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A comparison of the heart rate at different ambient temperatures during long-term hibernation in the garden dormouse, Eliomys quercinus L.

Heart rate in hibernating garden dormice, Eliomys quercinus, was studied by means of permanently implanted electrodes; ambient temperatures (TA's) were maintained at 0, 4, 6.5, and 9 degrees C during the 6-month test period in each winter study. The animals were kept under constant conditions in darkness and without food or water. Heart rate remained at a low level during deep hibernation at all TA's studied. There were no differences in midwinter values between the TA's of 6.5 and 9 degrees C: the means were 9-12 beats/min during apnea. Heart rate thus differs from other hibernation parameters studied simultaneously, which were strongly TA dependent. However, the optimal TA of 4 degrees C could be distinguished and heart rate was significantly lower, 8-10 beats/min. At 0 degree C the values were slightly higher: 12-13 beats/min. The TA of 0 degree C was exceptional for all parameters studied. At the beginning of the hibernation season was a transition period with elevated heart rate values. Respiratory-related heart-rate changes appeared during periodic respiration, heart rate being significantly higher during respiratory periods at all TA's. At 0, 6.5, and 9 degrees C tachycardia occurred also during apnea, very close to the respiratory period. There are responses that are comparable to hypoxic environmental conditions during hibernation, diving, and pregnancy and under high-altitude conditions. Parallel adaptations appear in heart rate and respiration, i.e., bradycardia and periodic respiration. In conclusion, heart-rate values were low during deep hibernation, and compared with other parameters measured at different TA's heart rate is maintained inside narrow limits during deep hibernation.

Animals↗

Viscosity of water in hibernating and nonhibernating mammals estimated by proton NMR relaxation times.

Longitudinal (T1) and transverse (T2) nuclear magnetic resonance relaxation times were measured in vitro at 37, 30, 25, 15, and 5 degrees C on serum, brain, liver, kidney, and heart samples from a hibernator, the European hamster, active in summer (SA), active in winter, or in the hibernating state in winter; from a less efficient hibernator, the golden hamster; and from a homeotherm, the rat. T1 and T2 relaxation times varied between species and in the European hamster between the active and hibernating subjects. Despite the major relaxation time differences between the organs, NMR relaxation time measurements showed a general trend to an increase in the viscosity of water for the European hamster in the active state. Although these modifications were not directly related to the process of hibernation itself, the relaxation times observed in the hibernating animals were closer to those seen in the rat. This evidenced that changes of physical properties of water reflect a better adaptation to low temperatures of the hamster, as compared to the nonhibernator, given that the low water viscosity of SA hamster allows the decrease of the viscosity with temperature during the hibernating state. These in vitro studies permit the study the viscosity which is an important physicochemical parameter involved in NMR longitudinal relaxation time of water proton. More detailed studies of other physiological parameters must be undertaken by further in vivo measurements.

Animals↗

Decreases in pineal melatonin content during the hibernation bout in the golden-mantled group squirrel, Spermophilus lateralis.

The role of the pineal gland in modulating the rhythmic bouts of hibernation in the golden-mantled ground squirrel (S. lateralis) was explored by comparing pineal melatonin content in hibernating animals with that of euthermic animals at the same time of year. Significant decreases in pineal melatonin content were found in hibernating versus euthermic animals. In addition, significantly lower values for pineal melatonin were observed in hibernating animals that were sacrificed in the late bout period, just prior to expected spontaneous arousal, as compared to hibernating animals that were sacrificed on the first day of their respective bouts. A strong correlation was evident between pineal melatonin content and the duration of the individual hibernation bout. These data suggest that pineal melatonin may be important in determining the duration of individual bouts of hibernation in this species.

Acclimatization↗

Involvement of brain tryptophan hydroxylase in the mechanism of hibernation.

