Measuring Arctic ground squirrel adult neurogenesis in caudal hippocampus using DAB/DAB-Ni enzyme-based immunohistochemistry

Abstract

Unconscious patients who are resuscitated after cardiac arrest often have a poor prognosis for neurological recovery and are routinely treated with therapeutic hypothermia as a standard of care. This intervention aims to reduce the risk of death and minimize brain damage. However, results from the Targeted Temperature Management 2 (TTM2) trial in 2021 indicate that cooling does not improve neurological outcomes in these patients. This finding raises questions about the neurological benefits, particularly regarding neurogenesis, of the rewarming phase of therapeutic hypothermia, as well as the effects of repeated cooling and rewarming cycles. Neurogenesis has been largely overlooked in therapeutic hypothermia research, leaving significant gaps in our understanding. Notably, hibernation and therapeutic hypothermia share physiological similarities, but hibernation is a more complex process that confers neurological protection against cardiac arrest in the Arctic ground squirrel (AGS), an extreme hibernator. Specifically, during hibernation, AGS repeatedly lower body temperature to as low as 1 to -3°C, along with reduced blood flow and metabolism during torpor, then periodically rewarm to normal levels during brief interbout arousals. These repeated cooling and rewarming cycles happened at least eight times during hibernation. Building on this knowledge, this thesis explores the possibility that cooling and rewarming could activate neurogenesis and thereby improve the benefits of therapeutic hypothermia by replacing dead and damaged neurons. As proof of concept, I developed a method for measuring hippocampal neurogenesis in the AGS across three seasonal states: summer active, hibernation torpor, and interbout arousal.