N6-cyclohexyladenosine is better than meperidine and buspirone at suppressing metabolism during TTM32 but does not improve outcome after cardiac arrest.

N6-clyclohexyladenosine (CHA) is an adenosine A1 receptor agonist that inhibits thermogenesis. Cardiovascular side effects however, limit use of CHA as a therapeutic. We and others have shown that this can be reversed by administering 8-p-(sulfophenyl)theophylline (8-SPT), a nonspecific antagonist that does not cross the BBB. Other evidence shows that CNS actions of CHA may contribute to bradycardia through enhanced vagal tone and other mechanisms. Here we test the hypothesis that 8-SPT pretreatment alone is sufficient to prevent hypotension caused by CHA. To test this hypothesis, we pretreated rats with 8-SPT alone, and in combination with other antagonists to test the hypothesis that direct action of CHA on the heart is the primary mechanism by which CHA induces bradycardia and hypotension. Results show that pretreatment with 8-SPT alone is not sufficient to prevent CHA-induced hypotension. Pretreatment with 8-SPT or atropine alone did not prevent the fall in mean arterial pressure (MAP) and heart rate (HR), however, pretreatment with 8-SPT (25 mg/kg) and atropine (1 mg/kg) 15 min before CHA (1 mg/kg) preserves MAP and HR baseline values after CHA administration. We next asked if blood pressure was managed during the transition into a hypometabolic state, would prolong CHA-mediated inhibition of metabolism after cardiac arrest improve outcome better than anti-shivering medications meperidine and buspirone. We found that CHA-mediated hypotension can be mitigated by pretreatment with atropine and 8-SPT. This combination administered after cardiac arrest facilitated temperature management and metabolic suppression better than meperidine and buspirone, however, did not improve survival.

Remodeling of skeletal muscle myosin metabolic states in hibernating mammals.

Hibernation is a period of metabolic suppression utilized by many small and large mammal species to survive during winter periods. As the underlying cellular and molecular mechanisms remain incompletely understood, our study aimed to determine whether skeletal muscle myosin and its metabolic efficiency undergo alterations during hibernation to optimize energy utilization. We isolated muscle fibers from small hibernators, Ictidomys tridecemlineatus and Eliomys quercinus and larger hibernators, Ursus arctos and Ursus americanus. We then conducted loaded Mant-ATP chase experiments alongside X-ray diffraction to measure resting myosin dynamics and its ATP demand. In parallel, we performed multiple proteomics analyses. Our results showed a preservation of myosin structure in U. arctos and U. americanus during hibernation, whilst in I. tridecemlineatus and E. quercinus, changes in myosin metabolic states during torpor unexpectedly led to higher levels in energy expenditure of type II, fast-twitch muscle fibers at ambient lab temperatures (20 °C). Upon repeating loaded Mant-ATP chase experiments at 8 °C (near the body temperature of torpid animals), we found that myosin ATP consumption in type II muscle fibers was reduced by 77-107% during torpor compared to active periods. Additionally, we observed Myh2 hyper-phosphorylation during torpor in I. tridecemilineatus, which was predicted to stabilize the myosin molecule. This may act as a potential molecular mechanism mitigating myosin-associated increases in skeletal muscle energy expenditure during periods of torpor in response to cold exposure. Altogether, we demonstrate that resting myosin is altered in hibernating mammals, contributing to significant changes to the ATP consumption of skeletal muscle. Additionally, we observe that it is further altered in response to cold exposure and highlight myosin as a potentially contributor to skeletal muscle non-shivering thermogenesis.

