• Mechanisms regulating the circannual rhythm of hibernation

      Frare, Carla; Drew, Kelly L.; Bult-Ito, Abel; Green, Thomas K.; Kuhn, Thomas B. (2019-08)
      Hibernation is a unique adaptation to conserve energy entering a hypometabolic (low metabolic rate) and hypothermic (low body temperature) state called torpor. Torpor is characterized by a drop in metabolism to 1-2% of basal metabolic rate and a decrease in body temperature to one to two degrees above ambient temperature. Metabolic rate is restored to basal metabolic rate and body temperature increases from 2-3⁰C to 36⁰C during the regularly timed arousal. The adenosine A1 receptor agonists promote the onset of hibernation and torpor in different species, through a yet undefined neuronal circuit. In the Arctic ground squirrel, CHA, an adenosine A1 receptor agonist, induces hibernation during the winter- hibernation season but not in summer even when the environmental conditions are kept constant (ambient temperature of 2⁰C and a light cycle of 4L:20D). Thus, the phenomenon of CHA-induced hibernation is entrained to an endogenous circannual rhythm. In this work, I aim to identify the changes in neuronal activation that reflect the circannual rhythm regulating the seasonal difference in response to CHA. Arctic ground squirrels, housed at constant ambient temperature (2°C) and light cycle (4L:20D), were implanted with body temperature transmitters. I collected tissue during Summer, Fall, Winter and Torpor conditions for seasonal analysis. For treatment analysis, I collected tissue form animals treated with CHA or vehicle in Summer and Winter. Primarily, I used immunohistochemistry to identify cell groups affected by season and treatment. I used cFos to identify neuronal activity and other immunohistochemical markers to identify neuronal phenotypes, based on specific cytoplasmic proteins. An overall seasonal decrease in thermogenesis, measured as reduced neuronal activity in the thermoregulatory pathways, and increase in vasoconstriction reflected the higher order processing necessary for CHA-induced hibernation. CHA inhibited the histaminergic neurons in the hypothalamus suppressing wakefulness and dis-inhibited the nucleus tractus solitarius, further suppressing thermogenesis. Preliminary data also suggested a seasonal change in the adenosine metabolic pathway, which may have increased adenosine receptor sensitivity during the hibernation season. Our results suggest that histaminergic neurons in the hypothalamus and the nucleus tractus solitarius are likely targets to manipulate metabolic demand in the clinical setting inducing therapeutic hypothermia or increasing metabolic rate.
    • Modulation of ischemia- reperfusion injury in mammalian hibernators and non-hibernators: a comparative study

      Bhowmick, Saurav; Drew, Kelly L.; Kuhn, Thomas B.; Duffy, Lawrence K.; Oliver, Scott R. (2017-12)
      Events characterized by ischemia/reperfusion (I/R), such as stroke and cardiac arrest, are among the most frequent causes of debilitating neurological injury and death worldwide. During ischemia, the brain experiences oxygen and nutrition deprivation due to lack of blood flow, and tissue damage ensues. Arctic ground squirrel (AGS; Urocitellus parryii), a hibernating species has the innate ability to survive profound decreases in blood flow (ischemia) during torpor and return of blood flow (reperfusion) during intermittent euthermic periods without any neurological deficit. However, the mechanisms by which AGS tolerate the extreme fluctuations in blood flow remain unclear. The main focus of this thesis is to investigate the modulation of I/R injury in mammalian hibernators and non-hibernators. The first study validates the microperfusion approach for studying in vitro I/R injury (oxygen glucose deprivation, OGD) modeled in acute hippocampal slices and investigates the complex interactions of glutamate-mediated excitotoxicity with acidosis-mediated acidotoxicity to understand the role of acid-sensing ion channels (ASIC1a) and pH in mediating cellular injury during OGD. Using an ischemic tolerant animal model, AGS, the second and third studies explore if hibernation season or state influences tolerance to I/R injury and tests hypotheses regarding mechanisms involving nitric oxide and superoxide radicals in mediating cellular damage during cerebral I/R. Together, this dissertation demonstrates that when OGD is combined with acidosis as occurs in vivo, acidotoxicity mediated via ASIC1a occurs but low pH abolishes NMDAR mediated excitotoxicity. This dissertation also presents evidence that AGS tolerate OGD injury independent of hibernation season and state. At the tissue level, when tissue temperature is normalized to 36°C despite ATP depletion, ionic derangement, tissue acidosis, and excitatory neurotransmitter efflux, AGS hippocampus resists OGD injury. Finally, the dissertation shows that AGS resist brain injury caused by ONOO- generated from NO or O2•− during OGD while rat brain tissue succumbs to this mechanism of injury.