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PubMed2005

Elevated plasma concentrations of haptoglobin in European brown bears during hibernation.

Haptoglobin (Hp), a hemoglobin-binding protein, is known as an acute phase protein and increases during the acute phase of inflammation in most mammals. We reported previously in brown bears that the mean Hp concentrations were higher in blood samples obtained in winter than those in spring. To examine a possible relation of the seasonal variations of Hp to hibernation, in the present study, we measured the plasma concentrations of Hp as well as some other acute phase proteins (alpha(2)-macroglobulin, alpha(1)-antitrypsin, C-reactive protein) in 6 European brown bears (Ursus arctos), from which blood samples were obtained at 5-6 different months of year including February, the time of hibernation. The Hp concentrations showed clear seasonal variations, being highest in February. The alpha(2)-macroglobulin concentrations also showed a similar but much smaller rise in February, but those of alpha(1)-antitrypsin and C-reactive protein did not show any seasonal variations. Our results suggest that the seasonal variation of plasma Hp concentration in brown bears is associated with a hibernation-specific mechanism more than that of acute phase response.

Animals↗
PubMed2006

Adrenergic receptor density in brown adipose tissue of active and hibernating hamsters and ground squirrels.

The ligand-binding characteristics (B(max) and K(D)) of alpha(1)- and beta(1)/beta(2)-adrenoceptors were investigated in membranes prepared from brown adipose tissue (BAT) of warm-acclimated, cold-acclimated, hibernating and arousing ground squirrels (Spermophillus undulatus) and hamsters (Mesocricetus auratus) by specific binding of [(3)H]prazosin and [(3)H]CGP-12177, respectively. The physiological state did not change the affinity for the adrenoceptors in the BAT of ground squirrels and hamsters. There was a significant decrease in alpha(1)-receptor density in arousing ground squirrels and a significant decrease in beta(1)/beta(2) density in hibernating ground squirrels. The level of alpha(1)-receptors was in all conditions higher than that of beta(1)/beta(2) receptors. The results indicate a possible change in balance of adrenoceptor density in the processes of cold acclimation, hibernation and arousal. The balance between the various adrenoceptor subtypes may be important for the final effect of catecholamines in BAT in different physiological states.

Acclimatization↗
PubMed2004

Ascorbate distribution during hibernation is independent of ascorbate redox state.

Distribution of ascorbate into tissues is an essential process in ascorbate antioxidant defense. Hibernating animals are studied as a model of tolerance to ischemia-reperfusion because of their tolerance to fluctuations in blood flow associated with prolonged torpor and periodic arousal episodes. Throughout hibernation, plasma ascorbate concentration ([Asc](p)) repetitively increases during torpor, then falls during periodic arousal bouts. We previously proposed that high [Asc](p) provides a ready source of antioxidant protection for distribution to the central nervous system and peripheral tissues during arousal. Here we tested whether deliberate oxidation of plasma ascorbate by intravenous administration of ascorbate oxidase (AO), prior to arousal, compromised tissue levels of ascorbate or the other water-soluble antioxidants, glutathione (GSH) and urate. Although AO decreased [Asc](p) to below the level of detection during torpor and after arousal, ascorbate oxidation did not decrease post-arousal tissue levels of reduced ascorbate, glutathione, or urate in any tissue examined, except liver. The data imply that ascorbate is taken up equally well into brain and other tissues as either ascorbate or its oxidized product dehydroascorbate, with subsequent intracellular reduction of dehydroascorbate. Lack of effect of ascorbate oxidation on tissue levels of GSH or urate indicates that dehydroascorbate uptake and reduction do not compromise tissue concentrations of these other water-soluble antioxidants. Thus, we show equal availability of reduced and oxidized plasma ascorbate during metabolically demanding thermogenesis and reperfusion associated with arousal from hibernation.

Animals↗
PubMed2006

Experimental infection of ground squirrels (Citellus pygmaeus Pallas) with Yersinia pestis during hibernation.

