Could Chronic Hypothermia in a Human Affect the Clock System?

AbstractHibernation-like episodes would be particularly interesting for clinical and spatial use if they could be observed and induced in humans. As animal hibernation differs from hypothermia with its control by a temperature-dependent clock, we undertook to find evidence that human hypothermia might affect the circadian clock system. We revisited Siffre's 1962 abyss experiment. Deprived of temporal information and showing signs of chronic hypothermia, Siffre underestimated his stay underground by 22 d. We show that the temperature-dependent clock equation for classical hibernators accurately predicts Siffre's subjective times, and we list potential conditions to be further explored for inducing hibernation-like bouts in humans.

Cool birds: first evidence of energy-saving nocturnal torpor in free-living common swifts Apus apus resting in their nests.

Daily torpor is a means of saving energy by controlled lowering of the metabolic rate (MR) during resting, usually coupled with a decrease in body temperature. We studied nocturnal daily torpor under natural conditions in free-living common swifts Apus apus resting in their nests as a family using two non-invasive approaches. First, we monitored nest temperature (Tnest) in up to 50 occupied nests per breeding season in 2010-2015. Drops in Tnest were the first indication of torpor. Among 16 673 observations, we detected 423 events of substantial drops in Tnest of on average 8.6°C. Second, we measured MR of the families inside nest-boxes prepared for calorimetric measurements during cold periods in the breeding seasons of 2017 and 2018. We measured oxygen consumption and carbon dioxide production using a mobile indirect respirometer and calculated the percentage reduction in MR. During six torpor events observed, MR was gradually reduced by on average 56% from the reference value followed by a decrease in Tnest of on average 7.6°C. By contrast, MR only decreased by about 33% on nights without torpor. Our field data gave an indication of daily torpor, which is used as a strategy for energy saving in free-living common swifts.

How much does it cost? Teaching physiology of energy metabolism in mice using an indirect calorimetry system in a practical course for veterinary students.

In endothermic mammals total energy expenditure (EE) is composed of basal metabolic rate (BMR), energy spent for muscle activity, thermoregulation, any kind of production (such as milk, meat, or egg production), and the thermic effect of feeding. The BMR is predominantly determined by body mass and the surface-to-volume ratio of the body. The EE can be quantified by either direct or indirect calorimetry. Direct calorimetry measures the rate of heat loss from the body, whereas indirect calorimetry measures oxygen consumption and carbon dioxide production and calculates heat production from oxidative nutrient combustion. A deep and sustainable understanding of EE in animals is crucial for veterinarians to properly calculate and evaluate feed rations during special circumstances such as anesthesia or in situations with increased energy demands as commonly seen in high-yielding livestock. The practical class described in this article provides an experimental approach to understanding how EE can be measured and calculated by indirect calorimetry. Two important factors that affect the EE of animals (the thermic effect of feeding and the effect of ambient temperature) are measured. A profound knowledge about the energy requirements of animal life and its measurement is also relevant for education in general biology, animal and human physiology, and nutrition. Therefore, this teaching unit can equally well be implemented in other areas of life sciences.

European space agency's hibernation (torpor) strategy for deep space missions: Linking biology to engineering.

Long-duration space missions to Mars will impose extreme stresses of physical and psychological nature on the crew, as well as significant logistical and technical challenges for life support and transportation. Main challenges include optimising overall mass and maintaining crew physical and mental health. These key scopes have been taken up as the baseline for a study by the European Space Agency (ESA) using its Concurrent Design Facility (CDF). It focussed on the biology of hibernation in reducing metabolism and hence stress, and its links to the infrastructure and life support. We concluded that torpor of crew members can reduce the payload with respect to oxygen, food and water but will require monitoring and artificial intelligence (AI) assisted monitoring of the crew. These studies additionally offer new potential applications for patient care on Earth. Keywords: Space flight, concurrent design facility, metabolic reduction.

Torpor at high ambient temperature in a neotropical didelphid, the grey short-tailed opossum (Monodelphis domestica).

