Blood transcriptomics mirror regulatory mechanisms during hibernation-a comparative analysis of the Djungarian hamster with other mammalian species.

Hibernation enables many species of the mammalian kingdom to overcome periods of harsh environmental conditions. During this physically inactive state metabolic rate and body temperature are drastically downregulated, thereby reducing energy requirements (torpor) also over shorter time periods. Since blood cells reflect the organism´s current condition, it was suggested that transcriptomic alterations in blood cells mirror the torpor-associated physiological state. Transcriptomics on blood cells of torpid and non-torpid Djungarian hamsters and QIAGEN Ingenuity Pathway Analysis (IPA) revealed key target molecules (TMIPA), which were subjected to a comparative literature analysis on transcriptomic alterations during torpor/hibernation in other mammals. Gene expression similarities were identified in 148 TMIPA during torpor nadir among various organs and phylogenetically different mammalian species. Based on TMIPA, IPA network analyses corresponded with described inhibitions of basic cellular mechanisms and immune system-associated processes in torpid mammals. Moreover, protection against damage to the heart, kidney, and liver was deduced from this gene expression pattern in blood cells. This study shows that blood cell transcriptomics can reflect the general physiological state during torpor nadir. Furthermore, the understanding of molecular processes for torpor initiation and organ preservation may have beneficial implications for humans in extremely challenging environments, such as in medical intensive care units and in space.

Role of glucose in daily torpor of Djungarian hamsters (Phodopus sungorus): challenge of continuous in vivo blood glucose measurements.

Djungarian hamsters use daily torpor to save energy during winter. This metabolic downstate is part of their acclimatization strategy in response to short photoperiod and expressed spontaneously without energy challenges. During acute energy shortage, torpor incidence, depth, and duration can be modulated. Torpor induction might rely on glucose availability as acute metabolic energy source. To investigate this, the present study provides the first continuous in vivo blood glucose measurements of spontaneous daily torpor in short photoperiod-acclimated and fasting-induced torpor in long photoperiod-acclimated Djungarian hamsters. Glucose levels were almost identical in both photoperiods and showed a decrease during resting phase. Further decreases appeared during spontaneous daily torpor entrance, parallel with metabolic rate but before body temperature, while respiratory exchange rates were rising. During arousal, blood glucose tended to increase, and pretorpor values were reached at torpor termination. Although food-restricted hamsters underwent a considerable energetic challenge, blood glucose levels remained stable during the resting phase regardless of torpor expression. The activity phase preceding a torpor bout did not reveal changes in blood glucose that might be used as torpor predictor. Djungarian hamsters show a robust, circadian rhythm in blood glucose irrespective of season and maintain appropriate levels throughout complex acclimation processes including metabolic downstates. Although these measurements could not reveal blood glucose as proximate torpor induction factor, they provide new information about glucose availability during torpor. Technical innovations like in vivo microdialysis and in vitro transcriptome or proteome analyses may help to uncover the connection between torpor expression and glucose metabolism.

Short Photoperiod-Dependent Enrichment of Akkermansia spec. as the Major Change in the Intestinal Microbiome of Djungarian Hamsters (Phodopus sungorus).

The Djungarian hamster (Phodopus sungorus) is a prominent model organism for seasonal acclimatization, showing drastic whole-body physiological adjustments to an energetically challenging environment, which are considered to also involve the gut microbiome. Fecal samples of hamsters in long photoperiod and again after twelve weeks in short photoperiod were analyzed by 16S-rRNA sequencing to evaluate seasonal changes in the respective gut microbiomes. In both photoperiods, the overall composition was stable in the major superordinate phyla of the microbiota, with distinct and delicate changes of abundance in phyla representing each <1% of all. Elusimicrobia, Tenericutes, and Verrucomicrobia were exclusively present in short photoperiod hamsters. In contrast to Elusimicrobium and Aneroplasma as representatives of Elusimicrobia and Tenericutes, Akkermansia muciniphila is a prominent gut microbiome inhabitant well described as important in the health context of animals and humans, including neurodegenerative diseases and obesity. Since diet was not changed, Akkermansia enrichment appears to be a direct consequence of short photoperiod acclimation. Future research will investigate whether the Djungarian hamster intestinal microbiome is responsible for or responsive to seasonal acclimation, focusing on probiotic supplementation.

