An hourglass mechanism controls torpor bout length in hibernating garden dormice

Hibernating mammals drastically lower their rate of oxygen consumption and body temperature (Tb) for up to several weeks, but regularly rewarm and stay euthermic for brief periods (< 30 h). It has been hypothesized that these periodic arousals are driven by the development of a metabolic imbalance during torpor, that is, the accumulation or the depletion of metabolites or the accrual of cellular damage that can be eliminated only in the euthermic state. We obtained oxygen consumption (as a proxy of metabolic rate) and Tb at 7-minute intervals over entire torpor-arousal cycles in the garden dormouse (Eliomys quercinus). Torpor bout duration was highly dependent on mean oxygen consumption during the torpor bout. Oxygen consumption during torpor, in turn, was elevated by Tb, which fluctuated only slightly in dormice kept at∼3-8°C. This corresponds to a well-known effect of higher Tb on shortening torpor bout lengths in hibernators. Arousal duration was independent from prior torpor length, but arousal mean oxygen consumption increased with prior torpor Tb. These results, particularly the effect of torpor oxygen consumption on torpor bout length, point to an hourglass mechanism of torpor control, i.e., the correction of a metabolic imbalance during arousal. This conclusion is in line with previous comparative studies providing evidence for significant interspecific inverse relationships between the duration of torpor bouts and metabolism in torpor. Thus, a simple hourglass mechanism is sufficient to explain torpor/arousal cycles, without the need to involve non-temperature-compensated circadian rhythms.

Humidity, huddling & the hibernation energetics of big brown bats (Eptesicus fuscus)

During winter, many mammals hibernate and lower their body temperature and metabolic rate (MR) in prolonged periods of torpor. Hibernators will use energetically expensive arousals (i.e., restore body temperature and MR) presumably to re-establish water balance. Some hibernating mammals however will huddle in groups, possibly to decrease energetic costs and total evaporative water loss (EWL), although the benefit is not fully understood. Research on the relationship between behaviour, physiology, water loss, and energy expenditure of bats during hibernation is especially important because of a fungal disease called white-nose syndrome (WNS). To date, 12 North American bat species are affected by WNS, however big brown bats (Eptesicus fuscus) appear resistant, although the underlying mechanism is poorly understood. The overall objective of my thesis was to understand the influence of humidity and huddling on the behavioural and physiological responses of hibernating big brown bats. To test my hypotheses, I used a captive colony of hibernating big brown bats (n = 20). Specifically, for Chapter 2, I first tested the hypothesis that big brown bats adjust huddling and drinking behaviour depending on humidity, to maintain a consistent pattern of periodic arousals, and therefore energy balance during hibernation. I found that bats hibernating in a dry environment did not differ in arousal/torpor bout frequency, or torpor bout duration throughout hibernation but drank at twice the rate as bats in a humid environment. Bats in the dry treatment also had shorter arousals, and huddled in a denser huddle, potentially to reduce rates of total EWL. During late hibernation, for Chapter 3, I used open-flow respirometry to test two additional hypotheses, first that phenotypic flexibility in total EWL helps explain the tolerance of hibernating big brown bats for a wide range of humidity relative to other bat species. I found that dry-acclimated bats had lower rates of total EWL, compared to bats acclimated to humid conditions. I then tested the second hypothesis that big brown bats can use huddling to mitigate the challenge of dry conditions. I found that, for humid-acclimated bats, rates of total EWL were reduced with huddling bats but there was no effect of huddling on EWL for bats acclimated to dry conditions. These results suggest that the ability of big brown bats to reduce rates of total EWL through acclimation may reduce the need to huddle with conspecifics to avoid water loss and thus dehydration. Overall, my thesis suggests that big brown bats use both behavioural and physiological mechanisms to reduce water loss which could allow them to exploit habitats for hibernation that are unavailable to other bat species and could also help explain their apparent resistance to WNS.

Liver Transcriptome Dynamics During Hibernation Are Shaped by a Shifting Balance Between Transcription and RNA Stability