Marked changes were revealed in the activity of the key enzyme in serotonin biosynthesis, tryptophan hydroxylase (TPH), during entry into hibernation, hibernation, and arousal in ground squirrels (Citellus erythrogenys). An increase in TPH activity was found in the midbrain, hippocampus, and striatum during the prehibernation period in euthermic ground squirrels. A further increase in TPH activity was observed during the entry into hibernation. Significant elevation was found not only in potential TPH activity measured at the incubation temperature of 37 degrees C but also at incubation temperature of 7 degrees C, approximating the body temperature in hibernation. Vmax in the midbrain of hibernating animals was about 50% higher than in active ones without significant changes in Km. Thus, brain TPH maintains functionality during torpidity and is activated before the entry into hibernation. The results support the idea that brain serotonin is crucially involved in the transition to and the maintenance of the hibernation state.

Animals↗

Relationship between structure and function of liver mitochondria from hibernating and active ground squirrels, Citellus undulatus.

1. Electron microscopy of liver tissue preparations, obtained from hibernating ground squirrels, reveals mitochondria in a condensed state. 2. When kept on ice, mitochondria isolated from the livers of hibernating and active ground squirrels are in a shrunken state. 3. Incubation of mitochondria isolated from the livers of active ground squirrels in the presence of succinate, at 27 degrees C, results in mitochondrial swelling, while mitochondria from hibernating ground squirrels under the same conditions remain relatively shrunken. 4. The swollen mitochondria from active ground squirrels show high oxidative activity, while the shrunken mitochondria from hibernating animals show low oxidative activity. 5. Swelling of mitochondria from hibernating ground squirrels in a hypo-osmolar medium is accompanied by a significant increase in oxidative activity. 6. It is inferred that the shrinkage of hibernating ground squirrel mitochondria is one of the main causes of the inhibition of oxidative activity and other mitochondrial functions during hibernation.

Animals↗

In vitro measurements of cholinergic activity in brain regions of hibernating ground squirrels.

High affinity choline uptake (HACU) and choline acetyltransferase (ChAT) activity were measured in synaptosomal P2 fractions from four brain regions in a mammalian hibernator, the golden-mantled ground squirrel. The 14CO2 evolution from [6-14C]glucose was also measured. Comparisons were made across the euthermic (not hibernating) and hibernation state in synaptosomes from cortex (CTX), preoptic area and hypothalamus (POA/HYP), olfactory apparatus (OA), and hippocampus (HPC). HACU was significantly increased in the CTX, from hibernating ground squirrels compared to euthermic animals. ChAT activity was significantly increased in the CTX and OA from hibernating animals. No change in either cholinergic marker was evident for the POA/HYP and HPC. The evolution of 14CO2 from [6-14C]glucose was generally, though not significantly, higher for synaptosomes from euthermic animals compared to hibernating animals. The results are discussed with reference to the involvement of cholinergic mechanisms in the control of hibernation.

Animals↗

State-dependent changes of brain endogenous opioids in mammalian hibernation.

Endogenous opioids belonging to three opioid families were measured in different states of the hibernation cycle in brain of the Columbian ground squirrels. Using high-performance liquid chromatography-EC detection, the hypothalamic and septal concentrations of met-enkephalin were found to be significantly higher (p < 0.05) in the hibernating state than that in the nonhibernating state. In contrast, met-enkephalin content in the medulla decreased significantly during hibernation. Leuenkephalin content was only increased in the hypothalamus of hibernating animals. Using radioimmunoassay, dynorphin A-like immunoreactivity was observed to increase in the claustrum and striatum, whereas beta-endorphin-like peptides showed a significant increase in the hypothalamus during hibernation. It is evident that the changes in endogenous opioids in brain during hibernation are state dependent, type specific and region specific. These specific alterations of various endogenous opioids may imply their different roles in hibernation.

Animals↗

Brain antioxidant levels in hamsters during hibernation, arousal and cenothermia.