Many animals use hibernation as a tactic to survive harsh winters. During this dormant, inactive state, animals reduce or limit body processes, such as heart rate and body temperature, to minimise their energy use. To conserve energy during hibernation, animals can use different approaches. For example, garden dormice undergo periodic states of extremely low core temperatures (down to 4­8^oC); whereas Eurasian brown bears see milder temperature drops (down to 23­25^oC). An important organ that changes during hibernation is skeletal muscle. Skeletal muscle typically uses large amounts of energy, making up around 50% of body mass. To survive, hibernating animals must change how their skeletal muscle uses energy. Traditionally, active myosin ­ a protein found in muscles that helps muscles to contract ­ was thought to be responsible for most of the energy use by skeletal muscle. But, more recently, resting myosin has also been found to use energy when muscles are relaxed. Lewis et al. studied myosin and skeletal muscle energy use changes during hibernation and whether they could impact the metabolism of hibernating animals. Lewis et al. assessed myosin changes in muscle samples from squirrels, dormice and bears during hibernation and during activity. Experiments showed changes in resting myosin in squirrels and dormice (whose temperature drops to 4­8^oC during hibernation) but not in bears. Further analysis revealed that cooling samples from non-hibernating muscle to 4­8^oC increased energy use in resting myosin, thereby generating heat. However, no increase in energy use was found after cooling hibernating muscle samples to 4­8^oC. This suggest that resting myosin generates heat at cool temperatures ­ a mechanism that is switched off in hibernating animals to allow them to cool their body temperature. These findings reveal key insights into how animals conserve energy during hibernation. In addition, the results show that myosin regulates energy use in skeletal muscles, which indicates myosin may be a potential drug target in metabolic diseases, such as obesity.

Characterization and Seasonal Modulation of Adenosine A1 Receptors in the Arctic Ground Squirrel Brain.

Hibernation is an adaptation that allows animals such as the Arctic ground squirrel (AGS) to survive the absence of food or water during the winter season. Understanding mechanisms of metabolic suppression during hibernation torpor promises new therapies for critical care. The activation of the Adenosine A1 receptor (A1AR) has been shown to be necessary and sufficient for entrance into hibernation with a winter season sensitization to the agonist, but the role of the A1AR in seasonal sensitization is unknown. In the current study, we characterize the A1AR in the forebrain, hippocampus and hypothalamus of summer and torpid AGS. For the first time, we define the pharmacological characteristics of the A1AR agonist, N6-cyclohexyladenosine and the A1AR antagonist dipropylcyclopentylxanthine (DPCPX) in the AGS brain. In addition, we test the hypothesis that increased A1AR agonist efficacy is responsible for sensitization of the A1AR during the torpor season. The resulting 35S-GTPγS binding data indicate an increase in agonist potency during torpor in two out of three brain regions. In addition to 35S-GTPγS binding, [3H]DPCPX saturation and competition assays establish for the first-time pharmacological characteristics for the A1AR agonist, N6-cyclohexyladenosine and the A1AR antagonist dipropylcyclopentylxanthine (DPCPX) in AGS brain.

Seasonal decrease in thermogenesis and increase in vasoconstriction explain seasonal response to N6 -cyclohexyladenosine-induced hibernation in the Arctic ground squirrel (Urocitellus parryii).

Hibernation is a seasonal phenomenon characterized by a drop in metabolic rate and body temperature. Adenosine A1 receptor agonists promote hibernation in different mammalian species, and the understanding of the mechanism inducing hibernation will inform clinical strategies to manipulate metabolic demand that are fundamental to conditions such as obesity, metabolic syndrome, and therapeutic hypothermia. Adenosine A1 receptor agonist-induced hibernation in Arctic ground squirrels is regulated by an endogenous circannual (seasonal) rhythm. This study aims to identify the neuronal mechanism underlying the seasonal difference in response to the adenosine A1 receptor agonist. Arctic ground squirrels were implanted with body temperature transmitters and housed at constant ambient temperature (2°C) and light cycle (4L:20D). We administered CHA (N6 -cyclohexyladenosine), an adenosine A1 receptor agonist in euthermic-summer phenotype and euthermic-winter phenotype and used cFos and phenotypic immunoreactivity to identify cell groups affected by season and treatment. We observed lower core and subcutaneous temperature in winter animals and CHA produced a hibernation-like response in winter, but not in summer. cFos-ir was greater in the median preoptic nucleus and the raphe pallidus in summer after CHA. CHA administration also resulted in enhanced cFos-ir in the nucleus tractus solitarius and decreased cFos-ir in the tuberomammillary nucleus in both seasons. In winter, cFos-ir was greater in the supraoptic nucleus and lower in the raphe pallidus than in summer. The seasonal decrease in the thermogenic response to CHA and the seasonal increase in vasoconstriction, assessed by subcutaneous temperature, reflect the endogenous seasonal modulation of the thermoregulatory systems necessary for CHA-induced hibernation. Cover Image for this issue: doi: 10.1111/jnc.14528.

