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With current plans for manned missions to Mars and beyond, the need to better understand, prevent, and counteract the harmful effects of long-duration spaceflight on the body is becoming increasingly important. In this study, an automated heart-on-a-chip platform was flown to the International Space Station on a 1-mo mission during which contractile cardiac function was monitored in real-time. Upon return to Earth, engineered human heart tissues (EHTs) were further analyzed with ultrastructural imaging and RNA sequencing to investigate the impact of prolonged microgravity on cardiomyocyte function and health. Spaceflight EHTs exhibited significantly reduced twitch forces, increased incidences of arrhythmias, and increased signs of sarcomere disruption and mitochondrial damage. Transcriptomic analyses showed an up-regulation of genes and pathways associated with metabolic disorders, heart failure, oxidative stress, and inflammation, while genes related to contractility and calcium signaling showed significant down-regulation. Finally, in silico modeling revealed a potential link between oxidative stress and mitochondrial dysfunction that corresponded with RNA sequencing results. This represents an in vitro model to faithfully reproduce the adverse effects of spaceflight on three-dimensional (3D)-engineered heart tissue., Competing Interests: Competing interests statement:D.-H.K. is a co-founder, scientific advisory board member, and equity holder of Curi Bio, Inc. N.J.S. is a co-founder with equity of Stasys Medical Corporation and is a scientific advisory board member and equity holder of Curi Bio, Inc. J.H.T. is an employee and equity holder of Tenaya Therapeutics, Inc. The other authors declare no competing interest.

Importance: Understanding potential predisposing factors associated with spaceflight-associated neuro-ocular syndrome (SANS) may influence its management., Objective: To describe a severe case of SANS associated with 2 potentially predisposing factors., Design, Setting, and Participants: Ocular testing of and blood collections from a female astronaut were completed preflight, inflight, and postflight in the setting of the International Space Station (ISS)., Exposure: Weightlessness throughout an approximately 6-month ISS mission. Mean carbon dioxide (CO2) partial pressure decreased from 2.6 to 1.3 mm Hg weeks before the astronaut's flight day (FD) 154 optical coherence tomography (OCT) session. In response to SANS, 4 B-vitamin supplements (vitamin B6, 100 mg; L-methylfolate, 5 mg; vitamin B12, 1000 μg; and riboflavin, 400 mg) were deployed, unpacked on FD153, consumed daily through FD169, and then discontinued due to gastrointestinal discomfort., Main Outcomes and Measures: Refraction, distance visual acuity (DVA), optic nerve, and macular assessment on OCT., Results: Cycloplegic refraction was -1.00 diopter in both eyes preflight and +0.50 - 0.25 × 015 in the right eye and +1.00 diopter in the left eye 3 days postflight. Uncorrected DVA was 20/30 OU preflight, 20/16 or better by FD90, and 20/15 OU 3 days postflight. Inflight peripapillary total retinal thickness (TRT) peaked between FD84 and FD126 (right eye, 401 μm preflight, 613 μm on FD84; left eye, 404 μm preflight, 636 μm on FD126), then decreased. Peripapillary choroidal folds, quantified by surface roughness, peaked at 12.7 μm in the right eye on FD154 and 15.0 μm in the left eye on FD126, then decreased. Mean choroidal thickness increased throughout the mission. Genetic analyses revealed 2 minor alleles for MTRR 66 and 2 major alleles for SHMT1 1420 (ie, 4 of 4 SANS risk alleles). One-week postflight, lumbar puncture opening pressure was normal, at 19.4 cm H2O., Conclusions and Relevance: To the authors' knowledge, no other report of SANS documented as large of a change in peripapillary TRT or hyperopic shift during a mission as in this astronaut, and this was only 1 of 4 astronauts to experience chorioretinal folds approaching the fovea. This case showed substantial inflight improvement greater than the sensitivity of the measure, possibly associated with B-vitamin supplementation and/or reduction in cabin CO2. However, as a single report, such improvement could be coincidental to these interventions, warranting further evaluation.