Warming from hibernation to cenothermia involves intense metabolic activity coincident with large fluxes in blood flow and is considered to be a period of oxidative stress during which utilization of endogenous antioxidants prevents pathology. Very slow flow brain microdialysis enabled temperature independent sampling of the brain striatal extracellular fluid (ECF) during hibernation, arousal and cenothermia in Syrian hamsters (Mesocricetus auratus). Brain tissue and dialysates were analyzed to provide the first profile of changes in ECF levels of ascorbate (AA), glutathione (GSH) and urate during hibernation and the transition to cenothermia. Brain tissue content of AA and GSH was unchanged between hibernation and cenothermia; however, arousal was associated with substantial oxidation of AA from the brain ECF and plasma compartments. ECF GSH increased during arousal. Brain tissue urate content was decreased 50% during hibernation. ECF urate levels were unchanged in hibernation and cenothermia but transiently increased 100% during arousal. These experiments demonstrate that arousal from hibernation is a suitable experimental model for examination of the mechanisms by which non-pathological tissue integrity is maintained in the face of the generation of free radicals during increasing metabolism, temperature and cerebral reperfusion.

Adaptation, Physiological↗

Organ arrest, protection and preservation: natural hibernation to cardiac surgery.

Cardiac surgery continues to be limited by an inability to achieve complete myocardial protection from ischemia-reperfusion injury. This paper considers the following questions: (1) what lessons can be learned from mammalian hibernators to improve current methods of human myocardial arrest, protection and preservation? and (2) can the human heart be pharmacologically manipulated during acute global ischemia to act more like the heart of a hibernating mammal? After reviewing the major entropy-slowing strategies of hibernation, a major player identified in the armortarium is maintenance of the membrane potential. The resting membrane potential of the hibernator's heart appears to be maintained close to its pre-torpid state of around -85 mV. In open-heart surgery, 99% of all surgical heart arrest solutions (cardioplegia) employ high potassium (>16 mM) which depolarises the membrane voltage from -85 to around -50 mV. However, depolarising potassium cardioplegia has been increasingly linked to myocyte and microvascular damage leading to functional loss during reperfusion. Our recent work has been borrowed from hibernation biology and is focused on a very different arrest strategy which 'clamps' the membrane near its resting potential and depresses O2 consumption from baseline by about 90%. The new 'polarising' cardioplegia incorporates adenosine and lidocaine (AL) as the arresting combination, not high potassium. Studies in the isolated rat heart show that AL cardioplegia delivered at 37 degrees C can arrest the heart for up to 4 h with 70-80% recovery of the cardiac output, 85-100% recovery of heart rate, systolic pressure and rate-pressure product and 70-80% of baseline coronary flows. Only 14% of hearts arrested with crystalloid St. Thomas' solution No. 2 cardioplegia survived after 4 h. In conclusion, maintenance of the myocardial membrane potential near or close to its resting state appears to be an important feature of the hibernator's heart that may find great utility in surgical arrest and cellular preservation strategies. Identifying and safely turning 'off' and 'on' the entropy-slowing genes to down-regulate the hibernator's heart and applying this to human organs and tissues remains a major challenge for future genomics and proteomics.

Adenosine Triphosphate↗

Enhanced antioxidant defense due to extracellular catalase activity in Syrian hamster during arousal from hibernation.

Mammalian hibernators are considered a natural model for resistance to ischemia-reperfusion injuries, and protective mechanisms against oxidative stress evoked by repeated hibernation-arousal cycles in these animals are increasingly the focus of experimental investigation. Here we show that extracellular catalase activity provides protection against oxidative stress during arousal from hibernation in Syrian hamster. To examine the serum antioxidant defense system, we first assessed the hibernation-arousal state-dependent change in serum attenuation of cytotoxicity induced by hydrogen peroxide. Serum obtained from hamsters during arousal from hibernation at a rectal temperature of 32 degrees C, concomitant with the period of increased oxidative stress, attenuated the cytotoxicity four-fold more effectively than serum from cenothermic control hamsters. Serum catalase activity significantly increased during arousal, whereas glutathione peroxidase activity decreased by 50%, compared with cenothermic controls. The cytoprotective effect of purified catalase at the concentration found in serum was also confirmed in a hydrogen peroxide-induced cytotoxicity model. Moreover, inhibition of catalase by aminotriazole led to an 80% loss of serum hydrogen peroxide scavenging activity. These results suggest that extracellular catalase is effective for protecting hibernators from oxidative stress evoked by arousal from hibernation.