OBJECTIVES: In this study, we investigated the capability remain latent infection of Yersinia pestis in ground squirrels (Citellus pygmaeus Pallas) after their challenge with Y. pestis over the hibernation period. METHODS: Ground squirrels were challenged with Y. pestis by either subcutaneous injection (n=106 animals) or by infected fleabite (n=33 rodents). The development of plague in hibernating rodents was monitored daily for the signs of illness; if the animal died, the spleen, liver, lung, lymph nodes, bone brain, kidney and blood were taken for bacteriological and histopathologic examinations. RESULTS: In 35.8% (38 of 106) of the ground squirrels the plague has been developed after the challenge with Y. pestis via the s.c. injection within eight months, i.e. winter, spring and summer the periods of the observation. Overall, in 48.5% (16 of 33) of the animals the plague was developed during three months after the challenge with infected fleabite. The infection with Y. pestis was confirmed by bacteriologically and histopathological changes, e.g. pneumonia and necrosis in tissues of the ground squirrels. CONCLUSIONS: The results support the hypothesis that Y. pestis can remain latent in winter-sleeping ground squirrels during their hibernation and this phenomenon plays an important role in the epizootics of rodents after reawakening in the spring.

Animals↗
PubMed2001

Mechanisms of metabolic defense against hypoxia in hibernating frogs.

The cold submerged frog (Rana temporaria) serves as a useful model for many hibernating ectotherms that take refuge in hypoxic ponds and lakes until more favourable conditions of climate and food availability return. In all such animals, entry into a hypometabolic state effectively extends their survival time by lessening the impact of ATP demands on endogenous substrates. At the cellular level, metabolic depression may be brought about by decreasing energy-consuming processes and/or by increasing the efficiency of energy-producing pathways. Since the mitochondrion is the major contributor to the total energy production during aerobic metabolism and frog survival during winter depends on entry into a hypometabolic state, this review focuses on the respiratory properties of mitochondria that serve to increase the efficiency of energy production in hibernation. Energy conservation during overwintering also occurs through decreases in the ATP demand of the energy-consuming processes. For example, hibernating frogs decrease their ATP demands for Na(+)/K(+)-ATPase activity as part of a coordinated process of energy conservation wherein O(2)-limitation initiates a generalised suppression of ion channel densities and/or channel leak activities. The net result is that cell membrane permeabilities are reduced, thereby lowering the energetic costs of maintaining transmembrane ion gradients.

Adaptation, Physiological↗
PubMed2002

Classical pathway serum complement activity throughout various stages of the annual cycle of a mammalian hibernator, the golden-mantled ground squirrel, Spermophilus lateralis.

Little is known about the changes in the immune system that coincide with the annual cycle of hibernating mammals. This study investigates classical pathway complement activity in the serum of the golden-mantled ground squirrel, a mammalian hibernator. Complement activity varied significantly among discreet stages of the annual cycle and is lowest during torpor and greatest during stages of arousal. C3 mRNA levels follow a pattern similar to that of complement-mediated hemolysis throughout the year but do not vary significantly among hibernation states. The classical pathway of the serum complement system is able to function in vitro at 5 degrees C, although at a slower rate than at 34 degrees C. The classical pathway of the serum complement system is active throughout all phases of the annual cycle of the golden-mantled ground squirrel.

Animals↗
PubMed2002

Microbial origin of glutamate, hibernation and tissue trauma: an in vivo microdialysis study.

Using quantitative microdialysis in hibernating Arctic ground squirrels (AGS), striatal glutamate concentrations ([glu](dia)) progressively increased to approximately 200 microM after 3 days of microdialysis in euthermic but not hibernating ground squirrels. Initially, the progressive increase in [glu](dia) was thought to be related to greater tissue response in euthermic animals. Alternatively, given the vastly different body temperatures between the two groups (37 vs. 3 degrees C), glutamate might have originated from microbes, replicating at a faster rate in the warmer animals. To test these hypotheses, microdialysis was repeated using sterile technique and tissue response surrounding the probe tract was assessed in hematoxylin and eosin stained sections. Using sterile microdialysis technique, traumatic tissue response was greater in euthermic compared to hibernating tissue. However, sterile microdialysis abolished the progressive increase in glutamate. To confirm the microbial origin of glutamate we monitored [glu](dia) collected in vitro from probes immersed in glutamine-rich liquid medium incubated at 37 degrees C. In vitro, [glu](dia) increased as much as in vivo. Two bacteria isolated from in vitro dialysate and liquid medium were both identified as Ralstonia pickettii. Growth of these isolates as well as glutamate release was enhanced when glutamine rather than NH(4)NO(3) was added to the medium suggesting the bacteria utilize glutamine preferentially over ammonium as a nitrogen source.

Animals↗
PubMed2001

Neuropeptide Y gene expression in the jerboa arcuate nucleus: modulation by food deprivation and relationship with hibernation.