The grey short-tailed opossum, Monodelphis domestica, has been an established research animal for more than five decades, but relatively, little is known about its thermophysiology. Here we studied core body temperature (T b) and metabolic rate (MR) of female adult M. domestica housed in the laboratory at an ambient temperature (T a) of 26 °C. In expanding previous reports, the average recorded core T b of M. domestica was 34.3 °C. The T b of an individual M. domestica can drop below 30 °C (minimal T b: 28.6 °C) accompanied by a reduction in MR of up to 52 % even while having ad libitum access to food. These findings demonstrate for the first time the presence of spontaneous torpor in M. domestica. Metabolic suppression at relatively high T a and T b furthermore broadens our perspective on the use of torpor as a metabolic strategy not just restricted to cold climates.

Metabolic depression during warm torpor in the Golden spiny mouse (Acomys russatus) does not affect mitochondrial respiration and hydrogen peroxide release.

Small mammals actively decrease metabolism during daily torpor and hibernation to save energy. Recently, depression of mitochondrial substrate oxidation in isolated liver mitochondria was observed and associated to hypothermic/hypometabolic states in Djungarian hamsters, mice and hibernators. We aimed to clarify whether hypothermia or hypometabolism causes mitochondrial depression during torpor by studying the Golden spiny mouse (Acomys russatus), a desert rodent which performs daily torpor at high ambient temperatures of 32°C. Notably, metabolic rate but not body temperature is significantly decreased under these conditions. In isolated liver, heart, skeletal muscle or kidney mitochondria we found no depression of respiration. Moderate cold exposure lowered torpor body temperature but had minor effects on minimal metabolic rate in torpor. Neither decreased body temperature nor metabolic rate impacted mitochondrial respiration. Measurements of mitochondrial proton leak kinetics and determination of P/O ratio revealed no differences in mitochondrial efficiency. Hydrogen peroxide release from mitochondria was not affected. We conclude that interspecies differences of mitochondrial depression during torpor do not support a general relationship between mitochondrial respiration, body temperature and metabolic rate. In Golden spiny mice, reduction of metabolic rate at mild temperatures is not triggered by depression of substrate oxidation as found in liver mitochondria from other cold-exposed rodents.

Brown adipose tissue dynamics in wild-type and UCP1-knockout mice: in vivo insights with magnetic resonance.

We used noninvasive magnetic resonance imaging (MRI) and magnetic resonance spectroscopy to compare interscapular brown adipose tissue (iBAT) of wild-type (WT) and uncoupling protein 1 (UCP1)-knockout mice lacking UCP1-mediated nonshivering thermogenesis (NST). Mice were sequentially acclimated to an ambient temperature of 30°C, 18°C, and 5°C. We detected a remodeling of iBAT and a decrease in its lipid content in all mice during cold exposure. Ratios of energy-rich phosphates (ATP/ADP, phosphocreatine/ATP) in iBAT were maintained stable during noradrenergic stimulation of thermogenesis in cold- and warm-adapted mice and no difference between the genotypes was observed. As free fatty acids (FFAs) serve as fuel for thermogenesis and activate UCP1 for uncoupling of oxidative phosphorylation, brown adipose tissue is considered to be a main acceptor and consumer of FFAs. We measured a major loss of FFAs from iBAT during noradrenergic stimulation of thermogenesis. This mobilization of FFAs was observed in iBAT of WT mice as well as in mice lacking UCP1. The high turnover and the release of FFAs from iBAT suggests an enhancement of lipid metabolism, which in itself contributes to the sympathetically activated NST and which is independent from uncoupled respiration mediated by UCP1. Our study demonstrates that MRI, besides its potential for visualizing and quantification of fat tissue, is a valuable tool for monitoring functional in vivo processes like lipid and phosphate metabolism during NST.

Brown fat in a protoendothermic mammal fuels eutherian evolution.

Endothermy has facilitated mammalian species radiation, but the sequence of events leading to sustained thermogenesis is debated in multiple evolutionary models. Here we study the Lesser hedgehog tenrec (Echinops telfairi), a phylogenetically ancient, 'protoendothermic' eutherian mammal, in which constantly high body temperatures are reported only during reproduction. Evidence for nonshivering thermogenesis is found in vivo during periodic ectothermic-endothermic transitions. Anatomical studies reveal large brown fat-like structures in the proximity of the reproductive organs, suggesting physiological significance for parental care. Biochemical analysis demonstrates high mitochondrial proton leak catalysed by an uncoupling protein 1 ortholog. Strikingly, bioenergetic profiling of tenrec uncoupling protein 1 reveals similar thermogenic potency as modern mouse uncoupling protein 1, despite the large phylogenetic distance. The discovery of functional brown adipose tissue in this 'protoendothermic' mammal links nonshivering thermogenesis directly to the roots of eutherian evolution, suggesting physiological importance prior to sustained body temperatures and migration to the cold.