The aerobic dive limit (ADL) and the hypothesis that most dives are aerobic in nature have become fundamental to the understanding of diving physiology and to the interpretation of diving behavior and foraging ecology of marine mammals and seabirds. An ADL, the dive duration associated with the onset of post-dive blood lactate accumulation, has only been documented with blood lactate analyses in five species. Applications to other species have involved behavioral estimates or use of an oxygen store / metabolic rate formula. Both approaches have limitations, but have proved useful to the evaluation of the dive behavior and ecology of many species., (Copyright © 2020. Published by Elsevier Inc.)

Among the physiological consequences of extended spaceflight are loss of skeletal muscle and bone mass. One signaling pathway that plays an important role in maintaining muscle and bone homeostasis is that regulated by the secreted signaling proteins, myostatin (MSTN) and activin A. Here, we used both genetic and pharmacological approaches to investigate the effect of targeting MSTN/activin A signaling in mice that were sent to the International Space Station. Wild type mice lost significant muscle and bone mass during the 33 d spent in microgravity. Muscle weights of Mstn -/- mice, which are about twice those of wild type mice, were largely maintained during spaceflight. Systemic inhibition of MSTN/activin A signaling using a soluble form of the activin type IIB receptor (ACVR2B), which can bind each of these ligands, led to dramatic increases in both muscle and bone mass, with effects being comparable in ground and flight mice. Exposure to microgravity and treatment with the soluble receptor each led to alterations in numerous signaling pathways, which were reflected in changes in levels of key signaling components in the blood as well as their RNA expression levels in muscle and bone. These findings have implications for therapeutic strategies to combat the concomitant muscle and bone loss occurring in people afflicted with disuse atrophy on Earth as well as in astronauts in space, especially during prolonged missions., Competing Interests: The authors declare no competing interest., (Copyright © 2020 the Author(s). Published by PNAS.)

The bar-headed goose is famed for migratory flight at extreme altitude. To better understand the physiology underlying this remarkable behavior, we imprinted and trained geese, collecting the first cardiorespiratory measurements of bar-headed geese flying at simulated altitude in a wind tunnel. Metabolic rate during flight increased 16-fold from rest, supported by an increase in the estimated amount of O 2 transported per heartbeat and a modest increase in heart rate. The geese appear to have ample cardiac reserves, as heart rate during hypoxic flights was not higher than in normoxic flights. We conclude that flight in hypoxia is largely achieved via the reduction in metabolic rate compared to normoxia. Arterial [Formula: see text] was maintained throughout flights. Mixed venous P O2 decreased during the initial portion of flights in hypoxia, indicative of increased tissue O 2 extraction. We also discovered that mixed venous temperature decreased during flight, which may significantly increase oxygen loading to hemoglobin., Competing Interests: JM, JY, BC, WJ, LH, WM No competing interests declared

Synopsis: Exercise at high altitude is extremely challenging, largely due to hypobaric hypoxia (low oxygen levels brought about by low air pressure). In humans, the maximal rate of oxygen consumption decreases with increasing altitude, supporting progressively poorer performance. Bar-headed geese (Anser indicus) are renowned high altitude migrants and, although they appear to minimize altitude during migration where possible, they must fly over the Tibetan Plateau (mean altitude 4800 m) for much of their annual migration. This requires considerable cardiovascular effort, but no study has assessed the extent to which bar-headed geese may train prior to migration for long distances, or for high altitudes. Using implanted loggers that recorded heart rate, acceleration, pressure, and temperature, we found no evidence of training for migration in bar-headed geese. Geese showed no significant change in summed activity per day or maximal activity per day. There was also no significant change in maximum heart rate per day or minimum resting heart rate, which may be evidence of an increase in cardiac stroke volume if all other variables were to remain the same. We discuss the strategies used by bar-headed geese in the context of training undertaken by human mountaineers when preparing for high altitude, noting the differences between their respective cardiovascular physiology., (© The Author 2017. Published by Oxford University Press on behalf of the Society for Integrative and Comparative Biology. All rights reserved. For permissions please email: journals.permissions@oup.com.)

