: The human ventilatory response to stress: rate or depth?

Authors: Tipton MJ, Harper A, Paton JFR, Costello JT.

Journal: J Physiol. 2017 Sep 1;595(17):5729-5752. doi: 10.1113/JP274596. Epub 2017 Jul 27. PMID: 28650070; PMCID: PMC5577533.

Link to full text: The human ventilatory response to stress: rate or depth?

Abstract: Many stressors cause an increase in ventilation in humans. This is predominantly reported as an increase in minute ventilation (V̇E). But, the same V̇E can be achieved by a wide variety of changes in the depth (tidal volume, VT ) and number of breaths (respiratory frequency, ƒR ). This review investigates the impact of stressors including: cold, heat, hypoxia, pain and panic on the contributions of ƒR and VT to V̇E to see if they differ with different stressors. Where possible we also consider the potential mechanisms that underpin the responses identified, and propose mechanisms by which differences in ƒR and VT are mediated. Our aim being to consider if there is an overall differential control of ƒR and VT that applies in a wide range of conditions. We consider moderating factors, including exercise, sex, intensity and duration of stimuli. For the stressors reviewed, as the stress becomes extreme V̇E generally becomes increased more by ƒR than VT . We also present some tentative evidence that the pattern of ƒR and VT could provide some useful diagnostic information for a variety of clinical conditions. In The Physiological Society's year of 'Making Sense of Stress', this review has wide-ranging implications that are not limited to one discipline, but are integrative and relevant for physiology, psychophysiology, neuroscience and pathophysiology.

Title: The physiological effects of slow breathing in the healthy human

Authors: Russo MA, Santarelli DM, O'Rourke D.

Journal: Breathe (Sheff). 2017 Dec;13(4):298-309. doi: 10.1183/20734735.009817. PMID: 29209423; PMCID: PMC5709795.

Link to full text: The physiological effects of slow breathing in the healthy human

Abstract: Slow breathing practices have been adopted in the modern world across the globe due to their claimed health benefits. This has piqued the interest of researchers and clinicians who have initiated investigations into the physiological (and psychological) effects of slow breathing techniques and attempted to uncover the underlying mechanisms. The aim of this article is to provide a comprehensive overview of normal respiratory physiology and the documented physiological effects of slow breathing techniques according to research in healthy humans. The review focuses on the physiological implications to the respiratory, cardiovascular, cardiorespiratory and autonomic nervous systems, with particular focus on diaphragm activity, ventilation efficiency, haemodynamics, heart rate variability, cardiorespiratory coupling, respiratory sinus arrhythmia and sympathovagal balance. The review ends with a brief discussion of the potential clinical implications of slow breathing techniques. This is a topic that warrants further research, understanding and discussion.

Title: Slow breathing for reducing stress: The effect of extending exhale

Authors: Birdee G, Nelson K, Wallston K, Nian H, Diedrich A, Paranjape S, Abraham R, Gamboa A.

Journal: Complement Ther Med. 2023 May;73:102937. doi: 10.1016/j.ctim.2023.102937. Epub 2023 Mar 5. PMID: 36871835; PMCID: PMC10395759.

Link to full text: Slow breathing for reducing stress: The effect of extending exhale

Abstract: Introduction: Slow breathing techniques are commonly used to reduce stress. While it is believed by mind-body practitioners that extending the exhale time relative to inhale increases relaxation, this has not been demonstrated.

Methods: We conducted a 12-week randomized, single-blinded trial among 100 participants to compare if yoga-based slow breathing with an exhale greater inhale versus an exhale equals inhale produces measurable differences in physiological and psychological stress among healthy adults.

Results: Participants mean individual instruction attendance was 10.7 ± 1.5 sessions out of 12 offered sessions. The mean weekly home practice was 4.8 ± 1.2 practices per week. There was no statistical difference between treatment groups for frequency of class attendance, home practice, or achieved slow breathing respiratory rate. Participants demonstrated fidelity to assigned breath ratios with home practice as measured by remote biometric assessments through smart garments (HEXOSKIN). Regular slow breathing practice for 12 weeks significantly reduced psychological stress as measured by PROMIS Anxiety (-4.85 S.D. ± 5.53, confidence interval [-5.60, -3.00], but not physiological stress as measured by heart rate variability. Group comparisons showed small effect size differences (d = 0.2) with further reductions in psychological stress and physiological stress from baseline to 12 weeks for exhale greater than inhale versus exhale equals inhale, however these differences were not statistically significant.

Conclusion: While slow breathing significantly reduces psychological stress, breath ratios do not have a significant differential effect on stress reduction among healthy adults.

Title: Respiratory alkalosis may impair the production of vitamin D and lead to significant morbidity, including the fibromyalgia syndrome

Authors: Lewis JM, Fontrier TH, Coley JL.

Journal: Med Hypotheses. 2017 May;102:99-101. doi: 10.1016/j.mehy.2017.03.013. Epub 2017 Mar 8. PMID: 28478843.

Link to PubMed: Respiratory alkalosis may impair the production of vitamin D and lead to significant morbidity, including the fibromyalgia syndrome

Abstract: Hyperventilation caused by physical and/or psychological stress may lead to significant respiratory alkalosis and an elevated systemic pH. The alkalotic pH may in turn suppress the normal renal release of phosphate into the urine, thereby interrupting the endogenous production of 1,25-dihydroxyvitamin D (calcitriol). This could cause a shortfall in its normal production, leading to a variety of adverse consequences. It might partially explain the pathogenesis of acute mountain sickness, a treatable disease characterized by severe hyperventilation secondary to the hypoxia of high altitude exposure. Milder degrees of hyperventilation due to different forms of stress may produce other conditions which share characteristics with acute mountain sickness. One of these may be the fibromyalgia syndrome, a chronic painful disorder for which no satisfactory treatment exists. Should fibromyalgia and acute mountain sickness have a common etiology, may they also share a common form of treatment? Evidence is presented to support this hypothesis.

Keywords: Acute mountain sickness; Fibromyalgia; Respiratory alkalosis; Vitamin D.

Title: Hyperventilation with Maintenance of Isocapnia. An "Old New" Method in Carbon Monoxide Intoxication

Authors: Sein Anand J, Schetz D, Waldman W, Wiśniewski M.

Journal: PLoS One. 2017 Jan 20;12(1):e0170621. doi: 10.1371/journal.pone.0170621. PMID: 28107437; PMCID: PMC5249161.

Link to full text: Hyperventilation with Maintenance of Isocapnia. An "Old New" Method in Carbon Monoxide Intoxication

Abstract: Introduction: Exposure to carbon monoxide (CO) is among the most common causes of acute and chronic poisonings worldwide. The crucial point of treatment of such acute poisonings is to eliminate CO from the body as fast as possible. There are currently two approaches to the management of the CO intoxication: hyperbaric oxygen therapy (HOT) and normobaric oxygen therapy (NOT). HOT is highly effective and capable of achieving the CO elimination half-time (T½) as low as 15 minutes. Unfortunately this method is expensive and not always readily available. The elimination of CO with the use of NOT (T½~70 min) is slower, but treatment can be started even on the site of the exposure and continued while the patient is transported to a hospital. The aim of the study was to evaluate the effectiveness of a method using therapeutic hyperventilation with maintenance of isocapnia (IH) in the elimination of CO in volunteers exposed to CO and to compare selected gasometric and respiratory parameters during IH with the values obtained during hyperventilation with pure oxygen ("non-isocapnic hyperventilation"-NIH).

Material and methods: The study involved 13 healthy, chronically-smoking volunteers. Each of them participated in two independent hyperventilation tests: IH and NIH. The levels of carboxyhemoglobin (COHb) and selected gasometric, cardiac and respiratory parameters were measured at 0, 10 and 20 minutes during both tests. Among 13 volunteers (8 women and 5 men) the initial COHb level was 5.0±1.5% (mean±SD) before the IH tests and 5.1±1.9% before the NIH tests (p>0.05). After 20 minutes of the procedures the mean COHb level was 2.9±0.9% for IH and 3.6±1.2% for NIH (p<0.01). The T½ of COHb was 29.6±12.2 min and 47.3±19.2 min respectively (p<0.01). After 10 minutes of NIH respiratory alkalosis was noted in 11 participants (84.6%). Such problem was not seen during the IH procedures. No serious adverse effects were recorded during either IH or NIH. Mild symptoms such as: dyspnea, headache and paresthesias were reported by 6 volunteers (46%) during both IH and NIH tests. It is worth noting that paresthesias were only reported during NIH, by 2 participants (15.4%).

Conclusions: The elimination T½ of CO during IH was comparable with the values reported during HOT, and lower than can be achieved with NOT or NIH. No serious adverse effects were reported during IH procedures. Further studies, especially direct comparisons with NOT and HOT, are necessary to evaluate the effectiveness of IH in the treatment of acute CO poisoning.

Title: Isocapnic hyperpnea accelerates carbon monoxide elimination

Authors: Fisher JA, Rucker J, Sommer LZ, Vesely A, Lavine E, Greenwald Y, Volgyesi G, Fedorko L, Iscoe S.

Journal: Am J Respir Crit Care Med. 1999 Apr;159(4 Pt 1):1289-92. doi: 10.1164/ajrccm.159.4.9804040. PMID: 10194179.

Link to full text: Isocapnic hyperpnea accelerates carbon monoxide elimination

Abstract: A major impediment to the use of hyperpnea in the treatment of CO poisoning is the development of hypocapnia or discomfort of CO2 inhalation. We examined the effect of nonrebreathing isocapnic hyperpnea on the rate of decrease of carboxyhemoglobin levels (COHb) in five pentobarbital-anesthetized ventilated dogs first exposed to CO and then ventilated with room air at normocapnia (control). They were then ventilated with 100% O2 at control ventilation, and at six times control ventilation without hypocapnia ("isocapnic hyperpnea") for at least 42 min at each ventilator setting. We measured blood gases and COHb. At control ventilation, the half-time for elimination of COHb (t1/2) was 212 +/- 17 min (mean +/- SD) on room air and 42 +/- 3 min on 100% O2. The t1/2 decreased to 18 +/- 2 min (p < 0.0005) during isocapnic hyperpnea. In two similarly prepared dogs treated with hyperbaric O2, the t1/2 were 20 and 28 min. We conclude that isocapnic hyperpnea more than doubles the rate of COHb elimination induced by normal ventilation with 100% O2. Isocapnic hyperpnea could improve the efficacy of the standard treatment of CO poisoning, 100% O2 at atmospheric or increased pressures.

Title: Accelerated ethanol elimination via the lungs

Authors: Klostranec JM, Vucevic D, Crawley AP, Venkatraghavan L, Sobczyk O, Duffin J, Sam K, Holmes R, Fedorko L, Mikulis DJ, Fisher JA.

Journal: Sci Rep. 2020 Nov 12;10(1):19249. doi: 10.1038/s41598-020-76233-9. PMID: 33184355; PMCID: PMC7665168.

Link to full text: Accelerated ethanol elimination via the lungs

Abstract: Ethanol poisoning is endemic the world over. Morbidity and mortality depend on blood ethanol levels which in turn depend on the balance between its rates of absorption and clearance. Clearance of ethanol is mostly at a constant rate via enzymatic metabolism. We hypothesized that isocapnic hyperpnea (IH), previously shown to be effective in acceleration of clearance of vapour anesthetics and carbon monoxide, would also accelerate the clearance of ethanol. In this proof-of-concept pilot study, five healthy male subjects were brought to a mildly elevated blood ethanol concentration (~ 0.1%) and ethanol clearance monitored during normal ventilation and IH on different days. IH increased elimination rate of ethanol in proportion to blood levels, increasing the elimination rate more than three-fold. Increased veno-arterial ethanol concentration differences during IH verified the efficacy of ethanol clearance via the lung. These data indicate that IH is a nonpharmacologic means to accelerate the elimination of ethanol by superimposing first order elimination kinetics on underlying zero order liver metabolism. Such kinetics may prove useful in treating acute severe ethanol intoxication.

Title: The treatment of acute alcoholism: with ten per cent carbon dioxide and ninety per cent oxygen inhalation

Authors: Robinson LJ, Selesnick S.

Journal: JAMA. 1935;105(22):1734–1738. doi:10.1001/jama.1935.02760480004002

Link to full text: The treatment of acute alcoholism: with ten per cent carbon dioxide and ninety per cent oxygen inhalation

Abstract: For many years the Fifth Medical Service of the Boston City Hospital has been in charge of almost all medical alcoholic patients admitted to this hospital. It is estimated that approximately 700 alcoholic patients are admitted to this service for treatment each year. The opportunity has been present, therefore, to study alcoholism in all its phases.
The acute alcoholic patient1 generally can be given a good prognosis as to recovery. For this reason, even when medical aid is sought for him, he receives little active therapy. In the paralytic stage of acute alcoholic poisoning, however, emergency treatment may be life -saving. Patients in this stage present a picture best described by Sollmann, who states that the symptoms are those of beginning medullary paralysis. Since carbon dioxide increased the respiratory excretion of alcohol, and since the oxidation of alcohol might be increased through the administration of oxygen, it is conceivable that the inhalation of a combination of 10 per cent carbon dioxide and 90 per cent oxygen mixture might accelerate recovery from acute alcoholic coma, especially of the paralytic state with respiratory depression and cyanosis. If it did no more than keep such a patient from dying of respiratory depression, it would, by keeping him alive, enable him to oxidize the alcohol himself. CONCLUSIONS: Acute alcoholic coma with dangerous respiratory depression, paralysis and cyanosis is a medical emergency. Death may be definitely prevented and recovery accelerated by inhalation of a mixture of 10 per cent carbon dioxide and 90 per cent oxygen for a length of time sufficient to reestablish and maintain normal respiration and color even after the inhalation is suspended.

Title: Carbon Dioixde in Anaesthesia

Authors: Wilson S.R.

Journal: BJA: British Journal of Anaesthesia, Volume 3, Issue 3, January 1926, Pages 112–118, https://doi.org/10.1093/bja/3.3.112

Link to full text: Carbon Dioixde in Anaesthesia

Abstract: All general anaesthetics are toxic agents in varying degrees and prone to produce undesirable after-effects on the patient. It is therefore advantageous to effect their elimination the moment their work is completed. Henderson and Haggard have investigated the rate of excretion of ether after general anaesthesia and provided us with accurate data on this subject. They found that half the total ether in the body was eliminated in the first half hour after the administration is stopped. Half the remainder is excreted during the next one to two hours, but complete elimination does not occur until one to two days. If it is possible to accelerate this process undesirable after-effects can be greatly reduced or completely avoided.
Now, if 5–6 per cent CO₂ be added to the oxygen pulmonary ventilation is increased two, three or four times. The writer has tested these experimental observations in actual clinical practice during the last two years with most gratifying results, both to himself and the patients concerned.

Title: Isocapnic hyperventilation provides early extubation after head and neck surgery: A prospective randomized trial.

Authors: Hallén K, Jildenstål P, Stenqvist O, Oras J, Ricksten SE, Lindgren S.

Journal: Acta Anaesthesiol Scand. 2018 Sep;62(8):1064-1071. doi: 10.1111/aas.13133. Epub 2018 Apr 19. PMID: 29671866.

Link to PubMed: Isocapnic hyperventilation provides early extubation after head and neck surgery: A prospective randomized trial.

Abstract: Background: Isocapnic hyperventilation (IHV) shortens recovery time after inhalation anaesthesia by increasing ventilation while maintaining a normal airway carbon dioxide (CO2)-level. One way of performing IHV is to infuse CO2 to the inspiratory limb of a breathing circuit during mechanical hyperventilation (HV). In a prospective randomized study, we compared this IHV technique to a standard emergence procedure (control).

Methods: Thirty-one adult ASA I-III patients undergoing long-duration (>3 hours) sevoflurane anaesthesia for major head and neck surgery were included and randomized to IHV-treatment (n = 16) or control (n = 15). IHV was performed at minute ventilation 13.6 ± 4.3 L/min and CO2 delivery, dosed according to a nomogram tested in a pilot study. Time to extubation and eye-opening was recorded. Inspired (FICO2) and expired (FETCO2) CO2 and arterial CO2 levels (PaCO2) were monitored. Cognition was tested preoperatively and at 20, 40 and 60 minutes after surgery.

Results: Time from turning off the vapourizer to extubation was 13.7 ± 2.5 minutes in the IHV group and 27.4 ± 6.5 minutes in controls (P < .001). Two minutes after extubation, PaCO2 was 6.2 ± 0.5 and 6.2 ± 0.6 kPa in the IHV and control group respectively. In 69% (IHV) vs 53% (controls), post-operative cognition returned to pre-operative values within 40 minutes after surgery (NS). Incidences of pain and nausea/vomiting did not differ between groups.

Conclusions: In this randomized trial comparing an IHV method with a standard weaning procedure, time to extubation was reduced with 50% in the IHV group. The described IHV method can be used to decrease emergence time from inhalation anaesthesia.

Title: Isocapnic hyperventilation shortens washout time for sevoflurane - an experimental in vivo study

Authors: Hallén K, Stenqvist O, Ricksten SE, Lindgren S.

Journal: Acta Anaesthesiol Scand. 2016 Oct;60(9):1261-9. doi: 10.1111/aas.12761. Epub 2016 Jul 10. PMID: 27396945.

Link to PubMed: Isocapnic hyperventilation shortens washout time for sevoflurane - an experimental in vivo study

Abstract: Background: Isocapnic hyperventilation (IHV) is a method that fastens weaning from inhalation anaesthesia by increasing airway concentration of carbon dioxide (CO2 ) during hyperventilation (HV). In an animal model, we evaluated a technique of adding CO2 directly to the breathing circuit of a standard anaesthesia apparatus.

Methods: Eight anaesthetised pigs weighing 28 ± 2 kg were intubated and mechanically ventilated. From a baseline ventilation of 5 l/min, HV was achieved by doubling minute volume and fresh gas flow. Respiratory rate was increased from 15 to 22/min. The CO2 absorber was disconnected and CO2 was delivered (DCO2 ) to the inspiratory limb of a standard breathing circuit via a mixing box. Time required to decrease end-tidal sevoflurane concentration from 2.7% to 0.2% was defined as washout time. Respiration and haemodynamics were monitored by blood gas analysis, spirometry, electric impedance tomography and pulse contour analysis.

Results: A DCO2 of 261 ± 19 ml/min was necessary to achieve isocapnia during HV. The corresponding FICO2 -level remained stable at 3.1 ± 0.3%. During IHV, washout of sevoflurane was three times faster, 433 ± 135 s vs. 1387 ± 204 s (P < 0.001). Arterial CO2 tension and end-tidal CO2 , was 5.2 ± 0.4 kPa and 5.6 ± 0.4%, respectively, before IHV and 5.1 ± 0.3 kPa and 5.7 ± 0.3%, respectively, during IHV.

Conclusions: In this experimental in vivo model of isocapnic hyperventilation, the washout time of sevoflurane anaesthesia was one-third compared to normal ventilation. The method for isocapnic hyperventilation described can potentially be transferred to a clinical setting with the intention to decrease emergence time from inhalation anaesthesia.

Title: A simple method for isocapnic hyperventilation evaluated in a lung model

Authors: Hallén K, Stenqvist O, Ricksten SE, Lindgren S.

Journal: Acta Anaesthesiol Scand. 2016 May;60(5):597-606. doi: 10.1111/aas.12674. Epub 2015 Dec 21. PMID: 26688296.

Link to PubMed: A simple method for isocapnic hyperventilation evaluated in a lung model

Abstract: Background: Isocapnic hyperventilation (IHV) has the potential to increase the elimination rate of anaesthetic gases and has been shown to shorten time to wake-up and post-operative recovery time after inhalation anaesthesia. In this bench test, we describe a technique to achieve isocapnia during hyperventilation (HV) by adding carbon dioxide (CO2) directly to the breathing circuit of a standard anaesthesia apparatus with standard monitoring equipment.

Methods: Into a mechanical lung model, carbon dioxide was added to simulate a CO2 production (V(CO2)) of 175, 200 and 225 ml/min. Dead space (V(D)) volume could be set at 44, 92 and 134 ml. From baseline ventilation (BLV), HV was achieved by doubling the minute ventilation and fresh gas flow for each level of V(CO2), and dead space. During HV, CO2 was delivered (D(CO2)) by a precision flow meter via a mixing box to the inspiratory limb of the anaesthesia circuit to achieve isocapnia.

Results: During HV, the alveolar ventilation increased by 113 ± 6%. Tidal volume increased by 20 ± 0.1% during IHV irrespective of V(D) and V(CO2) level. D(CO2) varied between 147 ± 8 and 325 ± 13 ml/min. Low V(CO2) and large V(D) demanded a greater D(CO2) administration to achieve isocapnia. The FICO2 level during IHV varied between 2.3% and 3.3%.

Conclusion: It is possible to maintain isocapnia during HV by delivering carbon dioxide through a standard anaesthesia circuit equipped with modern monitoring capacities. From alveolar ventilation, CO2 production and dead space, the amount of carbon dioxide that is needed to achieve IHV can be estimated.

Title: Evaluation of a method for isocapnic hyperventilation: a clinical pilot trial

Authors: Hallén K, Jildenstål P, Stenqvist O, Ricksten SE, Lindgren S.

Journal: Acta Anaesthesiol Scand. 2018 Feb;62(2):186-195. doi: 10.1111/aas.13008. Epub 2017 Oct 16. PMID: 29034967.

Link to PubMed: Evaluation of a method for isocapnic hyperventilation: a clinical pilot trial

Abstract: Background: Isocapnic hyperventilation (IHV) is a method that shortens time to extubation after inhalation anaesthesia using hyperventilation (HV) without lowering airway CO2 . In a clinical trial on patients undergoing long-duration sevoflurane anaesthesia for major ear-nose-throat (ENT) surgery, we evaluated the utility of a technique for CO2 delivery (DCO2 ) to the inspiratory limb of a closed breathing circuit, during HV, to achieve isocapnia.

Methods: Fifteen adult ASA 1-3 patients were included. After end of surgery, mechanical HV was started by doubling baseline minute ventilation. Simultaneously, CO2 was delivered and dosed using a nomogram developed in a previous experimental study. Time to extubation and eye opening was recorded. Inspired (FICO2 ) and expired (FETCO2 ) CO2 and arterial CO2 levels were monitored during IHV. Cognition was tested pre-operatively and at 20, 40 and 60 min after surgery.

Results: A DCO2 of 285 ± 45 ml/min provided stable isocapnia during HV (13.5 ± 4.1 l/min). The corresponding FICO2 level was 3.0 ± 0.3%. Time from turning off the vaporizer (1.3 ± 0.1 MACage) to extubation (0.2 ± 0.1 MACage) was 11.3 ± 1.8 min after 342 ± 131 min of anaesthesia. PaCO2 and FETCO2 remained at normal levels during and after IHV. In 85% of the patients, post-operative cognition returned to pre-operative values within 60 min.

Conclusions: In this cohort of patients, a DCO2 nomogram for IHV was validated. The patients were safely extubated shortly after discontinuing long-term sevoflurane anaesthesia. Perioperatively, there were no adverse effects on arterial blood gases or post-operative cognition. This technique for IHV can potentially be used to decrease emergence time from inhalation anaesthesia.

Title: Impact of breathing patterns on the quality of life of 9- to 10-year-old schoolchildren

Authors: Leal RB, Gomes MC, Granville-Garcia AF, Goes PS, de Menezes VA.

Journal: Am J Rhinol Allergy. 2016 Sep;30(5):147-52. doi: 10.2500/ajra.2016.30.4363. PMID: 27657891.

Link to PubMed: Impact of breathing patterns on the quality of life of 9- to 10-year-old schoolchildren

Abstract: Background: Mouth breathing can cause a set of changes in craniofacial growth and development, with esthetic, functional, and psychological repercussions.

Objective: To determine the impact of mouth breathing on the quality of life of schoolchildren.

Methods: A school-based, cross-sectional study was conducted with 1911 children ages 9 and 10 years in the city of Recife, Brazil. The children answered the Mouth Breather Quality of Life questionnaire and a questionnaire that addressed sociodemographic data and health-related aspects. Clinical examinations were performed by an examiner who had undergone a training and calibration process for the diagnosis of mouth breathing (kappa = 0.90). Descriptive statistics were conducted to characterize the sample. Statistical analysis involved the Student's t-test and the F test (analysis of variance) (alpha = 5%).

Results: The prevalence of mouth breathing was 54.81%. Children with oral breathing demonstrated a poorer quality of life in comparison with children with nasal breathing (p < 0.001). The following variables were significantly associated with a poorer quality of life among the children with mouth breathing: a younger age (p < 0.001) and the use of medication (p = 0.002).

Conclusion: Based on the present findings, children with the mouth-breathing pattern experience a greater negative impact on quality of life in comparison with those with the nose-breathing pattern. Thus, the early diagnosis and treatment of this clinical condition are fundamental to minimizing the consequences of mouth breathing on the quality of life of schoolchildren with respiration disorders.

Title: Structural and physiological age-associated changes in aging lungs

Authors: Miller MR.

Journal: Semin Respir Crit Care Med. 2010 Oct;31(5):521-7. doi: 10.1055/s-0030-1265893. Epub 2010 Oct 12. PMID: 20941653.

Link to PubMed: Structural and physiological age-associated changes in aging lungs

Abstract: The tissues of the lungs undergo changes with age that lead to an increase in alveolar size without any destruction of alveolar walls. This lowers the alveolar surface tension and so reduces the elastic recoil of the lungs, which in turn causes a reduction in maximum achievable flow in the airways during the breathing cycle. Muscle performance diminishes with age and the chest wall becomes stiffer, which together lead to an increased residual volume and also counter the possible increase in total lung capacity (TLC) from the reduced elastic recoil. Exercise performance falls with age, with a small decrease in arterial oxygenation that stabilizes over the age of 70 years. Predicting lung function for the elderly is not easy, and basing clinical decisions on percent of predicted may lead to age, sex, and height bias. Focusing on what function remains can allow survival predictions to guide decisions on treatment options.

Title: Relationship between breath-hold time and physical performance in patients with cystic fibrosis

Authors: Barnai M, Laki I, Gyurkovits K, Angyan L, Horvath G.

Journal: Eur J Appl Physiol. 2005 Oct;95(2-3):172-8. doi: 10.1007/s00421-005-1350-3. Epub 2005 Jul 9. PMID: 16007450.

Link to PubMed: Relationship between breath-hold time and physical performance in patients with cystic fibrosis

Abstract: Rehabilitation including physiotherapy is an important part of the treatment used to help improve the quality of life of patients with cystic fibrosis (CF). The aim of this study was to determine the value of the breath-hold time as an index of exercise tolerance in patients with CF. Eighteen patients in different states of CF were included. The breath-hold time was measured in all patients. The fitness level was assessed by means of a progressive exercise test on a cycle-ergometer. During the test, oxygen uptake (VO(2)) and carbon dioxide elimination (VCO(2)) were measured breath by breath. The VO(2) and working capacity (WC) were computed at the anaerobic threshold (AT) and at peak. Duration of breath-hold was 24.7 +/- 2.87 (mean +/- SEM) seconds, varying between 10 and 58. The breath-hold time (BHT) displayed a significant correlation with VO(2) (r = 0.898), WC (r = 0.899) at the AT, and the peak VO(2) (r = 0.895). Regression equations were: VO(2) at the AT (ml/kg) = 5.53 + 0.42 x BHT and WC at the AT (watt/kg) = 0.56 + 0.38 x BHT Our results suggest that the voluntary breath-hold time might be a useful index for prediction of the exercise tolerance of CF patients.

Title: Physiology, Pulmonary Circulatory System

Authors: Jain V, Bordes SJ, Bhardwaj A.

Journal: StatPearls [Internet]. Treasure Island (FL): StatPearls Publishing; 2025 Jan-.

Link to PubMed: Physiology, Pulmonary Circulatory System

Abstract: Pulmonary circulation includes a vast network of arteries, veins, and lymphatics that function to exchange blood and other tissue fluids between the heart, the lungs, and back. They are designed to perform certain specific functions that are unique to the pulmonary circulation, such as ventilation and gas exchange. The pulmonary circulation receives the entirety of the cardiac output from the right heart and is a low pressure, low resistance system due to its parallel capillary circulation. The system can be divided into the following components:

The arterial circuit arises from the main pulmonary artery arising from the right ventricle and runs a course of only 5 cm before dividing into right and left main branches and many subsequent branches to form an extensive network of small arteries, arterioles, and capillaries. The pulmonary arteries are thinner (one-third the thickness of their counterpart systemic vessels) and have a larger diameter. The combined effect makes them much more distensible and compliant (approximately 7mL/mmHg).
The venous circuit begins with the venules that drain the capillaries. They join to form smaller veins and eventually merge to form the main pulmonary veins draining into the left atrium. Like the arteries, the pulmonary veins are thinner and more distensible than the counterpart systemic veins and accommodate more blood because of their larger compliance.
Lymphatics play a crucial role in maintaining a dry alveolar membrane and preventing accumulation of tissue fluid around the pulmonary circulation. They can be found close to the terminal bronchioles and drain the mediastinal lymphatics before emptying into the right lymphatic duct.

Title: Physiology, Pulmonary Ventilation and Perfusion

Authors: Powers KA, Dhamoon AS.

Journal: StatPearls [Internet]. Treasure Island (FL): StatPearls Publishing; 2025 Jan-.

Link to PubMed: Physiology, Pulmonary Ventilation and Perfusion

Abstract: One of the major roles of the lungs is to facilitate gas exchange between the circulatory system and the external environment. The lungs are composed of branching airways that terminate in respiratory bronchioles and alveoli, which participate in gas exchange. Most bronchioles and large airways are part of the conducting zone of the lung, which delivers gas to sites of gas exchange in alveoli. Gas exchange occurs in the lungs between alveolar air and the blood of the pulmonary capillaries. For effective gas exchange to occur, alveoli must be ventilated and perfused. Ventilation (V) refers to the flow of air into and out of the alveoli, while perfusion (Q) refers to the flow of blood to alveolar capillaries. Individual alveoli have variable degrees of ventilation and perfusion in different regions of the lungs. Collective changes in ventilation and perfusion in the lungs are measured clinically using the ratio of ventilation to perfusion (V/Q). Changes in the V/Q ratio can affect gas exchange and can contribute to hypoxemia.

Title: Dynamic CO2 therapy in periodic breathing: a modeling study to determine optimal timing and dosage regimes

Authors: Mebrate Y, Willson K, Manisty CH, Baruah R, Mayet J, Hughes AD, Parker KH, Francis DP.

Journal: J Appl Physiol (1985). 2009 Sep;107(3):696-706. doi: 10.1152/japplphysiol.90308.2008. Epub 2009 Jul 23. PMID: 19628721; PMCID: PMC2755997.

Link to full text: Dynamic CO2 therapy in periodic breathing: a modeling study to determine optimal timing and dosage regimes

Abstract: We examine the potential to treat unstable ventilatory control (seen in periodic breathing, Cheyne-Stokes respiration, and central sleep apnea) with carefully controlled dynamic administration of supplementary CO(2), aiming to reduce ventilatory oscillations with minimum increment in mean CO(2). We used a standard mathematical model to explore the consequences of phasic CO(2) administration, with different timing and dosing algorithms. We found an optimal time window within the ventilation cycle (covering approximately 1/6 of the cycle) during which CO(2) delivery reduces ventilatory fluctuations by >95%. Outside that time, therapy is dramatically less effective: indeed, for more than two-thirds of the cycle, therapy increases ventilatory fluctuations >30%. Efficiency of stabilizing ventilation improved when the algorithm gave a graded increase in CO(2) dose (by controlling its duration or concentration) for more severe periodic breathing. Combining gradations of duration and concentration further increased efficiency of therapy by 22%. The (undesirable) increment in mean end-tidal CO(2) caused was 300 times smaller with dynamic therapy than with static therapy, to achieve the same degree of ventilatory stabilization (0.0005 vs. 0.1710 kPa). The increase in average ventilation was also much smaller with dynamic than static therapy (0.005 vs. 2.015 l/min). We conclude that, if administered dynamically, dramatically smaller quantities of CO(2) could be used to reduce periodic breathing, with minimal adverse effects. Algorithms adjusting both duration and concentration in real time would achieve this most efficiently. If developed clinically as a therapy for periodic breathing, this would minimize excess acidosis, hyperventilation, and sympathetic overactivation, compared with static treatment.

Title: The effects of carbon dioxide in champagne on psychometric performance and blood-alcohol concentration

Authors: Ridout F, Gould S, Nunes C, Hindmarch I.

Journal: Alcohol Alcohol. 2003 Jul-Aug;38(4):381-5. doi: 10.1093/alcalc/agg092. PMID: 12814909.

Link to PubMed: The effects of carbon dioxide in champagne on psychometric performance and blood-alcohol concentration

Abstract: Aims: To assess the effects of carbon dioxide (CO(2)) in champagne on psychomotor performance and blood-alcohol concentration (BAC).

Methods: Twelve subjects consumed ethanol (0.6 g/kg body weight) served as champagne or champagne with the CO(2) removed, in a crossover study.

Results: Champagne produced significantly greater BACs and significantly increased reaction times in a divided attention task, than degassed champagne.

Conclusions: The CO(2) in champagne may accelerate absorption of alcohol, leading to more rapid or severe intoxication.

Title: Alcohol concentration and carbonation of drinks: the effect on blood alcohol levels

Authors: Roberts C, Robinson SP.

Journal: J Forensic Leg Med. 2007 Oct;14(7):398-405. doi: 10.1016/j.jflm.2006.12.010. Epub 2007 May 16. PMID: 17720590.

Link to PubMed: Alcohol concentration and carbonation of drinks: the effect on blood alcohol levels

Abstract: Alcohol absorption and elimination vary considerably amongst individuals, and are subject to influences from a variety of factors. The effects of alcohol concentration and beverage mixer type on the rate of alcohol absorption, in a controlled environment was studied. 21 subjects (12 male, 9 female) consumed a solution containing alcohol, on three separate occasions. The three solutions were, A: Neat vodka (37.5 vol%), B: Vodka mixed with still water (18.75 vol%), C: Vodka mixed with carbonated water (18.75 vol%). The volume of alcohol each subject consumed was determined by Widmark's equation. The alcohol was drunk in a 5 min period following an overnight fast and breath alcohol concentrations were measured over a 4h period using a breathalyser. 20/21 subjects absorbed the dilute alcohol at a faster rate than the concentrated alcohol. The difference between the absorption rates was found to be significant (p<0.001). The use of a carbonated mixer had varying effects on the alcohol absorption rate. 14/21 subjects absorbed the alcohol with the carbonated mixer at a faster rate, with 7 subjects showing either no change or a decrease in rate. The mean absorption rate for solution C was 4.39+/-0.45 (mg/100ml/min), and the difference between this absorption rate and that with the still mixer (1.08+0.36) was significant (p=0.006).

Title: Intraoperative End-Tidal Carbon Dioxide Concentrations: What Is the Target?

Authors: Way M, Hill GE.

Journal: Anesthesiol Res Pract. 2011;2011:271539. doi: 10.1155/2011/271539. Epub 2011 Oct 25. PMID: 22110496; PMCID: PMC3202118.

Link to full text: Intraoperative End-Tidal Carbon Dioxide Concentrations: What Is the Target?

Abstract: Recent publications suggest that target end-tidal carbon dioxide concentrations should be higher than values currently considered as acceptable. This paper presents evidence that end-tidal carbon dioxide values higher than concentrations that are currently targeted result in improved patient outcomes and are associated with a reduced incidence of postoperative complications.

Title: Association between Mouth Breathing and Atopic Dermatitis in Japanese Children 2-6 years Old: A Population-Based Cross-Sectional Study

Authors: Yamaguchi H, Tada S, Nakanishi Y, Kawaminami S, Shin T, Tabata R, Yuasa S, Shimizu N, Kohno M, Tsuchiya A, Tani K.

Journal: PLoS One. 2015 Apr 27;10(4):e0125916. doi: 10.1371/journal.pone.0125916. PMID: 25915864; PMCID: PMC4411141.