Using in situ hybridization, the mRNA levels encoding neuropeptide Y (NPY) was investigated in the arcuate nucleus (ARC) of jerboas under three different states of energy balance. (1) normally feeding animals, (2) hibernating animals and finally (3) animals food deprived for 5 days. The hibernating and food deprived jerboas exhibited a significant increase (130%; P < 0.05 and 210%; P < 0.01, respectively) of mRNA expression as compared with controls. This elevated NPY mRNA expression supports the hypothesis that NPY may be implicated in abnormal feeding behaviour associated with eating deprivation. The stimulation of NPY gene expression in hibernating jerboas may be related to food deprivation and / or cold exposure since NPY is known to be an hypothermiant factor. It is thus envisaged that NPY within neurons of the ARC plays an integrative role in the control of energy metabolism.

Animals↗
PubMed2001

Neuropeptide Y gene expression in the jerboa arcuate nucleus: modulation by food deprivation and relationship with hibernation.

Using in situ hybridization, the mRNA levels encoding neuropeptide Y (NPY) was investigated in the arcuate nucleus (ARC) of jerboas under three different states of energy balance. (1) normally feeding animals, (2) hibernating animals and finally (3) animals food deprived for 5 days. The hibernating and food deprived jerboas exhibited a significant increase (130%; P<0.05 and 210%; P<0.01, respectively) of mRNA expression as compared with controls. This elevated NPY mRNA expression supports the hypothesis that NPY may be implicated in abnormal feeding behaviour associated with eating deprivation. The stimulation of NPY gene expression in hibernating jerboas may be related to food deprivation and / or cold exposure since NPY is known to be a hypothermiant factor. It is thus envisaged that NPY within neurons of the ARC plays an integrative role in the control of energy metabolism.

Animals↗
PubMed1997

Characterization of opioid peptides and opioid receptors in the brain of jerboa (Jaculus orientalis), a hibernating rodent.

The present study was undertaken to investigate the biochemical characteristics of the opioid receptors and opioid peptides in the jerboa (Jaculus orientalis) brain, a subdesert rodent of Morocco. We have demonstrated the presence of delta, mu, and kappa sites in the jerboa brain. The endogenous opioid peptides methionine-enkephalin, beta-endorphin, and dynorphin were evaluated in different physiological states of the animal (active and hibernating). The circulating methionine-enkephalin in different states of the animal (active, hibernating, exposure to cold conditions, and fasting) was evaluated in the plasma. Our results indicate that the hibernating state the opioid receptors level decreased, whereas the concentration of opioid peptides increased. These findings suggest that both opioid receptors and opioid peptides could be involved in the adaptation of the jerboa to survive under thermal stress.

Acclimatization↗
PubMed1997

Iodothyronine deiodinases in a mammalian hibernator, the chipmunk (Tamias asiaticus).

We examined the activities of iodothyronine deiodinase, a key enzyme for thyroid hormone metabolism, in selected tissues of the chipmunk (Tamias asiaticus), a mammalian hibernator, of both sexes in the summer season. Reverse T3 5'-deiodinase (5'-D) activity was the highest in the liver followed by the kidney; T4 5'-D activity was the highest in brown adipose tissue (BAT) and T3 5'-deiodinase (5-D) activity was the highest in the testes followed by the brain. Distributions of three types of deiodinase activities in liver kidney BAT, and brain were comparable to other mammals reported, except that the type III deiodinase was unique in testes. The 5'-D activity of liver and kidney of chipmunks was 52% and 24%, respectively, of male rats and the 5-D activity of brain and testes of chipmunks was 227% and 567%, respectively of male rats. In addition, the cold exposure increased BAT 5'-D activity in chipmunks as reported in the ground squirrels. Our results indicated that tissue distribution of deiodinases and response to cold exposure in BAT in hibernators are similar to nonhibernators. However, there was a quantitative difference of rT3 5'-D and T3 5-D activities in some tissues between chipmunks and rats, indicating different local thyroid hormone metabolisms in hibernators and nonhibernators.

Adipose Tissue, Brown↗
PubMed2000

The histaminergic system in the brain: structural characteristics and changes in hibernation.