While bar-headed geese are renowned for migration at high altitude over the Himalayas, previous work on captive birds suggested that these geese are unable to maintain rates of oxygen consumption while running in severely hypoxic conditions. To investigate this paradox, we re-examined the running performance and heart rates of bar-headed geese and barnacle geese (a low altitude species) during exercise in hypoxia. Bar-headed geese (n = 7) were able to run at maximum speeds (determined in normoxia) for 15 minutes in severe hypoxia (7% O2; simulating the hypoxia at 8500 m) with mean heart rates of 466±8 beats min-1. Barnacle geese (n = 10), on the other hand, were unable to complete similar trials in severe hypoxia and their mean heart rate (316 beats.min-1) was significantly lower than bar-headed geese. In bar-headed geese, partial pressures of oxygen and carbon dioxide in both arterial and mixed venous blood were significantly lower during hypoxia than normoxia, both at rest and while running. However, measurements of blood lactate in bar-headed geese suggested that anaerobic metabolism was not a major energy source during running in hypoxia. We combined these data with values taken from the literature to estimate (i) oxygen supply, using the Fick equation and (ii) oxygen demand using aerodynamic theory for bar-headed geese flying aerobically, and under their own power, at altitude. This analysis predicts that the maximum altitude at which geese can transport enough oxygen to fly without environmental assistance ranges from 6,800 m to 8,900 m altitude, depending on the parameters used in the model but that such flights should be rare.

Although energetics is fundamental to animal ecology, traditional methods of determining metabolic rate are neither direct nor instantaneous. Recently, continuous blood oxygen (O2) measurements were used to assess energy expenditure in diving elephant seals (Mirounga angustirostris), demonstrating that an exceptional hypoxemic tolerance and exquisite management of blood O2 stores underlie the extraordinary diving capability of this consummate diver. As the detailed relationship of energy expenditure and dive behavior remains unknown, we integrated behavior, ecology, and physiology to characterize the costs of different types of dives of elephant seals. Elephant seal dive profiles were analyzed and O2 utilization was classified according to dive type (overall function of dive: transit, foraging, food processing/rest). This is the first account linking behavior at this level with in vivo blood O2 measurements in an animal freely diving at sea, allowing us to assess patterns of O2 utilization and energy expenditure between various behaviors and activities in an animal in the wild. In routine dives of elephant seals, the blood O2 store was significantly depleted to a similar range irrespective of dive function, suggesting that all dive types have equal costs in terms of blood O2 depletion. Here, we present the first physiological evidence that all dive types have similarly high blood O2 demands, supporting an energy balance strategy achieved by devoting one major task to a given dive, thereby separating dive functions into distinct dive types. This strategy may optimize O2 store utilization and recovery, consequently maximizing time underwater and allowing these animals to take full advantage of their underwater resources. This approach may be important to optimizing energy expenditure throughout a dive bout or at-sea foraging trip and is well suited to the lifestyle of an elephant seal, which spends > 90% of its time at sea submerged making diving its most "natural" state.

The bar-headed goose (Anser indicus) crosses the Himalaya twice a year at altitudes where oxygen (O2) levels are less than half those at sea level and temperatures are below -20°C. Although it has been known for over three decades that the major hemoglobin (Hb) component of bar-headed geese has an increased affinity for O2, enhancing O2 uptake, the effects of temperature and interactions between temperature and pH on bar-headed goose Hb-O2 affinity have not previously been determined. An increase in breathing of the hypoxic and extremely cold air experienced by a bar-headed goose at altitude (due to the enhanced hypoxic ventilatory response in this species) could result in both reduced temperature and reduced levels of CO2 at the blood-gas interface in the lungs, enhancing O2 loading. In addition, given the strenuous nature of flapping flight, particularly in thin air, blood leaving the exercising muscle should be warm and acidotic, facilitating O2 unloading. To explore the possibility that features of blood biochemistry in this species could further enhance O2 delivery, we determined the P50 (the partial pressure of O2 at which Hb is 50% saturated) of whole blood from bar-headed geese under conditions of varying temperature and [CO2]. We found that blood-O2 affinity was highly temperature sensitive in bar-headed geese compared with other birds and mammals. Based on our analysis, temperature and pH effects acting on blood-O2 affinity (cold alkalotic lungs and warm acidotic muscle) could increase O2 delivery by twofold during sustained flapping flight at high altitudes compared with what would be delivered by blood at constant temperature and pH.