Precise Control of Target Temperature Using N6-Cyclohexyladenosine and Real-Time Control of Surface Temperature.

Targeted temperature management is standard of care for cardiac arrest and is in clinical trials for stroke. N6-cyclohexyladenosine (CHA), an A1 adenosine receptor (A1AR) agonist, inhibits thermogenesis and induces onset of hibernation in hibernating species. Despite promising thermolytic efficacy of CHA, prior work has failed to achieve and maintain a prescribed target core body temperature (Tb) between 32°C and 34°C for 24 hours. We instrumented Sprague-Dawley rats (n = 19) with indwelling arterial and venous cannulae and a transmitter for monitoring Tb and ECG, then administered CHA via continuous IV infusion or intraperitoneal (IP) injection. In the first experiment (n = 11), we modulated ambient temperature and increased the dose of CHA in an attempt to manage Tb. In the second experiment (n = 8), we administered CHA (0.25 mg/[kg·h]) via continuous IV infusion and modulated cage surface temperature to control Tb. We rewarmed animals by increasing surface temperature at 1°C h-1 and discontinued CHA after Tb reached 36.5°C. Tb, brain temperature (Tbrain), heart rate, blood gas, and electrolytes were also monitored. Results show that titrating dose to adjust for individual variation in response to CHA led to tolerance and failed to manage a prescribed Tb. Starting with a dose (0.25 mg/[kg·h]) and modulating surface temperature to prevent overcooling proved to be an effective means to achieve and maintain Tb between 32°C and 34°C for 24 hours. Increasing surface temperature to 37°C during CHA administration brought Tb back to normothermic levels. All animals treated in this way rewarmed without incident. During the initiation of cooling, we observed bradycardia within 30 minutes of the start of IV infusion, transient hyperglycemia, and a mild hypercapnia; the latter normalized via metabolic compensation. In conclusion, we describe an intravenous delivery protocol for CHA at 0.25 mg/(kg·h) that, when coupled with conductive cooling, achieves and maintains a prescribed and consistent target Tb between 32°C and 34°C for 24 hours.

The Raphe Pallidus and the Hypothalamic-Pituitary-Thyroid Axis Gate Seasonal Changes in Thermoregulation in the Hibernating Arctic Ground Squirrel (Urocitellus parryii).

Thermoregulation is necessary to maintain energy homeostasis. The novel discovery of brown adipose tissue (BAT) in humans has increased research interests in better understanding BAT thermogenesis to restore energy balance in metabolic disorders. The hibernating Arctic ground squirrel (AGS) offers a novel approach to investigate BAT thermogenesis. AGS seasonally increase their BAT mass to increase the ability to generate heat during interbout arousals. The mechanisms promoting the seasonal changes in BAT thermogenesis are not well understood. BAT thermogenesis is regulated by the raphe pallidus (rPA) and by thyroid hormones produced by the hypothalamic-pituitary-thyroid (HPT) axis. Here, we investigate if the HPT axis and the rPA undergo seasonal changes to modulate BAT thermogenesis in hibernation. We used histological analysis and tandem mass spectrometry to assess activation of the HPT axis and immunohistochemistry to measure neuronal activation. We found an increase in HPT axis activation in fall and in response to pharmacologically induced torpor when adenosine A1 receptor agonist was administered in winter. By contrast, the rPA neuronal activation was lower in winter in response to pharmacologically induced torpor. Activation of the rPA was also lower in winter compared to the other seasons. Our results suggest that thermogenic capacity develops during fall as the HPT axis is activated to reach maximum capacity in winter seen by increased free thyroid hormones in response to cooling. However, thermogenesis is inhibited during torpor as sympathetic premotor neuronal activation is lower in winter, until arousal when inhibition of thermogenesis is relieved. These findings describe seasonal modulation of thermoregulation that conserves energy through attenuated sympathetic drive, but retains heat generating capacity through activation of the HPT axis.

Optimization of Thermolytic Response to A1 Adenosine Receptor Agonists in Rats.