Animals↗

Hibernation, a model of neuroprotection.

Hibernation, a natural model of tolerance to cerebral ischemia, represents a state of pronounced fluctuation in cerebral blood flow where no brain damage occurs. Numerous neuroprotective aspects may contribute in concert to such tolerance. The purpose of this study was to determine whether hibernating brain tissue is tolerant to penetrating brain injury modeled by insertion of microdialysis probes. Guide cannulae were surgically implanted in striatum of Arctic ground squirrels before any of the animals began to hibernate. Microdialysis probes were then inserted in some animals after they entered hibernation and in others while they remained euthermic. The brain tissue from hibernating and euthermic animals was examined 3 days after implantation of microdialysis probes. Tissue response, indicated by examination of hematoxylin and eosin-stained tissue sections and immunocytochemical identification of activated microglia, astrocytes, and hemeoxygenase-1 immunoreactivity, was dramatically attenuated around probe tracks in hibernating animals compared to euthermic controls. No difference in tissue response around guide cannulae was observed between groups. Further study of the mechanisms underlying neuroprotective aspects of hibernation may lead to novel therapeutic strategies for stroke and traumatic brain injury.

Animals↗

The translation state of differentially expressed mRNAs in the hibernating 13-lined ground squirrel (Spermophilus tridecemlineatus).

The translation state of differentially expressed mRNAs were compared in kidney and brown adipose tissue of the hibernating ground squirrel, Spermophilus tridecemlineatus. Polysome analysis revealed a striking disaggregation of polyribosomes during hibernation and the redistribution of Cox4 (cytochrome c oxidase subunit 4) and Oct2 (organic cation transporter type 2) transcripts into monosome and mRNP fractions of kidney cytoplasmic extracts. Additionally, OCT2 protein levels decreased in kidney of hibernating animals in line with a strong decrease (85%) in translation rate compared with euthermic kidney. There was no translational depression in brown adipose tissue during hibernation and the H isoform of fatty-acid-binding protein (H-FABP), that is up-regulated during hibernation, was increasingly abundant in the heavy polyribosome fraction isolated from the brown adipose of hibernators. This may indicate the existence of a tissue-specific mechanism for the translational control of a subset of genes that are physiologically relevant to the survival of hibernation.

Adipose Tissue, Brown↗

Autoradiographic determination of changes in opioid receptor binding in the limbic system of the Columbian ground squirrel at different hibernation states.

To localize and quantify the state-dependent changes in various opioid receptor subtypes in the limbic system of non-hibernating and hibernating Columbian ground squirrels, quantitative receptor-binding autoradiography was used. Compared to the non-hibernating animals, the binding density of [3H]-[D-Pen2,5]-enkephalin (DPDPE) to the delta receptor in the lateral septum, CA3, and the hippocampal fissure of the hippocampal formation was significantly decreased in the hibernating ground squirrels. A significant reduction in the binding density of [3H]-[D-Ala2,N-Me-Phe4,Gly-ol5]-enkephalin (DAGO) to mu receptor was also observed in the medial septum and the CA3 region of the hippocampus of the hibernating animals. In contrast, a decrease in [3H]ethylketocyclazocine (EKC) binding to the kappa receptor was only observed in the claustrum and CA3 of the hippocampus during hibernation. The differential changes in binding to various opioid receptors suggest that different opioid subtypes may exert different physiological roles in regulating the specific states (entrance, maintenance and arousal) of a hibernation bout.