Link to full text: Association between Mouth Breathing and Atopic Dermatitis in Japanese Children 2-6 years Old: A Population-Based Cross-Sectional Study

Abstract: As mouth breathing is associated with asthma and otitis media, it may be associated with other diseases. Therefore, this population-based cross-sectional study evaluated the association of mouth breathing with the prevalences of various diseases in children. Preschool children older than 2 years were included. A questionnaire was given to parents/guardians at 13 nurseries in Tokushima City. There were 468 valid responses (45.2%). We defined a subject as a mouth breather in daytime (MBD) if they had 2 or more positive items among the 3 following items: "breathes with mouth ordinarily," "mouth is open ordinarily," and "mouth is open when chewing." We defined subjects as mouth breathers during sleep (MBS) if they had 2 or more positive items among the following 3 items: "snoring," "mouth is open during sleeping," and "mouth is dry when your child gets up." The prevalences of MBD and MBS were 35.5% and 45.9%, respectively. There were significant associations between MBD and atopic dermatitis (odds ratio [OR]: 2.4, 95% confidence interval [CI]: 1.4-4.2), MBS and atopic dermatitis (OR: 2.4, 95% CI: 1.3-4.2), and MBD and asthma (OR: 2.2, 95% CI: 1.2-4.0). After adjusting for history of asthma and allergic rhinitis; family history of atopic dermatitis, asthma, and allergic rhinitis; and nasal congestion; both MBD (OR: 2.6, 95% CI: 1.3-5.4) and MBS (OR: 4.1, 95% CI: 1.8-9.2) were significantly associated with atopic dermatitis. In preschool children older than 2 years, both MBD and MBS may be associated with the onset or development of atopic dermatitis.

Title: Carbon dioxide homeostasis and recovery after general anaesthesia

Authors: Hovorka J.

Journal: Acta Anaesthesiol Scand. 1982 Oct;26(5):498-504. doi: 10.1111/j.1399-6576.1982.tb01806.x. PMID: 6815976.

Link to PubMed: Carbon dioxide homeostasis and recovery after general anaesthesia

Abstract: The influence of different levels of carbon dioxide during general anaesthesia on postoperative recovery was studied. Sixty female patients were divided into two age groups. Thirty patients were over 60 years old and 30 patients were under 46 years old. Artificial ventilation with added carbon dioxide during general anaesthesia with thiopentone, nitrous oxide-oxygen, alcuronium and fentanyl was used. According to the arterial CO2 tension, patients were allocated to subgroups: hypercarbic, PaCO2 7.3 kPa, normocarbic, PaCO2 5.3 kPa and two different levels of hypocarbia: older patients PaCO2 3.7 kPa and younger patients PaCO2 2.9 kPa. As criteria for recovery, a battery of recovery tests and postoperative questionnaires were used. Regardless of age, patients subjected to hypercarbic ventilation scored better in the recovery tests than patients subjected to normo- or hypocarbia. Normocarbic ventilation also gave better results than hypocarbic ventilation. The level of hypocarbia used in the older patients and that used in the younger patients, though different, resulted in nearly the same deterioration of scoring in the recovery tests. This deterioration was seen in some patients up to 48 h postoperatively. No subjective differences were elicited from the questionnaires after various types of ventilation.

Title: Carbon dioxide tolerability and toxicity in rat and man: A translational study

Authors: van der Schrier R, van Velzen M, Roozekrans M, Sarton E, Olofsen E, Niesters M, Smulders C, Dahan A.

Journal: Front Toxicol. 2022 Oct 13;4:1001709. doi: 10.3389/ftox.2022.1001709. PMID: 36310693; PMCID: PMC9606673.

Link to full text: Carbon dioxide tolerability and toxicity in rat and man: A translational study

Abstract: Background: Due the increasing need for storage of carbon dioxide (CO2) more individuals are prone to be exposed to high concentrations of CO2 accidentally released into atmosphere, with deleterious consequences. Methods: We tested the effect of increasing CO2 concentrations in humans (6-12%) and rats (10-50%) at varying inhalation times (10-60 min). In humans, a continuous positive airway pressure helmet was used to deliver the gas mixture to the participants. Unrestrained rats were exposed to CO2 in a transparent chamber. In both species regular arterial blood gas samples were obtained. After the studies, the lungs of the animals were examined for macroscopic and microscopic abnormalities. Results: In humans, CO2 concentrations of 9% inhaled for >10 min, and higher concentrations inhaled for <10 min were poorly or not tolerated due to exhaustion, anxiety, dissociation or acidosis (pH < 7.2), despite intact oxygenation. In rats, concentrations of 30% and higher were associated with CO2 narcosis, epilepsy, poor oxygenation and, at 50% CO2, spontaneous death. Lung hemorrhage and edema were observed in the rats at inhaled concentrations of 30% and higher. Conclusion: This study provides essential insight into the occurrence of physiological changes in humans and fatalities in rats after acute exposure to high levels of CO2. Humans tolerate 9% CO2 and retain their ability to function coherently for up to 10 min. These data support reconsideration of the current CO2 levels (<7.5%) that pose a risk to exposed individuals (<7.5%) as determined by governmental agencies to ≤9%.

Keywords: CO2 storage; CO2 transport; carbon dioxide; tolerability; toxicity; translational study.

Title: Hyperventilation in panic disorder and asthma: empirical evidence and clinical strategies

Authors: Meuret AE, Ritz T.

Journal: Int J Psychophysiol. 2010 Oct;78(1):68-79. doi: 10.1016/j.ijpsycho.2010.05.006. Epub 2010 May 25. PMID: 20685222; PMCID: PMC2937087.

Link to full text: Hyperventilation in panic disorder and asthma: empirical evidence and clinical strategies

Abstract: Sustained or spontaneous hyperventilation has been associated with a variety of physical symptoms and has been linked to a number of organic illnesses and mental disorders. Theories of panic disorder hold that hyperventilation either produces feared symptoms of hypocapnia or protects against feared suffocation symptoms of hypercapnia. Although the evidence for both theories is inconclusive, findings from observational, experimental, and therapeutic studies suggest an important role of low carbon dioxide (CO2) levels in this disorder. Similarly, hypocapnia and associated hyperpnia are linked to bronchoconstriction, symptom exacerbation, and lower quality of life in patients with asthma. Raising CO2 levels by means of therapeutic capnometry has proven beneficial effects in both disorders, and the reversing of hyperventilation has emerged as a potent mediator for reductions in panic symptom severity and treatment success.

Title: Association of lung function with overall mortality is independent of inflammatory, cardiac, and functional biomarkers in older adults: the ActiFE-study

Authors: Weinmayr, G., Schulz, H., Klenk, J., Denkinger, M., Duran-Tauleria, E., Koenig, W., ... & ActiFE Study Group Böhm B. 12 13 Geiger H. 14 Laszlo R. 15 Steinacker JM 15 Ludolph A. 16 von Arnim C. 16 Lukas A. 17 Peter R. 18 Rapp K. 4 Riepe M. 19 Scharffetter-Kochanek K. 20 Stingl J. 21.

Journal: Scientific reports 10.1 (2020): 11862.

Link to full text: Association of lung function with overall mortality is independent of inflammatory, cardiac, and functional biomarkers in older adults: the ActiFE-study

Abstract: Reduced lung function is associated with overall and cardiovascular mortality. Chronic low grade systemic inflammation is linked to impaired lung function and cardiovascular outcomes.We assessed the association of lung function with overall 8-year mortality in 867 individuals of theActivity and Function in the Elderly study using confounder-adjusted Cox proportional hazards models (including gait speed and daily walking time as measures of physical function) without and with adjustment for inflammatory and cardiac markers. Forced expiratory volume in 1 s/forced vital capacity ­(FEV1/FVC) but not FVC was related to mortality after adjustment for physical function and other confounders. Additional adjustment for inflammatory and cardiac markers did not change the hazard ratios (HR) markedly, e.g. for a ­FEV1/FVC below 0.7 from 1.55 [95% confdence-interval (CI) 1.14–2.11] to 1.49 (95% CI 1.09–2.03).These independent associations were also observed in the apparently lung healthy subpopulation with even higher HRs up to 2.76 (95% CI 1.52–4.93).A measure of airfow limitation but not vital capacity was associated with overall mortality in this community-dwelling older population and in the subgroup classifed as lung healthy.These associations were independent of adjustment for inflammatory and cardiac markers and support the role of airfow limitation as independent predictor of mortality in older adults.

Title: The effect of pursed lips breathing in increasing oxygen saturation in patients with chronic obstructive pulmonary disease in internal ward 2 of the general hospital of dr. R. Soedarsono pasuruan

Authors: Budiono, B., Mustayah, M., & Aindrianingsih, A.

Journal: Public Health of Indonesia 3.3 (2017): 117-123.

Link to full text: The effect of pursed lips breathing in increasing oxygen saturation in patients with chronic obstructive pulmonary disease in internal ward 2 of the general hospital of dr. R. Soedarsono pasuruan

Abstract: Background: Chronic obstructive pulmonary disease is a leading cause of death and disability. Thus, the effort to reduce the symptoms, such as dyspnea is necessity. Pursed lip breathing is assumed increasing oxygen saturation.
Objective: This study aims to determine the effect of pursed lips breathing in increasing oxygen saturation in patients with COPD in internal ward 2 of the General Hospital of Dr. R. Soedarsono Pasuruan on May 16 - June 30, 2017.
Methods: This was a pre-experimental design with pre-posttest design. There were 24 patients with COPD selected using simple random sampling technique. Pulse oximetry (oximeter pulse fingertip) was used to measure oxygen saturation. Data were analyzed using Wilcoxon Sign Rank Test.
Results: Findings showed p-value 0.000 (<0.05), which indicated that there was a statistically significant difference in oxygen saturation before and after pursed lips breathing.
Conclusion: There was a significant effect of pursed lips breathing on oxygen saturation in patients with COPD in the internal ward 2 of the general hospital of Dr. R. Soedarsono Pasuruan. Thus, pursed lips breathing could be applied as a nursing intervention in patients with COPD.
Key words: Pursed lips breathing, COPD, oxygen saturation

Title: A comparison between the airway response to isocapnic hyperventilation and hypertonic saline in subjects with asthma

Authors: Smith CM, Anderson SD.

Journal: Eur Respir J. 1989 Jan;2(1):36-43. PMID: 2495982.

Link to PubMed: A comparison between the airway response to isocapnic hyperventilation and hypertonic saline in subjects with asthma

Abstract: We compared the response to isocapnic hyperventilation (ISH), where both cooling and drying of the mucosa occur, with the response to inhaling aerosols of hypertonic saline (HS), where airway osmolarity increases without airway cooling. We studied nine subjects on two days. For ISH, subjects ventilated at 70% of their estimated maximum voluntary ventilation (MVV). For HS, they inhaled aerosols of 2.7, 3.6, or 4.5% saline. The concentration that was used depended on the rate of ventilation during ISH. For both challenges the stimulus was given for one minute. Forced expiratory volume in one second (FEV1) was measured once between each minute, and the challenge ceased when the FEV1 did not change for two successive minutes. A plateau in FEV1 occurred after 8.1 +/- 2.4 (mean +/- 1 SD) min of ISH, and 8.3 +/- 2.4 min of HS. The lowest FEV1 (% predicted) after ISH was 45 +/- 16% and after HS was 51 +/- 18% (r = 0.93). However, the maximum responses occurred after the final challenge and were not the same as the plateau. For HS, the plateau represented 89 +/- 11% of the maximum response which developed within one minute of the final challenge. For ISH, the plateau was only 56 +/- 26% of the maximum response, which developed within 5.2 +/- 2.9 min after challenge. The similarities in the response to these challenges are consistent with the hypothesis that ISH induces asthma via hyperosmolarity. The delayed response to ISH suggests that cooling may delay the response to hyperosmolarity.

Title: Respiratory Abnormalities in Parkinson's Disease: What Do We Know from Studies in Humans and Animal Models?

Authors: Kaczyńska K, Orłowska ME, Andrzejewski K.

Journal: Int J Mol Sci. 2022 Mar 23;23(7):3499. doi: 10.3390/ijms23073499. PMID: 35408858; PMCID: PMC8998219.

Link to full text: Respiratory Abnormalities in Parkinson's Disease: What Do We Know from Studies in Humans and Animal Models?

Abstract: Parkinson's disease (PD) is the second most common progressive neurodegenerative disease characterized by movement disorders due to the progressive loss of dopaminergic neurons in the ventrolateral region of the substantia nigra pars compacta (SNpc). Apart from the cardinal motor symptoms such as rigidity and bradykinesia, non-motor symptoms including those associated with respiratory dysfunction are of increasing interest. Not only can they impair the patients' quality of life but they also can cause aspiration pneumonia, which is the leading cause of death among PD patients. This narrative review attempts to summarize the existing literature on respiratory impairments reported in human studies, as well as what is newly known from studies in animal models of the disease. Discussed are not only respiratory muscle dysfunction, apnea, and dyspnea, but also altered central respiratory control, responses to hypercapnia and hypoxia, and how they are affected by the pharmacological treatment of PD.

Keywords: Parkinson's disease; apnea; dyspnea; hypercapnia; hypoxia; respiratory dysfunction.

Title: Respiratory alkalosis in children with febrile seizures

Authors: Schuchmann S, Hauck S, Henning S, Grüters-Kieslich A, Vanhatalo S, Schmitz D, Kaila K.

Journal: Epilepsia. 2011 Nov;52(11):1949-55. doi: 10.1111/j.1528-1167.2011.03259.x. Epub 2011 Sep 12. PMID: 21910730.

Link to full text: Respiratory alkalosis in children with febrile seizures

Abstract: Purpose: Febrile seizures (FS) are the most common type of convulsive events in children. FS are suggested to result from a combination of genetic and environmental factors. However, the pathophysiologic mechanisms underlying FS remain unclear. Using an animal model of experimental FS, it was demonstrated that hyperthermia causes respiratory alkalosis with consequent brain alkalosis and seizures. Here we examine the acid-base status of children who were admitted to the hospital for FS. Children who were admitted because of gastroenteritis (GE), a condition known to promote acidosis, were examined to investigate a possible protective effect of acidosis against FS.

Methods: We enrolled 433 age-matched children with similar levels of fever from two groups presented to the emergency department. One group was admitted for FS (n = 213) and the other for GE (n = 220). In the FS group, the etiology of fever was respiratory tract infection (74.2%), otitis media (7%), GE (7%), tonsillitis (4.2%), scarlet fever (2.3%) chickenpox (1.4%), urinary tract infection (1.4%), postvaccination reaction (0.9%), or unidentified (1.4%). In all patients, capillary pH and blood Pco(2) were measured immediately on admission to the hospital.

Key findings: Respiratory alkalosis was found in children with FS (pH 7.46 ± 0.04, [mean ± standard deviation] Pco(2) 29.5 ± 5.5 mmHg), whereas a metabolic acidosis was seen in all children admitted for GE (pH 7.31 ± 0.03, Pco(2) 37.7 ± 4.3 mmHg; p < 0.001 for both parameters). No FS were observed in the latter group. A subgroup (n = 15; 7%) of the patients with FS had GE and, notably, their blood pH was more alkaline (pH 7.44 ± 0.04) than in the GE-admitted group. During the enrollment period, eight of the patients were admitted on separate occasions because of FS or GE. Consistent with the view that generation of FS requires a genetic susceptibility in addition to acute seizure triggering factors, each of these patients had an alkalotic blood pH when admitted because of FS, whereas they had an acidotic pH (and no FS) when admitted because of GE (pH 7.47 ± 0.05 vs. pH 7.33 ± 0.03, p < 0.005).

Significance: The results show that FS are associated with a systemic respiratory alkalosis, irrespective of the severity of the underlying infection as indicated by the level of fever. The lack of FS in GE patients is attributable to low pH, which also explains the fact that children with a susceptibility to FS do not have seizures when they have GE-induced fever that is associated with acidosis. The present demonstration of a close link between FS and respiratory alkalosis may pave the way for further clinical studies and attempts to design novel therapies for the treatment of FS by controlling the systemic acid-base status.

Title: Respiratory Dysfunction in Alzheimer's Disease-Consequence or Underlying Cause? Applying Animal Models to the Study of Respiratory Malfunctions

Authors: Wrzesień A, Andrzejewski K, Jampolska M, Kaczyńska K.

Journal: Int J Mol Sci. 2024 Feb 16;25(4):2327. doi: 10.3390/ijms25042327. PMID: 38397004; PMCID: PMC10888758.

Link to full text: Respiratory Dysfunction in Alzheimer's Disease-Consequence or Underlying Cause? Applying Animal Models to the Study of Respiratory Malfunctions

Abstract: Alzheimer's disease (AD) is a neurodegenerative brain disease that is the most common cause of dementia among the elderly. In addition to dementia, which is the loss of cognitive function, including thinking, remembering, and reasoning, and behavioral abilities, AD patients also experience respiratory disturbances. The most common respiratory problems observed in AD patients are pneumonia, shortness of breath, respiratory muscle weakness, and obstructive sleep apnea (OSA). The latter is considered an outcome of Alzheimer's disease and is suggested to be a causative factor. While this narrative review addresses the bidirectional relationship between obstructive sleep apnea and Alzheimer's disease and reports on existing studies describing the most common respiratory disorders found in patients with Alzheimer's disease, its main purpose is to review all currently available studies using animal models of Alzheimer's disease to study respiratory impairments. These studies on animal models of AD are few in number but are crucial for establishing mechanisms, causation, implementing potential therapies for respiratory disorders, and ultimately applying these findings to clinical practice. This review summarizes what is already known in the context of research on respiratory disorders in animal models, while pointing out directions for future research.

Keywords: Alzheimer's disease; animal models; hypercapnia; hypoxia; respiratory disorders; sleep obstructive apnea.

Title: Respiratory dysfunction in Parkinson's disease: a narrative review

Authors: D'Arrigo A, Floro S, Bartesaghi F, Casellato C, Sferrazza Papa GF, Centanni S, Priori A, Bocci T.

Journal: ERJ Open Res. 2020 Oct 5;6(4):00165-2020. doi: 10.1183/23120541.00165-2020. PMID: 33043046; PMCID: PMC7533305.

Link to full text: Respiratory dysfunction in Parkinson's disease: a narrative review

Abstract: The presence of respiratory symptoms in Parkinson's disease (PD) has been known since the first description of the disease, even though the prevalence and incidence of these disturbances are not well defined. Several causes have been reported, comprising obstructive and restrictive pulmonary disease and changes in the central ventilatory control, and different pathogenetic mechanisms have been postulated accordingly. In our review, we encompass the current knowledge about respiratory abnormalities in PD, as well as the impact of anti-Parkinsonian drugs as either risk or protective factors. A description of putative pathogenetic mechanisms is also provided, and possible treatments are discussed, focusing on the importance of recognising and treating respiratory symptoms as a key manifestation of the disease itself. A brief description of respiratory dysfunctions in atypical Parkinsonism, especially α-synucleinopathies, is also provided.

Title: Acetazolamide: a second wind for a respiratory stimulant in the intensive care unit?

Authors: Heming N, Urien S, Faisy C.

Journal: Crit Care. 2012 Aug 7;16(4):318. doi: 10.1186/cc11323. PMID: 22866939; PMCID: PMC3580678.

Link to full text: Acetazolamide: a second wind for a respiratory stimulant in the intensive care unit?

Abstract: Patients with chronic obstructive pulmonary disease (COPD) are affected by episodes of respiratory exacerbations, some of which can be severe and may necessitate respiratory support. Prolonged invasive mechanical ventilation is associated with increased mortality rates. Persistent failure to discontinue invasive mechanical ventilation is a major issue in patients with COPD. Pure or mixed metabolic alkalosis is a common finding in the intensive care unit (ICU) and is associated with a worse outcome. In patients with COPD, the condition is called post-hypercapnic alkalosis and is a complication of mechanical ventilation. Reversal of metabolic alkalosis may facilitate weaning from mechanical ventilation of patients with COPD. Acetazolamide, a non-specific carbonic anhydrase inhibitor, is one of the drugs employed in the ICU to reverse metabolic alkalosis. The drug is relatively safe, undesirable effects being rare. The compartmentalization of the different isoforms of the carbonic anhydrase enzyme may, in part, explain the lack of evidence of the efficacy of acetazolamide as a respiratory stimulant. Recent findings suggest that the usually employed doses of acetazolamide in the ICU may be insufficient to significantly improve respiratory parameters in mechanically ventilated patients with COPD. Randomized controlled trials using adequate doses of acetazolamide are required to address this issue.

Title: Carbon dioxide in the critically ill: too much or too little of a good thing?

Authors: Marhong J, Fan E.

Journal: Respir Care. 2014 Oct;59(10):1597-605. doi: 10.4187/respcare.03405. PMID: 25261559.

Link to full text: Carbon dioxide in the critically ill: too much or too little of a good thing?

Abstract: Hypercapnia and hypocapnia commonly complicate conditions that are present in critically ill patients. Both conditions have important physiologic effects that may impact the clinical management of these patients. For instance, hypercapnia results in bronchodilation and enhanced hypoxic vasoconstriction, leading to improved ventilation/perfusion matching. Hypocapnia reduces cerebral blood volume through arterial vasoconstriction. These effects have also been exploited for therapeutic aims. In patients with traumatic brain injury (TBI), hypocapnia is often utilized to control intracranial pressure. However, this effect is not sustained, and prolonged hypocapnia increases the risk of mortality and severe disability in patients with TBI. Hypercapnia and hypercapnic acidosis are common consequences of lung-protective ventilation in ARDS. Hypercapnic acidosis reduces ischemic lung injury and preserves lung compliance, but concern has arisen over hypercapnia-induced immunosuppression and the potential for bacterial proliferation in sepsis. Experimental studies suggest that buffering hypercapnic acidosis attenuates these effects, whereas hypocapnia appears to potentiate lung injury through increased capillary permeability and decreased lung compliance. Several areas of uncertainty surround the role of hypercapnia/hypocapnia in treating TBI and ARDS. Current data support recommendations to avoid hypocapnia in treating TBI, with the exception of emergent treatment of elevated intracranial pressure, while awaiting definitive management. Permissive hypercapnia is commonly accepted as a consequence of lung-protective ventilation in ARDS, but there is insufficient evidence to support the induction of hypercapnic acidosis in clinical practice. Buffering hypercapnic acidosis should be considered only for a specific clinical indication (eg, hemodynamic instability). For clinicians choosing to buffer hypercapnic acidosis, tris-hydroxymethyl aminomethane is recommended over sodium bicarbonate, as it is more effective in correcting pH and is not associated with increased carbon dioxide production. Future studies should aim to address these areas of uncertainty to help guide clinicians in the therapeutic use and management of hypercapnia/hypocapnia in critically ill patients.

Keywords: hypercapnia; hypocapnia; intensive care units; respiration, artificial; respiratory acidosis; respiratory distress syndrome, adult.

Title: Gaseous nitric oxide failed to inhibit the replication cycle of SARS-CoV-2 in vitro

Authors: Rousseaud A, Prot M, Loriere ES, Katz I, Ramirez-Gil JF, Farjot G.

Journal: Nitric Oxide. 2023 Mar 1;132:27-33. doi: 10.1016/j.niox.2023.01.004. Epub 2023 Jan 25. PMID: 36706864; PMCID: PMC9873364.

Link to full text: Gaseous nitric oxide failed to inhibit the replication cycle of SARS-CoV-2 in vitro

Abstract: Nitric oxide (NO) has been shown to have antimicrobial activity in vitro and in some in vivo models, while the virucidal activity of NO remains elusive. Some studies using NO donors have suggested that NO could be a potential candidate to treat SARS-CoV infection. The Covid-19 pandemic raised the hypothesis that NO gas might have an impact on Sars-CoV-2 replication cycle and might be considered as a candidate therapy to treat COVID-19. To our knowledge, there are no in vitro preclinical studies demonstrating a virucidal effect of gaseous NO on SARS-CoV-2. This study aims to determine whether gaseous NO has an impact on the replication cycle of SARS-CoV-2 in vitro. To that end, SARS-CoV-2 infected epithelial (VeroE6) and pulmonary (A549-hACE2) cells were treated with repeated doses of gaseous NO at different concentrations known to be efficient against bacteria. Our results show that exposing SARS-CoV-2 infected-cells to NO gas even at high doses (160 ppm, 6 h) does not influence the replication cycle of the virus in vitro. We report here that NO gas has no antiviral properties in vitro on SARS-COV-2. Therefore, there is no rationale for its usage in clinical settings to treat COVID-19 patients for direct antiviral purposes, which does not exclude other potential physiological benefits of this gas.

Keywords: COVID-19; Gaseous nitric oxide; SARS-CoV-2.

Title: Oxidative Stress in the Lung - The Essential Paradox

Authors: Rogers LK, Cismowski MJ.

Journal: Curr Opin Toxicol. 2018 Feb;7:37-43. doi: 10.1016/j.cotox.2017.09.001. Epub 2017 Sep 19. PMID: 29308441; PMCID: PMC5754020.

Link to full text: Oxidative Stress in the Lung - The Essential Paradox

Abstract: As eukaryotic life evolved, so too did the need for a source of energy that meets the requirements of complex organisms. Oxygen provides this vast potential energy source, but the same chemical reactivity which provides this potential also can have detrimental effects. The lung evolved as an organ that can efficiently promote gas exchange for the entire organism but as such, the lung is highly susceptible to its external environment. Oxygen can be transformed through both enzymatic and non-enzymatic processes into reactive oxygen species (ROS) and reactive nitrogen species (RNS), which can lead to protein, lipid, and DNA damage. Under normal conditions ROS/RNS concentrations are minimized through the activity of antioxidants located both intracellularly and in the epithelial lining fluid of the lung. Oxidative stress in the lung results when the antioxidant capacity is overwhelmed or depleted through external exposures, such as altered oxygen tension or air pollution, or internally. Internal sources of oxidative stress include systemic disease and the activation of resident cells and inflammatory cells recruited in response to an exposure or systemic response. Pulmonary responses to oxidative stress include activation of oxidases, lipid peroxidation, increases in nitric oxide, and autophagy. These internal and external exposures with the subsequent pulmonary responses contribute to development of diseases directly linked to oxidative stress. These include asthma, COPD, and lung cancers. While the vulnerability of the lung to oxidative stress is acknowledged, few effective preventative strategies or therapeutics are currently available.

Title: Topical cutaneous application of carbon dioxide via a hydrogel for improved fracture repair: results of phase I clinical safety trial

Authors: Niikura T, Iwakura T, Omori T, Lee SY, Sakai Y, Akisue T, Oe K, Fukui T, Matsushita T, Matsumoto T, Kuroda R.

Journal: BMC Musculoskelet Disord. 2019 Nov 25;20(1):563. doi: 10.1186/s12891-019-2911-7. PMID: 31766994; PMCID: PMC6878668.

Link to full text: Topical cutaneous application of carbon dioxide via a hydrogel for improved fracture repair: results of phase I clinical safety trial

Abstract: Background: Clinicians have very limited options to improve fracture repair. Therefore, it is critical to develop a new clinically available therapeutic option to assist fracture repair biologically. We previously reported that the topical cutaneous application of carbon dioxide (CO2) via a CO2 absorption-enhancing hydrogel accelerates fracture repair in rats by increasing blood flow and angiogenesis and promoting endochondral ossification. The aim of this study was to assess the safety and efficacy of CO2 therapy in patients with fractures.

Methods: Patients with fractures of the femur and tibia were prospectively enrolled into this study with ethical approval and informed consent. The CO2 absorption-enhancing hydrogel was applied to the fractured lower limbs of patients, and then 100% CO2 was administered daily into a sealed space for 20 min over 4 weeks postoperatively. Safety was assessed based on vital signs, blood parameters, adverse events, and arterial and expired gas analyses. As the efficacy outcome, blood flow at the level of the fracture site and at a site 5 cm from the fracture in the affected limb was measured using a laser Doppler blood flow meter.

Results: Nineteen patients were subjected to complete analysis. No adverse events were observed. Arterial and expired gas analyses revealed no adverse systemic effects including hypercapnia. The mean ratio of blood flow 20 min after CO2 therapy compared with the pre-treatment level increased by approximately 2-fold in a time-dependent manner.

Conclusions: The findings of the present study revealed that CO2 therapy is safe to apply to human patients and that it can enhance blood flow in the fractured limbs.

Title: Transcutaneous carbon dioxide application accelerates muscle injury repair in rat models

Authors: Akahane S, Sakai Y, Ueha T, Nishimoto H, Inoue M, Niikura T, Kuroda R.

Journal: Int Orthop. 2017 May;41(5):1007-1015. doi: 10.1007/s00264-017-3417-2. Epub 2017 Feb 16. PMID: 28210805.

Link to PubMed: Transcutaneous carbon dioxide application accelerates muscle injury repair in rat models

Abstract: Purpose: Skeletal muscle injuries are commonly observed in sports and traumatology medicine. Previously, we demonstrated that transcutaneous application of carbon dioxide (CO2) to lower limbs increased the number of muscle mitochondria and promoted muscle endurance. Therefore, we aimed to investigate whether transcutaneous CO2 application could enhance recovery from muscle injury.

Methods: Tibialis anterior muscle damage was induced in 27 Sprague Dawley rats via intramuscular injection of bupivacaine. After muscle injury, rats were randomly assigned to transcutaneous CO2-treated or -untreated groups. From each group, three rats were sacrificed at weeks one, two, four and six. At each time point, histology and immunofluorescence analyses were performed, and changes in muscle weight, muscle weight/body weight ratio, muscle fibre circumference, gene expression levels and capillary density were measured.

Results: Injured muscle fibres were completely repaired at week six in the CO2-treated group but only partially repaired in the untreated group. The repair of basement and plasma membranes did not differ significantly between groups. However, expression levels of genes and proteins related to muscle protein synthesis were significantly higher in the CO2-treated group and significantly more capillaries four weeks after injury.

Conclusion: Transcutaneous CO2 application can accelerate recovery after muscle injury in rats.

Title: Transcutaneous carbon dioxide application suppresses bone destruction caused by breast cancer metastasis

Authors: Takemori T, Kawamoto T, Ueha T, Toda M, Morishita M, Kamata E, Fukase N, Hara H, Fujiwara S, Niikura T, Kuroda R, Akisue T.

Journal: Oncology Reports, 40, 2079-2087. https://doi.org/10.3892/or.2018.6608

Link to full text: Transcutaneous carbon dioxide application suppresses bone destruction caused by breast cancer metastasis

Abstract: Hypoxia plays a significant role in cancer progression, including metastatic bone tumors. We previously reported that transcutaneous carbon dioxide (CO2) application could decrease tumor progression through the improvement of intratumor hypoxia. Therefore, we hypothesized that decreased hypoxia using transcutaneous CO2 could suppress progressive bone destruction in cancer metastasis. In the present study, we examined the effects of transcutaneous CO2 application on metastatic bone destruction using an animal model. The human breast cancer cell line MDA-MB-231 was cultured in vitro under three different oxygen conditions, and the effect of altered oxygen conditions on the expression of osteoclast-differentiation and osteolytic factors was assessed. An in vivo bone metastatic model of human breast cancer was created by intramedullary implantation of MDA-MB-231 cells into the tibia of nude mice, and treatment with 100% CO2 or a control was performed twice weekly for two weeks. Bone volume of the treated tibia was evaluated by micro-computed tomography (µCT), and following treatment, histological evaluation was performed by hematoxylin and eosin staining and immunohistochemical staining for hypoxia-inducible factor (HIF)-1α, osteoclast-differentiation and osteolytic factors, and tartrate-resistant acid phosphatase (TRAP) staining for osteoclast activity. In vitro experiments revealed that the mRNA expression of RANKL, PTHrP and IL-8 was significantly increased under hypoxic conditions and was subsequently reduced by reoxygenation. In vivo results by µCT revealed that bone destruction was suppressed by transcutaneous CO2, and that the expression of osteoclast-differentiation and osteolytic factors, as well as HIF-1α, was decreased in CO2-treated tumor tissues. In addition, multinucleated TRAP-positive osteoclasts were significantly decreased in CO2-treated tumor tissues. Hypoxic conditions promoted bone destruction in breast cancer metastasis, and reversal of hypoxia by transcutaneous CO2 application significantly inhibited metastatic bone destruction along with decreased osteoclast activity. The findings in this study strongly indicated that transcutaneous CO2 application could be a novel therapeutic strategy for treating metastatic bone destruction.

Title: Effects of the duration of transcutaneous CO2 application on the facilitatory effect in rat fracture repair

Authors: Oda T, Iwakura T, Fukui T, Oe K, Mifune Y, Hayashi S, Matsumoto T, Matsushita T, Kawamoto T, Sakai Y, Akisue T, Kuroda R, Niikura T.

Journal: J Orthop Sci. 2020 Sep;25(5):886-891. doi: 10.1016/j.jos.2019.09.017. Epub 2019 Oct 18. PMID: 31635930.

Link to full text: Effects of the duration of transcutaneous CO2 application on the facilitatory effect in rat fracture repair

Abstract: Background: Carbon dioxide therapy has been reported to be effective in treating certain cardiac diseases and skin problems. Although a previous study suggested that transcutaneous carbon dioxide application accelerated fracture repair in association with promotion of angiogenesis, blood flow, and endochondral ossification, the influence of the duration of carbon dioxide application on fracture repair is unknown. The aim of this study was to investigate the effect of the duration of transcutaneous carbon dioxide application on rat fracture repair.

Methods: A closed femoral shaft fracture was created in each rat. Animals were randomly divided into four groups: the control group; 1w-CO2 group, postoperative carbon dioxide treatment for 1 week; 2w-CO2 group, postoperative carbon dioxide treatment for 2 weeks; 3w-CO2 group, postoperative carbon dioxide treatment for 3 weeks. Transcutaneous carbon dioxide application was performed five times a week in the carbon dioxide groups. Sham treatment, where the carbon dioxide was replaced with air, was performed for the control group. Radiographic, histological, and biomechanical assessments were performed at 3 weeks after fracture.

Results: The fracture union rate was significantly higher in the 3w-CO2 group than in the control group (p < 0.05). Histological assessment revealed promotion of endochondral ossification in the 3w-CO2 group than in the control group. In the biomechanical assessment, all evaluation items related to bone strength were significantly higher in the 3w-CO2 group than in the control group (p < 0.05).

Conclusions: The present study, conducted using an animal model, demonstrated that continuous carbon dioxide application throughout the process of fracture repair was effective in enhancing fracture healing.

Title: Topical cutaneous application of CO2 accelerates bone healing in a rat femoral defect model

Authors: Kuroiwa Y, Fukui T, Takahara S, Lee SY, Oe K, Arakura M, Kumabe Y, Oda T, Matsumoto T, Matsushita T, Akisue T, Sakai Y, Kuroda R, Niikura T.

Journal: BMC Musculoskelet Disord. 2019 May 22;20(1):237. doi: 10.1186/s12891-019-2601-5. PMID: 31113412; PMCID: PMC6530028.

Link to full text: Topical cutaneous application of CO2 accelerates bone healing in a rat femoral defect model

Abstract: Background: Bone defects may occur because of severe trauma, nonunion, infection, or tumor resection. However, treatments for bone defects are often difficult and have not been fully established yet. We previously designed an efficient system of topical cutaneous application of carbon dioxide (CO2) using a novel hydrogel, which facilitates CO2 absorption through the skin into the deep area within a limb. In this study, the effect of topical cutaneous application of CO2 on bone healing was investigated using a rat femoral defect model.

Methods: In this basic research study, an in vivo bone defect model, fixed with an external fixator, was created using a rat femur. The affected limb was shaved, and CO2 was applied for 20 min/day, 5 days/week. In the control animals, CO2 gas was replaced with air. Radiographic, histological, biomechanical, and genetic assessments were performed to evaluate bone healing.

Results: Radiographically, bone healing rate was significantly higher in the CO2 group than in the control group at 4 weeks (18.2% vs. 72.7%). The degree of bone healing scored using the histopathological Allen grading system was significantly higher in the CO2 group than in the control group at 2 weeks (1.389 ± 0.334 vs. 1.944 ± 0.375). The ultimate stress, extrinsic stiffness, and failure energy were significantly greater in the CO2 group than in the control group at 4 weeks (3.2 ± 0.8% vs. 38.1 ± 4.8%, 0.6 ± 0.3% vs. 41.5 ± 12.2%, 2.6 ± 0.8% vs. 24.7 ± 5.9%, respectively.). The volumetric bone mineral density of the callus in micro-computed tomography analysis was significantly higher in the CO2 group than in the control group at 4 weeks (180.9 ± 43.0 mg/cm3 vs. 247.9 ± 49.9 mg/cm3). Gene expression of vascular endothelial growth factor in the CO2 group was significantly greater than that in the control group at 3 weeks (0.617 ± 0.240 vs. 2.213 ± 0.387).

Conclusions: Topical cutaneous application of CO2 accelerated bone healing in a rat femoral defect model. CO2 application can be a novel and useful therapy for accelerating bone healing in bone defects; further research on its efficacy in humans is warranted.