Histaminergic neurons in adult vertebrate brain are confined to the posterior hypothalamic area, where they are comprised of scattered groups of neurons referred to as the tuberomammillary nucleus. Histamine regulates hormonal functions, sleep, food intake, thermoregulation and locomotor activity, for example. In the zebrafish, Danio rerio, histamine was detected only in the brain, where also the histamine synthesizing enzyme L-histidine decarboxylase (HDC) was expressed. It is possible that histamine has first evolved as a neurotransmitter in the central nervous system. We established sensitive quantitative in situ hybridization methods for histamine H(1) and H(2) receptors and HDC, to study the modulation of brain histaminergic system under pathophysiological conditions. A transient increase in H(1) receptor expression was seen in the dentate gyrus and striatum after a single injection of kainic acid, a glutamate analog. H(1) antagonists are known to increase duration of convulsions, and increased brain histamine is associated with reduced convulsions in animal models of epilepsy. No HDC mRNA was detected in brain vessels by in situ hybridization, which suggests lack of histamine synthesis by brain endothelial cells. This was verified by lack of HDC mRNA in a rat brain endothelial cell line, RBE4 cells. Both H(1) and H(2) receptor mRNA was found in this cell line, and the expression of both receptors was downregulated by dexamethasone. The findings are in agreement with the concept that histamine regulates blood-brain barrier permeability through H(1) and H(2) receptor mediated mechanisms. Hibernation is characterized by a drastic reduction of central functions. The activity of most transmitter systems is maintained at a very low level. Surprisingly, histamine levels and turnover were clearly elevated in hibernating ground squirrels, and the density of histamine-containing fibers was higher than in euthermic animals. It is possible that histamine actively maintains the low activity of other transmitters during the hibernation state.

Amino Acid Sequence↗
PubMed2003

Energetics of hibernating yellow-bellied marmots (Marmota flaviventris).

Yellow-bellied marmots (Rodentia: Sciuridae) typically hibernate for eight months. This study explored energetic costs of hibernation in young and adults at 10 and 6 degrees C. Age significantly affected the percent time torpid, total and mass-specific VO(2), use of energy during torpor, and daily mass loss at 6 degrees C. Thus young had a higher mass-specific VO(2) during a torpor bout, which was attributed to higher metabolism during deep torpor. Total VO(2) during a bout was higher in young and there were significant temperature/age interactions; young had a higher VO(2) during torpor and deep torpor at 6 degrees C than at 10 degrees C. VO(2) increased at T(E)s below 6 degrees C. Young had a higher daily mass loss than adults at 6 degrees C. Euthermy increased energetic costs 19.3 times over those of torpor and 23.5 times over those of deep torpor. Energy costs are minimized by spending 88.6% of the hibernation period in torpor, by the rapid decline of VO(2) from euthermy to torpor and by allowing T(B) to decline at low T(E). Torpidity results in average energy savings during winter of 83.3% of the costs of maintaining euthermy. Energy savings are greater than those reported for Marmota marmota and M. monax.

Age Factors↗
PubMed2003

The paradoxically high reactivity of septal neurons in hibernating ground squirrels to endogenous neuropeptides is lost after chronic deafferentation of the septum from the preopticohypothalamic areas.

The effect of neuropeptides (TSKYR, TSKY and DY) and neurotransmitters (serotonin and noradrenaline) on the activity of medial septum (MS) neurons from the brain of summer wakening ground squirrels (WGS), hibernating ground squirrels (HGS), and hibernating ground squirrels with the undercut septum (UHGS) was studied. It was shown that in HGS, the neuropeptides were substantially more effective in modulating the spontaneous activity of MS neurons than in WGS. The undercutting of MS led to the disappearance of the increased responsiveness to the neuropeptides: in UHGS, neuropeptide-induced changes in the spontaneous activity became nearly identical to those in WGS. The decrease in MS responsiveness in UHGS is due mainly to pacemaker neurons, which cease to respond to the peptides. It was shown that the neuropeptides have a dual effect: they change the level of spontaneous activity through direct modulation of pacemaker potential and control responses to electrical stimulation by modulating the synaptic transmission. Contrary to neuropeptides, neurotransmitters were highly effective in neurons of all groups of animals. Presumably, the enhanced excitability of MS during hibernation, which is necessary for performing the 'sentry post' function, is formed under the influence of the preopticohypothalamic area, and this influence is mediated by peptides.

Action Potentials↗
PubMed2002

Cloning of hibernation-related genes of bullfrog (Rana catesbeiana) by cDNA subtraction.