Hibernators dramatically lower metabolism to save energy while fasting for months. Prolonged fasting challenges metabolic homeostasis, yet small-bodied hibernators emerge each spring ready to resume all aspects of active life, including immediate reproduction. The liver is the body’s metabolic hub, processing and detoxifying macromolecules to provide essential fuels to brain, muscle and other organs throughout the body. Here we quantify changes in liver gene expression across several distinct physiological states of hibernation in 13-lined ground squirrels, using RNA-seq to measure the steady-state transcriptome and GRO-seq to measure transcription for the first time in a hibernator. Our data capture key timepoints in both the seasonal and torpor-arousal cycles of hibernation. Strong positive correlation between transcription and the transcriptome indicates that transcriptional control dominates the known seasonal reprogramming of metabolic gene expression in liver for hibernation. During the torpor-arousal cycle, however, discordance develops between transcription and the steady-state transcriptome by at least two mechanisms: 1) although not transcribed during torpor, some transcripts are unusually stable across the torpor bout; and 2) unexpectedly, on some genes, our data suggest continuing, slow elongation with a failure to terminate transcription across the torpor bout. While the steady-state RNAs corresponding to these read through transcripts did not increase during torpor, they did increase shortly after rewarming despite their simultaneously low transcription. Both of these mechanisms would assure the immediate availability of functional transcripts upon rewarming. Integration of transcriptional, post-transcriptional and RNA stability control mechanisms, all demonstrated in these data, likely initiate a serial gene expression program across the short euthermic period that restores the tissue and prepares the animal for the next bout of torpor.

The evolution of a bat population with white-nose syndrome (WNS) reveals a shift from an epizootic to an enzootic phase

Background White-nose Syndrome (WNS) is a mycosis caused by a cutaneous infection with the fungus Pseudogymnoascus destructans (Pd). It produces hibernation mortality rates of 75–98% in 4 bats: Myotis lucifugus, M. septentrionalis, M. sodalis, and Perimyotis subflavus. These high mortality rates were observed during the first several years after the arrival of P. destructans at a hibernation site. Mortality is caused by a 60% decrease in torpor bout duration, which results in a premature depletion of depot fat prior to spring. Results Little is known about the long-term effects of Pd on torpor and mortality, thus we conducted a 9-year study on M. lucifugus at 5 of the hibernation sites where Pd first appeared in North America during the winter of 2007–08. The M. lucifugus hibernating at one of these sites one year after the arrival of Pd (2008–09) had: a) a mean torpor bout duration of 7.6 d, b) no depot fat reserves by March, and c) an apparent over-winter mortality rate of 88%. The M. lucifugus hibernating at this same site 6–9 years after the arrival of Pd, in contrast, had: a) a mean torpor bout duration of 14.7 d, b) depot fat remaining in March, and c) an apparent mortality rate of 50%. The number of M. lucifugus hibernating at 2 of these sites has consistently increased since 2010 and is now more than 3.0-fold higher than the number remaining after the winter of 2008–09. Conclusions These findings indicate that this population of M. lucifugus has evolved mechanisms to hibernate well in the presence of Pd, thus reducing over-winter mortality.

Implicating a Temperature-Dependent Clock in the Regulation of Torpor Bout Duration in Classic Hibernation

Syrian hamsters may present 2 types of torpor when exposed to ambient temperatures in the winter season, from 8°C to 22°C (short photoperiod). The first is daily torpor, which is controlled by the master circadian clock of the body, located in the SCN. In this paper, we show that daily torpor bout duration is unchanged over the 8°C to 22°C temperature range, as predicted from the thermal compensation of circadian clocks. These findings contrast with the second type of torpor: multi-day torpor or classic hibernation. In multi-day torpor, bout duration increases as temperature decreases, following Arrhenius thermodynamics. We found no evidence of hysteresis from metabolic inhibition and the process was thus reversible. As a confirmation, at any temperature, the arousal from multi-day torpor occurred at about the same subjective time given by this temperature-dependent clock. The temperature-dependent clock controls the reduced torpor metabolic rate while providing a reversible recovery of circadian synchronization on return to euthermy.

Torpor duration, more than temperature, is key to hummingbird nighttime energy savings

ABSTRACTTorpor is an important energy saving strategy in small endotherms, but it has been insufficiently studied in natural field conditions. Building on what we know from laboratory studies, we compared torpor use across hummingbird species and different natural temperature regimes to explore predominant hypotheses about torpor use and evaluate the possible effects of environmental variation on energy management.We found that the probability of an individual entering torpor was correlated with mass and unrelated to nighttime temperature, and that hummingbirds at both warm tropical and cooler temperate sites used torpor.Energy savings in torpor were maximized as ambient temperatures approached a species’ minimum body temperature consistent with laboratory studies; energy savings ranged between 65-92% of energy per hour in torpor compared to normothermy.However, variation in total nighttime energy expenditure was more significantly influenced by torpor bout duration than by the variation in energy savings in torpor.Our results show that a small endotherm’s nighttime energy management in its natural habitat is more affected by torpor bout duration, which is linked to photoperiod, than by temperature. This result suggests that in their natural environments, hummingbirds are able to save energy in torpor across a range nighttime temperature, indicating that they may have sufficient physiological flexibility to tolerate climatic variation.