Keywords: Bone defect; Bone healing; CO2.

Title: Transcutaneous carbon dioxide application with hydrogel prevents muscle atrophy in a rat sciatic nerve crush model

Authors: Nishimoto H, Inui A, Ueha T, Inoue M, Akahane S, Harada R, Mifune Y, Kokubu T, Nishida K, Kuroda R, Sakai Y.

Journal: J Orthop Res. 2018 Jun;36(6):1653-1658. doi: 10.1002/jor.23817. Epub 2017 Dec 19. PMID: 29193246.

Link to full text: Transcutaneous carbon dioxide application with hydrogel prevents muscle atrophy in a rat sciatic nerve crush model

Abstract: The acceleration of nerve regeneration remains a clinical challenge. We previously demonstrated that transcutaneous CO2 application using a novel hydrogel increases the oxygen concentration in local tissue via an "artificial Bohr effect" with the potential to prevent muscle atrophy. In this study, we investigated the effect of transcutaneous CO2 administration on limb function after peripheral nerve injury in a rat sciatic nerve injury model. In total, 73 Sprague-Dawley rats were divided into a sham group, a control group (crush injury to sciatic nerve and no treatment) or a CO2 group (crush injury with transcutaneous CO2 application). CO2 was administered percutaneously for 20 min five times per week. Scores for the sciatic function index and pinprick test were significantly higher in the CO2 group than control group. The muscle wet weight ratios of the tibialis anterior and soleus muscles were higher in the CO2 group than control group. Electrophysiological examination showed that the CO2 group had higher compound motor action potential amplitudes and shorter distal motor latency than the control group. Histological examination of the soleus muscle sections at postoperative week 2 showed shorter fiber diameter in the control group than in the CO2 group. The mRNA expression of Atrogin-1 and MuRF-1 was lower, mRNA expression of VEGF and myogenin and MyoD was higher in CO2 group at postoperative week 2 compared to the control group.

Clinical significance: Transcutaneous CO2 application has the therapeutic potential to accelerate the recovery of muscle atrophy in peripheral nerve injury. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 36:1653-1658, 2018.

Keywords: Bohr effect; muscle atrophy; peripheral nerve injury; transcutaneous carbon dioxide application.

Title: Carbon Dioxide Water Bathing Enhances Myogenin but Not MyoD Protein Expression after Skeletal Muscle Injury

Authors: Nonaka K, Akiyama J, Tatsuta N, Une S, Ito K, Ogaya S, Kataoka M, Iwata A, Okuda K.

Journal: J Phys Ther Sci. 2013 Jun;25(6):709-11. doi: 10.1589/jpts.25.709. Epub 2013 Jul 23. PMID: 24259835; PMCID: PMC3805014.

Link to full text: Carbon Dioxide Water Bathing Enhances Myogenin but Not MyoD Protein Expression after Skeletal Muscle Injury

Abstract: [Purpose] We reported that carbon dioxide (CO2) water bathing accelerates skeletal muscle regeneration; however, the underlying mechanism was unclear. MyoD and myogenin play roles in muscle regeneration, and the purpose of this study was to determine changes in MyoD and myogenin caused by CO2 water bathing after injury. [Subjects] Sixteen female Wistar rats (n = 4 per group) were used. [Methods] The rats were divided into four groups: no-injury (NI), injury (IC), injury + tap water bathing (ITW), and injury + CO2 water bathing (ICO2). Muscle injury was induced by injection of bupivacaine hydrochloride into the left tibial anterior (TA) muscles. Tap water and CO2 (1,000 ppm) water bathing were performed at 37 °C for 30 minutes once a day. The left TA muscles were removed 4 days after injury, and the expressions of MyoD and myogenin were measured. [Results] MyoD and myogenin were increased in the IC, ITW, and ICO2 groups compared with the NI group. Although the MyoD level was similar in the IC, ITW, and ICO2 groups, myogenin increased more in the ICO2 group than in the IC and ITW groups. [Conclusion] CO2 water bathing after muscle injury appears to induce an increase in the expression of myogenin.

Keywords: CO2 water bathing; Myogenic regulatory factors; Skeletal muscle injury.

Title: Local application of a transcutaneous carbon dioxide paste prevents excessive scarring and promotes muscle regeneration in a bupivacaine-induced rat model of muscle injury

Authors: Hirota J, Hasegawa T, Inui A, Takeda D, Amano-Iga R, Yatagai N, Saito I, Arimoto S, Akashi M.

Journal: Int Wound J. 2023 Apr;20(4):1151-1159. doi: 10.1111/iwj.13974. Epub 2022 Oct 17. PMID: 36250918; PMCID: PMC10031219.

Link to full text: Local application of a transcutaneous carbon dioxide paste prevents excessive scarring and promotes muscle regeneration in a bupivacaine-induced rat model of muscle injury

Abstract: In postoperative patients with head and neck cancer, scar tissue formation may interfere with the healing process, resulting in incomplete functional recovery and a reduced quality of life. Percutaneous application of carbon dioxide (CO2 ) has been reported to improve hypoxia, stimulate angiogenesis, and promote fracture repair and muscle damage. However, gaseous CO2 cannot be applied to the head and neck regions. Previously, we developed a paste that holds non-gaseous CO2 in a carrier and can be administered transdermally. Here, we investigated whether this paste could prevent excessive scarring and promote muscle regeneration using a bupivacaine-induced rat model of muscle injury. Forty-eight Sprague Dawley rats were randomly assigned to either a control group or a CO2 group. Both groups underwent surgery to induce muscle injury, but the control group received no treatment, whereas the CO2 group received the CO2 paste daily after surgery. Then, samples of the experimental sites were taken on days 3, 7, 14, and 21 post-surgery to examine the following: (1) inflammatory (interleukin [IL]-1β, IL-6), and transforming growth factor (TGF)-β and myogenic (MyoD and myogenin) gene expression by polymerase chain reaction, (2) muscle regeneration with haematoxylin and eosin staining, and (3) MyoD and myogenin protein expression using immunohistochemical staining. Rats in the CO2 group showed higher MyoD and myogenin expression and lower IL-1β, IL-6, and TGF-β expression than the control rats. In addition, treated rats showed evidence of accelerated muscle regeneration. Our study demonstrated that the CO2 paste prevents excessive scarring and accelerates muscle regeneration. This action may be exerted through the induction of an artificial Bohr effect, which leads to the upregulation of MyoD and myogenin, and the downregulation of IL-1β, IL-6, and TGF-β. The paste is inexpensive and non-invasive. Thus, it may be the treatment of choice for patients with muscle damage.

Title: Topical cutaneous CO2 application by means of a novel hydrogel accelerates fracture repair in rats

Authors: Koga T, Niikura T, Lee SY, Okumachi E, Ueha T, Iwakura T, Sakai Y, Miwa M, Kuroda R, Kurosaka M.

Journal: J Bone Joint Surg Am. 2014 Dec 17;96(24):2077-84. doi: 10.2106/JBJS.M.01498. PMID: 25520342.

Link to PubMed: Topical cutaneous CO2 application by means of a novel hydrogel accelerates fracture repair in rats

Abstract: Background: We previously demonstrated that topical cutaneous application of CO2, by means of a hydrogel in which the CO2 readily dissolves, increases blood flow and oxygen dissociation from hemoglobin in the soft tissues surrounding bone. In the present study, we utilized a rat fracture model to test the hypothesis that application of this treatment to fractured limbs would accelerate fracture repair.

Methods: A closed femoral shaft fracture was created in each rat. Topical cutaneous application of CO2 by means of a hydrogel was performed five times a week for up to four weeks in the CO2/hydrogel group (n = 60). Sham treatments were performed in the control group (n = 60). Radiographic, histological, immunohistochemical, laser Doppler perfusion imaging, real-time polymerase chain reaction, and biomechanical assessments were performed.

Results: Radiographic fracture union was evident at week 3 in twelve (86%) of fourteen animals in the CO2/hydrogel group compared with five (36%) of fourteen in the control group (p < 0.05; 95% CI [confidence interval] for the difference in union rate, 2.26% to 99.64%). Histological assessment revealed promotion of endochondral ossification in the CO2/hydrogel group. Immunohistochemical assessment at week 2 showed significantly greater capillary density in the CO2/hydrogel group (p < 0.05; 95% CI for the difference, 161 to 258 per mm(2)). Laser Doppler perfusion imaging demonstrated that the blood flow in the fractured limb was significantly greater at weeks 2 and 3 in the CO2/hydrogel group (p < 0.05; 95% CI for the difference, 8.4% to 22.4% and 6.7% to 19.0%, respectively). Gene expression of chondrogenic, osteogenic, and angiogenic markers was significantly greater in the CO2/hydrogel group at several time points. Ultimate stress, extrinsic stiffness, and failure energy (relative to the contralateral limb) were significantly greater in the CO2/hydrogel group at week 3 (p < 0.05; 95% CI for the difference, 24.8% to 67.5%, 4.0 % to 22.7%, and 9.6% to 58.8%, respectively). There were no significant differences between the groups with respect to any outcome measure at week 4.

Conclusions: Topical cutaneous application of CO2 by means of a hydrogel accelerated fracture repair in association with the promotion of angiogenesis, blood flow, and endochondral ossification.

Title: Transcutaneous carbon dioxide application inhibits muscle atrophy after fracture in rats

Authors: Inoue M, Sakai Y, Oe K, Ueha T, Koga T, Nishimoto H, Akahane S, Harada R, Lee SY, Niikura T, Kuroda R.

Journal: J Orthop Sci. 2020 Mar;25(2):338-343. doi: 10.1016/j.jos.2019.03.024. Epub 2019 Apr 25. PMID: 31031109.

Link to PubMed: Transcutaneous carbon dioxide application inhibits muscle atrophy after fracture in rats

Abstract: Background: Muscle atrophy causes difficulty in resuming daily activities after a fracture. Because transcutaneous carbon dioxide (CO2) application has previously upregulated oxygen pressure in the local tissue, thereby demonstrating its potential in preventing muscle atrophy, here we investigated effects of CO2 application on muscle atrophy after femoral shaft fracture.

Methods: Thirty fracture model rats were produced and randomly divided into a no treatment (control group) and treatment (CO2 group) groups. After treatment, the soleus muscle was dissected at post-fracture days 0, 14, and 21. Evaluations were performed by measuring muscle weight and performing histological examination and gene expression analysis.

Results: Muscle weight was significantly higher in the CO2 group than in the control group. Histological analysis revealed that the muscle fiber cross-sectional area was reduced in both groups. Nevertheless, the extent of atrophy was lesser in the CO2 group. Muscle fibers in the control group tended to change into fast muscle fibers. Vascular staining revealed that more capillary vessels surrounded the muscle fibers in the CO2 group than in the control group. Messenger RNA (mRNA) analysis revealed that the CO2 group had a significantly enhanced expression of genes that were related to muscle synthesis.

Conclusion: Transcutaneous CO2 application may be a novel therapeutic strategy for preventing skeletal muscle atrophy after fracture.

Title: The antioxidant action of СО2-one of the universal mechanisms of carboxytherapy

Authors: Drogovoz, S. M., K. O. Kalko, L. B. Ivantsyk, А. L. Shtroblya, Yu V. Stoletov, K. V. Drogovoz, and V. F. Ostasko.

Journal:

Link to full text: The antioxidant action of СО2-one of the universal mechanisms of carboxytherapy

Abstract: Carboxytherapy (carbon dioxide treatment) is a promising area and an alternative method of treating many diseases. Carbon dioxide (CO2) is constantly present in the body, being the end product of cellular metabolism. It accumulates in tissues, diffuses into the blood, and is transferred to the lungs in three forms: dissolved in plasma, in bicarbonates, and as oxyhemoglobin in erythrocytes. CO2 is totally indispensable for the normal course of biochemical, humoral, and tissue processes in the body, as it is a pacemaker of these processes.
Materials and research methods. Scientific publications in foreign and domestic journals on relevant topics over the past 5 years, Internet resources.
Research results and their discussion. Analysis of the above changes in the body under the action of CO2 and states of biochemical markers (ADMA and d-ROM, ROS, NO, NOS) under oxidative stress indicate that CO2 has antioxidant properties, and carboxytherapy is a pathogenetic therapy for diseases which pathogenesis includes oxidative stress. The ROS-inhibitory effect of CO2 is universal for different cell types and, obviously, for different species of living organisms.
Conclusions. Thus, it has been established that CO2 is a universal inhibitor of the reactive oxygen species generation by various human and animal cells. These data allow explaining the physiological and pathophysiological (medical) effects of CO2 in the mechanism of carboxytherapy in a new way from the standpoint of the biochemical role of ROS.
Key words: antioxidant action, carbon dioxide, carboxytherapy.

Title: Carbon dioxide-rich water bathing enhances collateral blood flow in ischemic hindlimb via mobilization of endothelial progenitor cells and activation of NO-cGMP system

Authors: Irie H, Tatsumi T, Takamiya M, Zen K, Takahashi T, Azuma A, Tateishi K, Nomura T, Hayashi H, Nakajima N, Okigaki M, Matsubara H.

Journal: Circulation. 2005 Mar 29;111(12):1523-9. doi: 10.1161/01.CIR.0000159329.40098.66. PMID: 15795362.

Link to full text: Carbon dioxide-rich water bathing enhances collateral blood flow in ischemic hindlimb via mobilization of endothelial progenitor cells and activation of NO-cGMP system

Abstract: Background: Carbon dioxide-rich water bathing has the effect of vasodilatation, whereas it remains undetermined whether this therapy exerts an angiogenic action associated with new vessel formation.

Methods and results: Unilateral hindlimb ischemia was induced by resecting the femoral arteries of C57BL/J mice. Lower limbs were immersed in CO2-enriched water (CO2 concentration, 1000 to 1200 mg/L) or freshwater (control) at 37 degrees C for 10 minutes once a day. Laser Doppler imaging revealed increased blood perfusion in ischemic limbs of CO2 bathing (38% increase at day 28, P<0.001), whereas N(G)-nitro-L-arginine methyl ester treatment abolished this effect. Angiography or immunohistochemistry revealed that collateral vessel formation and capillary densities were increased (4.1-fold and 3.7-fold, P<0.001, respectively). Plasma vascular endothelial growth factor (VEGF) levels were elevated at day 14 (18%, P<0.05). VEGF mRNA levels, phosphorylation of NO synthase, and cGMP accumulation in the CO2-bathed hindlimb muscles were increased (2.7-fold, 2.4-fold, and 3.4-fold, respectively) but not in forelimb muscles. The number of circulating Lin-/Flk-1+/CD34- endothelial-lineage progenitor cells was markedly increased by CO2 bathing (24-fold at day 14, P<0.001). The Lin-/Flk-1+/CD34- cells express other endothelial antigens (endoglin and VE-cadherin) and incorporated acetylated LDL.

Conclusions: Our present study demonstrates that CO2 bathing of ischemic hindlimb causes the induction of local VEGF synthesis, resulting in an NO-dependent neocapillary formation associated with mobilization of endothelial progenitor cells.

Title: Evidence-based hydro- and balneotherapy in Hungary--a systematic review and meta-analysis

Authors: Bender T, Bálint G, Prohászka Z, Géher P, Tefner IK.

Journal: Int J Biometeorol. 2014 Apr;58(3):311-23. doi: 10.1007/s00484-013-0667-6. Epub 2013 May 16. PMID: 23677421; PMCID: PMC3955132.

Link to full text: Evidence-based hydro- and balneotherapy in Hungary--a systematic review and meta-analysis

Abstract: Balneotherapy is appreciated as a traditional treatment modality in medicine. Hungary is rich in thermal mineral waters. Balneotherapy has been in extensive use for centuries and its effects have been studied in detail. Here, we present a systematic review and meta-analysis of clinical trials conducted with Hungarian thermal mineral waters, the findings of which have been published by Hungarian authors in English. The 122 studies identified in different databases include 18 clinical trials. Five of these evaluated the effect of hydro- and balneotherapy on chronic low back pain, four on osteoarthritis of the knee, and two on osteoarthritis of the hand. One of the remaining seven trials evaluated balneotherapy in chronic inflammatory pelvic diseases, while six studies explored its effect on various laboratory parameters. Out of the 18 studies, 9 met the predefined criteria for meta-analysis. The results confirmed the beneficial effect of balneotherapy on pain with weight bearing and at rest in patients with degenerative joint and spinal diseases. A similar effect has been found in chronic pelvic inflammatory disease. The review also revealed that balneotherapy has some beneficial effects on antioxidant status, and on metabolic and inflammatory parameters. Based on the results, we conclude that balneotherapy with Hungarian thermal-mineral waters is an effective remedy for lower back pain, as well as for knee and hand osteoarthritis.

Title: The Effectiveness of Manual Therapy in the Cervical Spine and Diaphragm, in Combination with Breathing Reeducation Exercises, in Patients with Non-Specific Chronic Neck Pain: Protocol for Development of Outcome Measures and a Randomized Controlled Trial

Authors: Tatsios PI, Grammatopoulou E, Dimitriadis Z, Koumantakis GA.

Journal: Diagnostics (Basel). 2022 Nov 4;12(11):2690. doi: 10.3390/diagnostics12112690. PMID: 36359533; PMCID: PMC9689657.

Link to full text: The Effectiveness of Manual Therapy in the Cervical Spine and Diaphragm, in Combination with Breathing Reeducation Exercises, in Patients with Non-Specific Chronic Neck Pain: Protocol for Development of Outcome Measures and a Randomized Controlled Trial

Abstract: Until now, non-specific chronic neck pain has mainly been considered as a musculoskeletal system dysfunction, with associated psychological involvement due to its prolonged or recurrent nature. However, patients with non-specific chronic neck pain frequently additionally exhibit respiratory dysfunction. Emerging evidence suggests that addressing the respiratory dysfunction in these patients will provide additional therapeutic benefits in musculoskeletal and respiratory-related outcomes for several reasons (biomechanical, biochemical, and psychological). Motor control dysfunction of the muscles surrounding the spine (diaphragm included) negatively affects the mechanics and biochemistry of breathing (pH-homeostasis). An impaired and ineffective breathing pattern has been recognized as the primary source of many unexplained symptoms (anxiety, depression, confusion, chest pain, hypocapnia, and breathlessness) in patients with non-specific chronic neck pain. The proposed protocol's purpose is dual: to assess the relative effectiveness of manual therapy in the cervical spine and the diaphragm, in combination with breathing reeducation exercises, along with cervical spine manual therapy or usual physical therapy care on the underlying dysfunctions in patients with non-specific chronic neck pain via a randomized controlled clinical trial, and to validate part of the outcome measures. Several musculoskeletal and respiratory dysfunction outcomes will be employed to delimit the initial extent and level of dysfunction and its resolution with the treatments under study.
Keywords: breathing exercises; breathing reeducation; chronic neck pain; diaphragm; manual therapy; respiratory dysfunction.

Title: The effect of transcutaneous application of carbon dioxide (CO₂) on skeletal muscle

Authors: Oe K, Ueha T, Sakai Y, Niikura T, Lee SY, Koh A, Hasegawa T, Tanaka M, Miwa M, Kurosaka M.

Journal: Biochem Biophys Res Commun. 2011 Apr 1;407(1):148-52. doi: 10.1016/j.bbrc.2011.02.128. Epub 2011 Mar 1. PMID: 21371433.

Link to PubMed: The effect of transcutaneous application of carbon dioxide (CO₂) on skeletal muscle

Abstract: In Europe, carbon dioxide therapy has been used for cardiac disease and skin problems for a long time. However there have been few reports investigating the effects of carbon dioxide therapy on skeletal muscle. Peroxisome proliferators-activated receptor (PPAR)-gamma coactivator-1 (PGC-1α) is up-regulated as a result of exercise and mediates known responses to exercise, such as mitochondrial biogenesis and muscle fiber-type switching, and neovascularization via up-regulation of vascular endothelial growth factor (VEGF). It is also known that silent mating type information regulation 2 homologs 1 (SIRT1) enhances PGC-1α-mediated muscle fiber-type switching. Previously, we demonstrated transcutaneous application of CO(2) increased blood flow and a partial increase of O(2) pressure in the local tissue known as the Bohr effect. In this study, we transcutaneously applied CO(2) to the lower limbs of rats, and investigated the effect on the fast muscle, tibialis anterior (TA) muscle. The transcutaneous CO(2) application caused: (1) the gene expression of PGC-1α, silent mating type information regulation 2 homologs 1 (SIRT1) and VEGF, and increased the number of mitochondria, as proven by real-time PCR and immunohistochemistry, (2) muscle fiber switching in the TA muscle, as proven by isolation of myosin heavy chain and ATPase staining. Our results suggest the transcutaneous application of CO(2) may have therapeutic potential for muscular strength recovery resulting from disuse atrophy in post-operative patients and the elderly population.

Title: Carbon Dioxide Therapy in the Treatment of Localized Adiposities: Clinical Study and Histopathological Correlations

Authors: Brandi C, D'Aniello C, Grimaldi L, Bosi B, Dei I, Lattarulo P, Alessandrini C.

Journal: Aesthetic Plast Surg. 2001 May-Jun;25(3):170-4. doi: 10.1007/s002660010116. PMID: 11426306.

Link to full text: Carbon Dioxide Therapy in the Treatment of Localized Adiposities: Clinical Study and Histopathological Correlations

Abstract: The authors report their experience using carbon dioxide (CO2) therapy for the treatment of 48 female patients presenting adipose accumulations, located on the thighs, knees, and/or abdomen; a Carbomed Programmable Automatic Carbon Dioxide Therapy apparatus was used. In light of the effects of CO2 on the microcirculation recently described in the literature, we expected this gas, which we administered subcutaneously, to positively affect the physiological oxidative lipolytic process. The aim of our study was to evaluate the effect of this therapy on localized adiposities. As such, we describe the method we used and report the results observed in the areas treated (in terms of reduction in maximum circumference) as well as side effects. Furthermore, we assessed the effect of subcutaneous administration of CO2 on the microcirculation by showing changes in the Laser Doppler signal and in the concentration of transcutaneous oxygen tension (tcPO2). Pre- and posttreatment biopsies of tissues were performed in seven patients in order to study the changes induced by the use of CO2 on both adipose and connective tissues. All data obtained were statistically analyzed; values of P < 0.05 were considered significant.

Title: An overview of the role of carboxytherapy in dermatology

Authors: Bagherani N, Smoller BR, Tavoosidana G, Ghanadan A, Wollina U, Lotti T.

Journal: J Cosmet Dermatol. 2023 Sep;22(9):2399-2407. doi: 10.1111/jocd.15741. Epub 2023 Mar 31. PMID: 36999460.

Link to full text: An overview of the role of carboxytherapy in dermatology

Abstract: Background: Carboxytherapy is defined as intradermal and/or subcutaneous microinjections of sterile purified carbon dioxide into different parts of the body for therapeutic aims. The vasodilatation effect and intradermal collagen reorganization associated with carboxytherapy have advantages for aesthetic dermatology and cosmetology.

Objective: In the current article, we have reviewed some of the most important indications of this modality in dermatology and aesthetic dermatology.

Method: Our review is a narrative one which has gathered some of the most important indications of carboxytherapy in dermatology and cosmetology.

Results: Carboxytherapy has successfully been applied for some dermatologic and cosmetic conditions among which skin aging, cellulite, localized fat deposits, striae distensae, infraorbital hyperpigmentation, scar, lymphedema, androgenetic alopecia, alopecia areata, psoriasis, morphea, and vitiligo are the most important.

Conclusion: Carboxytherapy can be considered as a safe, minimally-invasive modality used for rejuvenation, restoration, and recondition of the skin.

Title: Carboxytherapy: effects on microcirculation and its use in the treatment of severe lymphedema. A review

Authors: Varlaro V., Manzo G., Mugnaini F., Bisacci C., Fiorucci P., De Rango P., Bisacci R.

Journal: Acta Phlebologica 2007 August;8(2):79-91

Link to full text: Carboxytherapy: effects on microcirculation and its use in the treatment of severe lymphedema. A review

Abstract: Carboxytherapy refers to the administration of CO2 for therapeutic purposes. It has been shown that, because of the interaction between CO2 and regulating factors of tissue perfusion, Carboxytherapy acts on the microcirculation at the level of metarterioles, arterioles and precapillary shpincteres by increasing tissue flow velocity and consequently, by improving lymphatic drainage. Analysis of literature data shows a wide range of today applications for this treatment involving either phlebology or non-phlebology fields. Specifically, the positive effect on the increase of lymphatic drainage has more recently made Carboxytherapy useful for treatment of lymphatic stasis. Basic hemodynamic, histologyc and biochemical principles that explain the effects on microcirculation bed and lymphatic drainage are here analyzed to show how Carboxytherapy can be useful in the treatment of diseases such as severe lymphedema.

Title: The Influence of Carboxytherapy on Scar Reduction

Authors: Stolecka-Warzecha A, Chmielewski Ł, Deda A, Śmich A, Lebiedowska A, Wilczyński S.

Journal: Clin Cosmet Investig Dermatol. 2022 Dec 23;15:2855-2872. doi: 10.2147/CCID.S389380. PMID: 36582848; PMCID: PMC9793937.

Link to full text: The Influence of Carboxytherapy on Scar Reduction

Abstract: Introduction
Although it is not a new method, a carboxytherapy, which is based on intradermal or subcutaneous administration of controlled doses of CO2, is gaining more and more recognition among aesthetic medicine doctors, dermatologists and cosmetologists around the world. The consequence of applying carbon dioxide directly into tissues is associated with an immediate expansion of blood vessels, improvement of local blood supply, and, thus, tissue metabolism. It does also support natural regenerative processes. Oxygen and growth factors released from blood, within the area which undergoes treatment, stimulate fibroblasts to produce collagen and formation of new blood vessels also known as neovascularization. In addition to biochemical mechanisms, CO2 injection into the dermis or subcutaneous tissue leads to a mechanical effect exerted by pressure and flow of CO2 which is injected. It is of particular importance in scar treatments.

Methods
Twelve mature scars were subjected to the carboxytherapy which was performed in people aged 23–45 years. A small amount of heated, medical CO2 was injected till the moment a scar turns white. The applied flow rate equalled 100 mL/min (cc/min). Before and after a series of four treatments, the level of hydration, elasticity and colour of the skin were measured. A structured-light 3D scanner was used to determine an exact morphology of the examined scars. The 3D scanning device is seen as a sensitive and precise method of qualitative and quantitative assessment of a morphology of scars.

Results
The results of the measurements performed showed a reduction in the surface of the analyzed changes, as well as proved the ability of CO2 to rebuild collagen fibres. The decrease in value of parameters, which have been obtained thanks to the kutometric examination, indicates softening and loosening of connective tissue. It does confirm the effectiveness of carboxytherapy.

Title: Effect of leg immersion in mild warm carbonated water on skin and muscle blood flow

Authors: Ogoh S, Washio T, Suzuki K, Ikeda K, Hori T, Olesen ND, Muraoka Y.

Journal: Physiol Rep. 2018 Sep;6(18):e13859. doi: 10.14814/phy2.13859. PMID: 30221833; PMCID: PMC6139710.

Link to full text: Effect of leg immersion in mild warm carbonated water on skin and muscle blood flow

Abstract: Leg immersion in carbonated water improves endothelial-mediated vasodilator function and decreases arterial stiffness but the mechanism underlying this effect remains poorly defined. We hypothesized that carbonated water immersion increases muscle blood flow. To test this hypothesis, 10 men (age 21 ± 0 years; mean ± SD) underwent lower leg immersion in tap or carbonated water at 38°C. We evaluated gastrocnemius muscle oxyhemoglobin concentration and tissue oxygenation index using near-infrared spectroscopy, skin blood flow by laser Doppler flowmetry, and popliteal artery (PA) blood flow by duplex ultrasound. Immersion in carbonated, but not tap water elevated PA (from 38 ± 14 to 83 ± 31 mL/min; P < 0.001) and skin blood flow (by 779 ± 312%, P < 0.001). In contrast, lower leg immersion elevated oxyhemoglobin concentration and tissue oxygenation index with no effect of carbonation (P = 0.529 and P = 0.495). In addition, the change in PA blood flow in response to immersion in carbonated water correlated with those of skin blood flow (P = 0.005) but not oxyhemoglobin concentration (P = 0.765) and tissue oxygenation index (P = 0.136) while no relations was found for tap water immersion. These findings indicate that water carbonation has minimal effect on muscle blood flow. Furthermore, PA blood flow increases in response to lower leg immersion in carbonated water likely due to a large increase in skin blood flow.

Keywords: Carbonated water; Doppler ultrasound; near-infrared spectroscopy; popliteal artery.

Title: Carbonate ion-enriched hot spring water promotes skin wound healing in nude rats

Authors: Liang J, Kang D, Wang Y, Yu Y, Fan J, Takashi E.

Journal: PLoS One. 2015 Feb 11;10(2):e0117106. doi: 10.1371/journal.pone.0117106. PMID: 25671581; PMCID: PMC4324962.

Link to full text: Carbonate ion-enriched hot spring water promotes skin wound healing in nude rats

Abstract: Hot spring or hot spa bathing (Onsen) is a traditional therapy for the treatment of certain ailments. There is a common belief that hot spring bathing has therapeutic effects for wound healing, yet the underlying molecular mechanisms remain unclear. To examine this hypothesis, we investigated the effects of Nagano hot spring water (rich in carbonate ion, 42°C) on the healing process of the skin using a nude rat skin wound model. We found that hot spring bathing led to an enhanced healing speed compared to both the unbathed and hot-water (42°C) control groups. Histologically, the hot spring water group showed increased vessel density and reduced inflammatory cells in the granulation tissue of the wound area. Real-time RT-PCR analysis along with zymography revealed that the wound area of the hot spring water group exhibited a higher expression of matrix metalloproteinases-2 and -9 compared to the two other control groups. Furthermore, we found that the enhanced wound healing process induced by the carbonate ion-enriched hot spring water was mediated by thermal insulation and moisture maintenance. Our results provide the evidence that carbonate ion-enriched hot spring water is beneficial for the treatment of skin wounds.

Title: Carbon dioxide and MAPK signalling: towards therapy for inflammation

Authors: Gałgańska H, Jarmuszkiewicz W, Gałgański Ł.

Journal: Cell Commun Signal. 2023 Oct 10;21(1):280. doi: 10.1186/s12964-023-01306-x. PMID: 37817178; PMCID: PMC10566067.

Link to full text: Carbon dioxide and MAPK signalling: towards therapy for inflammation

Abstract: Inflammation, although necessary to fight infections, becomes a threat when it exceeds the capability of the immune system to control it. In addition, inflammation is a cause and/or symptom of many different disorders, including metabolic, neurodegenerative, autoimmune and cardiovascular diseases. Comorbidities and advanced age are typical predictors of more severe cases of seasonal viral infection, with COVID-19 a clear example. The primary importance of mitogen-activated protein kinases (MAPKs) in the course of COVID-19 is evident in the mechanisms by which cells are infected with SARS-CoV-2; the cytokine storm that profoundly worsens a patient's condition; the pathogenesis of diseases, such as diabetes, obesity, and hypertension, that contribute to a worsened prognosis; and post-COVID-19 complications, such as brain fog and thrombosis. An increasing number of reports have revealed that MAPKs are regulated by carbon dioxide (CO2); hence, we reviewed the literature to identify associations between CO2 and MAPKs and possible therapeutic benefits resulting from the elevation of CO2 levels. CO2 regulates key processes leading to and resulting from inflammation, and the therapeutic effects of CO2 (or bicarbonate, HCO3-) have been documented in all of the abovementioned comorbidities and complications of COVID-19 in which MAPKs play roles. The overlapping MAPK and CO2 signalling pathways in the contexts of allergy, apoptosis and cell survival, pulmonary oedema (alveolar fluid resorption), and mechanical ventilation-induced responses in lungs and related to mitochondria are also discussed. Video Abstract.

Keywords: ERK1/2; Hypercapnia; Hypercarbia; Hypocapnia; JNK; p38.

Title: The inflammatory reflex

Authors: Tracey KJ.

Journal: Nature. 2002 Dec 19-26;420(6917):853-9. doi: 10.1038/nature01321. PMID: 12490958.

Link to PubMed: The inflammatory reflex

Abstract: Inflammation is a local, protective response to microbial invasion or injury. It must be fine-tuned and regulated precisely, because deficiencies or excesses of the inflammatory response cause morbidity and shorten lifespan. The discovery that cholinergic neurons inhibit acute inflammation has qualitatively expanded our understanding of how the nervous system modulates immune responses. The nervous system reflexively regulates the inflammatory response in real time, just as it controls heart rate and other vital functions. The opportunity now exists to apply this insight to the treatment of inflammation through selective and reversible 'hard-wired' neural systems.

Title: Effects of repeated carbon dioxide-rich water bathing on core temperature, cutaneous blood flow and thermal sensation

Authors: Nishimura N, Sugenoya J, Matsumoto T, Kato M, Sakakibara H, Nishiyama T, Inukai Y, Okagawa T, Ogata A.

Journal: Eur J Appl Physiol. 2002 Aug;87(4-5):337-42. doi: 10.1007/s00421-002-0626-0. Epub 2002 Jun 7. PMID: 12172871.

Link to full text: Effects of repeated carbon dioxide-rich water bathing on core temperature, cutaneous blood flow and thermal sensation

Abstract: We examined the effects of repeated artificial CO(2) (1,000 ppm) bathing on tympanic temperature (T(ty)), cutaneous blood flow, and thermal sensation in six healthy males. Each subject was immersed in CO(2)-rich water at a temperature of 34 degrees C up to the level of the diaphragm for 20 min. The CO(2)-rich water was prepared using a multi-layered composite hollow-fiber membrane. The CO(2) bathing was performed consecutively for 5 days. As a control study, subjects bathed in fresh water at 34 degrees C under the same conditions. T(ty) was significantly lowered during CO(2) bathing (P < 0.05). Cutaneous blood flow in the immersed skin (right forearm) was significantly increased during CO(2) bathing compared with that during fresh-water bathing (P < 0.05), whereas cutaneous blood flow in the non-immersed skin (chest) was not different between CO(2) and fresh-water bathing. Subjects reported a "warm" sensation during the CO(2) bathing, whereas they reported a "neutral" sensation during the fresh-water bathing. The effects of the repeated CO(2) bathing were not obvious for core temperature and cutaneous blood flow, but the thermal sensation score during the CO(2) bathing was reduced sequentially by repeated CO(2) bathing (P < 0.05). These thermal effects of CO(2) bathing could be ascribed largely to the direct action of CO(2) on vascular smooth muscles and to the activity of thermoreceptors in the skin. Serial CO(2) bathing may influence the activity of thermoreceptors in the skin.

Title: Effect of carbon dioxide-enriched water and fresh water on the cutaneous microcirculation and oxygen tension in the skin of the foot

Authors: Hartmann BR, Bassenge E, Pittler M.

Journal: Angiology. 1997 Apr;48(4):337-43. doi: 10.1177/000331979704800406. PMID: 9112881.

Link to full text: Effect of carbon dioxide-enriched water and fresh water on the cutaneous microcirculation and oxygen tension in the skin of the foot

Abstract: The effects of immersion of the lower leg and foot in fresh water and in carbon dioxide (CO2)-enriched water (1200 mg CO2 per kg water; succinate+sodium bicarbonate: Actibath, KAO Tokyo) on cutaneous circulation, vasomotion and oxygen tension (PO2) were measured by laser Doppler flowmetry and transcutaneous oximetry. On the first of two consecutive days patients were randomly assigned to have the lower extremities immersed in either fresh water or CO2-enriched water under standardized conditions (temperature, 34 degrees C; depth, 35 cm; immersion time, twenty minutes) with concurrent measurement. On the second day patients were switched to the other bath type. For both sets of measurements probes were attached symmetrically to the dorsum of each foot. Included in the study were 18 patients with mild, bilateral, peripheral, occlusive arterial disease (intermittent claudication, femoral or iliac type). During immersion in CO2-enriched water the Doppler laser signal and vasomotion amplitude rose by 300%, while PO2 increased by 10%. These increases were still apparent during the latter part of the measurement period, following withdrawal of the limbs from the bath, while patients were seated and supine. During immersion in fresh water and thereafter the Doppler laser signal was unchanged and the PO2 increase was considerably less marked. The authors were thus able to demonstrate vasodilation and increased oxygen utilization (Bohr effect) resulting from topical CO2 application, and hence, that the use of topical CO2 has an objective basis.