Animals↗

Nitric oxide-dependent vasodilation maintains blood flow in true hibernating myocardium.

Patients with chronic coronary artery disease may exhibit chronically depressed regional myocardial function, which can be reversed by revascularization. This was called hibernating myocardium, since it was thought that myocardial blood flow was also chronically reduced and since animals during true hibernation are thought to have downregulated function and blood flow. The missing link in this syllogism, whose discovery became our goal, was to measure myocardial blood flow during true hibernation. Five woodchucks (Marmota monax) were anesthetized and chronically instrumented. Hemodynamics were recorded prior to placement in a hibernaculum and again after the animals displayed a stable period of hibernation. Body temperature had fallen from 35.7 +/- 0.5 to 13.4 +/- 0.6 degrees C. Heart rate, mean aortic pressure, and left ventricular dP/dt had decreased, P < 0.05, by 67 +/- 12% from 86 +/- 4 bpm, 36 +/- 11% from 76 +/- 4 mmHg, and 61 +/- 11% from 1924 +/- 222 mmHg/s, respectively. Although renal blood flow, as assessed by radioactive microspheres, had decreased by 95 +/- 2% and overall cardiac output fell by approximately 84%, surprisingly, transmural myocardial blood flow did not fall during hibernation (0.70 +/- 0.16 ml/min/g) relative to when awake (0.84 +/- 0.09 ml/min/g). Interestingly, during hibernation, nitric oxide (NO) synthase inhibition resulted in a significant decrease, P < 0.05, in myocardial blood flow (-41 +/- 5%) and increase in coronary vascular resistance (CVR, +90 +/- 17% from 77 +/- 13 mmHg min g/ml). Thus, true mammalian hibernation results in downregulation of blood flow to the visceral organs, e.g. to the kidneys, but a surprising maintenance of myocardial blood flow, which appears to be due to an NO mechanism.

Animals↗

Quantitative ultrastructural changes of hepatocyte constituents in euthermic, hibernating and arousing dormice (Muscardinus avellanarius).

Hibernating animals represent a suitable model for investigating the structural effects of drastic changes in cell activity under physiological conditions. In this study we investigated by means of electron microscopy and morphometrical analysis the fine structural counterpart of functional rest in hepatocytes of the hibernating dormouse, Muscardinus avellanarius, in comparison with arousing and euthermic dormice. Our observations demonstrate that during hibernation several structural constituents of the hepatocyte undergo modifications. In particular, during deep hibernation, the total cell and cytoplasm area significantly reduced, as well as the total and percent glycogen and residual body area, and the Golgi apparatus almost disappeared. Upon arousal, the amount of glycogen was minimal, whereas total cell and cytoplasm area significantly increased towards the euthermic value as well as total and percent residual body area. In comparison with the euthermic condition, the total and percent cell lipid area significantly increased in early hibernation, reduced in deep hibernation and almost disappeared during arousal. Taken together, our findings give quantitative ultrastructural support to the marked reduction found in hepatocyte functional activities during hibernation. Such a reduced activity involves profound rearrangement of the euthermic cell structure, which is rapidly resumed upon arousal.

Animals↗

Hibernation-induction trigger. II. In vitro effects of prairie dog plasma albumin on mouse vas deferens contractility.

Involvement of opioid molecules in hibernation is well established, with the delta opioid receptor implicated in hibernation induction. Previous studies have shown that plasma albumin fractions (PAFs) from hibernating mammals contain an uncharacterized ligand called "hibernation-induction trigger" (HIT), which causes inhibition of induced contractility in the guinea pig ileum (GPI). In part I of this study, we described effects of PAF from two species of prairie dogs on induced contractility of the GPI. In the present study (part II), we examine the response of the mouse vas deferens (MVD), which is populated with the delta receptor subtype, to increasing concentrations of PAF from the white-tailed prairie dog (WT) and the black-tailed prairie dog (BT). Dose-response curves of lyophilized PAF yielded IC50 values (mg) (mean dose that inhibits contractility to 50% of control) of 11.0 for summer WT, 10.6 for hibernating WT, 9.4 for summer BT, 12.2 for winter active BT, and 4.7 for winter hibernating BT. These results suggest that delta opioid (HIT) is present in both species throughout the calendar year and that the induction of hibernation may involve not only levels of opioid but also dynamic interactions between endogenous opioid and its receptors.