Cardiac arrest is a leading cause of death in the United States, and, currently, therapeutic hypothermia, now called targeted temperature management (TTM), is the only recent treatment modality proven to increase survival rates and reduce morbidity for this condition. Shivering and subsequent metabolic stress, however, limit application and benefit of TTM. Stimulating central nervous system A1 adenosine receptors (A1AR) inhibits shivering and nonshivering thermogenesis in rats and induces a hibernation-like response in hibernating species. In this study, we investigated the pharmacodynamics of two A1AR agonists in development as antishivering agents. To optimize body temperature (Tb) control, we evaluated the influence of every-other-day feeding, dose, drug, and ambient temperature (Ta) on the Tb-lowering effects of N6-cyclohexyladenosine (CHA) and the partial A1AR agonist capadenoson in rats. The highest dose of CHA (1.0 mg/kg, i.p.) caused all ad libitum-fed animals tested to reach our target Tb of 32°C, but responses varied and some rats overcooled to a Tb as low as 21°C at 17.0°C Ta Dietary restriction normalized the response to CHA. The partial agonist capadenoson (1.0 or 2.0 mg/kg, i.p.) produced a more consistent response, but the highest dose decreased Tb by only 1.6°C. To prevent overcooling after CHA, we studied continuous i.v. administration in combination with dynamic surface temperature control. Results show that after CHA administration control of surface temperature maintains desired target Tb better than dose or ambient temperature.

Arctic ground squirrel hippocampus tolerates oxygen glucose deprivation independent of hibernation season even when not hibernating and after ATP depletion, acidosis, and glutamate efflux.

Cerebral ischemia/reperfusion (I/R) triggers a cascade of uncontrolled cellular processes that perturb cell homeostasis. The arctic ground squirrel (AGS), a seasonal hibernator resists brain damage following cerebral I/R caused by cardiac arrest and resuscitation. However, it remains unclear if tolerance to I/R injury in AGS depends on the hibernation season. Moreover, it is also not clear if events such as depletion of ATP, acidosis, and glutamate efflux that are associated with anoxic depolarization are attenuated in AGS. Here, we employ a novel microperfusion technique to test the hypothesis that tolerance to I/R injury modeled in an acute hippocampal slice preparation in AGS is independent of the hibernation season and persists even after glutamate efflux. Acute hippocampal slices were harvested from summer euthermic AGS, hibernating AGS, and interbout euthermic AGS. Slices were subjected to oxygen glucose deprivation (OGD), an in vitro model of I/R injury to determine cell death marked by lactate dehydrogenase (LDH) release. ATP was assayed using ENLITEN ATP assay. Glutamate and aspartate efflux was measured using capillary electrophoresis. For acidosis, slices were subjected to pH 6.4 or ischemic shift solution (ISS). Acute hippocampal slices from rats were used as a positive control, susceptible to I/R injury. Our results indicate that when tissue temperature is maintained at 36°C, hibernation season has no influence on OGD-induced cell death in AGS hippocampal slices. Our data also show that tolerance to OGD in AGS hippocampal slices occurs despite loss of ATP and glutamate release, and persists during conditions that mimic acidosis and ionic shifts, characteristic of cerebral I/R. Read the Editorial Comment for this article on page 10.

Neural Signaling Metabolites May Modulate Energy Use in Hibernation.

Despite an epidemic in obesity and metabolic syndrome limited means exist to effect adiposity or metabolic rate other than life style changes. Here we review evidence that neural signaling metabolites may modulate thermoregulatory pathways and offer novel means to fine tune energy use. We extend prior reviews on mechanisms that regulate thermogenesis and energy use in hibernation by focusing primarily on the neural signaling metabolites adenosine, AMP and glutamate.

Translating drug-induced hibernation to therapeutic hypothermia.

Therapeutic hypothermia (TH) improves prognosis after cardiac arrest; however, thermoregulatory responses such as shivering complicate cooling. Hibernators exhibit a profound and safe reversible hypothermia without any cardiovascular side effects by lowering the shivering threshold at low ambient temperatures (Ta). Activation of adenosine A1 receptors (A1ARs) in the central nervous system (CNS) induces hibernation in hibernating species and a hibernation-like state in rats, principally by attenuating thermogenesis. Thus, we tested the hypothesis that targeted activation of the central A1AR combined with a lower Ta would provide a means of managing core body temperature (Tb) below 37 °C for therapeutic purposes. We targeted the A1AR within the CNS by combining systemic delivery of the A1AR agonist (6)N-cyclohexyladenosine (CHA) with 8-(p-sulfophenyl)theophylline (8-SPT), a nonspecific adenosine receptor antagonist that does not readily cross the blood-brain barrier. Results show that CHA (1 mg/kg) and 8-SPT (25 mg/kg), administered intraperitoneally every 4 h for 20 h at a Ta of 16 °C, induce and maintain the Tb between 29 and 31 °C for 24 h in both naïve rats and rats subjected to asphyxial cardiac arrest for 8 min. Faster and more stable hypothermia was achieved by continuous infusion of CHA delivered subcutaneously via minipumps. Animals subjected to cardiac arrest and cooled by CHA survived better and showed less neuronal cell death than normothermic control animals. Central A1AR activation in combination with a thermal gradient shows promise as a novel and effective pharmacological adjunct for inducing safe and reversible targeted temperature management.