Acclimation of intestinal morphology and function in Djungarian hamsters (Phodopus sungorus) related to seasonal and acute energy balance.

Small mammals exhibit seasonal changes in intestinal morphology and function via increased intestine size and resorptive surface and/or nutrient transport capacity to increase energy yield from food during winter. This study investigated whether seasonal or acute acclimation to anticipated or actual energetic challenges in Djungarian hamsters also resulted in higher nutrient resorption capacities owing to changes in small intestine histology and physiology. The hamsters show numerous seasonal energy-saving adjustments in response to short photoperiod. As spontaneous daily torpor represents one of these adjustments related to food quality and quantity, it was hypothesized that the hamsters' variable torpor expression patterns are influenced by their individual nutrient uptake capacity. Hamsters under short photoperiod showed longer small intestines and higher mucosal electrogenic transport capacities for glucose relative to body mass. Similar observations were made in hamsters under long photoperiod and food restriction. However, this acute energetic challenge caused a stronger increase of glucose transport capacity. Apart from that, neither fasting-induced torpor in food-restricted hamsters nor spontaneous daily torpor in short photoperiod-exposed hamsters clearly correlated with mucosal glucose transport capacity. Both seasonally anticipated and acute energetic challenges caused adjustments in the hamsters' small intestine. Short photoperiod appeared to induce an integration of these and other acclimation processes in relation to body mass to achieve a long-term adjustment of energy balance. Food restriction seemed to result in a more flexible, short-term strategy of maximizing energy uptake possibly via mucosal glucose transport and reducing energy consumption via torpor expression as an emergency response.

The Chemistry of Cold: Mechanisms of Torpor Regulation in the Siberian Hamster.

Siberian hamsters use spontaneous daily torpor, a state of hypometabolism and hypothermia, to save energy during winter. Multiple neuroendocrine signals set the scene for spontaneous torpor to occur, and several brain areas have been identified as potential sites for torpor regulation. Here, we summarize the known mechanisms of a fascinating physiological state in the Siberian hamster.

Thyroid hormone status affects expression of daily torpor and gene transcription in Djungarian hamsters (Phodopus sungorus).

Thyroid hormones (TH) play a key role in regulation of seasonal as well as acute changes in metabolism. Djungarian hamsters (Phodopus sungorus) adapt to winter by multiple changes in behaviour and physiology including spontaneous daily torpor, a state of hypometabolism and hypothermia. We investigated effects of systemic TH administration and ablation on the torpor behaviour in Djungarian hamsters adapted to short photoperiod. Hyperthyroidism was induced by giving T4 or T3 and hypothyroidism by giving methimazole (MMI) and sodium perchlorate via drinking water. T3 treatment increased water, food intake and body mass, whereas MMI had the opposite effect. Continuous recording of body temperature revealed that low T3 serum concentrations increased torpor incidence, lowered Tb and duration, whereas high T3 serum concentrations inhibited torpor expression. Gene expression of deiodinases (dio) and uncoupling proteins (ucp) were analysed by qPCR in hypothalamus, brown adipose tissue (BAT) and skeletal muscle. Expression of dio2, the enzyme generating T3 by deiodination of T4, and ucps, involved in thermoregulation, indicated a tissue specific response to treatment. Torpor per se decreased dio2 expression irrespective of treatment or tissue, suggesting low intracellular T3 concentrations during torpor. Down regulation of ucp1 and ucp3 during torpor might be a factor for the inhibition of BAT thermogenesis. Hypothalamic gene expression of neuropeptide Y, propopiomelanocortin and somatostatin, involved in feeding behaviour and energy balance, were not affected by treatment. Taken together our data indicate a strong effect of thyroid hormones on torpor, suggesting that lowered intracellular T3 concentrations in peripheral tissues promote torpor.

Low body temperature governs the decline of circulating lymphocytes during hibernation through sphingosine-1-phosphate.