Animals↗

Effect of hibernation on responses of hamster vas deferens to sympathetic nerve stimulation and exogenous neurotransmitters.

The present study investigated the responses of the vas deferens to sympathetic nerve stimulation and exogenous neurotransmitters taken from golden hamsters which had undergone 8 weeks of hibernation, 2 h of arousal from hibernation, those exposed to the cold but which failed to hibernate and age-matched control animals. Electrical field stimulation (EFS) of the vas deferens from each group produced frequency-dependent, tetrodotoxin-sensitive contractions. Contractions elicited by low frequencies of EFS in the hibernating group were significantly greater than in the other groups in the absence of any blocking agents. In the presence of the alpha1-adrenoceptor antagonist prazosin (3 microM) responses from all groups were reduced by approximately 40%, with the residual responses from the hibernating group being somewhat increased compared to the other groups. In the presence of the P2X receptor antagonist pyridoxalphosphate-6-azophenyl-2'4'-disulphonic acid (30 microM), there was no significant difference in responses from all 4 groups. Exogenously applied beta, gamma-methylene ATP (beta,gamma-meATP; 0.1-300 microM), a P2X receptor agonist, and noradrenaline (NA; 30 nM(-1) mM) both caused transient concentration-dependent contractions in all groups of animals. Contractions to beta,gamma-meATP at concentrations above 0.3 microM, and NA above 0.3 microM in the hibernating animals were statistically significantly greater than the cold- and age-matched control groups, although not significantly different from the aroused group. This study has shown postjunctional increases in responses to beta,gamma-meATP and NA as a result of hibernation, possible explanations for these increases are discussed.

Adenosine Triphosphate↗

Beta-adrenergic signal transduction in the hypothalamus of the European hamster: relation with the seasonal hibernation cycle and the diurnal activity cycle.

Mammalian hibernation, an adaptation to survive harsh winter conditions, is one of the most prominent seasonal rhythmic processes exactly regulated on a low metabolic level. Diurnal variations in vegetative physiology are missing during hibernation; however, a precisely working diurnal system is mandatory for both the proper initiation and termination of the annual hibernation phase and the periodical arousal reactions. Biorhythms and the vegetative physiological processes connected with hibernation are, among others, controlled by hypothalamic noradrenaline systems. In this study, the density, binding capacity, and relative proportions of beta 1- and beta 2-adrenergic receptors (AR) within the hypothalamus of: 1) motorically inactive summer; 2) motorically active summer; 3) aroused, motorically active winter; and 4) deeply hibernating winter European hamsters (Cricetus cricetus) were studied. For further analysis of the beta-adrenergic signal transduction cascade, the activity of adenylyl cyclase (AC) was measured by formation of cAMP in controls, after stimulation of G proteins, or after forskolin stimulation without or in presence of manganese ions. While beta 1- and beta 2-AR subtypes were nearly equally abundant (50% beta 1:50% beta 2) in active summer, inactive summer, and hibernating hamsters, a significant redistribution in favor of beta 2-AR occurred after arousal (40% beta 1:60% beta 2). The activity of AC was much higher in active summer hamsters than in inactive summer, aroused winter, and hibernating winter hamsters. When AC was stimulated by guanylylimidophosphate [Gpp(NH)p], MnCl2, forskolin, or by forskolin in presence of MnCl2 instead of MgCl2, the potency to stimulate AC was found to show the following rank order: basal < Gpp(NH)p < MnCl2 < or = forskolin + MnCl2 < forskolin.

1-Methyl-3-isobutylxanthine↗