Since the introduction of the aerobic dive limit (ADL) 30 years ago, the concept that most dives of marine mammals and sea birds are aerobic in nature has dominated the interpretation of their diving behavior and foraging ecology. Although there have been many measurements of body oxygen stores, there have been few investigations of the actual depletion of those stores during dives. Yet, it is the pattern, rate and magnitude of depletion of O(2) stores that underlie the ADL. Therefore, in order to assess strategies of O(2) store management, we review (a) the magnitude of O(2) stores, (b) past studies of O(2) store depletion and (c) our recent investigations of O(2) store utilization during sleep apnea and dives of elephant seals (Mirounga angustirostris) and during dives of emperor penguins (Aptenodytes forsteri). We conclude with the implications of these findings for (a) the physiological responses underlying O(2) store utilization, (b) the physiological basis of the ADL and (c) the value of extreme hypoxemic tolerance and the significance of the avoidance of re-perfusion injury in these animals.

The physiological basis of the aerobic dive limit (ADL), the dive duration associated with the onset of post-dive blood lactate elevation, is hypothesized to be depletion of the muscle oxygen (O(2)) store. A dual wavelength near-infrared spectrophotometer was developed and used to measure myoglobin (Mb) O(2) saturation levels in the locomotory muscle during dives of emperor penguins (Aptenodytes forsteri). Two distinct patterns of muscle O(2) depletion were observed. Type A dives had a monotonic decline, and, in dives near the ADL, the muscle O(2) store was almost completely depleted. This pattern of Mb desaturation was consistent with lack of muscle blood flow and supports the hypothesis that the onset of post-dive blood lactate accumulation is secondary to muscle O(2) depletion during dives. The mean type A Mb desaturation rate allowed for calculation of a mean muscle O(2) consumption of 12.4 ml O(2) kg(-1) muscle min(-1), based on a Mb concentration of 6.4 g 100 g(-1) muscle. Type B desaturation patterns demonstrated a more gradual decline, often reaching a mid-dive plateau in Mb desaturation. This mid-dive plateau suggests maintenance of some muscle perfusion during these dives. At the end of type B dives, Mb desaturation rate increased and, in dives beyond the ADL, Mb saturation often reached near 0%. Thus, although different physiological strategies may be used during emperor penguin diving, both Mb desaturation patterns support the hypothesis that the onset of post-dive lactate accumulation is secondary to muscle O(2) store depletion.

Hypothermia-induced reductions in metabolic rate have been proposed to suppress metabolism and prolong the duration of aerobic metabolism during dives of marine mammals and birds. To determine whether core hypothermia might contribute to the repetitive long-duration dives of the northern elephant seal Mirounga angustirostris, blood temperature profiles were obtained in translocated juvenile elephant seals equipped with a thermistor and backpack recorder. Representative temperature (the y-intercept of the mean temperature vs. dive duration relationship) was 37.2 degrees C +/- 0.6 degrees C (n=3 seals) in the extradural vein, 38.1 degrees C +/- 0.7 degrees C (n = 4 seals) in the hepatic sinus, and 38.8 degrees +/- 1.6 degrees C (n = 6 deals) in the aorta. Mean temperature was significantly though weakly negatively related to dive duration in all but one seal. Mean venous temperatures of all dives of individual seals ranged between 36 degrees and 38 degrees C, while mean arterial temperatures ranged between 35 degrees and 39 degrees C. Transient decreases in venous and arterial temperatures to as low as 30 degrees -33 degrees C occurred in some dives >30 min (0.1% of dives in the study). The lack of significant core hypothermia during routine dives (10-30 min) and only a weak negative correlation of mean temperature with dive duration do not support the hypothesis that a cold-induced Q(10) effect contributes to metabolic suppression of central tissues during dives. The wide range of arterial temperatures while diving and the transient declines in temperature during long dives suggest that alterations in blood flow patterns and peripheral heat loss contribute to thermoregulation during diving.