Title: Effect of Carbon Dioxide Therapy on Diabetic Foot Ulcer

Authors: Shalan N., Al-Bazzaz A., Al-Ani I., Najem F. and Al-Masri M.

Journal: Journal of Diabetes mellitus 5, no. 4 (2015): 284-289.

Link to full text: Effect of Carbon Dioxide Therapy on Diabetic Foot Ulcer

Abstract: Ulceration of diabetic foot represents one of the most concerning complications associated with uncontrolled blood sugar in diabetes mellitus. The aim of this study is to evaluate the condition of twenty-two diabetic patients with different degrees of ulceration in their feet after daily secessions of carbon dioxide therapy. Blood flow to the affected foot was measured by Doppler; also the size, color, degree of ulceration and sensation of the ulcerative area were all evaluated. Results showed improvement of blood flow to the affected foot as well as improvement in the sensation and color of the ulcerative area. It was concluded that carbon dioxide therapy of diabetic foot was promising and needed thorough investigation to be brought widely into application.
Keywords: Diabetic Foot Ulcer, Carbon Dioxide Therapy

Title: Carbon dioxide therapy in the treatment of cellulite: an audit of clinical practice

Authors: Lee GS.

Journal: Aesthetic Plast Surg. 2010 Apr;34(2):239-43. doi: 10.1007/s00266-009-9459-0. Epub 2010 Jan 29. PMID: 20111965; PMCID: PMC2847160.

Link to full text: Carbon dioxide therapy in the treatment of cellulite: an audit of clinical practice

Abstract: Background: The clinical practice of using carbon dioxide therapy for localized adiposities was audited over a 4-year period.

Methods: Patients receiving physical, dietary, or drug concurrent therapy were excluded from the audit. Original measurements in terms of mean +/- standard error of the mean (SEM) were compared with those obtained after five sessions.

Results: This series included 101 women who underwent abdominal therapy. Significant reduction (p < 0.05) in mean upper, mid, and lower abdomen circumference was experienced by all three age groups: respectively, 1.8 +/- 0.5, 1.6 +/- 0.4, and 2.1 +/- 0.3 cm in the 20- to 29-year- old group, 1.6 +/- 0.4, 2.3 +/- 0.3, and 2.1 +/- 0.3 cm in the 30- to 39-year-old group, and 2.0 +/- 0.4, 2.5 +/- 0.4, and 2.6 +/- 0.4 cm in the 40- to 50-year-old group. For 57 women who underwent localized thigh therapy (27 in the 20- to 29-year-old group, 18 in the 30- to 39-year-old group, and 12 in the 40- to 50-year-old group), thigh circumference was significantly reduced in the right versus left thigh: respectively, 1.6 +/- 0.3 versus 1.5 +/- 0.2 cm, 1.1 +/- 0.3 versus 1.1 +/- 0.3 cm, 1.6 +/- 0.3 versus 1.5 +/- 0.4. Weight loss was significant for older women who underwent abdominal therapy: 1.3 +/- 0.2 kg in the 30- to 39-year-old group (n = 43) and 1.3 +/- 0.2 kg in the 40- to 50 year-old group (n = 29). Older women who underwent thigh therapy also recorded significant weight reduction: 0.9 +/- 0.4 kg in the 30- to 39-year-old group (n = 18) and 1.6 +/- 0.3 kg in the 40- to 50-year-old group (n = 12). The results for 10 men were not significant.

Conclusion: These results agree with those reported originally and demonstrate that carboxytherapy is safe and effective.

Title: Carbon dioxide therapy: effects on skin irregularity and its use as a complement to liposuction

Authors: Brandi C, D'Aniello C, Grimaldi L, Caiazzo E, Stanghellini E.

Journal: Aesthetic Plast Surg. 2004 Jul-Aug;28(4):222-5. doi: 10.1007/s00266-004-2068-z. PMID: 15599534.

Link to PubMed: Carbon dioxide therapy: effects on skin irregularity and its use as a complement to liposuction

Abstract: For a successful conventional or superficial liposuction, it is necessary to consider the competence of the surgeon who is to administer the procedure necessary for this type of surgery as well as the physical and psychological evaluation of the determined patient. A poor result often is related to the persistence of adipose tissue irregularity in the form of fatty tissue accumulation. This complication, common to this type of surgery, has called for research to determine methods for its treatment. Carbon dioxide (CO2) therapy refers to the transcutaneous and subcutaneous administration of CO2 for therapeutic purposes. This treatment originated at the Royal Spas of France in 1932 with the treatment of patients affected by obliteration of arteriopathies. Recent studies have demonstrated the effect of subcutaneous CO2 therapy performed to improve local parameters of circulation (performed by Doppler, laser-Doppler, and trans-cutaneous partial pressure of oxygen determination), and to reduce localized adiposities (verified reporting variations in maximum circumference and performing histologic studies). With these results, the absence of toxicity, and the relevant side effects related to this treatment taken into consideration, the Plastic Surgery Unit of Siena has been committed to researching the role that CO2 therapy can play in the treatment of skin irregularity and as a complement to liposuction. The authors report their experience using Carbomed programmable automatic CO2 therapy apparatus and 30GA1/2 0,3X13 microlance needles for the treatment of patients with adipose tissue accumulations located on the thighs and knees. In their study, 42 patients were divided into three groups: A, B, and C. In Group A, only liposuction was performed. In group B 3 weeks after liposuction CO2 therapy was administered in two weekly subcutaneous applications of CO2 for 10 consecutive weeks. In group C, CO2 therapy alone was administered with the same contingencies used for group B (two weekly subcutaneous applications of CO2 for 10 consecutive weeks). The objective was to assess the effectiveness of CO2 therapy for skin irregularity and as a complement to liposuction for adipose tissue accumulation by reporting variations in circumference and skin elasticity monitored by the Cutometer SEM 474 in all treated areas. The data obtained were analyzed statistically. Values of p less than 0.05 were considered significant. The authors report their experience and the results achieved from the study.

Title: Immersing Feet in Carbon Dioxide-enriched Water Prevents Expansion and Formation of Ischemic Ulcers after Surgical Revascularization in Diabetic Patients with Critical Limb Ischemia

Authors: Hayashi H, Yamada S, Kumada Y, Matsuo H, Toriyama T, Kawahara H.

Journal: Ann Vasc Dis. 2008;1(2):111-7. doi: 10.3400/avd.AVDoa08001. Epub 2008 Oct 24. PMID: 23555347; PMCID: PMC3595724.

Link to full text: Immersing Feet in Carbon Dioxide-enriched Water Prevents Expansion and Formation of Ischemic Ulcers after Surgical Revascularization in Diabetic Patients with Critical Limb Ischemia

Abstract: Objective: We investigated the effect of immersion of feet in CO2-enriched water for preventing expansion and formation of ischemic ulcer in critical limb ischemia of diabetic patients after surgical revascularization.

Materials and methods: Eligible patients were allocated CO2 group (CO2 immersion plus standard care) or control group (standard care alone) and were followed up for 3 months after surgical revascularization. The end point is defined as an expansion of a target ulcer (more than 101% of original size) or the formation of new ulcers during the follow-up period.

Results: Fifty-nine patients out of originally enrolled 66 patients with type II diabetes were included in intention-to-treat population. The cumulative prevention rate for ischemic ulcer after 3 months was 97.1% in the CO2 group, while, in the control group, it was 77.8%, i.e., significantly lower than the CO2 group (P = 0.012, log-rank test). The transcutaneous oxygen pressure increased significantly only in the CO2 group, from 56 ± 14 to 63 ± 15 mmHg (P < 0.01, Wilcoxon signed rank test), in 3 months.

Conclusion: These results suggest that addition of CO2 immersion to standard care of critical limb ischemia in diabetic patients improves early postoperative outcome after vascular surgery.

Keywords: adjunctive treatments; carbon dioxide; critical limb ischemia; revascularization; transcutaneous oxygen pressure.

Title: A novel system for transcutaneous application of carbon dioxide causing an "artificial Bohr effect" in the human body

Authors: Sakai Y, Miwa M, Oe K, Ueha T, Koh A, Niikura T, Iwakura T, Lee SY, Tanaka M, Kurosaka M.

Journal: PLoS One. 2011;6(9):e24137. doi: 10.1371/journal.pone.0024137. Epub 2011 Sep 8. PMID: 21931656; PMCID: PMC3169585.

Link to full text: A novel system for transcutaneous application of carbon dioxide causing an "artificial Bohr effect" in the human body

Abstract: Background: Carbon dioxide (CO(2)) therapy refers to the transcutaneous administration of CO(2) for therapeutic purposes. This effect has been explained by an increase in the pressure of O(2) in tissues known as the Bohr effect. However, there have been no reports investigating the oxygen dissociation of haemoglobin (Hb) during transcutaneous application of CO(2)in vivo. In this study, we investigate whether the Bohr effect is caused by transcutaneous application of CO2 in human living body.

Methods: We used a novel system for transcutaneous application of CO(2) using pure CO(2) gas, hydrogel, and a plastic adaptor. The validity of the CO(2) hydrogel was confirmed in vitro using a measuring device for transcutaneous CO(2) absorption using rat skin. Next, we measured the pH change in the human triceps surae muscle during transcutaneous application of CO(2) using phosphorus-31 magnetic resonance spectroscopy ((31)P-MRS) in vivo. In addition, oxy- and deoxy-Hb concentrations were measured with near-infrared spectroscopy in the human arm with occulted blood flow to investigate O2 dissociation from Hb caused by transcutaneous application of CO(2).

Results: The rat skin experiment showed that CO(2) hydrogel enhanced CO(2) gas permeation through the rat skin. The intracellular pH of the triceps surae muscle decreased significantly 10 min. after transcutaneous application of CO(2). The NIRS data show the oxy-Hb concentration decreased significantly 4 min. after CO(2) application, and deoxy-Hb concentration increased significantly 2 min. after CO(2) application in the CO(2)-applied group compared to the control group. Oxy-Hb concentration significantly decreased while deoxy-Hb concentration significantly increased after transcutaneous CO(2) application.

Conclusions: Our novel transcutaneous CO(2) application facilitated an O(2) dissociation from Hb in the human body, thus providing evidence of the Bohr effect in vivo.

Title: Possibilities and limits of CO2 balneotherapy

Authors: Resch KL, Just U.

Journal: Wien Med Wochenschr. 1994;144(3):45-50. German. PMID: 8017066.

Link to PubMed: Possibilities and limits of CO2 balneotherapy

Abstract: CO2 baths are considered an integral feature of balneotherapy. Natural springs containing a concentration of CO2 probably sufficient to be clinically effective (> 1 g/l) as well as artificially enriched baths are applied. Phenomena like a marked erythema (at concentrations above 0.5 g/l) imply clinical efficacy beyond unspecific effects of baths (water immersion, thermal effects). Prolonged effects are postulated to exceed "direct" effects, thus forming the rationale for serial applications of CO2-baths. CO2 is believed to cause an acute induction of local vasodilation and a shift of the O2 binding curve, resulting in a facilitation of the delivery of O2 to the tissue. A positive impact of CO2-baths on the flow properties of blood has been reported. A wide variety of indications is mentioned in the literature, while clear evidence from controlled trials exists only for a minority of them, mainly for chronic circulatory disturbances based on atherosclerotic diseases such as peripheral arterial occlusive disease, trophic ulceration, microangiopathies of various origins, and mild hypertension. Some evidence supports the idea that CO2 baths might represent an efficient therapeutic means in the rehabilitation of coronary heart disease, myocardial infarction and stroke, and in the treatment of chronic venous insufficiency, certain inflammatory diseases, and functional disturbances.

Title: Comparison of the effects of carbonated baths with those of carbon dioxide baths, "dry baths"

Authors: Lecomte J, Lagneaux D.

Journal: Acta Belg Med Phys. 1990 Jan-Mar;13(1):11-4. French. PMID: 2110708.

Link to PubMed: Comparison of the effects of carbonated baths with those of carbon dioxide baths, "dry baths"

Abstract: Therapeutic activities of CO2 water bathes are explained by a synergism between many factors. The first ones are related to immersion it-self: hydrostatic pressure and Archimedes force inducing orthosympathetic inhibition and muscular relaxation. The second ones depend on the pharmacological properties of carbon dioxide acting directly on the blood vessels of the skin, causing vasodilation and heat sensation. As "dry" carbon dioxide bath is devoid of the physical factors which are necessary for a successful therapeutic activity, such an administration has no interest in balneotherapy.

Title: A Prospective Clinical and Instrumental Study on the Effects of a Transcutaneous Cosmeceutical Gel that is Claimed to Produce CO₂

Authors: Leibaschoff GH, Coll L, Roberts WE.

Journal: Surg Technol Int. 2018 Jun 1;32:33-45. PMID: 29566423.

Link to PubMed: A Prospective Clinical and Instrumental Study on the Effects of a Transcutaneous Cosmeceutical Gel that is Claimed to Produce CO₂

Abstract: Carboxytherapy is the therapeutic use of carbon dioxide (CO2) in its gaseous state. Since 1933, carboxytherapy has referred to either the subcutaneous injection of CO2 or percutaneous application in a warm bath. The present clinical study was performed to determine if there were any changes in the dermis after the application of a transcutaneous gel, which is claimed to produce CO2, and, if so, how these changes compared to those with CO2 injection. Ten patients received transcutaneous treatment with the gel on one side of the face and the other side without any product was used as a control. We used videocapillaroscopy with an optic probe (VCSO) to evaluate the changes in the microcirculation of the skin. VCSO was performed for the treated right and untreated left ear lobes in each patient. VCSO was performed before treatment was started (VCSO1) and after 7 days of treatment (VCSO2). A comparison of VCSO1 to VCSO2 showed an increase in the microcirculation, an increase in vertical and horizontal capillaries, and a reduction in the area of ischemia. These results are similar to those observed in other studies with CO2 injection. In conclusion, use of this transcutaneous CO2 gel produced changes in the dermis similar to those observed with subcutaneous injection of CO2.

Title: Effectiveness of carboxytherapy in the treatment of cellulite in healthy women: a pilot study

Authors: Pianez LR, Custódio FS, Guidi RM, de Freitas JN, Sant'Ana E.

Journal: Clin Cosmet Investig Dermatol. 2016 Aug 22;9:183-90. doi: 10.2147/CCID.S102503. PMID: 27578994; PMCID: PMC5001663.

Link to full text: Effectiveness of carboxytherapy in the treatment of cellulite in healthy women: a pilot study

Abstract: Background: Carbon dioxide therapy, better known as carboxytherapy, relates to percutaneous infusion of medical carbon dioxide with therapeutic approaches, and its use in the treatment of localized fat has demonstrated good results. Gynoid lipodystrophy, also known as cellulite, affects 80%-90% of women after puberty, especially in the buttocks and thighs. Its etiology is complex and involves multifactorial aspects. Its treatment and evaluation require the use of new technologies (more effective and low-cost approaches). The objective was to investigate the effectiveness of carboxytherapy in the treatment of cellulite in the areas of buttocks and posterior thigh.

Patients and methods: Ten women, 29±6.1 years, were selected and all of them received eight treatment sessions, with an interval of 7 days between sessions. Standardized digital photographs were used to assess the severity of cellulite, and panoramic images were collected by ultrasound diagnosis. The evaluations were performed before the first treatment (baseline) and 7 days after the last treatment session of carboxytherapy.

Results: After the treatment, there was a significant reduction (P=0.0025) of the cellulite from degree III to degree II, and this improvement had correlation with the improvement in the organization of the fibrous lines and the disposal of adipose tissue lines of the treated regions observed through the panoramic ultrasound images diagnosis.

Conclusion: Carboxytherapy is an effective technique of treatment of cellulite in the buttocks region and posterior thighs of healthy women.

Keywords: carbon dioxide; cellulite; gynoid lipodystrophy; localized fat; panoramic ultrasound; skin.

Title: Carboxytherapy in dermatology: A systematic review

Authors: Ahramiyanpour N, Shafie'ei M, Sarvipour N, Amiri R, Akbari Z.

Journal: J Cosmet Dermatol. 2022 May;21(5):1874-1894. doi: 10.1111/jocd.14834. Epub 2022 Feb 14. PMID: 35124882.

Link to PubMed: Carboxytherapy in dermatology: A systematic review

Abstract: Introduction: The overall effects of Carboxytherapy, defined as the administration of carbon dioxide, have been studied for many years. It has been suggested that by improving oxygenation, interacting with the tissue perfusion regulators, and disrupting the adipose cell membranes, the method can lead to notable improvements in different esthetic and pathological conditions. Therefore, we aimed to systematically review the available studies evaluating the potential benefits of carboxytherapy in dermatological conditions and how it objectively stands against scientific scrutiny.

Methods: We searched the PubMed, Scopus, Embase, and Web of Science databases, including the studies exploring the method's efficacy in managing any dermatological condition.

Results: A total 27 of studies were identified (with a pooled sample of over 700 cases), most of which were clinical trials. Facial wrinkles, periorbital hyperpigmentation, skin laxity deficiency, scars, striae distensae, localized lipolysis and cellulite, alopecia, chronic diabetic wounds, and psoriatic plaques comprised the package of the dermatological conditions that were studied. Except for a few studies, the method mainly demonstrated significant improvements on all of the mentioned conditions. The inter- and post-operational adverse events were mild and transient, including erythema, pain, crepitus, and ecchymoses.

Discussion: Carboxytherapy can provide those practicing in the field with sustainably favorable results. However, the numbers of cases on whom the fat-reducing capabilities of the method were studied and experienced varying degrees of recurrence caught our eye. In addition, we observed a notable disparity between the outcome measures utilized in the studies. The modest sample size in each condition also added to the injury, as the conditions on which the method was evaluated are pretty common in the general population. Therefore, for a definite conclusion, more randomized controlled trials with the shortcomings mentioned well addressed need to be conducted.

Keywords: carbon dioxide therapy; carboxytherapy; dermatology.

Title: Carboxytherapy in esthetic medicine – A review

Authors: Zelenkova, H. and Wollina, U.

Journal: A review." Cosmoderma 3 (2023).

Link to full text: Carboxytherapy in esthetic medicine – A review

Abstract: Carboxytherapy is a minimally invasive method used not only in esthetic medicine. It is defined as the transcutaneous or subcutaneous application of carbon dioxide gas. The treatment exerts effects on various levels including improvement of tissue oxygenation, vasodilatation, stimulation of fibroblasts, and antimicrobial activity. In this review, we will focus on facial and neck esthetics, localized adiposities and cellulite, and scars and stretch marks. We discuss reported evidence for these indications and safety issues. We also mention possible combinations with other treatments. In general, a repeated application is necessary to achieve desired results.
Keywords: Esthetic medicine, Carboxytherapy, Indications, Safety

Title: Carboxytherapy--a new non-invasive method in aesthetic medicine

Authors: Koutná N.

Journal: Cas Lek Cesk. 2006;145(11):841-3. Czech. PMID: 17168416.

Link to PubMed: Carboxytherapy--a new non-invasive method in aesthetic medicine

Abstract: Basic information about carboxytherapy and its new use in non-invasive aesthetic medicine and in dermatology is presented in the article.

Title: Comparison of the effects of carboxytherapy and radiofrequency on skin rejuvenation

Authors: Pinheiro NM, Crema VO, Millan BM, Carvalho FA, Mendonça AC.

Journal: J Cosmet Laser Ther. 2015 Jun;17(3):156-61. doi: 10.3109/14764172.2014.1003238. Epub 2015 Jan 30. PMID: 25549818.

Link to PubMed: Comparison of the effects of carboxytherapy and radiofrequency on skin rejuvenation

Abstract: Background: Carboxytherapy (CA) refers to the cutaneous and subcutaneous administration of CO(2) for therapeutic purposes. Radiofrequency (RF) is a method that uses electric current for heating layers of the skin. Both techniques are indicated for the treatment of skin laxity.

Objective: The aim of this study was to compare the effects of CA and RF on human skin.

Methods: After eight patients underwent abdominoplasty, each of them received a single treatment of CA and a single treatment of RF on the right and left infra-umbilical regions, respectively. In the infra-umbilical region, CA was performed on the right and RF was performed on the left side. Untreated skin was used as a control. The sample collection period lasted 120 days. CA was administered at a velocity of 40 mL/min, and the total quantity of CO(2) infused was approximately 20 mL. RF was carried out at a temperature higher than 40°C on the epidermis for 5 min.

Results: CA and RF led to collagen remodeling; however, this result was more evident and lasted longer with RF. With CA an increase in elastic fibers was observed, whereas with RF no alteration was observed.

Conclusion: Our results suggest that RF is more efficient than CA in stimulating collagen synthesis.

Keywords: carbon dioxide; carboxytherapy; radiofrequency; rejuvenation; skin aging.

Title: Increase in collagen turnover induced by intradermal injection of carbon dioxide in rats

Authors: Ferreira JC, Haddad A, Tavares SA.

Journal: J Drugs Dermatol. 2008 Mar;7(3):201-6. PMID: 18380201.

Link to PubMed: Increase in collagen turnover induced by intradermal injection of carbon dioxide in rats

Abstract: Introduction: Results from clinical observations have demonstrated that percutaneous infiltration of carbon dioxide improves the appearance of the skin in adjacent areas. No studies have been found in the literature that showed evidence of histological changes caused by carbon dioxide injections.

Objectives and methods: A blind cross-sectional pilot study was performed in the Departments of Pharmacology and Morphology of the Federal University of Ceará, with the aim of histologically investigating whether intradermal and/or subcutaneous injection of medicinal carbon dioxide would increase collagen turnover in rats. Ten male Wistar rats were used, aged 3 months (2 animals) and 14 months (8 animals). The 2 younger rats were used as controls. Four of the older rats received injections of saline solution (0.9%), and were also considered to be controls. In the remaining 4, carbon dioxide was injected into the subcutaneous cellular tissue and intradermally. Biopsy samples were collected before and after treatment with carbon dioxide.

Results: Collagen turnover increased in the treated animals in comparison with the controls. Compression of collagen bundles in the tissue samples where intradermal injection was used was more intense than in the subcutaneous treatment. The histological characteristics of the samples with carbon dioxide injected intradermally were similar to the characteristics of the younger rats (controls).

Conclusions: The results obtained corroborate clinical observations of aesthetic improvements in the facial skin with carbon dioxide injections. Future research should address the comparison between intradermal and subcutaneous injections, the volume of gas used, and the frequency of treatment sessions.

Title: CO2 release to the atmosphere from thermal springs of Sperchios Basin and northern Euboea (Greece): The contribution of "hidden" degassing

Authors: D'Alessandro W, Li Vigni L, Gagliano Antonina L, Calabrese S, Kyriakopoulos K, Daskalopoulou K.

Journal: Applied Geochemistry, Volume 119, 2020, 104660, ISSN 0883-2927, https://doi.org/10.1016/j.apgeochem.2020.104660.

Link to ScienceDirect: CO2 release to the atmosphere from thermal springs of Sperchios Basin and northern Euboea (Greece): The contribution of "hidden" degassing

Abstract: Greece is a region characterised by intense geodynamic activity that favours the circulation of hydrothermal fluids in the crust transporting volatiles from either the deep crust or the mantle to the surface. Elevated heat flow values are detectable at Sperchios Basin and North Euboea (central Greece), two areas defined by a system of deeply rooted extensional faults and Quaternary volcanic activity. This setting contributed to the formation of numerous hydrothermal systems, which are mostly expressed as CO2-rich thermal springs with intense bubbling. The CO2 output from six bubbling pools has been determined by flux measurements with the use of the floating chamber method. The highest bubbling CO2 output has been found at Thermopyles and Psoroneria (0.6 and 0.84 t/d, respectively), where the outgoing channels have an elevated flow (>250 l/s) of gas-charged water (>15 mmol/l of CO2). Although no bubbling is noticed along the stream, the CO2 content decreases by an order of magnitude after few hundreds of metres, indicating an intense degassing from the water. Taking into account the water flow and the amount of CO2 lost to the atmosphere, the CO2 output of the outgoing channels is quantified in 12.5 t/d for Thermopyles and 9.23 t/d for Psoroneria. Output estimation has also been made for the other springs of the area. The diffuse CO2 release from the outgoing channels has always been higher respect to that of the visible bubbling, suggesting that most of the degassing is "hidden". Furthermore, the loss of CO2 from the water has determined a shift in dissolved carbonate species as demonstrated by the pH increase along the channel that led eventually to an oversaturation in carbonate minerals and therefore travertine deposition. The total CO2 output to the atmosphere of the study area is estimated at ~27 t/d, with the major contribution deriving from the degassing along the outflow channels of the thermal springs. Such output is comparable to that of the single active volcanic systems along the South Aegean Active Volcanic Arc and highlights the importance of "hidden" degassing along CO2-oversaturated streams.

Title: Natural skin surface pH is on average below 5, which is beneficial for its resident flora

Authors: Lambers H, Piessens S, Bloem A, Pronk H, Finkel P.

Journal: Int J Cosmet Sci. 2006 Oct;28(5):359-70. doi: 10.1111/j.1467-2494.2006.00344.x. PMID: 18489300.

Link to full text: Natural skin surface pH is on average below 5, which is beneficial for its resident flora

Abstract: Variable skin pH values are being reported in literature, all in the acidic range but with a broad range from pH 4.0 to 7.0. In a multicentre study (N = 330), we have assessed the skin surface pH of the volar forearm before and after refraining from showering and cosmetic product application for 24 h. The average pH dropped from 5.12 +/- 0.56 to 4.93 +/- 0.45. On the basis of this pH drop, it is estimated that the 'natural' skin surface pH is on average 4.7, i.e. below 5. This is in line with existing literature, where a relatively large number of reports (c. 50%) actually describes pH values below 5.0; this is in contrast to the general assumption, that skin surface pH is on average between 5.0 and 6.0. Not only prior use of cosmetic products, especially soaps, have profound influence on skin surface pH, but the use of plain tap water, in Europe with a pH value generally around 8.0, will increase skin pH up to 6 h after application before returning to its 'natural' value of on average below 5.0. It is demonstrated that skin with pH values below 5.0 is in a better condition than skin with pH values above 5.0, as shown by measuring the biophysical parameters of barrier function, moisturization and scaling. The effect of pH on adhesion of resident skin microflora was also assessed; an acid skin pH (4-4.5) keeps the resident bacterial flora attached to the skin, whereas an alkaline pH (8-9) promotes the dispersal from the skin.

Title: The Shampoo pH can Affect the Hair: Myth or Reality?

Authors: Gavazzoni Dias MF, de Almeida AM, Cecato PM, Adriano AR, Pichler J.

Journal: Int J Trichology. 2014 Jul;6(3):95-9. doi: 10.4103/0974-7753.139078. PMID: 25210332; PMCID: PMC4158629.

Link to full text: The Shampoo pH can Affect the Hair: Myth or Reality?

Abstract: Aim: Dermatologists most frequently prescribe shampoos for the treatment of hair shed and scalp disorders. Prescription of hair care products is often focused on improving scalp hair density, whereas the over-the-counter products focus on hair damage prevention. Little is taught in medical schools about the hair cosmetics, so that the prescriptions are based only on the treatment of the scalp and usually disregards the hair fiber health.

Materials and methods: In this work, we review the current literature about the mode of action of a low-pH shampoo regarding the hair shaft's health and analyze the pH of 123 shampoos of international brands.

Results: All shampoo pH values ranged from 3.5 to 9.0. 38.21% of all 123 shampoos presented a pH ≤ 5.5 (IC: 29.9-47%) and 61.78% presented a pH > 5.5. 26 anti-dandruff shampoos were analyzed. About 19.23% presented pH ≤ 5.5.(IC: 7.4-37.6%). 80.77% of all anti-dandruffs shampoos presented a pH > 5.5. The dermatological shampoo group (n = 19) presented 42.10% with pH ≤ 5.5 (IC: 21.8-64.6%), and 57.90% with pH > 5.5. Among the commercial (popular) products (n = 96), 34.37% presented pH ≤ 5.5 (IC: 25.4-44.3%) and 65.62% presented pH > 5.5. 15 professional products (used in hair salons) were analyzed, of which 75% had a pH ≤ 5.5 (IC: 18-65, 4%), and 25% had a pH > 5.5. 100% of the children's shampoos presented a pH > 5.5.

Conclusions: Alkaline pH may increase the negative electrical charge of the hair fiber surface and, therefore, increase friction between the fibers. This may lead to cuticle damage and fiber breakage. It is a reality and not a myth that lower pH of shampoos may cause less frizzing for generating less negative static electricity on the fiber surface. Interestingly, only 38% of the popular brand shampoos against 75% of the salons shampoos presented a pH ≤ 5.0. Pediatric shampoos had the pH of 7.0 because of the "no-tear" concept. There is no standardized value for the final pH. The authors believe that it is important to reveal the pH value on the shampoo label, but studies are needed to establish the best pH range for both the scalp and the hair fiber's health.

Title: Evaluation of pH of Bathing Soaps and Shampoos for Skin and Hair Care

Authors: Tarun J, Susan J, Suria J, Susan VJ, Criton S.

Journal: Indian J Dermatol. 2014 Sep;59(5):442-4. doi: 10.4103/0019-5154.139861. PMID: 25284846; PMCID: PMC4171909.

Link to full text: Evaluation of pH of Bathing Soaps and Shampoos for Skin and Hair Care

Abstract: Background: Normal healthy skin has potential of hydrogen (pH) range of 5.4-5.9 and a normal bacterial flora. Use of soap with high pH causes an increase in skin pH, which in turn causes an increase in dehydrative effect, irritability and alteration in bacterial flora. The majority of soaps and shampoos available in the market do not disclose their pH.

Aims and objectives: The aim of this study was to assess the pH of different brands of bathing soaps and shampoos available in the market.

Materials and methods: The samples of soaps and shampoos were collected from shops in the locality. The samples of different brands are coded before the analysis of the pH. Solution of each sample was made and pH was measured using pH meter.

Results: Majority of the soaps have a pH within the range of 9-10. Majority of the shampoos have a pH within the range of 6-7.

Conclusions: The soaps and shampoos commonly used by the population at large have a pH outside the range of normal skin and hair pH values. Therefore, it is hoped that before recommending soap to patient especially those who have sensitive and acne prone skin, due consideration is given to the pH factor and also that manufacturers will give a thought to pH of soaps and shampoos manufactured by them, so that their products will be more skin and hair friendly.

Title: Effects of carboxytherapy on skin laxity

Authors: Oliveira SMD, Rocha LB, da Cunha MTR, Cintra MMM, Pinheiro NM, Mendonça AC.

Journal: J Cosmet Dermatol. 2020 Nov;19(11):3007-3013. doi: 10.1111/jocd.13337. Epub 2020 Feb 24. PMID: 32091181.

Link to PubMed: Effects of carboxytherapy on skin laxity

Abstract: Background: Carboxytherapy is capable of inducing tissue repair which results in an increase in elastic and collagen fibers.

Objectives: To evaluate the effects of carboxytherapy upon human skin collagen and elastic fiber synthesis.

Methods: Case study of nine volunteers who received a single application of carboxytherapy in the left infraumbilical region, with infusion rate of 100 mL/min and 0.6 mL/kg weight over an area of 25 cm2 . After 60 days on average, the skin was collected for histological analysis and stained with picrosirius red for collagen and Verhoeff for elastic fibers. The percentage of fibers found was marked by the Image J® program and recorded in a Microsoft Excel spreadsheet. Statistical analysis was performed using SPSS (version 20), with a significance level of 95%.

Results: An increase in the collagen and elastic fibers sample was observed in the treated group. Morphometrically, a significant increase in the percentage of collagen in the Carboxytherapy group (41.44 ± 4.50%) was observed compared with the control group (37.44 ± 3.87%) with P = .04; for elastic fibers, the percentage showed no significant difference between the control group (10.55 ± 4.33%) and the carboxytherapy group (10.44 ± 3.71%).

Conclusions: Carboxytherapy with the parameters used in this study was able to stimulate collagen and elastic fiber synthesis, with significant differences in the morphometry for collagen fibers.

Title: CO2 laser physics and tissue interactions in skin

Authors: Fulton JE, Shitabata PK.

Journal: Lasers Surg Med. 1999;24(2):113-21. doi: 10.1002/(sici)1096-9101(1999)24:2<113::aid-lsm6>3.0.co;2-7. PMID: 10100648.

Link to PubMed: CO2 laser physics and tissue interactions in skin

Abstract:
Background and objectives: The theoretical model of CO2 laser tissue interaction appeared to be too simplistic. To explain the reactions seen in skin, a more complex model was needed. We hoped to correlate the clinical-histologic patterns of CO2 laser tissue interactions.

Study design/materials and methods: The Ultrapulse CO2 laser was used on normal and pathologic skin conditions. Clinical observations were correlated with histologic examinations of biopsies.

Results: It was possible to demonstrate cavitation at the dermal-epidermal junction 2-3 diameters beyond the actual spot of CO2 laser contact with the skin. Dermal heat damage was seen as homogenization of collagen 1-2 diameters beyond the spot of laser contact. This flow of energy laterally at the dermal-epidermal junction and vertically down the skin follicles was both clinically beneficial and detrimental. Beneficially, superficial skin lesions separated at this junction and were easily removed. The heat coagulation of the dermis facilitated lesion removal without bleeding. The clinician had a better view of the pathology and could find focal zones of deeper pathology that could be easily re-treated. Detrimentally, this extended damage delayed wound healing and led to persistent erythema.

Conclusion: These clinical-histologic correlations have provided a better understanding of CO2 laser tissue interactions in skin. It has been possible to take advantage of these findings to remove pathologic skin conditions more efficiently.

Title: The Effects on Skin Blood Flow of Immersing the Ischemic Legs of Patients with Peripheral Arterial Disease into Artificially Carbonated Water

Authors: Makita, S., Ohira, A., Naganuma, Y. et al.

Journal: Int J Angiol 15, 12–15 (2006). https://doi.org/10.1007/s00547-006-2063-0

Link to Springer: The Effects on Skin Blood Flow of Immersing the Ischemic Legs of Patients with Peripheral Arterial Disease into Artificially Carbonated Water

Abstract: It is known that carbon dioxide causes vasodilatation. Therefore, immersing the extremities into carbonated water would be expected to increase skin blood flow, given that carbon dioxide can permeate through the skin surface. We immersed the ischemic legs of patients with peripheral arterial disease (PAD) into water artificially carbonated by a new device. Sixteen Fontaine grade II PAD patients were randomly divided into two groups: eight patients whose legs were immersed into artificially carbonated water (CO2 group), and eight patients whose legs were immersed into tap water as a control (CTL group). The water temperature was set at 35°C, and the immersion (10 min) was repeated daily for 3 weeks in both groups. During immersion, the average percent skin blood flow increase was larger in the CO2 group than in the CTL group (426 vs. 173%, p < 0.001), and increased gradually with each immersion in the CO2 group (272% at the first immersion, 343% after 2 weeks, and 359% after 3 weeks, p < 0.01). Leg symptoms at rest, such as a sensation of cold and/or numbness, were significantly alleviated in the CO2 group compared to the CTL group (2.5 points, CO2 group vs. 8.7 points, CTL group after 3-weeks based on a 10-point relative scale, p < 0.01). Immersion into artificially carbonated water appears to be a convenient treatment option for treating limb ischemia.

Title: The Effectiveness of Dry Carbon Dioxide Baths in Menopausal Syndrome: a Randomized Clinical Study

Authors: Anzhela N. Chekhoeva, Georgy E. Zangionov, Alina B. Bugulova, Alan S. Tsogoev, Olga O. Borisevich, Natalya V. Kotenko.

Journal: Вестник восстановительной медицины. 2024;23(4):55-61. doi:10.38025/2078-1962-2024-23-4-55-61

Link to full text: The Effectiveness of Dry Carbon Dioxide Baths in Menopausal Syndrome: a Randomized Clinical Study

Abstract:
INTRODUCTION: Climacteric syndrome is characterized by significant disorders in the nervous, musculoskeletal, urinary and cardiovascular systems in women of the perimenopausal and postmenopausal periods. The main reason is a sharp decrease in estrogen levels. Menopausal hormone therapy is often used for treatment, but due to contraindications and complications, it does not always completely solve the problem. This has sparked interest in alternative treatments such as dry carbon dioxide baths.

AIM: To study the effectiveness of using dry carbon dioxide baths in women with menopausal syndrome and evaluate their impact on the neurovegetative, metabolic and psycho-emotional state of these patients.