Hibernation is an energy-conserving behavior consisting of periods of inhibited metabolism ('torpor') with lowered body temperature. Torpor bouts are interspersed by arousal periods, in which metabolism increases and body temperature returns to euthermia. In deep torpor, the body temperature typically decreases to 2-10 °C, and major physiological and immunological changes occur. One of these alterations constitutes an almost complete depletion of circulating lymphocytes that is reversed rapidly upon arousal. Here we show that torpor induces the storage of lymphocytes in secondary lymphoid organs in response to a temperature-dependent drop in plasma levels of sphingosine-1-phosphate (S1P). Regulation of lymphocyte numbers was mediated through the type 1 S1P receptor (S1P(1)), because administration of a specific antagonist (W146) during torpor (in a Syrian hamster at ∼8 °C) precluded restoration of lymphocyte numbers upon subsequent arousal. Furthermore, S1P release from erythrocytes via ATP-binding cassette (ABC)-transporters was significantly inhibited at low body temperature (4 °C) but was restored upon rewarming. Reversible lymphopenia also was observed during daily torpor (in a Djungarian hamster at ± 25 °C), during forced hypothermia in anesthetized (summer-active) hamsters (at ± 9 °C), and in a nonhibernator (rat at ∼19 °C). Our results demonstrate that lymphopenia during hibernation in small mammals is driven by body temperature, via altered plasma S1P levels. S1P is recognized as an important bioactive lipid involved in regulating several other physiological processes as well and may be an important factor regulating additional physiological processes in hibernation as well as in mediating the effects of therapeutic hypothermia in patients.

Comparative transcriptomics of the garden dormouse hypothalamus during hibernation.

Torpor or heterothermy is an energy-saving mechanism used by endotherms to overcome harsh environmental conditions. During winter, the garden dormouse (Eliomys quercinus) hibernates with multiday torpor bouts and body temperatures of a few degrees Celsius, interrupted by brief euthermic phases. This study investigates gene expression within the hypothalamus, the key brain area controlling energy balance, adding information on differential gene expression potentially relevant to orchestrate torpor. A de novo assembled transcriptome of the hypothalamus was generated from garden dormice hibernating under constant darkness without food and water at 5 °C. Samples were collected during early torpor, late torpor, and interbout arousal. During early torpor, 765 genes were differentially expressed as compared with interbout arousal. Twenty-seven pathways were over-represented, including pathways related to hemostasis, extracellular matrix organization, and signaling of small molecules. Only 82 genes were found to be differentially expressed between early and late torpor, and no pathways were over-represented. During late torpor, 924 genes were differentially expressed relative to interbout arousal. Despite the high number of differentially expressed genes, only 10 pathways were over-represented. Of these, eight were also observed to be over-represented when comparing early torpor and interbout arousal. Our results are largely consistent with previous findings in other heterotherms. The addition of a transcriptome of a novel species may help to identify species-specific and overarching torpor mechanisms through future species comparisons.

Ultrasonic vocalisation rate tracks the diurnal pattern of activity in winter phenotype Djungarian hamsters (Phodopus sungorus).

Vocalisations are increasingly being recognised as an important aspect of normal rodent behaviour yet little is known of how they interact with other spontaneous behaviours such as sleep and torpor, particularly in a social setting. We obtained chronic recordings of the vocal behaviour of adult male and female Djungarian hamsters (Phodopus sungorus) housed under short photoperiod (8 h light, 16 h dark, square wave transitions), in different social contexts. The animals were kept in isolation or in same-sex sibling pairs, separated by a grid which allowed non-physical social interaction. On approximately 20% of days hamsters spontaneously entered torpor, a state of metabolic depression that coincides with the rest phase of many small mammal species in response to actual or predicted energy shortages. Animals produced ultrasonic vocalisations (USVs) with a peak frequency of 57 kHz in both social and asocial conditions and there was a high degree of variability in vocalisation rate between subjects. Vocalisation rate was correlated with locomotor activity across the 24-h light cycle, occurring more frequently during the dark period when the hamsters were more active and peaking around light transitions. Solitary-housed animals did not vocalise whilst torpid and animals remained in torpor despite overlapping with vocalisations in social-housing. Besides a minor decrease in peak USV frequency when isolated hamsters were re-paired with their siblings, changing social contexts did not influence vocalisation behaviour or structure. In rare instances, temporally overlapping USVs occurred when animals were socially-housed and were grouped in such a way that could indicate coordination. We did not observe broadband calls (BBCs) contemporaneous with USVs in this paradigm, corroborating their correlation with physical aggression which was absent from our experiment. Overall, we find little evidence to suggest a direct social function of hamster USVs. We conclude that understanding the effects of vocalisations on spontaneous behaviours, such as sleep and torpor, will inform experimental design of future studies, especially where the role of social interactions is investigated.