The emperor penguin (Aptenodytes forsteri) thrives in the Antarctic underwater environment, diving to depths greater than 500 m and for durations longer than 23 min. To examine mechanisms underlying the exceptional diving ability of this species and further describe blood oxygen (O2) transport and depletion while diving, we characterized the O2-hemoglobin (Hb) dissociation curve of the emperor penguin in whole blood. This allowed us to (1) investigate the biochemical adaptation of Hb in this species, and (2) address blood O2 depletion during diving, by applying the dissociation curve to previously collected partial pressure of O2 (PO2) profiles to estimate in vivo Hb saturation (SO2) changes during dives. This investigation revealed enhanced Hb-O2 affinity (P50=28 mmHg, pH 7.5) in the emperor penguin, similar to high-altitude birds and other penguin species. This allows for increased O2 at low blood PO2 levels during diving and more complete depletion of the respiratory O2 store. SO2 profiles during diving demonstrated that arterial SO2 levels are maintained near 100% throughout much of the dive, not decreasing significantly until the final ascent phase. End-of-dive venous SO2 values were widely distributed and optimization of the venous blood O2 store resulted from arterialization and near complete depletion of venous blood O2 during longer dives. The estimated contribution of the blood O2 store to diving metabolic rate was low and highly variable. This pattern is due, in part, to the influx of O2 from the lungs into the blood during diving, and variable rates of tissue O2 uptake.

Species that maintain aerobic metabolism when the oxygen (O(2)) supply is limited represent ideal models to examine the mechanisms underlying tolerance to hypoxia. The repetitive, long dives of northern elephant seals (Mirounga angustirostris) have remained a physiological enigma as O(2) stores appear inadequate to maintain aerobic metabolism. We evaluated hypoxemic tolerance and blood O(2) depletion by 1) measuring arterial and venous O(2) partial pressure (Po(2)) during dives with a Po(2)/temperature recorder on elephant seals, 2) characterizing the O(2)-hemoglobin (O(2)-Hb) dissociation curve of this species, 3) applying the dissociation curve to Po(2) profiles to obtain %Hb saturation (So(2)), and 4) calculating blood O(2) store depletion during diving. Optimization of O(2) stores was achieved by high venous O(2) loading and almost complete depletion of blood O(2) stores during dives, with net O(2) content depletion values up to 91% (arterial) and 100% (venous). In routine dives (>10 min) Pv(O(2)) and Pa(O(2)) values reached 2-10 and 12-23 mmHg, respectively. This corresponds to So(2) of 1-26% and O(2) contents of 0.3 (venous) and 2.7 ml O(2)/dl blood (arterial), demonstrating remarkable hypoxemic tolerance as Pa(O(2)) is nearly equivalent to the arterial hypoxemic threshold of seals. The contribution of the blood O(2) store alone to metabolic rate was nearly equivalent to resting metabolic rate, and mean temperature remained near 37 degrees C. These data suggest that elephant seals routinely tolerate extreme hypoxemia during dives to completely utilize the blood O(2) store and maximize aerobic dive duration.

In order to further define O(2) store utilization during dives and understand the physiological basis of the aerobic dive limit (ADL, dive duration associated with the onset of post-dive blood lactate accumulation), emperor penguins (Aptenodytes forsteri) were equipped with either a blood partial pressure of oxygen (P(O(2))) recorder or a blood sampler while they were diving at an isolated dive hole in the sea ice of McMurdo Sound, Antarctica. Arterial P(O(2)) profiles (57 dives) revealed that (a) pre-dive P(O(2)) was greater than that at rest, (b) P(O(2)) transiently increased during descent and (c) post-dive P(O(2)) reached that at rest in 1.92+/-1.89 min (N=53). Venous P(O(2)) profiles (130 dives) revealed that (a) pre-dive venous P(O(2)) was greater than that at rest prior to 61% of dives, (b) in 90% of dives venous P(O(2)) transiently increased with a mean maximum P(O(2)) of 53+/-18 mmHg and a mean increase in P(O(2)) of 11+/-12 mmHg, (c) in 78% of dives, this peak venous P(O(2)) occurred within the first 3 min, and (d) post-dive venous P(O(2)) reached that at rest within 2.23+/-2.64 min (N=84). Arterial and venous P(O(2)) values in blood samples collected 1-3 min into dives were greater than or near to the respective values at rest. Blood lactate concentration was less than 2 mmol l(-1) as far as 10.5 min into dives, well beyond the known ADL of 5.6 min. Mean arterial and venous P(N(2)) of samples collected at 20-37 m depth were 2.5 times those at the surface, both being 2.1+/-0.7 atmospheres absolute (ATA; N=3 each), and were not significantly different. These findings are consistent with the maintenance of gas exchange during dives (elevated arterial and venous P(O(2)) and P(N(2)) during dives), muscle ischemia during dives (elevated venous P(O(2)), lack of lactate washout into blood during dives), and arterio-venous shunting of blood both during the surface period (venous P(O(2)) greater than that at rest) and during dives (arterialized venous P(O(2)) values during descent, equivalent arterial and venous P(N(2)) values during dives). These three physiological processes contribute to the transfer of the large respiratory O(2) store to the blood during the dive, isolation of muscle metabolism from the circulation during the dive, a decreased rate of blood O(2) depletion during dives, and optimized loading of O(2) stores both before and after dives. The lack of blood O(2) depletion and blood lactate elevation during dives beyond the ADL suggests that active locomotory muscle is the site of tissue lactate accumulation that results in post-dive blood lactate elevation in dives beyond the ADL.