Depression of mitochondrial respiration during daily torpor of the Djungarian hamster, Phodopus sungorus, is specific for liver and correlates with body temperature.

Small mammals actively decrease metabolism during daily torpor and hibernation to save energy. Increasing evidence suggests depression of mitochondrial respiration during daily torpor of the Djungarian hamster but tissue-specificity and relation to torpor depth is unknown. We first confirmed a previous study by Brown and colleagues reporting on the depressed substrate oxidation in isolated liver mitochondria of the Djungarian hamster (Phodopus sungorus) during daily torpor. Next, we show that mitochondrial respiration is not depressed in kidneys, skeletal muscle and heart. In liver mitochondria, we found that state 3 and state 4 respirations correlate with body temperature, suggesting inhibition related to torpor depth and to metabolic rate. We conclude that molecular events leading to depression of mitochondrial respiration during daily torpor are specific to liver and linked to a decrease in body temperature. Different tissue-specificity of mitochondrial depression may assist to compare and identify the molecular nature of mitochondrial alterations during torpor.

The dietary flavonoids naringenin and quercetin acutely impair glucose metabolism in rodents possibly via inhibition of hypothalamic insulin signalling.

Secondary metabolites of herbs and spices are widely used as an alternative strategy in the therapy of various diseases. The polyphenols naringenin, quercetin and curcumin have been characterised as anti-diabetic agents. Conversely, in vitro, naringenin and quercetin are described to inhibit phosphoinositide-3-kinase (PI3K), an enzyme that is essential for the neuronal control of whole body glucose homoeostasis. Using both in vitro and in vivo experiments, we tested whether the inhibitory effect on PI3K occurs in neurons and if it might affect whole body glucose homoeostasis. Quercetin was found to inhibit basal and insulin-induced phosphorylation of Akt (Ser473), a downstream target of PI3K, in HT-22 cells, whereas naringenin and curcumin had no effect. In Djungarian hamsters (Phodopus sungorus) naringenin and quercetin (10 mg/kg administered orally) diminished insulin-induced phosphorylation of Akt (Ser473) in the arcuate nucleus, indicating a reduction in hypothalamic PI3K activity. In agreement with this finding, glucose tolerance in naringenin-treated hamsters (oral) and mice (oral and intracerebroventricular) was reduced compared with controls. Dietary quercetin also impaired glucose tolerance, whereas curcumin was ineffective. Circulating levels of insulin and insulin-like growth factor-binding protein were not affected by the polyphenols. Oral quercetin reduced the respiratory quotient, suggesting that glucose utilisation was impaired after treatment. These data demonstrate that low doses of naringenin and quercetin acutely and potently impair glucose homoeostasis. This effect may be mediated by inhibition of hypothalamic PI3K signalling. Whether chronic impairments in glucose homoeostasis occur after long-term application remains to be identified.

That's hot: golden spiny mice display torpor even at high ambient temperatures.

Golden spiny mice (Acomys russatus) living in the Judean desert are exposed to extended periods of food and water shortage. We investigated their thermal and metabolic response to three weeks of 50% food reduction at ambient temperatures of 23, 27, 32 and 35 °C by long term records of metabolic rate and body temperature in the laboratory. At all ambient temperatures, A. russatus responded to starvation by a reduction of daily energy expenditure. At 32 and 35 °C, this metabolic adjustment fully compensated the reduced food availability and they maintained their energy balance at a slightly reduced body mass. At lower ambient temperatures, they could not fully compensate for the reduced food availability and kept a negative energy balance. The reduction of daily energy expenditure was largely achieved by the occurrence of daily torpor. Torpor even occurred at high ambient temperatures of 32 and 35 °C during which metabolic depression was not associated with a marked decrease of body temperature. The results show that the occurrence of daily torpor is not necessarily linked to cold exposure and the development of a pronounced hypothermia, but may even occur as depression of metabolic rate in a hot environment.