MATERIALS AND METHODS: A prospective randomized study was conducted involving 70 patients with moderate climacteric syndrome aged from 48 to 60 years. Using a simple randomization method, the women were divided into 2 groups: 35 patients of the main group received 10 procedures of dry carbon dioxide baths with a CO2 concentration of 15–20 %, temperature 28–32 °C, lasting 15–20 minutes, 35 patients of the control group did not receive any treatment. To assess the effectiveness of treatment, a modified Kupperman-Uvarova menopausal index, an assessment of the effect of “hot flashes” on daily life on the HFRDIS scale, as well as daily blood pressure monitoring (SMAD) were used.

RESULTS AND DISCUSSION: After treatment, patients in the main group observed a significantly significant (p < 0.05) improvement in the patients’ condition in the form of a decrease in integral indicators of the modified menopausal index and an assessment of the impact of hot flashes on daily activity on the HFRDIS scale by 28.2 and 43.1 %, respectively, as well as in the form of a decrease in average daily systolic blood pressure by 13.7 %.

CONCLUSION: The use of dry carbon dioxide baths is recommended for use in women with moderate climacteric syndrome, since this method significantly improves the quality of life of patients and is an effective non-drug that can reduce the drug load on the body of a woman suffering from neurovegetative disorders

Title: Asthma at 8 years of age in children born by caesarean section

Authors: Roduit C, Scholtens S, de Jongste JC, Wijga AH, Gerritsen J, Postma DS, Brunekreef B, Hoekstra MO, Aalberse R, Smit HA.

Journal: Thorax. 2009 Feb;64(2):107-13. doi: 10.1136/thx.2008.100875. Epub 2008 Dec 3. PMID: 19052046.

Link to PubMed: Asthma at 8 years of age in children born by caesarean section

Abstract: Background: Caesarean section might be a risk factor for asthma because of delayed microbial colonisation, but the association remains controversial. A study was undertaken to investigate prospectively whether children born by caesarean section are more at risk of having asthma in childhood and sensitisation at the age of 8 years, taking into account the allergic status of the parents. Methods: 2917 children who participated in a birth cohort study were followed for 8 years. The definition of asthma included wheeze, dyspnoea and prescription of inhaled steroids. In a subgroup (n = 1454), serum IgE antibodies for inhalant and food allergens were measured at 8 years. Results: In the total study population, 12.4% (n = 362) of the children had asthma at the age of 8 years. Caesarean section, with a total prevalence of 8.5%, was associated with an increased risk of asthma (OR 1.79; 95% CI 1.27 to 2.51). This association was stronger among predisposed children (with two allergic parents: OR 2.91; 95% CI 1.20 to 7.05; with only one: OR 1.86; 95% CI 1.12 to 3.09) than in children with non-allergic parents (OR 1.36; 95% CI 0.77 to 2.42). The association between caesarean section and sensitisation at the age of 8 years was significant only in children of non-allergic parents (OR 2.14; 95% CI 1.16 to 3.98). Conclusions: Children born by caesarean section have a higher risk of asthma than those born by vaginal delivery, particularly children of allergic parents. Caesarean section increases the risk for sensitisation to common allergens in children with non-allergic parents only.

Title: Physiologic Changes in the Airway and the Respiratory System Affecting Management in Pregnancy

Authors: Izakson A, Cohen Y, Landau R.

Journal: Principles and practice of maternal critical care (2020): 271-283.

Link to Springer: Physiologic Changes in the Airway and the Respiratory System Affecting Management in Pregnancy

Abstract: Pregnancy is associated with significant anatomical and physiological changes in the airway and respiratory system. Reduction of functional residual capacity in parallel with increased oxygen consumption shortens the time available for airway manipulation before hypoxia becomes significant. Hormone-induced changes in respiratory drive cause a reduction of normal partial pressure of arterial carbon dioxide values during pregnancy, associated with compensatory metabolic acidosis. A thorough understanding of these changes is key for anesthesiologists and critical care providers managing obstetric patients with compromised oxygenation requiring airway manipulation and mechanical ventilation.

Title: Maternal carbon dioxide level during labor and its possible effect on fetal cerebral oxygenation: mini review

Authors: Tomimatsu T, Kakigano A, Mimura K, Kanayama T, Koyama S, Fujita S, Taniguchi Y, Kanagawa T, Kimura T.

Journal: J Obstet Gynaecol Res. 2013 Jan;39(1):1-6. doi: 10.1111/j.1447-0756.2012.01944.x. Epub 2012 Jul 6. PMID: 22765270.

Link to PubMed: Maternal carbon dioxide level during labor and its possible effect on fetal cerebral oxygenation: mini review

Abstract: During pregnancy, and especially during labor, the maternal carbon dioxide level declines considerably. Maternal carbon dioxide levels show a close relation with fetal carbon dioxide levels. The latter affects fetal cerebral oxygenation by regulating cerebral blood flow and shifting the oxyhemoglobin dissociation curve. In addition, maternal hypocapnia appears to impair placental oxygen transfer. Thus, maternal hyperventilation may interfere with optimal fetal cerebral oxygenation. Here, we provide a brief overview of the literature relevant to this issue.

Title: The effect of hyperventilation on maternal placental blood flow in pregnant rabbits

Authors: Leduc B.

Journal: J Physiol. 1972 Sep;225(2):339-48. doi: 10.1113/jphysiol.1972.sp009943. PMID: 4561482; PMCID: PMC1331109.

Link to full text: The effect of hyperventilation on maternal placental blood flow in pregnant rabbits

Abstract: 1. In anaesthetized pregnant rabbits near term, cardiac output and its distribution were measured by injection of isotope-labelled microspheres. Hypocapnia (mean arterial P(CO) (2) 18 mm Hg), induced by intermittent positive pressure hyperventilation, caused a 43% reduction in maternal placental blood flow, attributed mainly to vasoconstriction. Myometrial flow was not significantly changed.2. Moderate hypercapnia (mean arterial P(CO) (2) 46 mm Hg) caused no change in placental flow, compared with observations made while breathing air spontaneously (P(CO) (2) 31 mm Hg).3. Intravenous infusions of adrenaline or noradrenaline 1 mug/kg. min caused maternal placental vasoconstriction.4. During the especially warm summer of 1969, there was a mean 46% reduction in maternal placental blood flow in pregnant rabbits near term, breathing room air spontaneously with normal blood gas values and rectal temperatures. This was associated with an increase in the number of runts and dead foetuses.

Title: The effects of pregnancy on nasal physiology

Authors: Demir UL, Demir BC, Oztosun E, Uyaniklar OO, Ocakoglu G.

Journal: Int Forum Allergy Rhinol. 2015 Feb;5(2):162-6. doi: 10.1002/alr.21438. Epub 2014 Oct 27. PMID: 25348597.

Link to PubMed: The effects of pregnancy on nasal physiology

Abstract: Background: Nasal congestion that is not present before pregnancy represents a distinct clinical entity called pregnancy rhinitis. The aim of this study is to evaluate the clinical characteristics of nasal physiology over the course of pregnancy. Methods: The study was conducted with 85 pregnant women and 26 nonpregnant controls. We measured nasal airway patency objectively via acoustic rhinometry (ARM) and anterior rhinomanometry (RMM) and subjectively via the Nasal Obstruction Symptom Evaluation (NOSE) scale in each trimester and compared the results to those of the controls. Results: The NOSE scores of control and pregnant women showed no difference (p = 0.866). Minimal cross-sectional area (MCA1; minimal cross sectional area at nasal valve and MCA2; minimal cross sectional area at the level where the head of inferior turbinate is placed) decreased significantly between the first and third trimesters: first trimester 0.37 cm(2), third trimester 0.31 cm(2). There was no difference between each trimester with regard to total nasal resistance. The correlation analysis between the NOSE score and both total volume and MCA1 in all patients showed no significance (r = -0.10, p = 0.318; r = -0.04, p = 0.654, respectively). Conclusion: Pregnancy affects nasal physiology adversely and impairs nasal breathing in some women. However, based on the findings of this study, we concluded that this clinical entity may not be considered as a disease without complementary symptoms despite the presence of objective changes in nasal parameters. Keywords: acoustic rhinometry; nasal symptoms; pregnancy; rhinitis; rhinomanometry.

Title: Effects of human pregnancy on the ventilatory chemoreflex response to carbon dioxide

Authors: Jensen D, Wolfe LA, Slatkovska L, Webb KA, Davies GA, O'Donnell DE.

Journal: Am J Physiol Regul Integr Comp Physiol. 2005 May;288(5):R1369-75. doi: 10.1152/ajpregu.00862.2004. Epub 2005 Jan 27. PMID: 15677521.

Link to full text: Effects of human pregnancy on the ventilatory chemoreflex response to carbon dioxide

Abstract: This study examined the effects of human pregnancy on the central chemoreflex control of breathing. Subjects were two groups (n=11) of pregnant subjects (PG, gestational age, 36.5+/-0.4 wk) and nonpregnant control subjects (CG), equated for mean age, body height, prepregnant body mass, parity, and aerobic fitness. All subjects performed a hyperoxic CO2 rebreathing procedure, which includes prior hyperventilation and maintenance of iso-oxia. Resting blood gases and plasma progesterone and estradiol concentrations were measured. During rebreathing trials, end-tidal Pco2 increased, whereas end-tidal Po2 was maintained at a constant hyperoxic level. The point at which ventilation (Ve) began to rise as end-tidal Pco2 increased was identified as the central chemoreflex ventilatory recruitment threshold for CO2 (VRTco2). Ve levels below (basal Ve) and above (central chemoreflex sensitivity) the VRTco2 were determined. The VRTco2 was significantly lower in the PG vs. CG (40.5+/-0.8 vs. 45.8+/-1.6 Torr), and both basal Ve (14.8+/-1.1 vs. 9.3+/-1.6 l/min) and central chemoreflex sensitivity (5.07+/-0.74 vs. 3.16+/-0.29 l.min-1.Torr-1) were significantly higher in the PG vs. CG. Pooled data from the two groups showed significant correlations for resting arterial Pco2 with basal Ve, central chemoreflex sensitivity, and the VRTco2. The VRTco2 was also correlated with progesterone and estradiol concentrations. These data support the hypothesis that pregnancy decreases the threshold and increases the sensitivity of the central chemoreflex response to CO2. These changes may be due to the effects of gestational hormones on chemoreflex and/or nonchemoreflex drives to breathe.

Title: Maternal hyperventilation and the fetus

Authors: Huch R.

Journal: J Perinat Med. 1986;14(1):3-17. doi: 10.1515/jpme.1986.14.1.3. PMID: 3517286.

Link to PubMed: Maternal hyperventilation and the fetus

Abstract: Pregnant women experience hyperventilation during pregnancy for several reasons: It occurs regularly and spontaneously during pregnancy, it occurs because of the type of ventilation practiced during the actual hours of labor and delivery, and sometimes it is induced by the anesthesiologic technique during obstetrical operations. This often results in excessive hyperventilation and has significant effects on blood gases, the cardiovascular and neuro-psychometrical systems of the female organism.

Title: Phasic menstrual cycle effects on the control of breathing in healthy women

Authors: Slatkovska L, Jensen D, Davies GA, Wolfe LA.

Journal: Respir Physiol Neurobiol. 2006 Dec;154(3):379-88. doi: 10.1016/j.resp.2006.01.011. Epub 2006 Mar 15. PMID: 16542884.

Link to PubMed: Phasic menstrual cycle effects on the control of breathing in healthy women

Abstract: This study examined the effects of menstrual cycle phase on ventilatory control. Fourteen eumenorrheic women were studied in the early follicular (FP; 1-6 days) and mid-luteal (LP; 20-24 days) phase of the menstrual cycle. Blood for the determination of arterial PCO(2) (PaCO(2)) , plasma strong ion difference ([SID]), progesterone ([P(4)]), and 17beta-estradiol ([E(2)]) concentrations were obtained at rest. Subjects performed a CO(2) rebreathing procedure that included prior hyperventilation and maintenance of iso-oxia to evaluate central and peripheral chemoreflex, and nonchemoreflex drives to breathe. Resting PaCO(2) and [SID] were lower; minute ventilation (V (E)), [P(4)] and [E(2)] were higher in the LP versus FP. Within the LP, significant correlations were observed for PaCO(2) with [P(4)], [E(2)] and [SID]. Menstrual cycle phase had no effect on the threshold or sensitivity of the central and/or peripheral ventilatory chemoreflex response to CO(2). Both (V (E)) and the ventilatory response to hypocapnia (representing nonchemoreflex drives to breathe) were approximately 1L/min greater in the LP versus FP accounting for the reduction in PaCO(2) . These data support the hypothesis that phasic menstrual cycle changes in PaCO(2) may be due, at least in part, to the stimulatory effects of [P(4)], [E(2)] and [SID] on ventilatory drive.

Title: Nasal congestion during pregnancy

Authors: Ellegård E, Karlsson G.

Journal: Clin Otolaryngol Allied Sci. 1999 Aug;24(4):307-11. doi: 10.1046/j.1365-2273.1999.00264.x. PMID: 10472465.

Link to PubMed: Nasal congestion during pregnancy

Abstract: We define pregnancy rhinitis as nasal congestion in the last 6 or more weeks of pregnancy without other signs of respiratory tract infection and with no known allergic cause, disappearing completely within 2 weeks after delivery. In order to describe physiological variations of nasal obstruction during pregnancy, subjective scores and nasal as well as oral peak expiratory flow values were recorded daily in 23 pregnancies until 1 month after delivery. Scores were higher during early and late pregnancy than in the month after delivery. Objectively registered blockage increased during pregnancy in eight women only. Unexpectedly nine women showed declining blockage. Five of 23 women had pregnancy rhinitis.

Title: Nasal Congestion and Its Management in Pregnancy Rhinitis

Authors: Poerbonegoro N.L.

Journal: Indonesian Journal of Obstetrics and Gynecology (2019): 320-328.

Link to full text: Nasal Congestion and Its Management in Pregnancy Rhinitis

Abstract: Background: Pregnancy rhinitis occurs approximately in one-fifth of pregnancies, at almost any gestational week. The incidence rate of pregnancy rhinitis reaches up to 40%, with prevalence as high as 17%. Pathomechanism is still unclear, but it is suspected that estrogen and placental growth hormone (PGH) play roles in the development of disease. Objective: To elaborate the pathomechanism of pregnancy rhinitis and the proper management of rhinitis symptoms, particularly nasal obstruction. Methods: Literature review. Conclusion: Pregnancy rhinitis, manifested as nasal congestion, is considered a phenomenon and may become a serious condition. Persistent nasal congestion acts as a potential risk factor in affecting fetal growth and development through gradual hypoxia process. This condition can lead to various complications such as maternal hypertension, preeclampsia, impaired fetal growth, and low APGAR scores. Indepth knowledge of pathomechanism is essential as guidance to accurate treatment including conservative and pharmaca therapies, which will lead to optimal outcome for both mother and baby.

Title: Nasal lavage in pregnant women with seasonal allergic rhinitis: a randomized study

Authors: Garavello W, Somigliana E, Acaia B, Gaini L, Pignataro L, Gaini RM.

Journal: Int Arch Allergy Immunol. 2010;151(2):137-41. doi: 10.1159/000236003. Epub 2009 Sep 15. PMID: 19752567.

Link to PubMed: Nasal lavage in pregnant women with seasonal allergic rhinitis: a randomized study

Abstract: Background: Nasal rinsing appears particularly suitable in the management of pregnant women with seasonal allergic rhinitis since no deleterious effects on the fetus are to be expected. However, to date, no studies have specifically investigated this option. Methods: Pregnant women with seasonal allergic rhinitis were randomized to intranasal lavage with hypertonic saline solution 3 times daily (n = 22) versus no local therapy (n = 23) during a 6-week period corresponding to the pollen season. Patients were invited to keep a daily record of rhinitis symptoms (rhinorrea, obstruction, nasal itching and sneezing), to record consumption of oral antihistamine and to undergo rhinomanometry. Results: The rhinitis score was similar at study entry but a statistically significant improvement in this score was observed in the study group during all subsequent weeks (p < 0.001 for weeks 2-6). The mean number of daily antihistamines use per patient per week was significantly reduced at weeks 2, 3 and 6 (p < 0.001, p < 0.001 and p = 0.001, respectively). Baseline rhinomanometry performed at week 1 showed similar nasal resistance in the study and control groups. In contrast, a statistically significant difference emerged in the 2 following evaluations. At week 3, nasal resistance in the study and control groups was 0.96 +/- 0.44 and 1.38 +/- 0.52 Pa/ml/s, respectively (p = 0.006). At week 6, it was 0.94 +/- 0.38 and 1.35 +/- 0.60 Pa/ml/s, respectively (p = 0.006). No adverse effect was reported in the active group. Conclusions: Nasal rinsing is a safe and effective treatment option in pregnant women with seasonal allergic rhinitis.

Title: Oxygen and oxidative stress in the perinatal period

Authors: Torres-Cuevas I, Parra-Llorca A, Sánchez-Illana A, Nuñez-Ramiro A, Kuligowski J, Cháfer-Pericás C, Cernada M, Escobar J, Vento M.

Journal: Redox Biol. 2017 Aug;12:674-681. doi: 10.1016/j.redox.2017.03.011. Epub 2017 Mar 12. PMID: 28395175; PMCID: PMC5388914.

Link to full text: Oxygen and oxidative stress in the perinatal period

Abstract: Fetal life evolves in a hypoxic environment. Changes in the oxygen content in utero caused by conditions such as pre-eclampsia or type I diabetes or by oxygen supplementation to the mother lead to increased free radical production and correlate with perinatal outcomes. In the fetal-to-neonatal transition asphyxia is characterized by intermittent periods of hypoxia ischemia that may evolve to hypoxic ischemic encephalopathy associated with neurocognitive, motor, and neurosensorial impairment. Free radicals generated upon reoxygenation may notably increase brain damage. Hence, clinical trials have shown that the use of 100% oxygen given with positive pressure in the airways of the newborn infant during resuscitation causes more oxidative stress than using air, and increases mortality. Preterm infants are endowed with an immature lung and antioxidant system. Clinical stabilization of preterm infants after birth frequently requires positive pressure ventilation with a gas admixture that contains oxygen to achieve a normal heart rate and arterial oxygen saturation. In randomized controlled trials the use high oxygen concentrations (90% to 100%) has caused more oxidative stress and clinical complications that the use of lower oxygen concentrations (30-60%). A correlation between the amount of oxygen received during resuscitation and the level of biomarkers of oxidative stress and clinical outcomes was established. Thus, based on clinical outcomes and analytical results of oxidative stress biomarkers relevant changes were introduced in the resuscitation policies. However, it should be underscored that analysis of oxidative stress biomarkers in biofluids has only been used in experimental and clinical research but not in clinical routine. The complexity of the technical procedures, lack of automation, and cost of these determinations have hindered the routine use of biomarkers in the clinical setting. Overcoming these technical and economical difficulties constitutes a challenge for the immediate future since accurate evaluation of oxidative stress would contribute to improve the quality of care of our neonatal patients. Keywords: Biomarkers; High-risk pregnancy; Ischemia-reperfusion; Newborn; Oxidative stress; Oxygen.

Title: Respiratory physiology of pregnancy: Physiology masterclass

Authors: LoMauro A, Aliverti A.

Journal: Breathe (Sheff). 2015 Dec;11(4):297-301. doi: 10.1183/20734735.008615. PMID: 27066123; PMCID: PMC4818213.

Link to full text: Respiratory physiology of pregnancy: Physiology masterclass

Abstract: During healthy pregnancy, pulmonary function, ventilatory pattern and gas exchange are affected through both biochemical and mechanical pathways, as summarised in figure 1. During pregnancy, the physiological alteration of hormonal patterns is the main cause of ventilatory changes in respiratory function.

Title: Respiratory and Acid-Base changes during pregnancy

Authors: Prowse C.M, Gaensler E.A.

Journal: Anesthesiology. 1965 Jul-Aug;26:381-92. doi: 10.1097/00000542-196507000-00003. PMID: 14313450.

Link to PubMed: Respiratory and Acid-Base changes during pregnancy

Abstract: Much of our knowledge of respiratory and acid-base changes in pregnancy stems from studies made more than ten years ago and even that material is limited and incomplete. This is surprising when one considers that pregnancy constitutes one of the most severe states of physiologic adaptations. Incongruously, the effect upon lung function of a number of exceedingly rare diseases has been studied exhaustively from every aspect, and by the most modern techniques, whereas the effect of pregnancy, where we are confronted with an ever-growing and inexhaustible supply of clinical material, has been explored so incompletely. Actually, more attention has been paid to states of pathologic stress upon pregnancy such as heart disease, tuberculosis and following chest surgery than to the physiologic norm.

Title: Respiratory physiologic changes in pregnancy

Authors: Wise RA, Polito AJ, Krishnan V.

Journal: Immunol Allergy Clin North Am. 2006 Feb;26(1):1-12. doi: 10.1016/j.iac.2005.10.004. PMID: 16443140.

Link to PubMed: Respiratory physiologic changes in pregnancy

Abstract: In summary, the major physiologic changes that occur in pregnancy are the increased minute ventilation, which is caused by increased respiratory center sensitivity and drive; a compensated respiratory alkalosis; and a low expiratory reserve volume. The vital capacity and measures of forced expiration are well preserved. Patients who have many lung diseases tolerate pregnancy well, with the exception of those who have pulmonary hypertension or chronic respiratory insufficiency from parenchymal or neuromuscular disease.

Title: Sleep-disordered breathing in pregnancy

Authors: Balserak BI.

Journal: Am J Respir Crit Care Med. 2014 Aug 15;190(4):P1-2. doi: 10.1164/rccm.1904P1. PMID: 25127314.

Link to PubMed: Sleep-disordered breathing in pregnancy

Abstract: Sleep disordered breathing (SDB) is very common during pregnancy, and is most likely explained by hormonal, physiological and physical changes. Maternal obesity, one of the major risk factors for SDB, together with physiological changes in pregnancy may predispose women to develop SDB. SDB has been associated with poor maternal and fetal outcomes. Thus, early identification, diagnosis and treatment of SDB are important in pregnancy. This article reviews the pregnancyrelated changes affecting the severity of SDB, the epidemiology and the risk factors of SDB in pregnancy, the association of SDB with adverse pregnancy outcomes, and screening and management options specific for this population

Title: The effect on the fetus of maternal hyperventilation during labour

Authors: Saling E, Ligdas P.

Journal: J Obstet Gynaecol Br Commonw. 1969 Oct;76(10):877-80. doi: 10.1111/j.1471-0528.1969.tb15724.x. PMID: 5386830.

Link to PubMed: The effect on the fetus of maternal hyperventilation during labour

Abstract: ONLY a few years ago it was difficult for midwives and obstetricians to imagine that an increase in the frequency and depth of respiration of women in labour might be too great and constitute a hazard for the fetus. It is still the practice in many places to encourage as much as possible deep breathing during labour so that "a good oxygen supply is ensured for the baby", although such a policy was proved wrong many years ago. It is also suggested that the majority of women in labour are tense and cannot relax because they are afraid of pain, and that, therefore, they do not breathe properly. Whilst tension is not rare during labour, the assumption that this is always associated with an inadequate breathing technique is erroneous. Hasselbach (1912) showed that pregnant women have a decreased blood carbon dioxide tension (Pco2) during pregnancy, and later studies (Sjosted, 1962) proved that this also applied to women in labour. An increase in blood progesterone levels is thought to account for hyperventilation.

Title: Blood flow and oxygenation of tumors in mice. I. Effects of breathing gases containing carbon dioxide at atmospheric pressure

Authors: Kruuv JA, Inch WR, McCredie JA.

Journal: Cancer. 1967 Jan;20(1):51-9. doi: 10.1002/1097-0142(1967)20:1<51::aid-cncr2820200108>3.0.co;2-d. PMID: 6016872.

Link to PubMed: Blood flow and oxygenation of tumors in mice. I. Effects of breathing gases containing carbon dioxide at atmospheric pressure

Abstract: Inhalation of 5% carbon dioxide (CO,) and 95% oxygen (0,)or 10% CO, and90% 0, at one a t m pressure increased blood flow of the C3HBA isotransplanta n d C3H isoimplant in C3H mice compared with animals breathing 100% 0,or air. Oxygen concentration, measured b y a platinum microelectrode, was im-proved in anoxic areas of the tumor when the animals inhaled the gases con-taining CO, compared with those breathing 100% 0, or air; the relative increasewas less when the electrode was placed in a well-vascularized area. Rectal tem-perature, heart a n d respiration rates were decreased in animals breathing thegases containing CO, compared with air; rectal temperature was increased b yinhalation of 100% 0,. The results suggest that inhalation of a gas mixturecontaining CO, a t atmospheric pressure during radiotherapy or chemotherapymay be of value i n treating cancer in humans

Title: Transcutaneous application of carbon dioxide (CO2) induces mitochondrial apoptosis in human malignant fibrous histiocytoma in vivo

Authors: Onishi Y, Kawamoto T, Ueha T, Kishimoto K, Hara H, Fukase N, Toda M, Harada R, Minoda M, Sakai Y, Miwa M, Kurosaka M, Akisue T.

Journal: PLoS One. 2012;7(11):e49189. doi: 10.1371/journal.pone.0049189. Epub 2012 Nov 15. PMID: 23166610; PMCID: PMC3499556.

Link to full text: Transcutaneous application of carbon dioxide (CO2) induces mitochondrial apoptosis in human malignant fibrous histiocytoma in vivo

Abstract: Mitochondria play an essential role in cellular energy metabolism and apoptosis. Previous studies have demonstrated that decreased mitochondrial biogenesis is associated with cancer progression. In mitochondrial biogenesis, peroxisome proliferator-activated receptor gamma coactivator-1 alpha (PGC-1α) regulates the activities of multiple nuclear receptors and transcription factors involved in mitochondrial proliferation. Previously, we showed that overexpression of PGC-1α leads to mitochondrial proliferation and induces apoptosis in human malignant fibrous histiocytoma (MFH) cells in vitro. We also demonstrated that transcutaneous application of carbon dioxide (CO(2)) to rat skeletal muscle induces PGC-1α expression and causes an increase in mitochondrial proliferation. In this study, we utilized a murine model of human MFH to determine the effect of transcutaneous CO(2)) exposure on PGC-1α expression, mitochondrial proliferation and cellular apoptosis. PGC-1α expression was evaluated by quantitative real-time PCR, while mitochondrial proliferation was assessed by immunofluorescence staining and the relative copy number of mitochondrial DNA (mtDNA) was assessed by real-time PCR. Immunofluorescence staining and DNA fragmentation assays were used to examine mitochondrial apoptosis. We also evaluated the expression of mitochondrial apoptosis related proteins, such as caspases, cytochorome c and Bax, by immunoblot analysis. We show that transcutaneous application of CO(2)) induces PGC-1α expression, and increases mitochondrial proliferation and apoptosis of tumor cells, significantly reducing tumor volume. Proteins involved in the mitochondrial apoptotic cascade, including caspase 3 and caspase 9, were elevated in CO(2)) treated tumors compared to control. We also observed an enrichment of cytochrome c in the cytoplasmic fraction and Bax protein in the mitochondrial fraction of CO(2)) treated tumors, highlighting the involvement of mitochondria in apoptosis. These data indicate that transcutaneous application of CO(2)) may represent a novel therapeutic tool in the treatment of human MFH.

Title: Transcutaneous carbon dioxide application suppresses bone destruction caused by breast cancer metastasis

Authors: Takemori T, Kawamoto T, Ueha T, Toda M, Morishita M, Kamata E, Fukase N, Hara H, Fujiwara S, Niikura T, Kuroda R, Akisue T.

Journal: Oncol Rep. 2018 Oct;40(4):2079-2087. doi: 10.3892/or.2018.6608. Epub 2018 Jul 30. PMID: 30066936.

Link to full text: Transcutaneous carbon dioxide application suppresses bone destruction caused by breast cancer metastasis

Abstract: Hypoxia plays a significant role in cancer progression, including metastatic bone tumors. We previously reported that transcutaneous carbon dioxide (CO2) application could decrease tumor progression through the improvement of intratumor hypoxia. Therefore, we hypothesized that decreased hypoxia using transcutaneous CO2 could suppress progressive bone destruction in cancer metastasis. In the present study, we examined the effects of transcutaneous CO2 application on metastatic bone destruction using an animal model. The human breast cancer cell line MDA-MB-231 was cultured in vitro under three different oxygen conditions, and the effect of altered oxygen conditions on the expression of osteoclast-differentiation and osteolytic factors was assessed. An in vivo bone metastatic model of human breast cancer was created by intramedullary implantation of MDA-MB-231 cells into the tibia of nude mice, and treatment with 100% CO2 or a control was performed twice weekly for two weeks. Bone volume of the treated tibia was evaluated by micro-computed tomography (µCT), and following treatment, histological evaluation was performed by hematoxylin and eosin staining and immunohistochemical staining for hypoxia-inducible factor (HIF)-1α, osteoclast-differentiation and osteolytic factors, and tartrate-resistant acid phosphatase (TRAP) staining for osteoclast activity. In vitro experiments revealed that the mRNA expression of RANKL, PTHrP and IL-8 was significantly increased under hypoxic conditions and was subsequently reduced by reoxygenation. In vivo results by µCT revealed that bone destruction was suppressed by transcutaneous CO2, and that the expression of osteoclast-differentiation and osteolytic factors, as well as HIF-1α, was decreased in CO2-treated tumor tissues. In addition, multinucleated TRAP-positive osteoclasts were significantly decreased in CO2-treated tumor tissues. Hypoxic conditions promoted bone destruction in breast cancer metastasis, and reversal of hypoxia by transcutaneous CO2 application significantly inhibited metastatic bone destruction along with decreased osteoclast activity. The findings in this study strongly indicated that transcutaneous CO2 application could be a novel therapeutic strategy for treating metastatic bone destruction.

Title: Optimization of antitumor treatment conditions for transcutaneous CO2 application: An in vivo study

Authors: Ueha T, Kawamoto T, Onishi Y, Harada R, Minoda M, Toda M, Hara H, Fukase N, Kurosaka M, Kuroda R, Akisue T, Sakai Y.

Journal: Oncol Rep. 2017 Jun;37(6):3688-3694. doi: 10.3892/or.2017.5591. Epub 2017 Apr 20. PMID: 28440513.

Link to full text: Optimization of antitumor treatment conditions for transcutaneous CO2 application: An in vivo study

Abstract: Carbon dioxide (CO2) therapy can be applied to treat a variety of disorders. We previously found that transcutaneous application of CO2 with a hydrogel decreased the tumor volume of several types of tumors and induced apoptosis via the mitochondrial pathway. However, only one condition of treatment intensity has been tested. For widespread application in clinical antitumor therapy, the conditions must be optimized. In the present study, we investigated the relationship between the duration, frequency, and treatment interval of transcutaneous CO2 application and antitumor effects in murine xenograft models. Murine xenograft models of three types of human tumors (breast cancer, osteosarcoma, and malignant fibrous histiocytoma/undifferentiated pleomorphic sarcoma) were used to assess the antitumor effects of transcutaneous CO2 application of varying durations, frequencies, and treatment intervals. In all human tumor xenografts, apoptosis was significantly induced by CO2 treatment for ≥10 min, and a significant decrease in tumor volume was observed with CO2 treatments of >5 min. The effect on tumor volume was not dependent on the frequency of CO2 application, i.e., twice or five times per week. However, treatment using 3- and 4-day intervals was more effective at decreasing tumor volume than treatment using 2- and 5-day intervals. The optimal conditions of transcutaneous CO2 application to obtain the best antitumor effect in various tumors were as follows: greater than 10 min per application, twice per week, with 3- and 4-day intervals, and application to the site of the tumor. The results suggest that this novel transcutaneous CO2 application might be useful to treat primary tumors, while mitigating some side effects, and therefore could be safe for clinical trials.

Title: Transcutaneous carbon dioxide induces mitochondrial apoptosis and suppresses metastasis of oral squamous cell carcinoma in vivo

Authors: Takeda D, Hasegawa T, Ueha T, Imai Y, Sakakibara A, Minoda M, Kawamoto T, Minamikawa T, Shibuya Y, Akisue T, Sakai Y, Kurosaka M, Komori T.

Journal: PLoS One. 2014 Jul 2;9(7):e100530. doi: 10.1371/journal.pone.0100530. PMID: 24988190; PMCID: PMC4079455.

Link to full text: Transcutaneous carbon dioxide induces mitochondrial apoptosis and suppresses metastasis of oral squamous cell carcinoma in vivo

Abstract: Squamous cell carcinoma (SCC) is the main histological type of oral cancer. Its growth rate and incidence of metastasis to regional lymph nodes is influenced by various factors, including hypoxic conditions. We have previously reported that transcutaneous CO2 induces mitochondrial apoptosis and decreases lung metastasis by reoxygenating sarcoma cells. However, previous studies have not determined the sequential mechanism by which transcutaneous CO2 suppresses growth of epithelial tumors, including SCCs. Moreover, there is no report that transcutaneous CO2 suppresses lymphogenous metastasis using human cell lines xenografts. In this study, we examined the effects of transcutaneous CO2 on cancer apoptosis and lymphogenous metastasis using human SCC xenografts. Our results showed that transcutaneous CO2 affects expressions of PGC-1α and TFAM and protein levels of cleavage products of caspase-3, caspase-9 and PARP, which relatives mitochondrial apoptosis. They also showed that transcutaneous CO2 significantly inhibits SCC tumor growth and affects expressions of HIF-1α, VEGF, MMP-2 and MMP-9, which play essential roles in tumor angiogenesis, invasion and metastasis. In conclusion, transcutaneous CO2 suppressed tumor growth, increased mitochondrial apoptosis and decreased the number of lymph node metastasis in human SCC by decreasing intra-tumoral hypoxia and suppressing metastatic potential with no observable effect in vivo. Our findings indicate that transcutaneous CO2 could be a novel therapeutic tool for treating human SCC.

Title: Carbogen breathing increases prostate cancer oxygenation: a translational MRI study in murine xenografts and humans

Authors: Alonzi R, Padhani AR, Maxwell RJ, Taylor NJ, Stirling JJ, Wilson JI, d'Arcy JA, Collins DJ, Saunders MI, Hoskin PJ.

Journal: Br J Cancer. 2009 Feb 24;100(4):644-8. doi: 10.1038/sj.bjc.6604903. Epub 2009 Feb 3. PMID: 19190629; PMCID: PMC2653742.

Link to full text: Carbogen breathing increases prostate cancer oxygenation: a translational MRI study in murine xenografts and humans

Abstract: Hypoxia has been associated with poor local tumour control and relapse in many cancer sites, including carcinoma of the prostate. This translational study tests whether breathing carbogen gas improves the oxygenation of human prostate carcinoma xenografts in mice and in human patients with prostate cancer. A total of 23 DU145 tumour-bearing mice, 17 PC3 tumour-bearing mice and 17 human patients with prostate cancer were investigated. Intrinsic susceptibility-weighted MRI was performed before and during a period of carbogen gas breathing. Quantitative R(2)* pixel maps were produced for each tumour and at each time point and changes in R(2)* induced by carbogen were determined. There was a mean reduction in R(2)* of 6.4% (P=0.003) for DU145 xenografts and 5.8% (P=0.007) for PC3 xenografts. In all, 14 human subjects were evaluable; 64% had reductions in tumour R(2)* during carbogen inhalation with a mean reduction of 21.6% (P=0.0005). Decreases in prostate tumour R(2)* in both animal models and human patients as a result of carbogen inhalation suggests the presence of significant hypoxia. The finding that carbogen gas breathing improves prostate tumour oxygenation provides a rationale for testing the radiosensitising effects of combining carbogen gas breathing with radiotherapy in prostate cancer patients.

Title: CO2 bubbling-based 'Nanobomb' System for Targetedly Suppressing Panc-1 Pancreatic Tumor via Low Intensity Ultrasound-activated Inertial Cavitation

Authors: Zhang K, Xu H, Chen H, Jia X, Zheng S, Cai X, Wang R, Mou J, Zheng Y, Shi J.

Journal: Theranostics. 2015 Sep 12;5(11):1291-302. doi: 10.7150/thno.12691. PMID: 26379793; PMCID: PMC4568455.