Daily changes in food availability, but not long-term unpredictability, determine daily torpor-bout occurrences and frequency in stripe-faced dunnarts (Sminthopsis macroura)

Daily torpor, a short-term reduction in body temperature and metabolism, is an energy-saving strategy that has been interpreted as an adaptation to unpredictable resource availability. However, the effect of food-supply variability on torpor, separately from consistent food restriction, remains largely unexamined. In this study, we investigated the effect of unpredictable food availability on torpor in stripe-faced dunnarts (Sminthopsis macroura). After a control period of ad libitum feeding, dunnarts were offered 65% of their average daily ad libitum intake over 31 days, either as a constant restriction (i.e. as equal amount of food offered each day) or as an unpredictable schedule of feed offered, varied daily as 0%, 30%, 60%, 100% or 130% of ad libitum. Both feeding groups had increased torpor-bout occurrences (as a proportion of all dunnarts on a given day) and torpor-bout frequency (average number of bouts each day) when on a restricted diet compared with ad libitum feeding, but torpor frequency did not differ between the consistently restricted and unpredictably restricted groups. Most importantly, torpor occurrence and daily bout frequency by the unpredictably restricted group appeared to change in direct association with the amount of food offered on each day; torpor frequency was higher on days of low food availability. Our data do not support the interpretation that torpor is a response to unpredictable food availability per se, but rather that torpor allowed a rapid adjustment of energy expenditure to manage daily fluctuations in food availability.

Basking behaviour and torpor use in free-ranging Planigale gilesi

We investigated the importance of energy-conserving strategies for free-ranging Planigale gilesi in arid Australia. We monitored torpor use and basking behaviour using internal temperature-sensitive transmitters. Torpor was used every day; the maximum torpor bout duration was 18.2 h and the minimum body temperature was 10.5°C. Basking behaviour was observed during rewarming from torpor as well as during normothermia. The use of torpor and basking is likely to reduce the energy requirements of P. gilesi, thus helping it to survive in a harsh and unpredictable environment.

Influence of torpor on cardiac expression of genes involved in the circadian clock and protein turnover in the Siberian hamster (Phodopus sungorus)

The Siberian hamster exhibits the key winter adaptive strategy of daily torpor, during which metabolism and heart rate are slowed for a few hours and body temperature declines by up to 20°C, allowing substantial energetic savings. Previous studies of hibernators in which temperature drops by >30°C for many days to weeks have revealed decreased transcription and translation during hypometabolism and identified several key physiological pathways involved. Here we used a cDNA microarray to define cardiac transcript changes over the course of a daily torpor bout and return to normothermia, and we show that, in common with hibernators, a relatively small proportion of the transcriptome (<5%) exhibited altered expression over a torpor bout. Pathways exhibiting significantly altered gene expression included transcriptional regulation, RNA stability and translational control, globin regulation, and cardiomyocyte function. Remarkably, gene representatives of the entire ubiquitylation pathway were significantly altered over the torpor bout, implying a key role for cardiac protein turnover and translation during a low-temperature torpor bout. The circadian clock maintained rhythmic transcription during torpor. Quantitative PCR profiling of heart, liver, and lung and in situ hybridization studies of clock genes in the hypothalamic circadian clock in the suprachiasmatic nucleus revealed that many circadian regulated transcripts exhibited synchronous alteration in expression during arousal. Our data highlight the potential importance of genes involved in protein turnover as part of the adaptive strategy of low-temperature torpor in a seasonal mammal.

mRNA Stability and Polysome Loss in Hibernating Arctic Ground Squirrels (Spermophilus parryii)

ABSTRACT All small mammalian hibernators periodically rewarm from torpor to high, euthermic body temperatures for brief intervals throughout the hibernating season. The functional significance of these arousal episodes is unknown, but one suggestion is that rewarming may be related to replacement of gene products lost during torpor due to degradation of mRNA. To assess the stability of mRNA as a function of the hibernation state, we examined the poly(A) tail lengths of liver mRNA from arctic ground squirrels sacrificed during four hibernation states (early and late during a torpor bout and early and late following arousal from torpor) and from active ground squirrels sacrificed in the summer. Poly(A) tail lengths were not altered during torpor, suggesting either that mRNA is stabilized or that transcription continues during torpor. In mRNA isolated from torpid ground squirrels, we observed a pattern of 12 poly(A) residues at greater densities approximately every 27 nucleotides along the poly(A) tail, which is a pattern consistent with binding of poly(A)-binding protein. The intensity of this pattern was significantly reduced following arousal from torpor and undetectable in mRNA obtained from summer ground squirrels. Analyses of polysome profiles revealed a significant reduction in polyribosomes in torpid animals, indicating that translation is depressed during torpor.