Seasonal and fasting induced changes in iron metabolism in Djungarian hamsters.

Djungarian hamsters are small rodents that show pronounced physiological acclimations in response to changes in photoperiod, and unfavorable environmental conditions such as reduced food availability and low external temperature. These include substantial adjustments, such as severe body weight loss and the use of daily torpor. Torpor is a state of decreased physiological activity in eutherms, usually marked by low metabolic rate and a reduced body temperature. In this study, we investigated the effects of photoperiodic acclimation and food deprivation on systemic iron metabolism in Djungarian hamsters. Our study illustrates the association between liver iron levels and the incidence of torpor expression during the course of the experiment. Moreover, we show that both, acclimation to short photoperiods and long-term food restriction, associated with iron sequestration in the liver. This effect was accompanied with hypoferremia and mild reduction in the expression of principal iron-hormone, hepcidin. In addition to iron, the levels of manganese, selenium, and zinc were increased in the liver of hamsters under food restriction. These findings may be important factors for regulating physiological processes in hamsters, since iron and other trace elements are essential for many metabolic and physiological processes.

Comparative transcriptomics of the Djungarian hamster hypothalamus during short photoperiod acclimation and spontaneous torpor.

The energy-saving strategy of Djungarian hamsters (Phodopus sungorus, Cricetidae) to overcome harsh environmental conditions comprises of behavioral, morphological, and physiological adjustments, including spontaneous daily torpor, a metabolic downstate. These acclimatizations are triggered by short photoperiod and orchestrated by the hypothalamus. Key mechanisms of long-term photoperiodic acclimatizations have partly been described, but specific mechanisms that acutely control torpor remain incomplete. Here, we performed comparative transcriptome analysis on hypothalamus of normometabolic hamsters in their summer- and winter-like state to enable us to identify changes in gene expression during photoperiodic acclimations. Comparing nontorpid and torpid hamsters may also be able to pin down mechanisms relevant for torpor control. A de novo assembled transcriptome of the hypothalamus was generated from hamsters acclimated to long photoperiod or to short photoperiod. The hamsters were sampled either during long photoperiod normothermia, short photoperiod normothermia, or short photoperiod-induced spontaneous torpor with a body temperature of 24.6 ± 1.0 °C, or. The mRNA-seq analysis revealed that 32 and 759 genes were differentially expressed during photoperiod or torpor, respectively. Biological processes were not enriched during photoperiodic acclimatization but were during torpor, where transcriptional and metabolic processes were reinforced. Most extremely regulated genes (those genes with |log2(FC)| > 2.0 and padj < 0.05 of a pairwise group comparison) underpinned the role of known key players in photoperiodic comparison, but these genes exhibit adaptive and protective adjustments during torpor. Targeted analyses of genes from potentially involved hypothalamic systems identified gene regulation of previously described torpor-relevant systems and a potential involvement of glucose transport.

Mitochondrial RNA stimulates beige adipocyte development in young mice.

Childhood obesity is a serious public health crisis and a critical factor that determines future obesity prevalence. Signals affecting adipocyte development in early postnatal life have a strong potential to trigger childhood obesity; however, these signals are still poorly understood. We show here that mitochondrial (mt)RNA efflux stimulates transcription of nuclear-encoded genes for mitobiogenesis and thermogenesis in adipocytes of young mice and human infants. While cytosolic mtRNA is a potential trigger of the interferon (IFN) response, young adipocytes lack such a response to cytosolic mtRNA due to the suppression of IFN regulatory factor (IRF)7 expression by vitamin D receptor signalling. Adult and obese adipocytes, however, strongly express IRF7 and mount an IFN response to cytosolic mtRNA. In turn, suppressing IRF7 expression in adult adipocytes restores mtRNA-induced mitobiogenesis and thermogenesis and eventually mitigates obesity. Retrograde mitochondrion-to-nucleus signalling by mtRNA is thus a mechanism to evoke thermogenic potential during early adipocyte development and to protect against obesity.