Link to full text: CO2 bubbling-based 'Nanobomb' System for Targetedly Suppressing Panc-1 Pancreatic Tumor via Low Intensity Ultrasound-activated Inertial Cavitation

Abstract: Noninvasive and targeted physical treatment is still desirable especially for those cancerous patients. Herein, we develop a new physical treatment protocol by employing CO2 bubbling-based 'nanobomb' system consisting of low-intensity ultrasound (1.0 W/cm(2)) and a well-constructed pH/temperature dual-responsive CO2 release system. Depending on the temperature elevation caused by exogenous low-intensity therapeutic ultrasound irradiation and the low pH caused by the endogenous acidic-environment around/within tumor, dual-responsive CO2 release system can quickly release CO2 bubbles, and afterwards, the generated CO2 bubbles waves will timely explode before dissolution due to triggering by therapeutic ultrasound waves. Related bio-effects (e.g., cavitation, mechanical, shock waves, etc) caused by CO2 bubbles' explosion effectively induce instant necrosis of panc-1 cells and blood vessel destruction within panc-1 tumor, and consequently inhibit the growth of panc-1 solid tumor, simultaneously minimizing the side effects to normal organs. This new physiotherapy employing CO2 bubbling-based 'nanobomb' system promises significant potentials in targetedly suppressing tumors, especially for those highly deadly cancers. Keywords: Nanobomb; dual-responsive CO2 release; inertial cavitation; low intensity ultrasound; panc-1 pancreatic cancer.

Title: Intra-arterially infused carbon dioxide-saturated solution for sensitizing the anticancer effect of cisplatin in a rabbit VX2 liver tumor model

Authors: Katayama N, Sugimoto K, Okada T, Ueha T, Sakai Y, Akiyoshi H, Mie K, Ueshima E, Sofue K, Koide Y, Tani R, Gentsu T, Yamaguchi M.

Journal: Int J Oncol. 2017 Aug;51(2):695-701. doi: 10.3892/ijo.2017.4056. Epub 2017 Jun 26. PMID: 28656217.

Link to full text: Intra-arterially infused carbon dioxide-saturated solution for sensitizing the anticancer effect of cisplatin in a rabbit VX2 liver tumor model

Abstract: The present study aimed to evaluate the efficacy of an intra-arterially infused carbon dioxide (CO2)-saturated solution in sensitizing the anticancer effect of cisplatin in a rabbit VX2 liver tumor model. Forty VX2 liver tumor-bearing Japanese white rabbits were randomly divided into four groups and infused via the proper hepatic artery with a saline solution (control group), CO2-saturated solution (CO2 group), cisplatin solution (cisplatin group), or CO2-saturated solution and cisplatin solution (combined group). The tumor volume (TV) and the relative tumor volume (RTV), RTV = (TV on day 3 or 7)/(TV on day 0) x 100, were calculated using contrast-enhanced computed tomography. Hypoxia-inducible factor-1α (HIF‑1α) and carbonic anhydrase IX (CA IX) staining were used to evaluate cellular hypoxia. Cleaved caspase-3 and cleaved caspase-9 were analyzed to assess tumor apoptosis. The mean RTV on days 3 and 7 were 202.6±23.7 and 429.2±94.8%, respectively, in the control group; 172.2±38.1 and 376.5±61.1% in the CO2 group; 156.1±15.1 and 269.6±45.2% in the cisplatin group; and 118.3±28.1 and 210.3±55.1% in the combined group. RTV was significantly lower in the CO2 group than in the control group (day 3; P<0.05), and in the combined group than in the cisplatin group (days 3 and 7; P<0.05). HIF-1α and CA IX suppression, and increased cleaved caspase-3 and cleaved caspase-9 expression, were detected in the CO2 and combined groups, compared with the other two groups. An intra-arterially infused CO2-saturated solution inhibits liver VX2 tumor growth and sensitizes the anticancer effect of cisplatin.

Title: Transcutaneous Carbon Dioxide Decreases Immunosuppressive Factors in Squamous Cell Carcinoma In Vivo

Authors: Yatagai N, Hasegawa T, Amano R, Saito I, Arimoto S, Takeda D, Kakei Y, Akashi M.

Journal: Biomed Res Int. 2021 Jul 2;2021:5568428. doi: 10.1155/2021/5568428. PMID: 34307656; PMCID: PMC8270696.

Link to full text: Transcutaneous Carbon Dioxide Decreases Immunosuppressive Factors in Squamous Cell Carcinoma In Vivo

Abstract: Introduction: In recent years, the tumour immunosuppressive mechanism has attracted attention as a cause of tumour chemoresistance. Although chemoresistance and immunosuppression of tumours have been reported to be associated with a hypoxic environment, effective treatments to improve hypoxia in tumours have not yet been established. We have previously applied carbon dioxide (CO2) to squamous cell carcinoma and have shown that improvement in local oxygenation has an antitumour effect. However, the effects of local CO2 administration on tumour immunosuppression, chemoresistance, and combination with chemotherapy are unknown. In this study, we investigated the effects of local CO2 administration on squamous cell carcinoma and the effects of combined use with chemotherapy, focusing on the effects on tumour immunosuppressive factors. Methods: Human oral squamous cell carcinoma (HSC-3) was transplanted subcutaneously into the back of a nude mouse, and CO2 and cisplatin were administered. After administration twice a week for a total of 4 times, tumours were collected and the expression of tumour immunosuppressive factors (PD-L1, PD-L2, and galectin-9) was evaluated using real-time polymerase chain reaction and immunostaining. Results: Compared with the control group, a significant decrease in the mRNA expression of PD-L1 was observed in both, CO2-treated and combination groups. Similarly, the expression of PD-L2 and galectin-9 decreased in the CO2-treated and combination groups. Furthermore, immunostaining also showed a significant decrease in the protein expression of tumour immunosuppressive factors in the CO2-treated and combination groups. Conclusion: It was confirmed that the tumour immunosuppressive factors decreased due to local CO2 administration to the mouse model. CO2 administration has the potential to improve the hypoxic environment in tumours, and combined use with chemotherapy may also improve tumour immunosuppression.

Title: Reoxygenation using a novel CO2 therapy decreases the metastatic potential of osteosarcoma cells

Authors: Harada R, Kawamoto T, Ueha T, Minoda M, Toda M, Onishi Y, Fukase N, Hara H, Sakai Y, Miwa M, Kuroda R, Kurosaka M, Akisue T.

Journal: Exp Cell Res. 2013 Aug 1;319(13):1988-1997. doi: 10.1016/j.yexcr.2013.05.019. Epub 2013 May 29. PMID: 23727023.

Link to PubMed: Reoxygenation using a novel CO2 therapy decreases the metastatic potential of osteosarcoma cells

Abstract: Osteosarcoma is the most common primary solid malignant bone tumor. Despite substantial improvements in surgery and chemotherapy, metastasis remains a major cause of fatal outcomes, and the molecular mechanisms of metastasis are still poorly understood. Hypoxia, which is common in malignant tumors including osteosarcoma, increases expressions of hypoxia inducible factor (HIF)-1α, matrix metalloproteinase (MMP)-2 and MMP-9, and can induce invasiveness. As we previously showed a novel transcutaneous CO2 application to decrease HIF-1α expression and induce apoptosis in malignant fibrous histiocytoma, we hypothesize that transcutaneous CO2 application could suppress metastatic potential of osteosarcoma by improving hypoxic conditions. Here, we examined the effects of transcutaneous CO2 application on apoptosis, and development of pulmonary metastasis using a highly metastatic osteosarcoma cell line, LM8. Transcutaneous CO2 application significantly decreased tumor growth and pulmonary metastasis in LM8 cells. Apoptotic activity increased, and intratumoral hypoxia was improved with decreased expressions of HIF-1α, MMP-2 and MMP-9, significantly, in the CO2-treated tumors. In conclusion, we found that transcutaneous CO2 application can induce tumor cell apoptosis and might suppress pulmonary metastasis by improvement of hypoxic conditions with decreased expressions of HIF-1α and MMPs in highly metastatic osteosarcoma cell. These findings strongly indicate that this novel transcutaneous CO2 therapy could be a therapeutic breakthrough for osteosarcoma patients.

Keywords: Hypoxia; Matrix metalloproteinase (MMP); Metastasis; Osteosarcoma; Transcutaneous CO(2) application.

Title: Transcutaneous carbon dioxide suppresses skeletal muscle atrophy in a mouse model of oral squamous cell carcinoma

Authors: Sasaki A, Takeda D, Kawai H, Tadokoro Y, Murakami A, Yatagai N, Arimoto S, Nagatsuka H, Akashi M, Hasegawa T.

Journal: PLoS One. 2024 Apr 17;19(4):e0302194. doi: 10.1371/journal.pone.0302194. PMID: 38630690; PMCID: PMC11023300.

Link to full text: Transcutaneous carbon dioxide suppresses skeletal muscle atrophy in a mouse model of oral squamous cell carcinoma

Abstract: Cancer cachexia causes skeletal muscle atrophy, impacting the treatment and prognosis of patients with advanced cancer, but no treatment has yet been established to control cancer cachexia. We demonstrated that transcutaneous application of carbon dioxide (CO2) could improve local blood flow and reduce skeletal muscle atrophy in a fracture model. However, the effects of transcutaneous application of CO2 in cancer-bearing conditions are not yet known. In this study, we calculated fat-free body mass (FFM), defined as the skeletal muscle mass, and evaluated the expression of muscle atrophy markers and uncoupling protein markers as well as the cross-sectional area (CSA) to investigate whether transcutaneous application of CO2 to skeletal muscle could suppress skeletal muscle atrophy in cancer-bearing mice. Human oral squamous cell carcinoma was transplanted subcutaneously into the upper dorsal region of nude mice, and 1 week later, CO2 gas was applied to the legs twice a week for 4 weeks and FFM was calculated by bioimpedance spectroscopy. After the experiment concluded, the quadriceps were extracted, and muscle atrophy markers (muscle atrophy F-box protein (MAFbx), muscle RING-finger protein 1 (MuRF-1)) and uncoupling protein markers (uncoupling protein 2 (UCP2) and uncoupling protein 3 (UCP3)) were evaluated by real-time polymerase chain reaction and immunohistochemical staining, and CSA by hematoxylin and eosin staining. The CO2-treated group exhibited significant mRNA and protein expression inhibition of the four markers. Furthermore, immunohistochemical staining showed decreased MAFbx, MuRF-1, UCP2, and UCP3 in the CO2-treated group. In fact, the CSA in hematoxylin and eosin staining and the FFM revealed significant suppression of skeletal muscle atrophy in the CO2-treated group. We suggest that transcutaneous application of CO2 to skeletal muscle suppresses skeletal muscle atrophy in a mouse model of oral squamous cell carcinoma.

Title: Transcutaneous application of CO2 enhances the antitumor effect of radiation therapy in human malignant fibrous histiocytoma

Authors: Onishi Y, Akisue T, Kawamoto T, Ueha T, Hara H, Toda M, Harada R, Minoda M, Morishita M, Sasaki R, Nishida K, Kuroda R, Kurosaka M.

Journal: Int J Oncol. 2014 Aug;45(2):732-8. doi: 10.3892/ijo.2014.2476. Epub 2014 May 29. PMID: 24889546.

Link to full text: Transcutaneous application of CO2 enhances the antitumor effect of radiation therapy in human malignant fibrous histiocytoma

Abstract: Sarcomas are relatively resistant because of hypoxia. We previously demonstrated that the transcutaneous CO(2) therapy reduced hypoxic conditions in human malignant fibrous histiocytoma (MFH). Therefore, we hypothesized that transcutaneous CO(2) therapy could enhance the antitumor effect of radiation therapy in human MFH. Our purpose was to evaluate the effects of transcutaneous CO(2) therapy on the antitumor efficacy of X-ray irradiation using MFH. First, in an in vitro study, we assessed apoptotic activity and reactive oxygen species (ROS) production using flow cytometric and immunoblot analysis at 24 h after X-ray irradiation under three different oxygen conditions (normoxic, reoxygenated and hypoxic). In addition, in the in vivo study, 24 male athymic BALB/c nude mice with MFH tumors that were inoculated in the dorsal subcutaneous area were randomized into four groups: control, CO(2), X-ray irradiation and combination (CO(2) and X-ray irradiation). Treatments were performed twice weekly for 2 weeks, four times in total. Tumor volume was calculated. All tumors were excised and apoptotic activity, ROS production, related proteins and HIF-1α expression were assessed using flow cytometric and immunoblot analysis. The in vitro study revealed that X-ray irradiation induced increased apoptosis and ROS production in MFH cells under normoxic and reoxygenated conditions relative to hypoxic conditions (P<0.01). In the in vivo study, tumor volume in the combination group was reduced to 28, 42 and 47% of that in the control, CO(2), and X-ray groups, respectively (P<0.05). Apoptotic activity and ROS production in the combination group were strongly increased with decreasing HIF-1α expression relative to the control, CO(2) and X-ray groups. The transcutaneous CO(2) system enhanced the antitumor action of X-ray irradiation and could be a novel therapeutic tool for overcoming radio-resistance in human malignancies.

Title: Perfluorocarbon emulsions radiosensitise brain tumors in carbogen breathing mice with orthotopic GL261 gliomas

Authors: Feldman LA, Fabre MS, Grasso C, Reid D, Broaddus WC, Lanza GM, Spiess BD, Garbow JR, McConnell MJ, Herst PM.

Journal: PLoS One. 2017 Sep 5;12(9):e0184250. doi: 10.1371/journal.pone.0184250. PMID: 28873460; PMCID: PMC5584944.

Link to full text: Perfluorocarbon emulsions radiosensitise brain tumors in carbogen breathing mice with orthotopic GL261 gliomas

Abstract: Background: Tumour hypoxia limits the effectiveness of radiation therapy. Delivering normobaric or hyperbaric oxygen therapy elevates pO2 in both tumour and normal brain tissue. However, pO2 levels return to baseline within 15 minutes of stopping therapy.

Aim: To investigate the effect of perfluorocarbon (PFC) emulsions on hypoxia in subcutaneous and intracranial mouse gliomas and their radiosensitising effect in orthotopic gliomas in mice breathing carbogen (95%O2 and 5%CO2).

Results: PFC emulsions completely abrogated hypoxia in both subcutaneous and intracranial GL261 models and conferred a significant survival advantage orthotopically (Mantel Cox: p = 0.048) in carbogen breathing mice injected intravenously (IV) with PFC emulsions before radiation versus mice receiving radiation alone. Carbogen alone decreased hypoxia levels substantially and conferred a smaller but not statistically significant survival advantage over and above radiation alone.

Conclusion: IV injections of PFC emulsions followed by 1h carbogen breathing, radiosensitises GL261 intracranial tumors.

Title: Gas-Mediated Cancer Bioimaging and Therapy

Authors: Chen L, Zhou SF, Su L, Song J.

Journal: ACS Nano. 2019 Oct 22;13(10):10887-10917. doi: 10.1021/acsnano.9b04954. Epub 2019 Sep 27. PMID: 31538764.

Link to PubMed: Gas-Mediated Cancer Bioimaging and Therapy

Abstract: Gas-involving cancer theranostics have attracted considerable attention in recent years due to their high therapeutic efficacy and biosafety. We have reviewed the recent significant advances in the development of stimuli-responsive gas releasing molecules (GRMs) and gas nanogenerators for cancer bioimaging, targeted and controlled gas therapy, and gas-sensitized synergistic therapy. We have focused on gases with known anticancer effects, such as oxygen (O2), carbon monoxide (CO), nitric oxide (NO), hydrogen sulfide (H2S), hydrogen (H2), sulfur dioxide (SO2), carbon dioxide (CO2), and heavy gases that act via the gas-generating process. The GRMs and gas nanogenerators for each gas have been described in terms of the stimulation method, followed by their applications in ultrasound and multimodal imaging, and finally their primary and synergistic actions with other cancer therapeutic modalities. The current challenges and future possibilities of gas therapy and imaging vis-à-vis clinical translation have also been discussed.

Keywords: cancer imaging; cancer therapy; controlled release; endogenous/exogenous stimulation; gas nanogenerators; gas releasing system; gas therapy; synergetic therapy; theranostic.

Title: Gas-mediated cancer therapy combined with starvation therapy, ultrasound therapy, chemotherapy, radiotherapy, and photodynamic therapy: a review

Authors: Thakar, S. B., Ghorpade, P. N., Shaker, B., Lee, J., & Na, D.

Journal: Environmental Chemistry Letters 19 (2021): 2981-2993.

Link to Springer: Gas-mediated cancer therapy combined with starvation therapy, ultrasound therapy, chemotherapy, radiotherapy, and photodynamic therapy: a review

Abstract: Gas therapy appears promising in treating cancer, ischemia-reperfusion injury, heart attack, and acute situations of shock. Gas therapy is also becoming a good choice in preventive medicine against dementia, reducing cancer occurrence, cardiovascular diseases, and delaying aging. On-demand gas release allows precision therapy with few risks of gas poisoning. Here, we review targeted and controlled gas therapy, gas nanogenerators for cancer bioimaging, and gas-sensitized synergistic therapy. We focus on gases with anticancer effects such as hydrogen, carbon monoxide, carbon dioxide, nitric oxide, oxygen sulfur dioxide, hydrogen sulfide, and heavy gases. We discuss the efficiency of gas therapy combined with other therapies methods such as starvation therapy, ultrasound therapy, chemotherapy, radiotherapy, and photodynamic therapy, and applications in multimodal and ultrasound imaging.

Title: Impact of laparoscopic CO2-insufflation on tumor-associated molecules in cultured colorectal cancer cells

Authors: Kim ZG, Mehl C, Lorenz M, Gutt CN.

Journal: Surg Endosc. 2002 Aug;16(8):1182-6. doi: 10.1007/s00464-001-9194-3. Epub 2002 Apr 9. PMID: 12189480.

Link to PubMed: Impact of laparoscopic CO2-insufflation on tumor-associated molecules in cultured colorectal cancer cells

Abstract: Background: Laparoscopic CO2-insufflation is believed to stimulate proliferation and metastatic potential of gastrointestinal carcinomas. E-cadherin, I-CAM1, I-CAM2, and CD44 are involved into the metastatic process of different cancer cell lines. The current study investigates the influence of CO2-insufflation on the expression of E-cadherin, I-CAM1, I-CAM2, and CD44 in vitro.

Methods: CX-2 and CC531 colon carcinoma cells (human/rat) were exposed to pneumoperitoneal CO2-insufflation. E-cadherin, I-CAM1, I-CAM2, and CD44 were measured 0, 12, 24, 48, and 72 h after CO2-insufflation using flowcytometry. Control groups were exposed to room air. Data were analyzed by the Wilcoxon-Mann-Whitney U-test.

Results: Both cell lines showed significant alteration in E-cadherin, I-CAM1, and CD44 expression after CO2 exposure (p <0.05). No significant differences were found regarding I-CAM2 expression.

Conclusion: The present study demonstrates CO2-insufflation to influence the expression of E-cadherin, I-CAM1 and CD44. Whether these changes increases the metastatic potential of colorectal cancer cells in vivo needs further investigation.

Title: Improved respiratory delivery of the anticancer drugs, camptothecin and paclitaxel, with 5% CO2-enriched air: pharmacokinetic studies

Authors: Koshkina NV, Knight V, Gilbert BE, Golunski E, Roberts L, Waldrep JC.

Journal: Cancer Chemother Pharmacol. 2001 May;47(5):451-6. doi: 10.1007/s002800000230. PMID: 11391862.

Link to PubMed: Improved respiratory delivery of the anticancer drugs, camptothecin and paclitaxel, with 5% CO2-enriched air: pharmacokinetic studies

Abstract: Purpose: To increase pulmonary deposition of anticancer liposome aerosols in mice by modulation of respiratory physiology through the addition of 5% CO2 to the air source used to generate the aerosols. Breathing CO2-enriched aerosol increases pulmonary ventilation with concurrent increased deposition of inhaled particles.

Methods: Dilauroylphosphatidylcholine liposome formulations of two anticancer drugs, paclitaxel (PTX) and camptothecin (CPT), were investigated. The aerosol droplet size was measured using an Andersen cascade impactor. Drug concentrations in aerosol droplet fractions and tissues were determined by HPLC analysis. ICR mice were exposed to each liposome aerosol for 30 min. For each drug, one group of mice inhaled the drug-liposome aerosol generated with a mixture of 5% CO2 in air and another group inhaled the drug-liposome aerosols produced with normal air. Tissue distribution and pharmacokinetics were determined for both drug delivery systems.

Results: Significantly higher concentrations of PTX and CPT were found in organs of mice exposed to 5% CO2-air aerosols compared to organs of mice exposed to normal air aerosols. The highest concentrations of drug were detected in the lungs and were two- to fourfold higher with 5% CO2-air aerosols than with aerosols generated with normal air. Higher concentrations were also detected in liver, spleen, kidneys, blood, and brain.

Conclusion: 5% CO2 enrichment of air increased respiratory tract deposition of inhaled aerosol particles containing PTX and CPT.

Title: Influence of CO2 pneumoperitoneum during laparoscopic surgery on cancer cell growth

Authors: Takiguchi S, Matsuura N, Hamada Y, Taniguchi E, Sekimoto M, Tsujinaka M, Shiozaki H, Monden M, Ohashi S.

Journal: Surg Endosc. 2000 Jan;14(1):41-4. doi: 10.1007/s004649900008. PMID: 10653234.

Link to PubMed: Influence of CO2 pneumoperitoneum during laparoscopic surgery on cancer cell growth

Abstract: Background: CO2 pneumoperitoneum provides a new surgical environment to treat malignant disease. The purpose of this study was to investigate the influence of CO2 pneumoperitoneum during laparoscopic surgery on cancer cell growth.

Methods: WiDr human colon cancer cells were incubated for 3 h under the following two conditions: 100% CO2 at 10 mmHg, and 95% air/5% CO2 (control). Cell proliferation was assessed by the WST-1 assay and BrdU assay. Tumor growth was assessed by subcutaneous injection into 20 nude mice. Cellular damage was measured by lactate dehydrogenase (LDH) assay.

Results: The number of WiDr cells under pneumoperitoneal conditions decreased in the first 24 h. However, no significant difference was observed in the proliferation rate and tumor growth of the viable cells. LDH release of the CO2 pneumoperitoneal group was higher than that of the controls.

Conclusions: Our data indicate that CO2 pneumoperitoneum does not promote cancer cell proliferation but instead has a toxic effect on cancer cells.

Title: Cancer Cell Metabolism in Hypoxia: Role of HIF-1 as Key Regulator and Therapeutic Target

Authors: Infantino V, Santarsiero A, Convertini P, Todisco S, Iacobazzi V.

Journal: Int J Mol Sci. 2021 May 27;22(11):5703. doi: 10.3390/ijms22115703. PMID: 34071836; PMCID: PMC8199012.

Link to full text: Cancer Cell Metabolism in Hypoxia: Role of HIF-1 as Key Regulator and Therapeutic Target

Abstract: In order to meet the high energy demand, a metabolic reprogramming occurs in cancer cells. Its role is crucial in promoting tumor survival. Among the substrates in demand, oxygen is fundamental for bioenergetics. Nevertheless, tumor microenvironment is frequently characterized by low-oxygen conditions. Hypoxia-inducible factor 1 (HIF-1) is a pivotal modulator of the metabolic reprogramming which takes place in hypoxic cancer cells. In the hub of cellular bioenergetics, mitochondria are key players in regulating cellular energy. Therefore, a close crosstalk between mitochondria and HIF-1 underlies the metabolic and functional changes of cancer cells. Noteworthy, HIF-1 represents a promising target for novel cancer therapeutics. In this review, we summarize the molecular mechanisms underlying the interplay between HIF-1 and energetic metabolism, with a focus on mitochondria, of hypoxic cancer cells.

Keywords: HIF-1; TCA cycle; cancer metabolism; hypoxia; mitochondria.

Title: Relationship between tumour growth rate and carbogen-based functional MRI for a chemically induced HCC in mice

Authors: Thomas CD, Chenu E, Walczak C, Plessis MJ, Perin F, Volk A.

Journal: MAGMA. 2004 Dec;17(3-6):271-80. doi: 10.1007/s10334-004-0087-z. Epub 2004 Dec 20. PMID: 15614512.

Link to PubMed: Relationship between tumour growth rate and carbogen-based functional MRI for a chemically induced HCC in mice

Abstract: We previously performed MRI studies of HCC (hepatocellular carcinomas) in mice showing the feasibility of measuring a carbogen effect. In the present study carbogen response of the whole tumour was compared with growth characteristics during longitudinal follow-up. HCC were chemically induced. The imaging protocol at 4.7 T comprised a fast spin-echo sequence for high-resolution screening and measurement of growth curves, and a fast gradient echo sequence allowing an entire T2*w image acquisition per respiratory cycle to perform fMRI under carbogen breathing. A new parameter, T+, the fraction of tumour voxels with increased intensity under carbogen was measured on manually defined ROIs. Twenty-two HCC were followed for 3-10 weeks. Tumours were divided into two groups, "regularly" and "irregularly" growing tumours. A linear correlation between T+ and tumour growth rate was observed only for "regularly" growing HCC. These results suggest a link between tumour growth rates and tumour fractions exhibiting signal increase upon carbogen breathing. They are compatible with observations by others that rapidly growing tumours are more hypoxic than slowly growing ones. Combined measurement of T+ and tumour growth may become a useful noninvasive follow-up approach for assessment and/or management of therapies involving vasculature-targeting and anti-proliferative drugs.

Title: Effect of carbogen breathing on the physiological profile of human glioma xenografts

Authors: van der Sanden BP, Heerschap A, Hoofd L, Simonetti AW, Nicolay K, van der Toorn A, Colier WN, van der Kogel AJ.

Journal: Magn Reson Med. 1999 Sep;42(3):490-9. doi: 10.1002/(sici)1522-2594(199909)42:3<490::aid-mrm11>3.0.co;2-h. PMID: 10467293.

Link to full text: Effect of carbogen breathing on the physiological profile of human glioma xenografts

Abstract: The aim of this study was to evaluate the effect of carbogenbreathing on the physiological profile of human glioma xeno-grafts. Near infrared spectroscopy was used to investigatechanges in oxy- and deoxyhemoglobin concentrations in tumorblood. Oxygen tension changes in tumor tissue were evaluatedby 19F-MR relaxometry, using perfluoro-15-crown-5-ether, andmodifications of tumor blood perfusion (TBP) were analyzed byfast dynamic 1H-MR imaging of Gd-DTPA uptake. Finally, changesof the bioenergetic status and pH of tumor cells were analyzedby 31P-MRS. After 5 to 8 min of carbogen breathing, the averageoxygen tension increase in tumor tissue was 4.6 6 1.3 mm Hg,which is in agreement with an increase of the oxyhemoglobinconcentration in tumor blood (D[O 2Hb] 5 9.2 6 3 mM). However,simultaneously the TBP was reduced, the bioenergetic statuswas diminished, and pH was decreased. As 100% O 2 breathingalone did not result in a detectable increase of oxyhemoglobinin tumor blood, the increase of the tumor oxygenation bycarbogen appears to be mediated by its CO 2 content. Thiscomponent may cause a nutrient-limited decrease of oxidativeenergy metabolism, indirectly via a steal-effect and/or by inhibi-tion of the glycolytic rate resulting from tissue acidification.Magn Reson Med 42:490–499, 1999. r 1999 Wiley-Liss, Inc.Key words: gliomas; carbogen; perfusion; oxygenation; bioen-ergetic status

Title: Effects of a Simulated CO2 Pneumoperitoneum Environment on the Proliferation, Apoptosis, and Metastasis of Cervical Cancer Cells In Vitro

Authors: Lin F, Pan L, Li L, Li D, Mo L.

Journal: Med Sci Monit. 2014 Dec 1;20:2497-503. doi: 10.12659/MSM.891179. PMID: 25436974; PMCID: PMC4260668.

Link to full text: Effects of a Simulated CO2 Pneumoperitoneum Environment on the Proliferation, Apoptosis, and Metastasis of Cervical Cancer Cells In Vitro

Abstract: Background: This study aimed to investigate the growth curve, cell colony formation, cell cycle, apoptosis, anti-anoikis, and ability of invasion, adhesion, and migration of cervical cancer cells after exposure to a model of a simulated CO2 pneumoperitoneum environment with different pressures and at different times.

Material and methods: The cervical cancer cells were cultured in groups with 8 and 16 mmHg of 100% CO2 for 1, 2, 3, and 4 h in a model of a simulated environment of CO2 pneumoperitoneum. The cells in the control group were cultured in a standard environment. The growth curve was drawn through constant survival cell counts for 7 days, and the group with most obvious change was selected for subsequent experiments to detect cell colony formation, cell cycle apoptosis, and anti-anoikis, and the ability of invasion, adhesion, and migration.

Results: After a brief inhibition, the proliferation of cervical cancer cells was markedly increased and had no relationship with different CO2 pressures. Compared with the control group, the early apoptosis rate in the experimental group was higher, and the ability of invasion, migration, and adhesion decreased significantly.

Conclusions: Cervical cancer cells stimulated by a CO2 pneumoperitoneum environment in vitro have an increased the ability to proliferate after a short period of inhibition and have reduced abilities of invasion, migration, and adhesion.

Title: Effects of Fluosol DA 20% and carbogen on the radioresponse of SCK tumors and skin of A/J mice

Authors: Lee I, Levitt SH, Song CW.

Journal: Radiat Res. 1987 Oct;112(1):173-82. PMID: 3116597.

Link to PubMed: Effects of Fluosol DA 20% and carbogen on the radioresponse of SCK tumors and skin of A/J mice

Abstract: The effect of Fluosol DA 20%, an emulsion of perfluorochemicals, in combination with carbogen (95% O2 and 5% CO2) breathing on the response of mouse tumors to radiation was studied. When A/J mice bearing SCK tumors in the right hind limb were injected iv with Fluosol DA 20% at 12 ml/kg and exposed to carbogen for 1 h before and during the irradiation of tumors, the response of tumors to a single dose of X irradiation was significantly enhanced. The dose modification factors for growth delay and cure of SCK tumors were 2.10 +/- 0.01 (SE) and 1.86 +/- 0.18 (SE), respectively. Such a treatment slightly increased the radiation-induced skin damage by a factor of 1.17 +/- 0.02 (SE), resulting in a therapeutic gain of 1.79 +/- 0.01 (SE) for the growth delay and 1.59 +/- 0.09 (SE) for the curability. Carbogen breathing alone also increased the response of tumor and skin to radiation, but it was far less effective than the combination of Fluosol DA 20% and carbogen breathing. It was concluded that iv injection of Fluosol DA 20% in conjunction with carbogen breathing significantly increased the O2 transport to hypoxic areas in the SCK tumors and thus significantly enhanced the tumoricidal effect of radiation on SCK tumors.

Title: Heated and humidified CO₂ pneumoperitoneum inhibits tumour cell proliferation, migration and invasion in colon cancer

Authors: Cai W, Dong F, Wang Z, Yang X, Zheng M, Che X.

Journal: Int J Hyperthermia. 2014 May;30(3):201-9. doi: 10.3109/02656736.2014.898339. Epub 2014 Apr 10. PMID: 24720439.

Link to full text: Heated and humidified CO₂ pneumoperitoneum inhibits tumour cell proliferation, migration and invasion in colon cancer

Abstract: Background: Peritoneal carcinomatosis (PC) arising from colorectal cancer is associated with poor prognosis and few treatment options are currently available. Laparoscopic CO2 insufflation stimulates the progression and metastatic potential of gastrointestinal carcinomas. However, heated and humidified CO2 pneumoperitoneum (HH-CO2) is a promising treatment for PC, although its effects and mechanism of action in human colon cancer cells remain unclear. This study evaluated the anti-tumour effects of HH-CO2 on human colon cancer in vitro.

Methods: Cell viability was assessed using the WST-8 assay in two colon cancer cell lines. Apoptosis was assessed by Annexin V PI flow cytometry, and migration and invasion were examined using wound healing and Transwell® invasion assays. The expressions of Bcl-2, Bax, matrix metalloproteinase-2 (MMP-2), E-cadherin, ICAM-1, and CD44 were detected by western blotting.

Results: HH-CO2 significantly inhibited cell proliferation, migration, invasion and adhesion. HH-CO2 induced apoptosis and significantly inhibited the expression of Bcl-2, MMP-2, ICAM-1 and CD44, and increased Bax and E-cadherin expression in colon cancer cells.

Conclusions: HH-CO2 induces apoptosis and inhibits proliferation, migration, invasion and adhesion of human colon cancer cells. Our results suggest that HH-CO2 may serve as a potential candidate for the treatment and/or prevention of peritoneal carcinomatosis from colorectal cancer and warrant further in vivo investigation.

Title: Hypoxic microenvironment in cancer: molecular mechanisms and therapeutic interventions

Authors: Chen Z, Han F, Du Y, Shi H, Zhou W.

Journal: Signal Transduct Target Ther. 2023 Feb 17;8(1):70. doi: 10.1038/s41392-023-01332-8. PMID: 36797231; PMCID: PMC9935926.

Link to full text: Hypoxic microenvironment in cancer: molecular mechanisms and therapeutic interventions

Abstract: Having a hypoxic microenvironment is a common and salient feature of most solid tumors. Hypoxia has a profound effect on the biological behavior and malignant phenotype of cancer cells, mediates the effects of cancer chemotherapy, radiotherapy, and immunotherapy through complex mechanisms, and is closely associated with poor prognosis in various cancer patients. Accumulating studies have demonstrated that through normalization of the tumor vasculature, nanoparticle carriers and biocarriers can effectively increase the oxygen concentration in the tumor microenvironment, improve drug delivery and the efficacy of radiotherapy. They also increase infiltration of innate and adaptive anti-tumor immune cells to enhance the efficacy of immunotherapy. Furthermore, drugs targeting key genes associated with hypoxia, including hypoxia tracers, hypoxia-activated prodrugs, and drugs targeting hypoxia-inducible factors and downstream targets, can be used for visualization and quantitative analysis of tumor hypoxia and antitumor activity. However, the relationship between hypoxia and cancer is an area of research that requires further exploration. Here, we investigated the potential factors in the development of hypoxia in cancer, changes in signaling pathways that occur in cancer cells to adapt to hypoxic environments, the mechanisms of hypoxia-induced cancer immune tolerance, chemotherapeutic tolerance, and enhanced radiation tolerance, as well as the insights and applications of hypoxia in cancer therapy.

Title: Intraperitoneal chemotherapy hyperthermia (HIPEC) for peritoneal carcinomatosis of ovarian cancer origin by fluid and CO2 recirculation using the closed abdomen technique (PRS-1.0 Combat): A clinical pilot study

Authors: Sánchez-García S, Villarejo-Campos P, Padilla-Valverde D, Amo-Salas M, Martín-Fernández J.

Journal: Int J Hyperthermia. 2016 Aug;32(5):488-95. doi: 10.3109/02656736.2016.1152515. Epub 2016 Apr 7. PMID: 27056558.

Link to full text: Intraperitoneal chemotherapy hyperthermia (HIPEC) for peritoneal carcinomatosis of ovarian cancer origin by fluid and CO2 recirculation using the closed abdomen technique (PRS-1.0 Combat): A clinical pilot study

Abstract: Background This paper reports a study of 21 patients with peritoneal carcinomatosis from ovarian cancer who underwent cytoreductive surgery and HIPEC by means of PRS-1.0 Combat®, a new model for closed abdomen HIPEC aimed at improving fluid distribution with assistance from a CO2 recirculation system. This new technology has been previously shown to be successful in an experimental study (pig model) performed by our group, and has been approved for use in our hospital. Methods Twenty-one patients with peritoneal carcinomatosis of ovarian cancer origin were included in the study. Cytoreductive surgery and HIPEC were performed by a closed abdomen fluid and CO2 recirculation technique using the PRS-1.0 Combat(®) model. We analysed the intraoperative safety tolerance and post-operative morbidity and mortality during the first 30 days. Results Between November 2011 and March 2014 21 patients with epithelial ovarian cancer, International Federation of Gynecology and Obstetrics stage II-IV, were included in the study. During the procedure there were no significant haemodynamic or analytical disturbances. Complication rates were 38.1% and 57.14% for grade III/IV and minor (grade I/II) complications, respectively. Post-operative mortality was 4.76% (one patient). Complete cytoreductive surgery and intraperitoneal chemotherapy improved overall survival and disease-free survival in women with advanced ovarian cancer. The association of intra-abdominal hyperthermia with chemotherapy (HIPEC) increased the therapeutic benefit. Conclusions This study has shown that closed abdomen intraperitoneal chemohyperthermia by a fluid and CO2 recirculation system (PRS-1.0 Combat(®)) can be a safe and feasible model for the treatment of peritoneal carcinomatosis of ovarian cancer origin.

Keywords: Ovarian cancer; closed abdomen HIPEC.