Circannual rhythm in body temperature, torpor, and sensitivity to A1 adenosine receptor agonist in arctic ground squirrels.

A1 adenosine receptor (A1AR) activation within the central nervous system induces torpor, but in obligate hibernators such as the arctic ground squirrel (AGS; Urocitellus parryii), A1AR stimulation induces torpor only during the hibernation season, suggesting a seasonal increase in sensitivity to A1AR signaling. The purpose of this research was to investigate the relationship between body temperature (Tb) and sensitivity to an adenosine A1 receptor agonist in AGS. We tested the hypothesis that increased sensitivity in A1AR signaling would lead to lower Tb in euthermic animals during the hibernation season when compared with the summer season. We further predicted that if a decrease in euthermic Tb reflects increased sensitivity to A1AR activation, then it should likewise predict spontaneous torpor. We used subcutaneous IPTT-300 transponders to monitor Tb in AGS housed under constant ambient conditions (12:12 L:D, 18 °C) for up to 16 months. These animals displayed an obvious rhythm in euthermic Tb that cycled with a period of approximately 8 months. Synchrony in the Tb rhythm within the group was lost after several months of constant L:D conditions; however, individual rhythms in Tb continued to show clear sine wave-like waxing and waning. AGS displayed spontaneous torpor only during troughs in euthermic Tb. To assess sensitivity to A1AR activation, AGS were administered the A1AR agonist N(6)-cyclohexyladenosine (CHA, 0.1 mg/kg, ip), and subcutaneous Tb was monitored. AGS administered CHA during a seasonal minimum in euthermic Tb showed a greater drug-induced decrease in Tb (1.6 ± 0.3 °C) than did AGS administered CHA during a peak in euthermic Tb (0.4 ± 0.3 °C). These results provide evidence for a circannual rhythm in Tb that is associated with increased sensitivity to A1AR signaling and correlates with the onset of torpor.

Season primes the brain in an arctic hibernator to facilitate entrance into torpor mediated by adenosine A(1) receptors.

Torpor in hibernating mammals defines the nadir in mammalian metabolic demand and body temperature that accommodates seasonal periods of reduced energy availability. The mechanism of metabolic suppression during torpor onset is unknown, although the CNS is a key regulator of torpor. Seasonal hibernators, such as the arctic ground squirrel (AGS), display torpor only during the winter, hibernation season. The seasonal character of hibernation thus provides a clue to its regulation. In the present study, we delivered adenosine receptor agonists and antagonists into the lateral ventricle of AGSs at different times of the year while monitoring the rate of O(2) consumption and core body temperature as indicators of torpor. The A(1) antagonist cyclopentyltheophylline reversed spontaneous entrance into torpor. The adenosine A(1) receptor agonist N(6)-cyclohexyladenosine (CHA) induced torpor in six of six AGSs tested during the mid-hibernation season, two of six AGSs tested early in the hibernation season, and none of the six AGSs tested during the summer, off-season. CHA-induced torpor within the hibernation season was specific to A(1)AR activation; the A(3)AR agonist 2-Cl-IB MECA failed to induce torpor, and the A(2a)R antagonist MSX-3 failed to reverse spontaneous onset of torpor. CHA-induced torpor was similar to spontaneous entrance into torpor. These results show that metabolic suppression during torpor onset is regulated within the CNS via A(1)AR activation and requires a seasonal switch in the sensitivity of purinergic signaling.

Altered thermoregulation via sensitization of A1 adenosine receptors in dietary-restricted rats.