Photoperiodic regulation of glycogen metabolism, glycolysis, and glutamine synthesis in tanycytes of the Siberian hamster suggests novel roles of tanycytes in hypothalamic function.

The objective of this study is to investigate the impact of photoperiod on the temporal and spatial expression of genes involved in glucose metabolism in the brain of the seasonal mammal Phodopus sungorus (Siberian hamster). In situ hybridization was performed on brain sections obtained from male hamsters held in long photoperiod (high body weight and developed testes) or short photoperiod (reduced body weight with testicular regression). This analysis revealed upregulation in expression of genes involved in glycogen and glucose metabolism in short photoperiod and localized to the tanycyte layer of the third ventricle. On the basis of these data and a previously identified photoperiod-dependent increase in activity of neighboring hypothalamic neurons, we hypothesized that the observed expression changes may reflect alteration in either metabolic fuel or precursor neurotransmitter supply to surrounding neurons. Gene expression analysis was performed for genes involved in lactate and glutamate transport. This analysis showed that the gene for the lactate transporter MCT2 and glutamate transporter GLAST was decreased in the tanycyte layer in short photoperiod. Expression of mRNA for glutamine synthetase, the final enzyme in the synthesis of the neuronal neurotransmitter precursor, glutamine, was also decreased in short photoperiod. These data suggest a role for tanycytes in modulating glutamate concentrations and neurotransmitter supply in the hypothalamic environment.

Body Temperature and Activity Adaptation of Short Photoperiod-Exposed Djungarian Hamsters (Phodopus sungorus): Timing, Traits, and Torpor.

To survive the Siberian winter, Djungarian hamsters (Phodopus sungorus) adjust their behavior, morphology, and physiology to maintain energy balance. The reduction of body mass and the improvement of fur insulation are followed by the expression of spontaneous daily torpor, a state of reduced metabolism during the resting phase to save additional energy. Since these complex changes require time, the upcoming winter is anticipated via decreasing photoperiod. Yet, the extent of adaptation and torpor use is highly individual. In this study, adaptation was triggered by an artificially changed light regime under laboratory conditions with 20°C ambient temperature and food and water ad libitum. Two approaches analyzed data on weekly measured body mass and fur index as well as continuously recorded core body temperature and activity during: (1) the torpor period of 60 hamsters and (2) the entire adaptation period of 11 hamsters, aiming to identify parameters allowing (1) a better prediction of torpor expression in individuals during the torpor period as well as (2) an early estimation of the adaptation extent and torpor proneness. In approach 1, 46 torpor-expressing hamsters had a median torpor incidence of 0.3, covering the spectrum from no torpor to torpor every day within one representative week. Torpor use reduced the body temperature during both photo- and scotophase. Torpor was never expressed by 14 hamsters. They could be identified by a high, constant body temperature during the torpor period and a low body mass loss during adaptation to a short photoperiod. Already in the first week of short photoperiod, approach 2 revealed that the hamsters extended their activity over the prolonged scotophase, yet with reduced scotophase activity and body temperature. Over the entire adaptation period, scotophase activity and body temperature of the scoto- and photophases were further reduced, later accompanied by a body mass decline and winter fur development. Torpor was expressed by those hamsters with the most pronounced adaptations. These results provide insights into the preconditions and proximate stimuli of torpor expression. This knowledge will improve experimental planning and sampling for neuroendocrine and molecular research on torpor regulation and has the potential to facilitate acute torpor forecasting to eventually unravel torpor regulation processes.

What can seasonal models teach us about energy balance?

Torpid states are used by many endotherms to save energy during winter. During torpor, metabolic rate is downregulated to fractions of resting metabolic rate and often associated with a severe drop in body temperature that challenges mammalian physiology. Understanding the mechanisms regulating this extreme depression of metabolism bears enormous potential for biomedical research. Torpor behavior has been extensively studied in the Djungarian hamster, also known as Siberian hamster. It is dependent on many preparatory adaptations of physiological and endocrine systems that are likely to be integrated by the hypothalamus eventually controlling metabolism. Although substantial knowledge exists about prerequisites and characteristics of torpor in this species, the cascade of events and their mechanisms of action are not well understood. This review summarizes the current state of knowledge about mechanisms of metabolic regulation in the Djungarian hamster focusing on the potential roles of thyroid hormone and glucose metabolism.