To investigate the diving heart rate (f(H)) response of the emperor penguin (Aptenodytes forsteri), the consummate avian diver, birds diving at an isolated dive hole in McMurdo Sound, Antarctica were outfitted with digital electrocardiogram recorders, two-axis accelerometers and time depth recorders (TDRs). In contrast to any other freely diving bird, a true bradycardia (f(H) significantly

Blood gas analyses from emperor penguins (Aptenodytes forsteri) at rest, and intravascular P(O(2)) profiles from free-diving birds were obtained in order to examine hypoxemic tolerance and utilization of the blood O(2) store during dives. Analysis of blood samples from penguins at rest revealed arterial P(O(2))s and O(2) contents of 68+/-7 mmHg (1 mmHg= 133.3 Pa) and 22.5+/-1.3 ml O(2) dl(-1) (N=3) and venous values of 41+/-10 mmHg and 17.4+/-2.9 ml O(2) dl(-1) (N=9). Corresponding arterial and venous Hb saturations for a hemoglobin (Hb) concentration of 18 g dl(-1) were >91% and 70%, respectively. Analysis of P(O(2)) profiles obtained from birds equipped with intravascular P(O(2)) electrodes and backpack recorders during dives revealed that (1) the decline of the final blood P(O(2)) of a dive in relation to dive duration was variable, (2) final venous P(O(2)) values spanned a 40-mmHg range at the previously measured aerobic dive limit (ADL; dive duration associated with onset of post-dive blood lactate accumulation), (3) final arterial, venous and previously measured air sac P(O(2)) values were indistinguishable in longer dives, and (4) final venous P(O(2)) values of longer dives were as low as 1-6 mmHg during dives. Although blood O(2) is not depleted at the ADL, nearly complete depletion of the blood O(2) store occurs in longer dives. This extreme hypoxemic tolerance, which would be catastrophic in many birds and mammals, necessitates biochemical and molecular adaptations, including a shift in the O(2)-Hb dissociation curve of the emperor penguin in comparison to those of most birds. A relatively higher-affinity Hb is consistent with blood P(O(2)) values and O(2) contents of penguins at rest.

In order to determine the rate and magnitude of respiratory O2 depletion during dives of emperor penguins (Aptenodytes forsteri), air sac O2 partial pressure (PO2) was recorded in 73 dives of four birds at an isolated dive hole. These results were evaluated with respect to hypoxic tolerance, the aerobic dive limit (ADL; dive duration beyond which there is post-dive lactate accumulation) and previously measured field metabolic rates (FMRs). 55% of dives were greater in duration than the previously measured 5.6-min ADL. PO2 and depth profiles revealed compression hyperoxia and gradual O2 depletion during dives. 42% of final PO2s during the dives (recorded during the last 15 s of ascent) were <20 mmHg (<2.7 kPa). Assuming that the measured air sac PO2 is representative of the entire respiratory system, this implies remarkable hypoxic tolerance in emperors. In dives of durations greater than the ADL, the calculated end-of-dive air sac O2 fraction was <4%. The respiratory O2 store depletion rate of an entire dive, based on the change in O2 fraction during a dive and previously measured diving respiratory volume, ranged from 1 to 5 ml O2 kg(-1) min(-1) and decreased exponentially with diving duration. The mean value, 2.1+/-0.8 ml O2 kg(-1) min(-1), was (1) 19-42% of previously measured respiratory O(2) depletion rates during forced submersions and simulated dives, (2) approximately one-third of the predicted total body resting metabolic rate and (3) approximately 10% of the measured FMR. These findings are consistent with a low total body metabolic rate during the dive.