Title: Measuring tumor cycling hypoxia and angiogenesis using a side-firing fiber optic probe

Authors: Yu B, Shah A, Wang B, Rajaram N, Wang Q, Ramanujam N, Palmer GM, Dewhirst MW.

Journal: J Biophotonics. 2014 Jul;7(7):552-64. doi: 10.1002/jbio.201200187. Epub 2012 Dec 14. PMID: 23242854; PMCID: PMC3702687.

Link to full text: Measuring tumor cycling hypoxia and angiogenesis using a side-firing fiber optic probe

Abstract: Hypoxia and angiogenesis can significantly influence the efficacy of cancer therapy and the behavior of surviving tumor cells. There is a growing demand for technologies to measure tumor hypoxia and angiogenesis temporally in vivo to enable advances in drug development and optimization. This paper reports the use of frequency-domain photon migration with a side-firing probe to quantify tumor oxygenation and hemoglobin concentrations in nude rats bearing human head/neck tumors administered with carbogen gas, cycling hypoxic gas or just room air. Significant increase (with carbogen gas breathing) or decrease (with hypoxic gas breathing) in tumor oxygenation was observed. The trend in tumor oxygenation during forced cycling hypoxia (CH) followed that of the blood oxygenation measured with a pulse oximeter. Natural CH was also observed in rats under room air. The studies demonstrated the potential of the technology for longitudinal monitoring of tumor CH during tumor growth or in response to therapy.

Keywords: Diffuse optical spectroscopy; cancer therapy; fiber optic sensor; tumor hypoxia and angiogenesis.

Title: Reactive oxygen species in cancer: Current findings and future directions

Authors: Nakamura H, Takada K.

Journal: Cancer Sci. 2021 Oct;112(10):3945-3952. doi: 10.1111/cas.15068. Epub 2021 Aug 2. PMID: 34286881; PMCID: PMC8486193.

Link to full text: Reactive oxygen species in cancer: Current findings and future directions

Abstract: Reactive oxygen species (ROS), a class of highly bioactive molecules, have been widely studied in various types of cancers. ROS are considered to be normal byproducts of numerous cellular processes. Typically, cancer cells exhibit higher basal levels of ROS compared with normal cells as a result of an imbalance between oxidants and antioxidants. ROS have a dual role in cell metabolism: At low to moderate levels, ROS act as signal transducers to activate cell proliferation, migration, invasion, and angiogenesis. In contrast, high levels of ROS cause damage to proteins, nucleic acids, lipids, membranes, and organelles, leading to cell death. Extensive studies have revealed that anticancer therapies that manipulate ROS levels, including immunotherapies, show promising in vitro as well as in vivo results. In this review, we summarize molecular mechanisms and oncogenic functions that modulate ROS levels and are useful for the development of cancer therapeutic strategies. This review also provides insights into the future development of effective agents that regulate the redox system for cancer treatment.

Keywords: cell death; neoplasms; oxidative stress; reactive oxygen species; therapeutics.

Title: The mechanisms by which hyperbaric oxygen and carbogen improve tumour oxygenation

Authors: Brizel DM, Lin S, Johnson JL, Brooks J, Dewhirst MW, Piantadosi CA.

Journal: Br J Cancer. 1995 Nov;72(5):1120-4. doi: 10.1038/bjc.1995.474. PMID: 7577456; PMCID: PMC2033965.

Link to full text: The mechanisms by which hyperbaric oxygen and carbogen improve tumour oxygenation

Abstract: Hyperbaric oxygen (HBO) has been proposed to reduce tumour hypoxia by increasing the amount of dissolved oxygen in the plasma. That this actually occurs has not been verified experimentally. This study was performed to explore changes in tumour oxygenation induced by treatment with normobaric and hyperbaric oxygen and carbogen. R3230Ac mammary adenocarcinomas were implanted into Fisher 344 rats. Arterial blood gases, blood pressure and heart rate were monitored. Tumour oxygenation was measured polarographically in five sets of animals. They received either normobaric 100% oxygen, hyperbaric (3 atmospheres; atm) 100% oxygen, normobaric carbogen or hyperbaric (3 atm) carbogen (HBC) +/- bretylium. HBO reduced the mean level of low pO2 values (< 5 mmHg) from 0.49 to 0.07 (P = 0.0003) and increased the average median pO2 from 8 mmHg to 55 mmHg (P = 0.001). HBC reduced the level of low pO2 values from 0.82 to 0.51 (P = 0.002) an increased median pO2 from 2 mmHg to 6 mmHg (P = 0.05). Normobaric oxygen and carbogen did not change tumour oxygenation significantly. Sympathetic blockade with bretylium before HBC exposure improved oxygenation significantly more than HBC alone (low pO2 0.55-0.17, median pO2 4-17 mmHg). HBO and hyperbaric carbogen improved tumour oxygenation in this model, while normobaric oxygen or carbogen had no effect. Sympathetic-mediated vasoconstriction during hyperbaric carbogen caused it to be less effective than HBO. This mechanism also appeared to operate during normobaric carbogen breathing.

Title: Inhalation of low (0.5%–1.5%) CO2 as a potential treatment for apnea of prematurity

Authors: Al-Aif S, Alvaro R, Manfreda J, Kwiatkowski K, Cates D, Rigatto H.

Journal: Semin Perinatol. 2001 Apr;25(2):100-6. doi: 10.1053/sper.2001.23199. PMID: 11339662.

Link to PubMed: Inhalation of low (0.5%–1.5%) CO2 as a potential treatment for apnea of prematurity

Abstract: Apnea of prematurity is common and none of the treatments being used are fully effective and free of significant adverse side effects. We hypothesized that low concentrations of CO2 (< or = 1.5%) may reduce apnea without causing discomfort from an increase in ventilation. We studied 10 preterm infants at a gestational age of 32+/-1 wk (mean +/- SEM) and birthweight 1.8+/-0.2 kg. After a control period of 1 hour, concentrations of CO2 were given (0.5%, 1%, and 1.5%) for 1 hour each, followed by a recovery period of 1 hour. Apnea number significantly decreased from 2.0+/-0.3 apneas/min during control to 1.0+/-0.1 apneas/min (0.5% CO2; P < .05), 1.1+/-0.2 (1% CO2; P < .05), and to 0.7+/-0.2 (1.5% CO2; P < .01). The apnea time significantly decreased from 14.2+/-2.5 s/min during control to 5.2+/-0.8 (0.5% CO2; P < .01), 5.8+/-0.7 (1% CO2; P < .01), and to 3.7+/-0.9 (1.5% CO2; P < .01). Minute ventilation significantly increased with CO2 without evidence of respiratory discomfort. TcPCO2 did not change and TcPO2 increased slightly. These findings suggest that inhalation of low concentrations of CO2 in preterm infants with apnea 1) decreases the number and time of apneas, 2) improves oxygenation, 3) increases ventilation, and 4) is effective even in such low concentrations as 0.5%. We speculate that inhalation of CO2 (< 1%) is more effective and safer than methylxanthines for the treatment of apnea of prematurity.

Title: Carbon Dioxide in Sleep Medicine: The Next Frontier for Measurement, Manipulation, and Research

Authors: Thomas RJ.

Journal: J Clin Sleep Med. 2014 May 15;10(5):523-6. doi: 10.5664/jcsm.3702. PMID: 24910554; PMCID: PMC4046364.

Link to full text: Carbon Dioxide in Sleep Medicine: The Next Frontier for Measurement, Manipulation, and Research

Abstract: Carbon dioxide (CO2) is the most important regulator of respiration and blood pH. Papers are published with scant new information every so often that largely focus on clinical descriptions of hypercapnic individuals, often obese. While there is intense basic science interest in CO2 sensitive neurons in the brainstem, including the Phox2b/neurokinin-1 receptor (NK1R)-expressing neurons in the pre-Bötzinger complex (pre-BötzC),1 and exposing the carotid bodies to hypocapnia induces periodic breathing,2,3 the science and industry of sleep-breathing medicine has generally neglected CO2. Much of the information on chronic exposure to elevated but low levels of CO2 comes from submarine research, targeting sustained ambient concentrations in the low single digits.4–8 Acute exposure to high concentrations of CO2 results in extreme dyspnea and death; the gas has anesthetic properties and can induce a reversible isoelectric EEG.9 Demonstrating slowing of EEG rhythms with hypercapnia10 and power loss in classic oscillatory bands11 (besides increased slow wave sleep associated with respiratory failure12) is nonspecific and not informative on pathobiological mechanisms. Increased inhaled CO2 does suppress cerebral metabolic rate of oxygen13 and reduces resting state functional connectivity.14 While sympathetic drive is reliably increased acutely by hypercapnia, acclimatization mechanisms at the cellular and neural circuit levels, which can be remarkably potent enabling life at otherwise lethal CO2 levels, remain to be elucidated. We are not sure if there is a true CO2 sensor equivalent to hypoxia-inducible factors vs. simply pH mediated changes, or if there are profound direct effects of CO2 on inflammatory responses, the metabolome, the transcriptome, or epigenetic regulation. Widespread systemic dysregulation is plausible, and some data is suggestive. Hypercapnia can induce mitochondrial dysfunction through increased levels on microRNA-183, which decreases expression of isocitrate dehydrogenase.15 Hypoxia-hypercapnia cycles are neurotoxic.16 Obesity hypoventilation (vs. obese controls) was reported to show an increase in the pro-atherosclerotic RANTES chemokine, a decrease in the anti-inflammatory adipokine adiponectin, and impaired endothelial function.17

Title: Allergic rhinitis-induced nasal congestion: its impact on sleep quality

Authors: Storms W.

Journal: Prim Care Respir J. 2008 Mar;17(1):7-18. doi: 10.3132/pcrj.2008.00001. PMID: 18253678; PMCID: PMC6619860.

Link to full text: Allergic rhinitis-induced nasal congestion: its impact on sleep quality

Abstract: Allergic rhinitis (AR) is an extremely common health problem affecting 20 to 40 million Americans and between 10-25% of the world's population. Patients with AR suffer from both nasal symptoms (congestion, rhinorrhea, itching, and sneezing) and ocular symptoms (itching, redness, and tearing). The negative impact on sleep quality and quantity, and consequently on various aspects of the patient's life, is an under-recognised and under-treated component of AR morbidity. Nasal congestion, which is one of the most bothersome and prevalent symptoms of AR, is thought to be the leading symptom responsible for rhinitis-related sleep problems. In addition to reducing clinical symptoms, pharmacologic therapies for AR that specifically reduce inflammatory cells and mediators - and therefore nasal congestion and other symptoms - should also improve sleep quality and overall quality of life (QOL). Intranasal corticosteroids (INS) are the current mainstay of therapy for AR. Results of a number of clinical trials demonstrate that INS effectively reduce nasal congestion and ocular symptoms, improve sleep quality, and decrease daytime somnolence. Intranasal corticosteroids have also proved to be effective in reducing symptoms of acute rhinosinusitis and nasal polyposis, both of which also negatively impact on sleep quality. Intranasal corticosteroids are considered safe due to their low systemic bioavailability.

Title: Alternative approaches to treatment of Central Sleep Apnea

Authors: Thomas RJ.

Journal: Sleep Med Clin. 2014 Mar 1;9(1):87-104. doi: 10.1016/j.jsmc.2013.10.008. PMID: 24772053; PMCID: PMC3998090.

Link to full text: Alternative approaches to treatment of Central Sleep Apnea

Abstract: Divergent approaches to treatment of hypocapnic central sleep apnea syndromes reflect the difficulties in taming a hyperactive respiratory chemoreflex. As both sleep fragmentation and a narrow CO2 reserve or increased loop gain drive the disease, sedatives (to induce longer periods of stable non-rapid eye movement (NREM) sleep and reduce the destabilizing effects of arousals in NREM sleep) and CO2-based stabilization approaches are logical. Adaptive ventilation reduces mean hyperventilation yet can induce ventilator-patient dyssynchrony, while enhanced expiratory rebreathing space (EERS, dead space during positive pressure therapy) and CO2 manipulation directly stabilize respiratory control by moving CO2 above the apnea threshold. Carbonic anhydrase inhibition can provide further adjunctive benefits. Provent and Winx may be less likely to trigger central apneas or periodic breathing in those with a narrow CO2 reserve. An oral appliance can meaningfully reduce positive pressure requirements and thus enable treatment of complex apnea. Novel pharmacological approaches may target mediators of carotid body glomus cell excitation, such as the balance between gas neurotransmitters. In complex apnea patients, single mode therapy is not always successful, and multi-modality therapy might need to be considered. Phenotyping of sleep apnea beyond conventional scoring approaches is the key to optimal management.

Keywords: Winx; acetazolamide Provent; carbon dioxide; central apnea; multimodal complex; oxygen rebreathing; periodic breathing.

Title: Better quality of life when nasal breathing of snoring men is improved at night

Authors: Löth S, Petruson B, Wirén L, Wilhelmsen L.

Journal: Arch Otolaryngol Head Neck Surg. 1999 Jan;125(1):64-7. doi: 10.1001/archotol.125.1.64. PMID: 9932590.

Link to PubMed: Better quality of life when nasal breathing of snoring men is improved at night

Abstract: Objective: To evaluate whether improved nasal breathing changes the quality of life in snoring men and improves the female sleeping partners' well-being in the morning.

Design: During 1 month, 42 heavily snoring men slept with a nostril dilator. Before and after 1 month, the snorers rated their daytime tiredness and completed the Nottingham Health Profile questionnaire. Female sleeping partners rated the snoring, the quality of their sleep, and their sense of well-being in the morning. A population sample was used for comparison.

Setting: The Central Hospital, Skövde, Sweden.

Results: The snorers' quality of life before the study was significantly worse (P<.001) than that of the comparison population and improved significantly (P = .001). The men were significantly (P<.001) less tired during the day when their nasal airflow was increased. Female sleeping partners had significantly (P = .005) better sleep and an improved sense of well-being in the morning during the test period. Both were correlated with a significant reduction in the snoring (P<.001).

Conclusions: When nasal breathing of snoring men was improved at night, their quality of life was significantly improved. The female sleeping partners had a reduction in sleep disturbance that correlated well with an improvement in their own sleep and feelings of well-being in the morning.

Title: Blowing past the apneic threshold

Authors: Johal A, Lenet A, Sigua NL.

Journal: J Clin Sleep Med. 2022 Oct 1;18(10):2515-2518. doi: 10.5664/jcsm.10162. PMID: 35859324; PMCID: PMC9516585.

Link to full text: Blowing past the apneic threshold

Abstract: An 81-year-old man with severe obstructive sleep apnea, hypertension, and obesity (body mass index 32 kg/m2) presented to the sleep clinic for an initial evaluation. He has been on auto-titrating bilevel positive airway pressure (BPAP) with 2 L/min nocturnal oxygen for more than 10 years and has been adherent to therapy. He has no known cardiopulmonary disorder and was not on opiates, benzodiazepines, or other respiratory depressants. He denied tobacco, drug, or alcohol use. His initial sleep study, the indication for nocturnal oxygen supplementation, and serum bicarbonate level were not available for review.

Title: Central sleep apnea: pathophysiologic classification

Authors: Javaheri S, Badr MS.

Journal: Sleep. 2023 Mar 9;46(3):zsac113. doi: 10.1093/sleep/zsac113. PMID: 35551411; PMCID: PMC9995798.

Link to full text: Central sleep apnea: pathophysiologic classification

Abstract: Central sleep apnea is not a single disorder; it can present as an isolated disorder or as a part of other clinical syndromes. In some conditions, such as heart failure, central apneic events are due to transient inhibition of ventilatory motor output during sleep, owing to the overlapping influences of sleep and hypocapnia. Specifically, the sleep state is associated with removal of wakefulness drive to breathe; thus, rendering ventilatory motor output dependent on the metabolic ventilatory control system, principally PaCO2. Accordingly, central apnea occurs when PaCO2 is reduced below the "apneic threshold". Our understanding of the pathophysiology of central sleep apnea has evolved appreciably over the past decade; accordingly, in disorders such as heart failure, central apnea is viewed as a form of breathing instability, manifesting as recurrent cycles of apnea/hypopnea, alternating with hyperpnea. In other words, ventilatory control operates as a negative-feedback closed-loop system to maintain homeostasis of blood gas tensions within a relatively narrow physiologic range, principally PaCO2. Therefore, many authors have adopted the engineering concept of "loop gain" (LG) as a measure of ventilatory instability and susceptibility to central apnea. Increased LG promotes breathing instabilities in a number of medical disorders. In some other conditions, such as with use of opioids, central apnea occurs due to inhibition of rhythm generation within the brainstem. This review will address the pathogenesis, pathophysiologic classification, and the multitude of clinical conditions that are associated with central apnea, and highlight areas of uncertainty.

Keywords: Adaptive-Servo Ventilation (ASV); apneic threshold; bi-level positive pressure therapy (BPAP); central apnea; continuous positive pressure therapy (CPAP); controller gain; hypocapnia; loop gain; plant gain.

Title: CO2 homeostasis during periodic breathing in obstructive sleep apnea

Authors: Berger KI, Ayappa I, Sorkin IB, Norman RG, Rapoport DM, Goldring RM.

Journal: J Appl Physiol (1985). 2000 Jan;88(1):257-64. doi: 10.1152/jappl.2000.88.1.257. PMID: 10642388.

Link to full text: CO2 homeostasis during periodic breathing in obstructive sleep apnea

Abstract: The contribution of apnea to chronic hypercapnia in obstructive sleep apnea (OSA) has not been clarified. Using a model (D. M. Rapoport, R. G. Norman, and R. M. Goldring. J. Appl. Physiol. 75: 2302-2309, 1993), we previously illustrated failure of CO2 homeostasis during periodic breathing resulting from temporal dissociation between ventilation and perfusion ("temporal V/Q mismatch"). This study measures acute kinetics of CO2 during periodic breathing and addresses interapnea ventilatory compensation for maintenance of CO2 homeostasis in 11 patients with OSA during daytime sleep (37-171 min). Ventilation and expiratory CO2 and O2 fractions were measured on a breath-by-breath basis by means of a tight-fitting full facemask. Calculations included CO2 excretion, metabolic CO2 production, and CO2 balance (metabolic CO2 production - exhaled CO2). CO2 balance was tabulated for each apnea/hypopnea event-interevent cycle and as a cumulative value during sleep. Cumulative CO2 balance varied (-3,570 to +1,388 ml). Positive cumulative CO2 balance occurred in the absence of overall hypoventilation during sleep. For each cycle, positive CO2 balance occurred despite increased interevent ventilation to rates as high as 45 l/min. This failure of CO2 homeostasis was dependent on the event-to-interevent duration ratio. The results demonstrate that 1) periodic breathing provides a mechanism for acute hypercapnia in OSA, 2) acute hypercapnia during periodic breathing may occur without a decrease in average minute ventilation, supporting the presence of temporal V/Q mismatch, as predicted from our model, and 3) compensation for CO2 accumulation during apnea/hypopnea may be limited by the duration of the interevent interval. The relationship of this acute hypercapnia to sustained chronic hypercapnia in OSA remains to be further explored.

Title: Effect of Breathe Right nasal strip on snoring

Authors: Ulfberg J, Fenton G.

Journal: Rhinology. 1997 Jun;35(2):50-2. PMID: 9299650.

Link to PubMed: Effect of Breathe Right nasal strip on snoring

Abstract: Snoring is a significant problem both for the patient and for the bedpartner. It is well known that nasal stuffiness can contribute to snoring, and sleep quality may deteriorate because of the snoring. Nasal dilation can reduce snoring and improve sleep. Thirty-five habitual snorers (18 female, 17 male) and their bedpartners participated in an open label study. The patients were diagnosed as heavy snorers after they underwent overnight polysomnography showing that their apnoea indexes were below 5, thus sleep apnoea patients were not included in the study. The participants and their partners filled out evaluations concerning snoring intensity, mouth dryness and Epworth Sleepiness Scale prior to and after using Breath Right nasal strips for 14 consecutive nights. The Breathe Right external nasal dilator is a simple, nonpharmaceutical method to decrease nasal airway resistance and thus potentially reduce or eliminate snoring. After using the strips there were statistically significant decreases in snoring (p < 0.001) as graded by the bed partner, and in mouth dryness (p = 0.025) and in the Epworth Sleepiness Scale scores (p = 0.001), as graded by the patient. The results of this study indicate that Breathe Right nasal strips may be used to reduce snoring, mouth dryness and sleepiness in patients presenting with symptoms of snoring.

Title: Effect of hypoxia on the hypopnoeic and apnoeic threshold for CO(2) in sleeping humans

Authors: Xie A, Skatrud JB, Dempsey JA.

Journal: J Physiol. 2001 Aug 15;535(Pt 1):269-78. doi: 10.1111/j.1469-7793.2001.00269.x. PMID: 11507176; PMCID: PMC2278764.

Link to full text: Effect of hypoxia on the hypopnoeic and apnoeic threshold for CO(2) in sleeping humans

Abstract: 1. Rhythmic breathing during sleep requires that P(CO2) be maintained above a sensitive hypocapnic apnoeic threshold. Hypoxia causes periodic breathing during sleep that can be prevented or eliminated with supplemental CO(2). The purpose of this study was to determine the effect of hypoxia in changing the difference between the eupnoeic P(CO2) and the P(CO2) required to produce hypopnoea or apnoea (hypopnoea/apnoeic threshold) in sleeping humans. 2. The effect of hypoxia on eupnoeic end-tidal partial pressure of CO(2) (P(ET,CO2)) and hypopnoea/apnoeic threshold P(ET,CO2) was examined in seven healthy, sleeping human subjects. A bilevel pressure support ventilator in a spontaneous mode was used to reduce P(ET,CO2) in small decrements by increasing the inspiratory pressure level by 2 cmH2O every 2 min until hypopnoea (failure to trigger the ventilator) or apnoea (no breathing effort) occurred. Multiple trials were performed during both normoxia and hypoxia (arterial O(2) saturation, S(a,O2) = 80 %) in a random order. The hypopnoea/apnoeic threshold was determined by averaging P(ET,CO2) of the last three breaths prior to each hypopnoea or apnoea. 3. Hypopnoeas and apnoeas were induced in all subjects during both normoxia and hypoxia. Hypoxia reduced the eupnoeic P(ET,CO2) compared to normoxia (42.4 +/- 1.3 vs. 45.0 +/- 1.1 mmHg, P < 0.001). However, no change was observed in either the hypopnoeic threshold P(ET,CO2) (42.1 +/- 1.4 vs. 43.0 +/- 1.2 mmHg, P > 0.05) or the apnoeic threshold P(ET,CO2) (41.3 +/- 1.2 vs. 41.6 +/- 1.0 mmHg, P > 0.05). Thus, the difference in P(ET,CO2) between the eupnoeic and threshold levels was much smaller during hypoxia than during normoxia (-0.2 +/- 0.2 vs. -2.0 +/- 0.3 mmHg, P < 0.01 for the hypopnoea threshold and -1.1 +/- 0.2 vs. -3.4 +/- 0.3 mmHg, P < 0.01 for the apnoeic threshold). We concluded that hypoxia causes a narrowing of the difference between the baseline P(ET,CO2) and the hypopnoea/apnoeic threshold P(ET,CO2), which could increase the likelihood of ventilatory instability.

Title: Effect of nasal dilation on snoring and apneas during different stages of sleep

Authors: Hoffstein V, Mateika S, Metes A.

Journal: Sleep. 1993 Jun;16(4):360-5. doi: 10.1093/sleep/16.4.360. PMID: 8141871

Link to PubMed: Effect of nasal dilation on snoring and apneas during different stages of sleep

Abstract: This study was designed to test the hypothesis that nasal dilation reduces snoring. To achieve this we performed nocturnal polysomnography, including measurement of snoring, in 15 patients without nasal pathology before and after insertion of a nasal dilator (NOZOVENT). Snoring was quantified for each sleep stage by recording the number of snores per minute of sleep, number of snores per minute of snoring time and nocturnal sound intensities (maximum, average and minimum). We found that nasal dilation had no effect on the number of apneas, hypopneas or oxygen saturation. Snoring parameters were unaffected by NOZOVENT during stages I, II and REM sleep, but were all significantly reduced during slow wave sleep. We conclude that dilation of the anterior nares in patients without nasal pathology has a relatively weak effect on snoring, and routine use of nasal dilating appliances is not recommended for treatment of snoring.

Title: Hypocapnia is associated with increased upper airway expiratory resistance during sleep

Authors: Sankri-Tarbichi AG, Richardson NN, Chowdhuri S, Rowley JA, Safwan Badr M.

Journal: Respir Physiol Neurobiol. 2011 Jul 31;177(2):108-13. doi: 10.1016/j.resp.2011.04.004. Epub 2011 Apr 13. PMID: 21513820; PMCID: PMC3113471.

Link to full text: Hypocapnia is associated with increased upper airway expiratory resistance during sleep

Abstract: We hypothesized that hypocapnia is responsible for increased expiratory resistance during NREM sleep. Hypocapnia was induced by hypoxic hyperventilation in 21 subjects (aged 29.4 ± 7.8 yrs, 10 women, BMI 24.4 ± 4.3 kg/m(2)). Isocapnic hypoxia was induced in 12 subjects of whom, 6 underwent hypocapnic hypoxia in the same night. Upper airway resistance (R(UA)) was measured at the linear pressure-flow relationship during inspiration and expiration. Inspiratory flow limitation (IFL) was defined as the dissociation in pressure-flow relationship. (1) Expiratory R(UA) increased during hypocapnic but not isocapnic hypoxia relative to control (11.0 ± 5.6 vs. 8.2 ± 3.6 cm H(2)O/L/s; p < 0.05, and 11.45.0 vs. 10.94.4 cm H(2)O/L/s; p = NS, respectively). (2) No gender difference was found in R(UA) (p = NS). (3) Increased expiratory R(UA) correlated with the IFL change during hypocapnic but not isocapnic hypoxia. (4) No changes were noted in inspiratory R(UA) or IFL. Expiratory R(UA) increased during hypocapnia and was associated with IFL, indicating upper airway narrowing. Gender does not influence the upper airway response to hypocapnic hypoxia.

Title: Improved nasal breathing reduces snoring and morning tiredness. A 6-month follow-up study

Authors: Löth S, Petruson B.

Journal: Arch Otolaryngol Head Neck Surg. 1996 Dec;122(12):1337-40. doi: 10.1001/archotol.1996.01890240045010. PMID: 8956746.

Link to PubMed: Improved nasal breathing reduces snoring and morning tiredness. A 6-month follow-up study

Abstract: Background: Dilation of the nasal valve region can increase the ability to breathe through the nose and reduce the negative intrathoracic pressure required for inspiration. Vibrations of the palate and soft tissues of the throat, which generate snoring sounds, can be prevented when patients inhale less heavily.

Objectives: To evaluate the effect a nostril dilator has on patient snoring and tiredness in the morning and to determine how many patients would continue to use the device for half a year.

Design: For 6 months, 42 men who were heavy snorers graded their average tiredness in the morning and the patient's sleeping partner graded the average snoring using a visual analog scale from 0 to 100.

Setting: All patients were examined at the Department of Otorhinolaryngology, Central Hospital, in Skövde, Sweden.

Results: When the nostril dilator was used there was a significant decrease in snoring after both 1- and 6-month reports. There was a significant correlation between diminution of snoring and less tiredness in patients in the morning. The compliance was good since 60% of patients continued to use the device during the 6-month test period.

Conclusions: This study illustrates the benefits that reducing nasal airway resistance during sleep has on snoring and morning tiredness in patients.

Title: Influence of longterm CPAP therapy on CO2 drive in patients with obstructive sleep apnea

Authors: Verbraecken J, Willemen M, De Cock W, Wittesaele W, Govaert K, Van de Heyning P, De Backer W.

Journal: Respir Physiol. 2000 Oct;123(1-2):121-30. doi: 10.1016/s0034-5687(00)00140-7. PMID: 10996193.

Link to PubMed: Influence of longterm CPAP therapy on CO2 drive in patients with obstructive sleep apnea

Abstract: According to recent publications, the values of the hypercapnic ventilatory responses (HCVR) in normocapnic obstructive sleep apnea (OSA) patients are highly variable, but they are usually within the normal range. In our study, CPAP therapy during 1 month did not seem to influence the HCVR [Verbraecken, J., De Backer, W., Willemen, M., De Cock, W., Wittesaele, W., Van de Heyning, P., 1995. Respir. Physiol. 101, 279-287]. It is, however, not well studied whether long term (1 year) CPAP therapy can influence HCVR in normocapnic patients. Therefore, we evaluated the effect of 1 year CPAP therapy on CO(2) drive in 20 OSA patients. The slope of HCVR (SHCVR) changed from 2.35+/-0.21 L min(-1) mmHg(-1) (control) to 1.66+/-0.16 L min(-1) mm Hg(-1) (P=0.04), but the mean within subject coefficient of variation in repeated measurements of SHCVR in treated and untreated OSA patients was not statistically different. Pa(O(2)) increased from 72+/-2 mm Hg to 80+/-2 (1 year) mm Hg. We conclude that CPAP therapy improves daytime gas exchange in normocapnic OSA and may possibly decrease CO(2) drive (slope) after a treatment period of 1 year.

Title: Low-concentration carbon dioxide is an effective adjunct to positive airway pressure in the treatment of refractory mixed central and obstructive sleep-disordered breathing

Authors: Thomas RJ, Daly RW, Weiss JW.

Journal: Sleep. 2005 Jan;28(1):69-77. doi: 10.1093/sleep/28.1.69. PMID: 15700722.

Link to PubMed: Low-concentration carbon dioxide is an effective adjunct to positive airway pressure in the treatment of refractory mixed central and obstructive sleep-disordered breathing

Abstract: Objectives: To assess the efficacy of added carbon dioxide as adjunctive therapy to positive airway pressure-refractory mixed obstructive and central sleep-disordered breathing, using a prototype device-the positive airway pressure gas modulator.

Design: Open-label evaluation of low concentrations of carbon dioxide added to a positive airway pressure circuit.

Setting: Physician-attended polysomnographic titration in a free-standing sleep laboratory with end-tidal and transcutaneous carbon-dioxide monitoring.

Patients: Six adult men (age 54 +/- 5.7 years) with severe poorly controlled mixed sleep-disordered breathing in the absence of renal or heart failure.

Interventions: Flow-independent addition of incremental concentrations of carbon dioxide during sleep.

Measurements and results: The respiratory disturbance index before treatment was 66 +/- 14.5 events per hour of sleep, with a nocturnal desaturation low of 84.6% +/- 10.1%. Residual respiratory disturbance index on best treatment was 43 +/- 9 events per hour of sleep. There was an immediate (<1 minute) response to the addition of 0.5% to 1% carbon dioxide, and minimal changes were required to be made across the night. There was no discomfort, shortness of breath, palpitations, headache, or significant increase in respiratory or heart rate. The residual respiratory disturbance index on carbon dioxide, scored irrespective of desaturations, was in the normal range (< 5 / hour of sleep). Two subjects had a second night at the concentration of carbon dioxide determined to be efficacious, with no required concentration change. No adverse effects on overall sleep architecture were noted.

Conclusions: Low concentrations of carbon dioxide added to conventional positive airway pressure effectively control severe treatment-resistant mixed obstructive and central sleep-disordered breathing.

Title: Objective assessment of nasal obstruction in snoring and obstructive sleep apnea patients: experience of a Police Authority Hospital

Authors: Kalam I.

Journal: Ann Saudi Med. 2002 May-Jul;22(3-4):158-62. doi: 10.5144/0256-4947.2002.158. PMID: 17159386.

Link to full text: Objective assessment of nasal obstruction in snoring and obstructive sleep apnea patients: experience of a Police Authority Hospital

Abstract: Background: The role of nasal airflow resistance in the pathogenesis of obstructive sleep apnea (OSA) syndrome remains contentious. The aim of this study was to investigate the changes in apnea index in response to nasal surgery, as guided by acoustic rhinometry.

Patients and methods: Forty-five patients were referred for complaints of snoring and nasal obstruction. The patients were divided into two groups according to the severity of nasal obstruction: group 1 (nasal obstruction was the predominant symptom) included 21 patients, and group 2 (snoring was the predominant symptom) included 24 patients. In group 1, nasal surgery was performed as a first surgical procedure and was followed after 10-12 weeks by palatal surgery. In group 2, palatal surgery was performed as the primary procedure and was followed after 10-12 weeks by nasal surgery.

Results: The results showed a clear relationship between the correction of nasal obstruction and the severity of OSA, as indicated by measuring the apnea index (AI). However, in none of the group 1 cases was nasal surgery alone capable of reducing the AI by 50%. In group 2, palatal surgery effectively reduced AI, and when followed by nasal surgery, the reduction in AI was again statistically significant.

Conclusion: This indicates that nasal obstruction could be considered a contributing factor to the severity of OSA, but not a causative factor in the production of OSA. It also indicates that nasal surgery and palatal surgery combined are effective in improving snoring and OSA in properly selected patients.

Title: A review of preventing central sleep apnea by inspired CO2

Authors: Mulchrone A, Shokoueinejad M, Webster J.

Journal: Physiol Meas. 2016 May;37(5):R36-45. doi: 10.1088/0967-3334/37/5/R36. Epub 2016 Apr 19. PMID: 27093535.

Link to PubMed: A review of preventing central sleep apnea by inspired CO2

Abstract: Although almost completely unknown half a century ago, sleep disorders are gaining recognition as major issues to public health due to their growing prevalence and dire societal consequences. Despite being linked to several infamous catastrophic events such as Chernobyl, it is estimated that 90% of sufferers fail to get diagnosed and receive treatment, and a significant portion of the ones that do are often non-compliant due to the side effects of current treatments. This article presents a review of the current standard treatment for central sleep apnea, and investigates the advantages and possible consequences of using inspired carbon dioxide (CO2) as an alternative treatment option.

Title: Effects of inhaled CO2 and added dead space on idiopathic central sleep apnea

Authors: Xie A, Rankin F, Rutherford R, Bradley TD.

Journal: J Appl Physiol (1985). 1997 Mar;82(3):918-26. doi: 10.1152/jappl.1997.82.3.918. PMID: 9074983.

Link to full text: Effects of inhaled CO2 and added dead space on idiopathic central sleep apnea

Abstract: We hypothesized that reductions in arterial PCO2 (PaCO2) below the apnea threshold play a key role in the pathogenesis of idiopathic central sleep apnea syndrome (ICSAS). If so, we reasoned that raising PaCO2 would abolish apneas in these patients. Accordingly, patients with ICSAS were studied overnight on four occasions during which the fraction of end-tidal CO2 and transcutaneous PCO2 were measured: during room air breathing (N1), alternating room air and CO2 breathing (N2), CO2 breathing all night (N3), and addition of dead space via a face mask all night (N4). Central apneas were invariably preceded by reductions in fraction of end-tidal CO2. Both administration of a CO2-enriched gas mixture and addition of dead space induced 1- to 3-Torr increases in transcutaneous PCO2, which virtually eliminated apneas and hypopneas; they decreased from 43.7 +/- 7.3 apneas and hypopneas/h on N1 to 5.8 +/- 0.9 apneas and hypopneas/h during N3 (P < 0.005), from 43.8 +/- 6.9 apneas and hypopneas/h during room air breathing to 5.9 +/- 2.5 apneas and hypopneas/h of sleep during CO2 inhalation during N2 (P < 0.01), and to 11.6% of the room air level while the patients were breathing through added dead space during N4 (P < 0.005). Because raising PaCO2 through two different means virtually eliminated central sleep apneas, we conclude that central apneas during sleep in ICSA are due to reductions in PaCO2 below the apnea threshold.

Title: Effect of CO2 inhalation on central sleep apnea and arousals from sleep

Authors: Szollosi I, Jones M, Morrell MJ, Helfet K, Coats AJ, Simonds AK.

Journal: Respiration. 2004 Sep-Oct;71(5):493-8. doi: 10.1159/000080634. PMID: 15467327.

Link to PubMed: Effect of CO2 inhalation on central sleep apnea and arousals from sleep

Abstract: Background: CO(2) inhalation reduces central sleep apnea (CSA) in patients with congestive heart failure (CHF) and idiopathic CSA. CO(2) is also a stimulus for cortical arousal, which has been linked to increased sympathetic nerve activity (SNA) and increased mortality in CHF patients with CSA.

Objective: We have tested the hypothesis that during sleep, inhalation of CO(2) sufficient to reduce the apnea-hypopnea index (AHI) would not reduce the arousal index (AroI).