Seven genes specifically expressed during hibernation in the bullfrog (Rana catesbeiana) were cloned from a subtracted cDNA library constructed from livers of winter bullfrogs. Those genes were fibrinogen alpha-subunit, fibrinogen gamma-subunit, complement component C3, alpha-1-microglobulin/bikunin precursor (AMBP), transferrin, apoferritin middle subunit and one novel gene. Northern hybridization has indicated that these seven genes were specifically induced or enhanced in winter. Above all, expression of the novel gene was specifically induced in winter in liver, though the expression of that was neither induced in bullfrog nor Xenopus laevis by cold treatment. The novel gene, which was designated as rc-hirp (Rana catesbeiana hibernation-related protein), encoded 420 base pairs length and a putative protein of 139 amino acid residues. Annual analyses of the expression of these genes have suggested that the seven winter-specific genes are playing an important role in hibernation processes.

Amphibian Proteins↗
PubMed2006

Identification of proteins from non-model organisms using mass spectrometry: application to a hibernating mammal.

A major challenge in the life sciences is the extraction of detailed molecular information from plants and animals that are not among the handful of exhaustively studied "model organisms." As a consequence, certain species with novel phenotypes are often ignored due to the lack of searchable databases, tractable genetics, stock centers, and more recently, a sequenced genome. Characterization of phenotype at the molecular level commonly relies on the identification of differentially expressed proteins by combining database searching with tandem mass spectrometry (MS) of peptides derived from protein fragmentation. However, the identification of short peptides from nonmodel organisms can be hampered by the lack of sufficient amino acid sequence homology with proteins in existing databases; therefore, a database search strategy that encompasses both identity and homology can provide stronger evidence than a single search alone. The use of multiple algorithms for database searches may also increase the probability of correct protein identification since it is unlikely that each program would produce false negative or positive hits for the same peptides. In this study, four software packages, Mascot, Pro ID, Sequest, and Pro BLAST, were compared in their ability to identify proteins from the thirteen-lined ground squirrel (Spermophilus tridecemlineatus), a hibernating mammal that lacks a completely sequenced genome. Our results show similarities as well as the degree of variability among different software packages when the identical protein database is searched. In the process of this study, we identified the up-regulation of succinyl CoA-transferase (SCOT) in the heart of hibernators. SCOT is the rate-limiting enzyme in the catabolism of ketone bodies, an important alternative fuel source during hibernation.

Algorithms↗
PubMed2001

Reassessment of the cold-labile nature of phosphofructokinase from a hibernating ground squirrel.

This study reassesses the proposal that cellular conditions of low temperature and relative acidosis during hibernation contribute to a suppression of phosphofructokinase (PFK) activity which, in turn, contributes to glycolytic rate suppression during torpor. To test the proposal that a dilution effect during in vitro assay of PFK was the main reason for activity loss (tetramer dissociation) at lower pH values, the influence of the macromolecular crowding agent, polyethylene glycol 8000 (PEG), on purified skeletal muscle PFK from Spermophilus lateralis was evaluated at different pH values (6.5, 7.2 and 7.5) and assay temperatures (5, 25 and 37degrees C). A 78 +/- 2.5% loss of PFK activity during 1 h incubation at 5 degrees C and pH 6.5 was virtually eliminated when 10% PEG was present (only 7.0 +/- 1.5% activity lost). The presence of PEG also largely reversed PFK inactivation at pH 6.5 at warmer assay temperatures and reversed inhibitory effects by high urea (50 or 400 mM). Analysis of pH curves at 5 degrees C also indicated that approximately 70% of activity would remain at intracellular pH values in hibernator muscle. The data suggest that under high protein concentrations in intact cells that the conditions of relative acidosis, low temperature or elevated urea during hibernation would not have substantial regulatory effects on PFK.

Animals↗
PubMed2001

Ca-ATPase activity and protein composition of sarcoplasmic reticulum membranes isolated from skeletal muscles of typical hibernator, the ground squirrel Spermophilus undulatus.

Ca-ATPase activity in sarcoplasmic reticulum (SR) membranes isolated from skeletal muscles of the typical hibernator, the ground squirrel Spermophilus undulatus, is about 2-fold lower than that in SR membranes of rats and rabbits and is further decreased 2-fold during hibernation. The use of carbocyanine anionic dye Stains-All has revealed that Ca-binding proteins of SR membranes, histidine-rich Ca-binding protein and sarcalumenin, in ground squirrel, rat, and rabbit SR have different electrophoretic mobility corresponding to apparent molecular masses 165, 155, and 170 kDa and 130, 145, and 160 kDa, respectively; the electrophoretic mobility of calsequestrin (63 kDa) is the same in all preparations. The content of these Ca-binding proteins in SR membranes of the ground squirrels is decreased 3-4 fold and the content of 55, 30, and 22 kDa proteins is significantly increased during hibernation.

Animals↗