Breast milk alkylglycerols sustain beige adipocytes through adipose tissue macrophages.

Prevalence of obesity among infants and children below 5 years of age is rising dramatically, and early childhood obesity is a forerunner of obesity and obesity-associated diseases in adulthood. Childhood obesity is hence one of the most serious public health challenges today. Here, we have identified a mother-to-child lipid signaling that protects from obesity. We have found that breast milk-specific lipid species, so-called alkylglycerol-type (AKG-type) ether lipids, which are absent from infant formula and adult-type diets, maintain beige adipose tissue (BeAT) in the infant and impede the transformation of BeAT into lipid-storing white adipose tissue (WAT). Breast milk AKGs are metabolized by adipose tissue macrophages (ATMs) to platelet-activating factor (PAF), which ultimately activates IL-6/STAT3 signaling in adipocytes and triggers BeAT development in the infant. Accordingly, lack of AKG intake in infancy leads to a premature loss of BeAT and increases fat accumulation. AKG signaling is specific for infants and is inactivated in adulthood. However, in obese adipose tissue, ATMs regain their ability to metabolize AKGs, which reduces obesity. In summary, AKGs are specific lipid signals of breast milk that are essential for healthy adipose tissue development.

Hypothalamic control systems show differential gene expression during spontaneous daily torpor and fasting-induced torpor in the Djungarian hamster (Phodopus sungorus).

Djungarian hamsters are able to use spontaneous daily torpor (SDT) during the winter season as well as fasting-induced torpor (FIT) at any time of the year to cope with energetically challenging environmental conditions. Torpor is a state of severely reduced metabolism with a pronounced decrease in body temperature, which enables animals to decrease their individual energy requirements. Despite sharing common characteristics, such as reduced body mass before first torpor expression and depressed metabolism and body temperature during the torpid state, FIT and SDT differ in several physiological properties including torpor bout duration, minimal body temperature, fuel utilization and circadian organization. It remains unclear, whether SDT and FIT reflect the same phenomenon or two different physiological states. The hypothalamus has been suggested to play a key role in regulating energy balance and torpor. To uncover differences in molecular control mechanisms of torpor expression, we set out to investigate hypothalamic gene expression profiles of genes related to orexigenic (Agrp/Npy), circadian clock (Bmal1/Per1) and thyroid hormone (Dio2/Mct8) systems of animals undergoing SDT and FIT during different torpor stages. Orexigenic genes were mainly regulated during FIT and remained largely unaffected by SDT. Expression patterns of clock genes showed disturbed circadian clock rhythmicity in animals undergoing FIT, but not in animals undergoing SDT. During both, SDT and FIT, decreased Dio2 expression was detected, indicating reduced hypothalamic T3 availability in both types of torpor. Taken together, our results provide evidence that SDT and FIT also differ in certain central control mechanisms and support the observation that animals undergoing SDT are in energetical balance, whereas animals undergoing FIT display a negative energy balance. This should be carefully taken into account when interpreting data in torpor research, especially from animal models of fasting-induced hypometabolism such as mice.

Transcriptome Analysis of Hypothalamic Gene Expression during Daily Torpor in Djungarian Hamsters (Phodopus sungorus).

Animals living at high or temperate latitudes are challenged by extensive changes in environmental conditions over seasons. Djungarian hamsters (Phodopus sungorus) are able to cope with extremely cold ambient temperatures and food scarcity in winter by expressing spontaneous daily torpor. Daily torpor is a circadian controlled voluntary reduction of metabolism that can reduce energy expenditure by up to 65% when used frequently. In the past decades it has become more and more apparent, that the hypothalamus is likely to play a key role in regulating induction and maintenance of daily torpor, but the molecular signals, which lead to the initiation of daily torpor, are still unknown. Here we present the first transcriptomic study of hypothalamic gene expression patterns in Djungarian hamsters during torpor entrance. Based on Illumina sequencing we were able to identify a total number of 284 differentially expressed genes, whereby 181 genes were up- and 103 genes down regulated during torpor entrance. The 20 most up regulated group contained eight genes coding for structure proteins, including five collagen genes, dnha2 and myo15a, as well as the procoagulation factor vwf. In a proximate approach we investigated these genes by quantitative real-time PCR (qPCR) analysis over the circadian cycle in torpid and normothermic animals at times of torpor entrance, mid torpor, arousal and post-torpor. These qPCR data confirmed up regulation of dnah2, myo15a, and vwf during torpor entrance, but a decreased mRNA level for all other investigated time points. This suggests that gene expression of structure genes as well as the procoagulation factor are specifically initiated during the early state of torpor and provides evidence for protective molecular adaptions in the hypothalamus of Djungarian hamsters including changes in structure, transport of biomolecules and coagulation.