Seasonal changes in thermogenesis of a free-ranging afrotherian small mammal, the Western rock elephant shrew (Elephantulus rupestris).

We report on the seasonal metabolic adjustments of a small-sized member of the phylogenetically ancient Afrotheria, the Western rock elephant shrew (Elephantulus rupestris). We recorded body temperature (T (b)) patterns and compared the capacity for adrenergically induced nonshivering thermogenesis (NST) in E. rupestris captured in the wild in summer and winter. Noradrenaline (NA) treatment (0.4-0.5 mg/kg, s.c.) induced a pronounced elevation in oxygen consumption compared to controls (saline), and the increase in oxygen consumption following injection of NA was 1.8-fold higher in winter compared to summer. This suggests that the smaller members of Afrotheria possess functional brown adipose tissue, which changes in thermogenic capacity depending on the season. Torpor was recorded in both seasons, but in winter the incidence of torpor was higher (n = 205 out of 448 observations) and minimal T (b) during torpor was lower (T (b)min: 11.9°C) than in summer (n = 24 out of 674 observations; T (b)min: 26°C). In addition to cold, high air humidity emerged as a likely predictor for torpor entry. Overall, E. rupestris showed a high degree of thermoregulatory plasticity, which was mainly reflected in a variable timing of torpor entry and arousal. We conclude that E. rupestris exhibits seasonal metabolic adjustments comparable to what has been long known for many Holarctic rodents.

Induction of torpor: mimicking natural metabolic suppression for biomedical applications.

Mammalian hibernation consists of periods of depressed metabolism and reduced body temperature called "torpor" that are interspersed by normothermic arousal periods. Numerous cellular processes are halted during torpor, including transcription, translation, and ion homeostasis. Hibernators are able to survive long periods of low blood flow and body temperature followed by rewarming and reperfusion without overt signs of organ injury, which makes these animals excellent models for application of natural protective mechanisms to human medicine. This review examines efforts to induce torpor-like states in non-hibernating species using pharmacological compounds. Elucidating the underlying mechanisms of natural and pharmacologically induced torpor will speed the development of new clinical approaches to treat a variety of trauma and stress states in humans.

Adaptive thermogenesis and thermal conductance in wild-type and UCP1-KO mice.

We compared maximal cold-induced heat production (HPmax) and cold limits between warm (WA; 27°C), moderate cold (MCA; 18°C), or cold acclimated (CA; 5°C) wild-type and uncoupling-protein 1 knockout (UCP1-KO) mice. In wild-type mice, HPmax was successively increased after MCA and CA, and the cold limit was lowered to -8.3°C and -18.0°C, respectively. UCP1-KO mice also increased HPmax in response to MCA and CA, although to a lesser extent. Direct comparison revealed a maximal cold-induced recruitment of heat production by +473 mW and +227 mW in wild-type and UCP1-KO mice, respectively. The increase in cold tolerance of UCP1-KO mice from -0.9°C in MCA to -10.1°C in CA could not be directly related to changes in HPmax, indicating that UCP1-KO mice used the dissipated heat more efficiently than wild-type mice. As judged from respiratory quotients, acutely cold-challenged UCP1-KO mice showed a delayed transition toward lipid oxidation, and 5-h cold exposure revealed diminished physical activity and less variability in the control of metabolic rate. We conclude that BAT is required for maximal adaptive thermogenesis but also allows metabolic flexibility and a rapid switch toward sustained lipid-fuelled thermogenesis as an acute response to cold. In both CA groups, expression of contractile proteins (myosin heavy-chain isoforms) showed minor training effects in skeletal muscles, while cardiac muscle of UCP1-KO mice had novel expression of beta cardiac isoform. Neither respiration nor basal proton conductance of skeletal muscle mitochondria were different between genotypes. In subcutaneous white adipose tissue of UCP1-KO mice, cold exposure increased cytochrome-c oxidase activity and expression of the cell death-inducing DFFA-like effector A by 3.6-fold and 15-fold, respectively, indicating the recruitment of mitochondria-rich brown adipocyte-like cells. Absence of functional BAT leads to remodeling of white adipose tissue, which may significantly contribute to adaptive thermogenesis during cold acclimation.

Uncoupling protein 1 decreases superoxide production in brown adipose tissue mitochondria.