RATIONALE: Evidence links longevity to dietary restriction (DR). A decrease in body temperature (T(b)) is thought to contribute to enhanced longevity because lower T(b) reduces oxidative metabolism and oxidative stress. It is as yet unclear how DR decreases T(b). OBJECTIVE: Here, we test the hypothesis that prolonged DR decreases T(b) by sensitizing adenosine A(1) receptors (A(1)AR) and adenosine-induced cooling. METHODS AND RESULTS: Sprague-Dawley rats were dietary restricted using an every-other-day feeding protocol. Rats were fed every other day for 27 days and then administered the A(1)AR agonist, N(6)-cyclohexyladenosine (CHA; 0.5 mg/kg, i.p.). Respiratory rate (RR) and subcutaneous T(b) measured using IPTT-300 transponders were monitored every day and after drug administration. DR animals displayed lower RR on day 20 and lower T(b) on day 22 compared to animals fed ad libitum and displayed a larger response to CHA. In all cases, RR declined before T(b). Contrary to previous reports, a higher dose of CHA (5 mg/kg, i.p.) was lethal in both dietary groups. We next tested the hypothesis that sensitization to the effects of CHA was due to increased surface expression of A(1)AR within the hypothalamus. We report that the abundance of A(1)AR in the membrane fraction increases in hypothalamus, but not cortex of DR rats. CONCLUSION: These results suggest that every-other-day feeding lowers T(b) via sensitization of thermoregulatory effects of endogenous adenosine by increasing surface expression of A(1)AR. DISCUSSION: Evidence that diet can modulate purinergic signaling has implications for the treatment of stroke, brain injury, epilepsy, and aging.

Protein kinase C epsilon activation delays neuronal depolarization during cardiac arrest in the euthermic arctic ground squirrel.

During the pre-hibernation season, arctic ground squirrels (AGS) can tolerate 8 min of asphyxial cardiac arrest (CA) without detectable brain pathology. Better understanding of the mechanisms regulating innate ischemia tolerance in AGS has the potential to facilitate the development of novel prophylactic agents to induce ischemic tolerance in patients at risk of stroke or CA. We hypothesized that neuroprotection in AGS involves robust maintenance of ion homeostasis similar to anoxia-tolerant turtles. Ion homeostasis was assessed by monitoring ischemic depolarization (ID) in cerebral cortex during CA in vivo and during oxygen glucose deprivation in vitro in acutely prepared hippocampal slices. In both models, the onset of ID was significantly delayed in AGS compared with rats. The epsilon protein kinase C (epsilonPKC) is a key mediator of neuroprotection and inhibits both Na+/K+-ATPase and voltage-gated sodium channels, primary mediators of the collapse of ion homeostasis during ischemia. The selective peptide inhibitor of epsilonPKC (epsilonV1-2) shortened the time to ID in brain slices from AGS but not in rats despite evidence that epsilonV1-2 decreased activation of epsilonPKC in brain slices from both rats and AGS. These results support the hypothesis that epsilonPKC activation delays the collapse of ion homeostasis during ischemia in AGS.

Arctic ground squirrel (Spermophilus parryii) hippocampal neurons tolerate prolonged oxygen-glucose deprivation and maintain baseline ERK1/2 and JNK activation despite drastic ATP loss.

Oxygen-glucose deprivation (OGD) initiates a cascade of intracellular responses that culminates in cell death in sensitive species. Neurons from Arctic ground squirrels (AGS), a hibernating species, tolerate OGD in vitro and global ischemia in vivo independent of temperature or torpor. Regulation of energy stores and activation of mitogen-activated protein kinase (MAPK) signaling pathways can regulate neuronal survival. We used acute hippocampal slices to investigate the role of ATP stores and extracellular signal-regulated kinase (ERK)1/2 and Jun NH(2)-terminal kinase (JNK) MAPKs in promoting survival. Acute hippocampal slices from AGS tolerated 30 mins of OGD and showed a small but significant increase in cell death with 2 h OGD at 37 degrees C. This tolerance is independent of hibernation state or season. Neurons from AGS survive OGD despite rapid ATP depletion by 3 mins in interbout euthermic AGS and 10 mins in hibernating AGS. Oxygen-glucose deprivation does not induce JNK activation in AGS and baseline ERK1/2 and JNK activation is maintained even after drastic depletion of ATP. Surprisingly, inhibition of ERK1/2 or JNK during OGD had no effect on survival, whereas inhibition of JNK increased cell death during normoxia. Thus, protective mechanisms promoting tolerance to OGD by AGS are downstream from ATP loss and are independent of hibernation state or season. Journal of Cerebral Blood Flow & Metabolism (2008) 28, 1307-1319; doi:10.1038/jcbfm.2008.20; published online 9 April 2008.