Gene expression analysis and microdialysis suggest hypothalamic triiodothyronine (T3) gates daily torpor in Djungarian hamsters (Phodopus sungorus).

Thyroid hormones play an important role in regulating seasonal adaptations of mammals. Several studies suggested that reduced availability of 3,3',5-triiodothyronine (T3) in the hypothalamus is required for the physiological adaptation to winter in Djungarian hamsters. We have previously shown that T3 is involved in the regulation of daily torpor, but it remains unclear, whether T3 affects torpor by central or peripheral mechanisms. To determine the effect of T3 concentrations within the hypothalamus in regulating daily torpor, we tested the hypothesis that low hypothalamic T3 metabolism would favour torpor and high T3 concentrations would not. In experiment 1 gene expression in torpid hamsters was assessed for transporters carrying thyroid hormones between cerebrospinal fluid and hypothalamic cells and for deiodinases enzymes, activating or inactivating T3 within hypothalamic cells. Gene expression analysis suggests reduced T3 in hypothalamic cells during torpor. In experiment 2, hypothalamic T3 concentrations were altered via microdialysis and torpor behaviour was continuously monitored by implanted body temperature transmitters. Increased T3 concentrations in the hypothalamus reduced expression of torpor as well as torpor bout duration and depth. Subsequent analysis of gene expression in the ependymal layer of the third ventricle showed clear up-regulation of T3 inactivating deiodinase 3 but no changes in several other genes related to photoperiodic adaptations in hamsters. Finally, serum analysis revealed that increased total T3 serum concentrations were not necessary to inhibit torpor expression. Taken together, our results are consistent with the hypothesis that T3 availability within the hypothalamus significantly contributes to the regulation of daily torpor via a central pathway.

Daily torpor alters multiple gene expression in the suprachiasmatic nucleus and pineal gland of the Djungarian hamster (Phodopus sungorus).

Circadian rhythms are still expressed in animals that display daily torpor, implying a temperature compensation of the pacemaker. Nevertheless, it remains unclear how the clock works in hypothermic states and whether torpor itself, as a temperature pulse, affects the circadian system. To reveal changes in the clockwork during torpor, we compared clock gene and neuropeptide expression by in situ hybridization in the suprachiasmatic nucleus (SCN) and pineal gland of normothermic and torpid Djungarian hamsters (Phodopus sungorus). Animals from light-dark (LD) 8ratio16 were sacrificed at 8 time points throughout 24 h. To investigate the effect of a previous torpor episode on the clock, we sacrificed a group of normothermic hamsters 1 day after torpor. In normothermic animals, Per1 peaked at zeitgeber time (ZT)4; whereas, Bmal1 reached maximal expression between ZT16 and ZT19. AVP mRNA in the SCN showed highest levels at ZT7. On the day of torpor, the levels of all mRNAs investigated, except for AVP mRNA, were increased during the torpor bout. Moreover, the Bmal1 rhythm was advanced. On the day after the hypothermia, Bmal1 and AVP rhythms showed severely depressed amplitude. Those distinct amplitude changes of Bmal1 and AVP on the day after a torpor episode expression suggests that torpor affects the circadian system, probably by altered translational processes that might lead to a modified protein feedback on gene expression. In the pineal gland, an important clock output, Aanat expression, peaked between ZT16 and ZT22 in normothermic animals. Aanat levels were significantly advanced on the day of hypothermia, an effect which was still visible 1 day afterward. In summary, this study showed that daily torpor affects the phase and amplitude of rhythmic clock gene and clock-controlled gene expression in the SCN. Furthermore, the rhythmic gene expression in a peripheral oscillator, the pineal gland, is also affected.