In thermogenic brown adipose tissue, uncoupling protein 1 (UCP1) catalyzes the dissipation of mitochondrial proton motive force as heat. In a cellular environment of high oxidative capacity such as brown adipose tissue (BAT), mitochondrial uncoupling could also reduce deleterious reactive oxygen species, but the specific involvement of UCP1 in this process is disputed. By comparing brown adipose tissue mitochondria of wild type mice and UCP1-ablated litter mates, we show that UCP1 potently reduces mitochondrial superoxide production after cold acclimation and during fatty acid oxidation. We address the sites of superoxide production and suggest diminished probability of "reverse electron transport" facilitated by uncoupled respiration as the underlying mechanism of reactive oxygen species suppression in BAT. Furthermore, ablation of UCP1 represses the cold-stimulated increase of substrate oxidation normally seen in active BAT, resulting in lower superoxide production, presumably avoiding deleterious oxidative damage. We conclude that UCP1 allows high oxidative capacity without promoting oxidative damage by simultaneously lowering superoxide production.

CB(1) signaling in forebrain and sympathetic neurons is a key determinant of endocannabinoid actions on energy balance.

The endocannabinoid system (ECS) plays a critical role in obesity development. The pharmacological blockade of cannabinoid receptor type 1 (CB(1)) has been shown to reduce body weight and to alleviate obesity-related metabolic disorders. An unsolved question is at which anatomical level CB(1) modulates energy balance and the mechanisms involved in its action. Here, we demonstrate that CB(1) receptors expressed in forebrain and sympathetic neurons play a key role in the pathophysiological development of diet-induced obesity. Conditional mutant mice lacking CB(1) expression in neurons known to control energy balance, but not in nonneuronal peripheral organs, displayed a lean phenotype and resistance to diet-induced obesity. This phenotype results from an increase in lipid oxidation and thermogenesis as a consequence of an enhanced sympathetic tone and a decrease in energy absorption. In conclusion, CB(1) signaling in the forebrain and sympathetic neurons is a key determinant of the ECS control of energy balance.

UCP1 ectopically expressed in murine muscle displays native function and mitigates mitochondrial superoxide production.

Mitochondrial uncoupling in skeletal muscle has raised a major interest as a therapeutic target for treatment of obesity, insulin sensitivity, and age-related disease. These physiological effects could be demonstrated in several mouse models ectopically expressing uncoupling protein 1 (UCP1). Here, we investigated whether UCP1 expressed under the control of the human skeletal actin (HSA) promoter in mouse skeletal muscle can be regulated, and whether it affects mitochondrial superoxide production. We show that the skeletal muscle UCP1 can be fully inhibited by a purine nucleotide (GDP) and reactivated by fatty acids (palmitate). During mitochondrial resting state (State 4), mitochondrial superoxide production is about 76% lower in transgenic mice. We suggest that this reduction is due to uncoupling activity as the administration of GDP restores superoxide production to wildtype levels. Our study confirms native behaviour of UCP1 in skeletal muscle and demonstrates beneficial effects on prevention of mitochondrial reactive oxygen species production which may reduce age-related deleterious processes.

Torpor and ultradian rhythms require an intact signalling of the sympathetic nervous system.

During entrance into torpor heart and respiration rates are greatly reduced in parallel with the reduction of metabolic rate, suggesting an involvement of parasympathetic control. We compared the effect of parasympathetic inhibition with the effect of sympathetic inhibition on spontaneous torpor behaviour in the Djungarian hamster. Hamsters were acclimated to short photoperiod and displayed their standard torpor pattern as observed from T(b) records. Parasympathetic inhibition was achieved by a subcutaneous implant of 21-day release pellets with Atropine and the sympathetic noradrenergic pathway was inhibited with a single injection of 6-Hydroxydopamine. Atropine treatment did not affect the occurrence and quality of spontaneous daily torpor at all. However, the reversible sympathetic inhibition by 6-Hydroxydopamine injection resulted in a complete disappearance of torpor for about 6 days. These results conclude that the onset of daily torpor requires an intact noradrenergic signalling of the sympathetic nervous system. We further observed that parasympathetic as well as sympathetic blockade resulted in an immediate abolishment of ultradian rhythms of body temperature. This suggests that the expression of ultradian oscillations in body temperature require a continued interaction of sympathetic and parasympathetic activity.