Central nervous system regulation of mammalian hibernation: implications for metabolic suppression and ischemia tolerance.

Torpor during hibernation defines the nadir of mammalian metabolism where whole animal rates of metabolism are decreased to as low as 2% of basal metabolic rate. This capacity to decrease profoundly the metabolic demand of organs and tissues has the potential to translate into novel therapies for the treatment of ischemia associated with stroke, cardiac arrest or trauma where delivery of oxygen and nutrients fails to meet demand. If metabolic demand could be arrested in a regulated way, cell and tissue injury could be attenuated. Metabolic suppression achieved during hibernation is regulated, in part, by the central nervous system through indirect and possibly direct means. In this study, we review recent evidence for mechanisms of central nervous system control of torpor in hibernating rodents including evidence of a permissive, hibernation protein complex, a role for A1 adenosine receptors, mu opiate receptors, glutamate and thyrotropin-releasing hormone. Central sites for regulation of torpor include the hippocampus, hypothalamus and nuclei of the autonomic nervous system. In addition, we discuss evidence that hibernation phenotypes can be translated to non-hibernating species by H(2)S and 3-iodothyronamine with the caveat that the hypothermia, bradycardia, and metabolic suppression induced by these compounds may or may not be identical to mechanisms employed in true hibernation.

Persistent tolerance to oxygen and nutrient deprivation and N-methyl-D-aspartate in cultured hippocampal slices from hibernating Arctic ground squirrel.

Hibernating Arctic ground squirrel (hAGS), Spermophilus parryii, survive profound decreases in cerebral perfusion during torpor and return to normal blood flow during intermittent rewarming periods without neurologic damage. Hibernating AGS tolerate traumatic brain injury in vivo, and acute hippocampal slices from hibernating animals tolerate oxygen and glucose deprivation. It remains unclear, however, if neuroprotection results from intrinsic tissue properties or from differences in response to acute trauma associated with slice preparation. The goal of this work was therefore to determine whether an intrinsic tissue tolerance persists in chronic culture of AGS hippocampal slices at 37 degrees C. A second goal was to address N-methyl-D-aspartate (NMDA) receptor involvement and channel arrest as potential mechanisms of intrinsic tissue tolerance. Baseline neuronal survival and tolerance to oxygen and nutrient deprivation (OND), an in vitro model of ischemia-reperfusion, were assessed in the CA1 region of hippocampal slices from juvenile, hAGS and interbout euthermic AGS (ibeAGS). Early in culture (insult onset at 3 h), slices from both hAGS and ibeAGS tolerate OND (4 h deprivation followed by 20 h recovery) and 500 micromol/L NMDA plus 20 mmol/L KCl. Later in culture (insult onset at 24 h), tolerance persists in slices from hAGS but not in slices from ibeAGS. Ouabain (Na(+)K(+)ATPase inhibitor) administered 24 h in culture enhances survival of slices from hAGS (assessed 24 h later). Thus, tolerance to OND in slices from hAGS is due to intrinsic tissue properties likely involving NMDA receptors and ion channel arrest.

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.

La función del estrés oxidativo en la patogénesis de la enfermedad de Alzheimer / The role of oxidative stress in the pathogenesis of Alzheimer´s disease

La presencia de estrés oxidativo es la característica más temprana de la Enfermedad de Alzheimer (EA), lo cual proporciona un atractivo objetivo para intervenciones terapéuticas. Entre los mayores retos que se presentan actualmente están el establecimiento de la fuente de estrés oxidativo y la determinación de cómo este proceso puede influir en la etiología de la Enfermedad de Alzheimer. Este es un tema complejo, pues varios procesos, enzimáticos y no-enzimáticos, están implicados en la formación de oxígeno reactivo y otras moléculas tóxicas. En este artículo discutimos el progreso en el entendimiento de estos procesos.