Methods: In 10 male patients with CSA (7 with CHF and 3 with idiopathic CSA), the inspired CO(2) concentration was increased to raise the sleeping end-tidal CO(2) by 2-4 mm Hg during established stage 2 sleep. Each intervention was maintained for a 10-min period. Sleep stage was monitored with electroencephalograms, electrooculograms, submental electromyogram, airflow with pneumotachometer and respiratory effort and blood gases with oxygen saturation and end-tidal CO(2). During periods of air and CO(2) breathing, AHI and AroI were compared with paired t tests; patients acted as their own controls.

Results: Inhalation of CO(2) produced a significant reduction in AHI (mean +/- SEM) from 74.4 +/- 12.4 events/h during air breathing to 25.8 +/- 7.8 events/h with CO(2) inhalation (p = 0.002). However, the AroI was not significantly different between the two conditions, air 67.8 +/- 12.3 events/h and CO(2) inhalation 52.8 +/- 12.4 events/h (p = 0.264).

Conclusion: CO(2) inhalation reverses CSA but not arousals from sleep. Our findings highlight the need for treatment options that reduce both respiratory events and decrease arousals from sleep, with their associated SNA sequelae.

Title: Carbon dioxide in sleep medicine: the next frontier for measurement, manipulation, and research

Authors: Thomas RJ.

Journal: J Clin Sleep Med. 2014 May 15;10(5):523-6. doi: 10.5664/jcsm.3702. PMID: 24910554; PMCID: PMC4046364.

Link to full text: Carbon dioxide in sleep medicine: the next frontier for measurement, manipulation, and research

Abstract: Carbon dioxide (CO2) is the most important regulator of respiration and blood pH. Papers are published with scant new information every so often that largely focus on clinical descriptions of hypercapnic individuals, often obese. While there is intense basic science interest in CO2 sensitive neurons in the brainstem, including the Phox2b/neurokinin-1 receptor (NK1R)-expressing neurons in the pre-Bötzinger complex (pre-BötzC),1 and exposing the carotid bodies to hypocapnia induces periodic breathing,2,3 the science and industry of sleep-breathing medicine has generally neglected CO2.

Title: Obstructive sleep apnoea and breathing retraining

Authors: Birch M.

Journal: Aust Nurs J. 2004 Aug;12(2):27-9. PMID: 19160562.

Link to PubMed: Obstructive sleep apnoea and breathing retraining

Abstract: Obstructive sleep apnoea (OSA) is a sleep disorder where repeated upper airway obstruction during sleep leads to a decrease in blood oxygen saturation and disrupted sleep. Current treatment options include oral appliances, surgery, and/or the use of a continuous positive airway pressure (CPAP) machine. However, breathing retraining with the Buteyko Institute Method (BIM) is a safe, effective and convenient approach to OSA that could help eliminate the need for surgery or CPAP. This paper examines the role of breathing retraining in OSA and presents a case study to illustrate its effectiveness.

Title: Pathogenesis of upper airway occlusion during sleep

Authors: Remmers JE, deGroot WJ, Sauerland EK, Anch AM.

Journal: J Appl Physiol Respir Environ Exerc Physiol. 1978 Jun;44(6):931-8. doi: 10.1152/jappl.1978.44.6.931. PMID: 670014.

Link to full text: Pathogenesis of upper airway occlusion during sleep

Abstract: Ten patients with daytime somnolence and obesity were found to have periodic airway occlusion (AO) during nocturnal sleep. The cyclical ventilatory pattern consisted of a series of regular inspiratory efforts against an occluded airway (occlusive phase) alternating with a period of regular breathing (ventilatory phase). Significant periods of central respiratory apnea were observed only in one case. The effects of pharyngeal intubation and pharyngeal pressure recordings showed that the locus of airway closure lay in the oropharynx. The genioglossal electromyogram (EMG) consistently revealed periodicity: low level activity at the onset of occlusion and prominent discharge at the instant of pharyngeal opening. In one case, this activity was closely related to pharyngeal patency, whereas in other cases there was considerable overlap between EMG values recorded during occluded and ventilatory phases. In these cases, the relationship of genioglossal discharge to pharyngeal pressure correlated with the presence or absence of pharyngeal occlusion. We speculate that genioglossal force acts to open the oropharynx and that negative pharyngeal pressure promotes pharyngeal closure. The results are consistent with the idea that, once the pharvnx has collapsed, relative recruitment of genioglossal and inspiratory muscle act.ivity is such that the latter influence outstrips the former, so that pharyngeal transmural pressure increases more than genioglossal force. Pharyngeal opening occurs coincident with arousal and preferential activation of the genioglossus muscle of the tongue.

Title: Rethinking O2, CO2 and breathing during wakefulness and sleep

Authors: Dempsey JA, Gibbons TD.

Journal: J Physiol. 2024 Nov;602(21):5571-5585. doi: 10.1113/JP284551. Epub 2023 Sep 26. PMID: 37750243.

Link to PubMed: Rethinking O2, CO2 and breathing during wakefulness and sleep

Abstract: We have examined the importance of three long-standing questions concerning chemoreceptor influences on cardiorespiratory function which are currently experiencing a resurgence of study among physiologists and clinical investigators. Firstly, while carotid chemoreceptors (CB) are required for hypoxic stimulation of breathing, use of an isolated, extracorporeally perfused CB preparation in unanaesthetized animals with maintained tonic input from the CB, reveals that extra-CB hypoxaemia also provides dose-dependent ventilatory stimulation sufficient to account for 40-50% of the total ventilatory response to steady-state hypoxaemia. Extra-CB hyperoxia also provides a dose- and time-dependent hyperventilation. Extra-CB sites of O2-driven ventilatory stimulation identified to date include the medulla, kidney and spinal cord. Secondly, using the isolated or denervated CB preparation in awake animals and humans has demonstrated a hyperadditive effect of CB sensory input on central CO2 sensitivity, so that tonic CB activity accounts for as much as 35-40% of the normal, air-breathing eupnoeic drive to breathe. Thirdly, we argue for a key role for CO2 chemoreception and the neural drive to breathe in the pathogenesis of upper airway obstruction during sleep (OSA), based on the following evidence: (1) removal of the wakefulness drive to breathe enhances the effects of transient CO2 changes on breathing instability; (2) oscillations in respiratory motor output precipitate pharyngeal obstruction in sleeping subjects with compliant, collapsible airways; and (3) in the majority of patients in a large OSA cohort, a reduced neural drive to breathe accompanied reductions in both airflow and pharyngeal airway muscle dilator activity, precipitating airway obstruction.

Keywords: extra‐carotid chemoreceptor O2 sensing; obstructive sleep apnoea; peripheral‐central chemoreceptor interdependence.

Title: Respiratory alkalosis may impair the production of vitamin D and lead to significant morbidity, including the fibromyalgia syndrome

Authors: Lewis JM, Fontrier TH, Coley JL.

Journal: Med Hypotheses. 2017 May;102:99-101. doi: 10.1016/j.mehy.2017.03.013. Epub 2017 Mar 8. PMID: 28478843.

Link to PubMed: Respiratory alkalosis may impair the production of vitamin D and lead to significant morbidity, including the fibromyalgia syndrome

Abstract: Hyperventilation caused by physical and/or psychological stress may lead to significant respiratory alkalosis and an elevated systemic pH. The alkalotic pH may in turn suppress the normal renal release of phosphate into the urine, thereby interrupting the endogenous production of 1,25-dihydroxyvitamin D (calcitriol). This could cause a shortfall in its normal production, leading to a variety of adverse consequences. It might partially explain the pathogenesis of acute mountain sickness, a treatable disease characterized by severe hyperventilation secondary to the hypoxia of high altitude exposure. Milder degrees of hyperventilation due to different forms of stress may produce other conditions which share characteristics with acute mountain sickness. One of these may be the fibromyalgia syndrome, a chronic painful disorder for which no satisfactory treatment exists. Should fibromyalgia and acute mountain sickness have a common etiology, may they also share a common form of treatment? Evidence is presented to support this hypothesis.

Title: Breathing evaluation and retraining as an adjunct to manual therapy

Authors: McLaughlin L, Goldsmith CH, Coleman K.

Journal: Man Ther. 2011 Feb;16(1):51-2. doi: 10.1016/j.math.2010.08.006. Epub 2010 Oct 8. PMID: 20933458.

Link to PubMed: Breathing evaluation and retraining as an adjunct to manual therapy

Abstract: Back and neck pain are extremely common reasons for patients seeking manual therapy treatment. Epidemiological evidence supports a link between breathing difficulties and back pain. Since trunk muscles perform both postural and breathing functions, it is theorized that disruption in one function can negatively impact the other. Altered breathing mechanics can change respiratory chemistry and therefore pH causing smooth muscle constriction, altered electrolyte balance and decreased tissue oxygenation. These changes can profoundly impact any body system. Increased excitability in the muscular and nervous systems may be most relevant to a manual therapist. Respiratory function can be tested via capnography which measures CO₂ at the end of exhale known as End Tidal CO₂ (ETCO₂). ETCO₂ closely reflects arterial CO₂ in people with normal cardiopulmonary function. A case series of twenty nine outpatients with neck or back pain who had plateaued with manual therapy and exercise were identified all of whom were found to have low ETCO₂. Breathing retraining improved ETCO₂, pain and function in all patients with 93% achieving at least a clinically important change in either pain or function. Screening for breathing dysfunction using capnography may improve patient outcomes in those patients where manual therapy, exercise and education do not provide full resolution of symptoms.

Title: Hypocapnia in women with fibromyalgia

Authors: Jonsson K, Pikwer A, Olsson EMG, Peterson M.

Journal: Scand J Pain. 2024 Jun 21;24(1). doi: 10.1515/sjpain-2024-0003. PMID: 38907689.

Link to full text: Hypocapnia in women with fibromyalgia

Abstract: Objectives: The purpose of this study was to investigate whether people with fibromyalgia (FM) have dysfunctional breathing by examining acid-base balance and comparing it with healthy controls. Methods: Thirty-six women diagnosed with FM and 36 healthy controls matched for age and gender participated in this cross-sectional study. To evaluate acid-base balance, arterial blood was sampled from the radial artery. Carbon dioxide, oxygen, bicarbonate, base excess, pH and lactate were analysed for between-group differences. Blood gas analyses were performed stepwise on each individual to detect acid-base disturbance, which was categorized as primary respiratory and possible compensation indicating chronicity. A three-step approach was employed to evaluate pH, carbon dioxide and bicarbonate in this order. Results: Women with FM had significantly lower carbon dioxide pressure (p = 0.013) and higher lactate (p = 0.038) compared to healthy controls at the group level. There were no significant differences in oxygen pressure, bicarbonate, pH and base excess. Employing a three-step acid-base analysis, 11 individuals in the FM group had a possible renally compensated mild chronic hyperventilation, compared to only 4 among the healthy controls (p = 0.042). Conclusions: In this study, we could identify a subgroup of individuals with FM who may be characterized as mild chronic hyperventilators. The results might point to a plausible dysfunctional breathing in some women with FM.

Title: The role of carbon dioxide therapy in the treatment of chronic wounds

Authors: Brandi C, Grimaldi L, Nisi G, Brafa A, Campa A, Calabrò M, Campana M, D'Aniello C.

Journal: In Vivo. 2010 Mar-Apr;24(2):223-6. PMID: 20363999.

Link to full text: The role of carbon dioxide therapy in the treatment of chronic wounds

Abstract: A wound is defined as chronic when it does not heal according to the normal repair times and mechanisms. This particular condition may be principally due to local hypoxia. Carbon dioxide (CO(2)) therapy refers to the transcutaneous or subcutaneous administration of CO(2) for therapeutical effects on both microcirculation and tissue oxygenation. In this study, we report the clinical and instrumental results of the application of CO(2) in the therapy of chronic wounds. The study included 70 patients affected by chronic ulcers. The patients were selected by aetiology and wound extension and equally divided into two homogeneous groups. In group A, CO(2) therapy was used in addition to the routine methods of treatment for such lesions (surgical and/or chemical debridement, advanced dressings according to the features of each lesion). In group B, patients were treated using routine methods alone. Both groups underwent to instrumental (laser doppler flow, measurement of TcPO(2)), clinical and photographic evaluation. In the group that underwent subcutaneous treatment with CO(2) therapy, the results highlighted a significant increase in tissue oxygenation values, which was confirmed by greater progress of the lesions in terms of both healing and reduction of the injured area. Considering the safety, efficacy and reliability of this method, even if further studies are necessary, we believe that it is useful to include subcutaneous carbon dioxide therapy in the treatment of wounds involving hypoxia-related damage.

Title: Carboxytherapy: Controls the inflammation and enhances the production of fibronectin on wound healing under venous insufficiency

Authors: Brochado TMM, de Carvalho Schweich L, Di Pietro Simões N, Oliveira RJ, Antoniolli-Silva ACMB.

Journal: Int Wound J. 2019 Apr;16(2):316-324. doi: 10.1111/iwj.13031. Epub 2018 Nov 22. PMID: 30467979; PMCID: PMC7949331.

Link to full text: Carboxytherapy: Controls the inflammation and enhances the production of fibronectin on wound healing under venous insufficiency

Abstract: To examine the influence of carboxytherapy on wound healing under venous insufficiency, full‐thickness excisional wounds were created on Wistar rats. We used three groups with 32 rats each: Group (I): daily cleaning with 0.9% saline solution; Group Sulfadiazine (II): 1% silver sulfadiazine; and Carboxytherapy (III): subcutaneous application of 0.3 mL of carbon dioxide. The predetermined periods of analysis were the 3rd, 7th, 14th, and 30th day. The slides were stained with haematoxylin and eosin and Picrosirius red and submitted for immunohistochemistry. Groups II and III presented a statistically significant decrease in relation to the presence of neutrophilic and lymphocytic infiltrates. The presence of collagen significant increased in groups II and III. However, group III presented better organisation. Only group I maintained the neovascularisation until the 30th day. The new epithelium statistically significantly increased in groups II and III. On immunohistochemistry, regarding fibronectin expression, only group III demonstrated a statistically significant increase since the beginning of the healing process. Thus, the use of carboxytherapy promotes the formation of a tissue better structured and that may be an important resource for the treatment of wounds under venous insufficiency, especially those of recurrent re‐openings.

Title: Transdermal CO2 application in chronic wounds

Authors: Wollina U, Heinig B, Uhlemann C.

Journal: Int J Low Extrem Wounds. 2004 Jun;3(2):103-6. doi: 10.1177/1534734604265142. PMID: 15866797.

Link to PubMed: Transdermal CO2 application in chronic wounds

Abstract: Chronic wounds are a challenge to treatment. In this retrospective study, the effect of transdermal CO2 application on wound healing in chronic ulcers was investigated and compared to the effect of CO2 on acute surgical wounds. Eighty-six patients (52 females and 34 males) with chronic wounds of different origin except arterial occlusive disease were included. In addition, 17 patients (5 females, 12 males) with wide excision wounds after surgical therapy of acne inversa were considered. The indication for CO2 application was a wound at risk for infection. Treatment was performed with a Carboflow device once daily for 30 to 60 minutes. There was clinical evidence of improvement of granulation and reduction of discharge and malodor within 1 week of treatment in both chronic and acute wounds. Only 9 patients, all diabetics, needed an additional systemic antibiosis. The treatment was well tolerated. No adverse effects have been noted. Transdermal CO2 application is a useful method to reduce the risk of infection and improve wound healing in both chronic and certain acute wounds. Systematic prospective trials are needed.

Title: The impact of CO2 dry baths on subpopulations of NK and NK-T lymphocytes, cytotoxic activity, level of inflammation and pain management in elderly with musculosceletal syndromes - a pilot study

Authors: Kallistratos, E., Toliopoulos, I., Fragkoraptis, D., Gerou, S., & Fragkoraptis, E.

Journal: Experimental Medicine 1, no. 1 (2009).

Link to EBSCO: The impact of CO2 dry baths on subpopulations of NK and NK-T lymphocytes, cytotoxic activity, level of inflammation and pain management in elderly with musculosceletal syndromes - a pilot study

Abstract: Background - Aim of the study The effects of carbon dioxide-rich water bathing have been investigated in clinical research; CO2 is assumed to exert a direct action on vascular smooth muscles, induce vasodilatation and activate thermoreceptors of the skin. Our overall aim is to test our hypothesis that dry CO2 treatment's rehabilitative effect is achieved by using its immune boosting, anti-inflammatory and anti-depressive elements to encage the elderly's pathophysiologic mechanisms capable of supporting a repaired and significantly improved function. In particular, the present study compared the post-therapy effects with pre-therapy baseline of the treatment measuring immune mediated response, the levels of biochemical markers for chronic inflammatory conditions and assessing clinically the short - term effectiveness of pain management in a group of elderly with musculoskeletal syndromes. Material and Methods Repeated artificial CO2 dry treatments were performed on 37 volunteers with musculoskeletal pain disorders. Group A (intervention) comprised 27 patients (12 men and 15 women) and group B (placebo) tracked 10 patients (5 men and 5 women). Each patient was covered with a specific plastic hypoallergic bag up to neck level and underwent the treatment for 45 min. In group A the bag was filled with CO2 gas and in the placebo group B the bag was filled with fresh air only. The DCDB( Dry Carbon Dioxide Bath) treatment was performed for 17 consecutive days. Blood samples were drawn before and after the intervention period. The selected parameters included white blood cells, cytotoxicity, lymphocytes, NK, NK-T cells, β-endorphin, substance P and neurokinin A. Repeated measurements were made in heart frequency, arterial blood pressure and body temperature at the beginning and after all sessions. Results In the intervention group, a significant decrease was detected in the diastolic arterial pressure (only in women) and in the heart rate. Significant increase was measured in the white blood cells count, lymphocytes count (in men), NK-T cells count, the cytotoxicity measurements, β-endorphin and substance P levels as well as the neurokinin A levels in men. Interestingly, the intervention group members reported a kind of profound euphoria which remained during the intervention period and lasted for at least one month. The increase of Substance P and neurokinin A in the intervention group was not followed by any increase of pain intensity. In the control group, the parameters related to the arousal of the immune system remained unaffected but the levels of substance P and neurokinin A rose more sharply than in group A. Conclusions Dry CO2 bath seems to stimulate short-term immuneboosting responses that last during and after the intervention period. The rise in the endorphin levels explains the euphoric sensation reported by the intervention group patients. The effects of dry CO2 on the parasympathetic nervous system are induced probably via hemodynamic mechanisms; for the hemodynamic responses triggering, a direct action of dry CO2 on specific CO2-receptors in the skin is a possible physiologic pathway.

Title: The effect of transcutaneous application of gaseous CO2 on diabetic chronic wound healing-A double-blind randomized clinical trial

Authors: Macura M, Ban Frangez H, Cankar K, Finžgar M, Frangez I.

Journal: Int Wound J. 2020 Dec;17(6):1607-1614. doi: 10.1111/iwj.13436. Epub 2020 Jul 7. PMID: 32633896; PMCID: PMC7949258.

Link to full text: The effect of transcutaneous application of gaseous CO2 on diabetic chronic wound healing-A double-blind randomized clinical trial

Abstract: Chronic wounds in diabetics are difficult to treat, therefore, adjuvant therapies have been investigated. Bathing in CO2 -rich water (spa therapy) has been known in Europe for decades for its positive effect on peripheral vascular disorders. Recently, much effort has been invested in developing optimal application methods of CO2 . Uses include subcutaneous injections of CO2 , bathing in CO2 -enriched water, and transcutaneous application of CO2 . To verify the effect of transcutaneous application of gaseous CO2 on the healing of chronic diabetic wounds, a randomized double-blind clinical research was designed. The research included 30 and 27 wounds in the study and control groups, respectively. In addition to standard treatment, patients in the study group received 20 therapies with medical-grade CO2 gas and the control group received the same treatment with air. Results showed significantly faster healing in the study group: 20 of the 30 wounds in the study group were healed compared with none in the control group. Mean wound surface and volume in the study group was reduced significantly (surface: 96%, P = .001, volume: 99%, P = .003) compared with a small reduction in the control group (surface: 25%, P = .383, volume: 27%, P = .178). Considering our results, transcutaneous application of gaseous CO2 is an effective adjuvant therapy in diabetic chronic wound treatment.

Keywords: CO2 therapy; carboxy therapy; chronic wound treatment; diabetic chronic wound; diabetic foot ulcer; transcutaneous CO2 application.

Title: Effect of Transcutaneous Application of Carbon Dioxide on Wound Healing, Wound Recurrence Rate and Diabetic Polyneuropathy in Patients with Neuropathic, Ischemic and Neuroischemic Diabetes-Related Foot Ulcers

Authors: Bulum T, Poljičanin T, Badanjak A, Držič J, Metelko Ž.

Journal: Life (Basel). 2025 Apr 7;15(4):618. doi: 10.3390/life15040618. PMID: 40283172; PMCID: PMC12028706.

Link to full text: Effect of Transcutaneous Application of Carbon Dioxide on Wound Healing, Wound Recurrence Rate and Diabetic Polyneuropathy in Patients with Neuropathic, Ischemic and Neuroischemic Diabetes-Related Foot Ulcers

Abstract: (1) Background: Diabetes-related foot ulcers (DFUs) are a severe complication of diabetes mellitus (DM), with a five-year mortality rate of around 40%. Our study aimed to explore the effects of transcutaneous application of carbon dioxide (CO2 therapy) on DFU healing and recurrence rate, as well as diabetic polyneuropathy. (2) Methods: Adults with at least one chronic DFU were invited to undergo 20 50-min-long CO2 therapies within 4 weeks. After the completion of the last CO2 therapy, the effect of the therapies on wound healing and diabetic polyneuropathy was assessed, and 1 year later, we evaluated the incidence rate of DFU recurrence. (3) Results: Thirty-five subjects with DM and forty DFUs (ischemic: 15, neuropathic: 8, neuroischemic: 17) participated in our trial. After 4 weeks, 67.5% of all DFUs healed, whereas the reduction of the surface area of the unhealed wounds (74.0% ± 22.3%) was statistically significant. The restoration of protective sensations was also statistically significant. All unhealed wounds received standard care and healed within 2 additional weeks. The recurrence rate after 1 year was 17.5%. None of the patients required antibiotic treatment, hospitalization, or amputation. (4) Conclusion: CO2 therapy is a promising therapeutic intervention for treating DFUs and improving diabetic polyneuropathy.

Keywords: CO2 therapy; diabetes-related foot ulcer; diabetic foot; distal symmetrical polyneuropathy; loss of protective sensation; transcutaneous CO2 application; wound healing.

Title: Pain in the elderly: Prospective study of hyperbaric CO2 cryotherapy (neurocryostimulation)

Authors: Chatap G, De Sousa A, Giraud K, Vincent JP; Acute Pain in the Elderly Study Group.

Journal: Joint Bone Spine. 2007 Dec;74(6):617-21. doi: 10.1016/j.jbspin.2006.10.011. Epub 2007 Jul 19. PMID: 17897861.

Link to PubMed: Pain in the elderly: Prospective study of hyperbaric CO2 cryotherapy (neurocryostimulation)

Abstract: Objective: To evaluate the analgesic effects of hyperbaric CO(2) cryotherapy in elderly inpatients.

Methods: An open-label prospective study was conducted in two geriatrics departments in patients with a broad range of pain characteristics. Each patient underwent a physical evaluation followed by hyperbaric CO(2) cryotherapy sessions, whose spacing and number were at the discretion of the physiotherapist. Patients completed a 100-mm visual analog scale for pain severity before and after the sessions.

Results: We included 51 patients, who were treated between May 2 and June 30, 2005. Mean age was 83.7years, and the female-to-male ratio was 4/1. The patients had acute or chronic pain whose origin was usually musculoskeletal (80.3%) or neurological (18.6%). Pain scores decreased significantly after four sessions, from 52mm to 13mm (P<0.001) in patients with acute pain and from 45mm to 13mm (P<0.001) in those with chronic pain.

Conclusion: Hyperbaric CO(2) cryotherapy is an innovative tool that should be incorporated within the non-pharmacological armamentarium for achieving pain relief in older patients.

Title: Synergistic effect of pressurized carbon dioxide and sodium hypochlorite on the inactivation of Enterococcus sp. in seawater

Authors: Dang TT, Imai T, Le TV, Nguyen DK, Higuchi T, Kanno A, Yamamoto K, Sekine M.

Journal: Water Res. 2016 Dec 1;106:204-213. doi: 10.1016/j.watres.2016.10.003. Epub 2016 Oct 3. PMID: 27721172.

Link to PubMed: Synergistic effect of pressurized carbon dioxide and sodium hypochlorite on the inactivation of Enterococcus sp. in seawater

Abstract: This study investigated the effect of combined treatments using pressurized carbon dioxide (PCD) and sodium hypochlorite (NaOCl) on the inactivation of Enterococcus sp. in artificial seawater. Bacterial inactivation was conducted in a liquid-film-forming apparatus with various pressure conditions, CO2 supply rates, and chlorine dosages. Combined PCD/chlorine treatments resulted in greater disinfection efficiency than those for the two individual treatments. Synergy values were correlated with pressure and CO2 concentrations (p < 0.001). Combination of 0.9 MPa PCD (various CO2 supply rates: 25% CO2 + 75% N2, 50% CO2 + 50% N2, and 100% CO2) and chlorine (0.20 mg L-1) yielded average synergy values of 4.9, 5.2, and 4.4 log, respectively, within 3 min. Combined treatment with PCD (100% CO2, 0.3 MPa, and 20 °C) and chlorine (0.20-0.22 mg L-1) achieved an average synergy value of 4.6 log and complete inactivation (5.2-5.5 log reductions) of Enterococcus sp. within 4 min. In contrast, when the two individual treatments (PCD and chlorine) were used, only 3.7 and 1.8-2.3 log reductions, respectively, were achieved after 25 min. These findings suggest that the combined PCD/chlorine treatment has synergistic benefits and provides a promising method for the disinfection of ballast water.

Title: Antimicrobial Effect of Pressurized Carbon Dioxide on Staphylococcus aureus in Broth and Milk

Authors: Osman Erkmen

Journal: LWT - Food Science and Technology, Volume 30, Issue 8, 1997, Pages 826-829, ISSN 0023-6438, https://doi.org/10.1006/fstl.1997.0277.

Link to ScienceDirect: Antimicrobial Effect of Pressurized Carbon Dioxide on Staphylococcus aureus in Broth and Milk

Abstract: Studies were carried out to assess the use of high pressure CO2 treatment to inhibit Staphylococcus aureus in broth and raw cow's milk at 25 °C. Two phases in the survival curves were observed. The early stage was characterized by a slow rate of decrease in number of S. aureus; this rate increased sharply at the later stage. Staphylococcus aureus suspended in broth was completely inactivated after CO2 treatment at 7 MPa for 100 min and 8 MPa for 60 min. The sterilization effects of CO2 on both S. aureus and aerobic bacteria were observed at 14.6 MPa for 5 h and 9 MPa for 2 h in whole and skim milk, respectively. Staphylococcus aureus and aerobic bacteria were more difficult to inactivate when they were suspended in whole milk that may have protected the cell from penetration by CO2. Reduction rates of S. aureus and aerobic bacteria were sensitive to pressure, exposure time and the suspending medium.

Title: Humidified Warmed CO2 Treatment Therapy Strategies Can Save Lives With Mitigation and Suppression of SARS-CoV-2 Infection: An Evidence Review

Authors: El-Betany AMM, Behiry EM, Gumbleton M, Harding KG.

Journal: Front Med (Lausanne). 2020 Dec 11;7:594295. doi: 10.3389/fmed.2020.594295. PMID: 33425942; PMCID: PMC7793941.

Link to full text: Humidified Warmed CO2 Treatment Therapy Strategies Can Save Lives With Mitigation and Suppression of SARS-CoV-2 Infection: An Evidence Review

Abstract: The coronavirus disease (COVID-19) outbreak has presented enormous challenges for healthcare, societal, and economic systems worldwide. There is an urgent global need for a universal vaccine to cover all SARS-CoV-2 mutant strains to stop the current COVID-19 pandemic and the threat of an inevitable second wave of coronavirus. Carbon dioxide is safe and superior antimicrobial, which suggests it should be effective against coronaviruses and mutants thereof. Depending on the therapeutic regime, CO2 could also ameliorate other COVID-19 symptoms as it has also been reported to have antioxidant, anti-inflammation, anti-cytokine effects, and to stimulate the human immune system. Moreover, CO2 has beneficial effects on respiratory physiology, cardiovascular health, and human nervous systems. This article reviews the rationale of early treatment by inhaling safe doses of warmed humidified CO2 gas, either alone or as a carrier gas to deliver other inhaled drugs may help save lives by suppressing SARS-CoV-2 infections and excessive inflammatory responses. We suggest testing this somewhat counter-intuitive, but low tech and safe intervention for its suitability as a preventive measure and treatment against COVID-19. Overall, development and evaluation of this therapy now may provide a safe and economical tool for use not only during the current pandemic but also for any future outbreaks of respiratory diseases and related conditions.

Title: Virus and bacteria inactivation by CO2 bubbles in solution

Authors: Garrido Sanchis, A., Pashley, R. & Ninham, B.

Journal: npj Clean Water 2, 5 (2019). https://doi.org/10.1038/s41545-018-0027-5

Link to full text: Virus and bacteria inactivation by CO2 bubbles in solution

Abstract: The availability of clean water is a major problem facing the world. In particular, the cost and destruction caused by viruses in water remains an unresolved challenge and poses a major limitation on the use of recycled water. Here, we develop an environmentally friendly technology for sterilising water. The technology bubbles heated un-pressurised carbon dioxide or exhaust gases through wastewater in a bubble column, effectively destroying both bacteria and viruses. The process is extremely cost effective, with no concerning by-products, and has already been successfully scaled-up industrially.

Title: Carbon Dioxide as a Microbial Toxicity Enhancer of Some Antibacterial Agents: A New Potential Water Purification Tool

Authors: Varsik Martirosyan, Karlen Hovnanyan, Sinerik Ayrapetyan

Journal: International Scholarly Research Notices, 22 April 2012, https://doi.org/10.5402/2012/906761

Link to full text: Carbon Dioxide as a Microbial Toxicity Enhancer of Some Antibacterial Agents: A New Potential Water Purification Tool

Abstract: The aim of current paper was to investigate the possibility of increasing the toxicity of calcium hypochlorite (Ca(ClO)2) and hydrogen peroxide (H2O2) on Escherichia coli K-12 by preliminary enrichment of culture media by carbon dioxide (CO2). For this purpose, the microbes sensitivity to H2O2 or/and Ca(ClO)2 at normal and CO2-enriched medium was studied by spectrophotometric, radioisotopic, and electronmicroscopic methods. Ten-minute preincubation in CO2-enriched medium enhanced the toxic effect of both H2O2 or/and Ca(ClO)2 on bacteria as a result of induced growth inhibition, compared to no-CO2 enriched group. Additionally, changes in cell morphology and proliferation were observed. It was demonstrated that the preliminary incubation of microbes in CO2-enriched culture media in no supercritical concentration elevate the toxic effect of H2O2 or/and Ca(ClO)2 on microbes. This can serve as a novel, effective, inexpensive, and environmentally friendly approach for water purification from bacteria, further improving the protection of the environment and human health.

Title: Carbon dioxide inhibits the growth rate of Staphylococcus aureus at body temperature

Authors: Persson M, Svenarud P, Flock JI, van der Linden J.

Journal: Surg Endosc. 2005 Jan;19(1):91-4. doi: 10.1007/s00464-003-9334-z. Epub 2004 Nov 11. PMID: 15529188.

Link to PubMed: Carbon dioxide inhibits the growth rate of Staphylococcus aureus at body temperature

Abstract:
Background: Since the 1930s, carbon dioxide (CO(2)) has been combined with cold storage for the preservation of food. However, its use for the prevention of surgical wound infection was long considered to be impractical. Now CO(2) is widely used during laparoscopic procedures, and a method has been developed to create a CO(2) atmosphere in an open wound. The aim of this study was to investigate the effect of CO(2) on the growth of Staphylococcus aureus at body temperature.

Methods: First, S. aureus inoculated on blood agar were exposed to pure CO(2) (100%), standard anaerobic gas (5% CO(2), 10% hydrogen, 85% nitrogen), or air at 37 degrees C for a period of 24 h; then a viable count of the bacteria was made. Second, S. aureus inoculated in brain-heart infusion broth and kept at 37 degrees C were exposed to CO(2) or air for 0, 2, 4, 6, and 8 h; then the optical density of the bacteria was measured.

Results: After 24 h, the number of S. aureus on blood agar was about 100 times lower in CO(2) than in anaerobic gas (p = 0.001) and about 1,000 times lower than in air (p = 0.001). Also, in broth, there were fewer bacteria with CO(2) than with air (p < 0.01). After 2 h, the number of bacteria was increased with air (p < 0.001) but not with CO(2) (p = 0.13). After 8 h, the optical density had increased from zero to 1.2 with air but it had increased only to 0.01 with CO(2) (p = 0.001).

Conclusion: Pure CO(2) significantly decreased the growth rate of S. aureus at body temperature. The inhibitory effect of CO(2) increased exponentially with time. Its bacteriostatic effect may help to explain the low infection rates in patients who undergo laparoscopic procedures.

Title: Intraoperative CO2 insufflation can decrease the risk of surgical site infection

Authors: Persson M, van der Linden J.

Journal: Med Hypotheses. 2008;71(1):8-13. doi: 10.1016/j.mehy.2007.12.016. Epub 2008 Mar 4. PMID: 18304752.

Link to PubMed: Intraoperative CO2 insufflation can decrease the risk of surgical site infection

Abstract: Surgical wound infections may ruin otherwise successful operations, and are associated with extended hospital stay, extra costs, and high mortality rates. In open surgery the wound's exposure to ambient air increases the risk of wound infection via several independent factors. The open surgical wound is subjected to airborne bacterial contamination, desiccation, and heat loss that increase the bacterial load, cause superficial necrosis, and impair tissue oxygenation and cellular immune functions, respectively. The present hypothesis is that topically applied carbon dioxide in the open surgical wound can be used intraoperatively to avoid these risks, and thus help to prevent postoperative wound infection. We also criticize existing methods and describe the theoretical background and supporting evidence for our suggested method. If the hypothesis would prove to be correct in a clinical trial, the new method may be an effective complement, or even an alternative, to antibiotics in preventing surgical site infection.

Title: Temperature-dependent innate defense against the common cold virus limits viral replication at warm temperature in mouse airway cells

Authors: Foxman EF, Storer JA, Fitzgerald ME, Wasik BR, Hou L, Zhao H, Turner PE, Pyle AM, Iwasaki A.

Journal: Proc Natl Acad Sci U S A. 2015 Jan 20;112(3):827-32. doi: 10.1073/pnas.1411030112. Epub 2015 Jan 5. PMID: 25561542; PMCID: PMC4311828.

Link to full text: Temperature-dependent innate defense against the common cold virus limits viral replication at warm temperature in mouse airway cells

Abstract: Most isolates of human rhinovirus, the common cold virus, replicate more robustly at the cool temperatures found in the nasal cavity (33-35 °C) than at core body temperature (37 °C). To gain insight into the mechanism of temperature-dependent growth, we compared the transcriptional response of primary mouse airway epithelial cells infected with rhinovirus at 33 °C vs. 37 °C. Mouse airway cells infected with mouse-adapted rhinovirus 1B exhibited a striking enrichment in expression of antiviral defense response genes at 37 °C relative to 33 °C, which correlated with significantly higher expression levels of type I and type III IFN genes and IFN-stimulated genes (ISGs) at 37 °C. Temperature-dependent IFN induction in response to rhinovirus was dependent on the MAVS protein, a key signaling adaptor of the RIG-I-like receptors (RLRs). Stimulation of primary airway cells with the synthetic RLR ligand poly I:C led to greater IFN induction at 37 °C relative to 33 °C at early time points poststimulation and to a sustained increase in the induction of ISGs at 37 °C relative to 33 °C. Recombinant type I IFN also stimulated more robust induction of ISGs at 37 °C than at 33 °C. Genetic deficiency of MAVS or the type I IFN receptor in infected airway cells permitted higher levels of viral replication, particularly at 37 °C, and partially rescued the temperature-dependent growth phenotype. These findings demonstrate that in mouse airway cells, rhinovirus replicates preferentially at nasal cavity temperature due, in part, to a less efficient antiviral defense response of infected cells at cool temperature.

Keywords: RIG-I; airway; common cold; innate immunity; rhinovirus.