Anoxia-induced changes in reactive oxygen species and cyclic nucleotides in the painted turtle

Abstract

The Western painted turtle survives months without oxygen. A key adaptation is a coordinated reduction of cellular ATP production and utilization that may be signaled by changes in the concentrations of reactive oxygen species (ROS) and cyclic nucleotides (cAMP and cGMP). Little is known about the involvement of cyclic nucleotides in the turtle's metabolic arrest and ROS have not been previously measured in any facultative anaerobes. The present study was designed to measure changes in these second messengers in the anoxic turtle. ROS were measured in isolated turtle brain sheets during a 40-min normoxic to anoxic transition. Changes in cAMP and cGMP were determined in turtle brain, pectoralis muscle, heart and liver throughout 4 h of forced submergence at 20-22 degrees C. Turtle brain ROS production decreased 25% within 10 min of cyanide or N(2)-induced anoxia and returned to control levels upon reoxygenation. Inhibition of electron transfer from ubiquinol to complex III caused a smaller decrease in [ROS]. Conversely, inhibition of complex I increased [ROS] 15% above controls. In brain [cAMP] decreased 63%. In liver [cAMP] doubled after 2 h of anoxia before returning to control levels with prolonged anoxia. Conversely, skeletal muscle and heart [cAMP] remained unchanged; however, skeletal muscle [cGMP] became elevated sixfold after 4 h of submergence. In liver and heart [cGMP] rose 41 and 127%, respectively, after 2 h of anoxia. Brain [cGMP] did not change significantly during 4 h of submergence. We conclude that turtle brain ROS production occurs primarily between mitochondrial complexes I and III and decreases during anoxia. Also, cyclic nucleotide concentrations change in a manner suggestive of a role in metabolic suppression in the brain and a role in increasing liver glycogenolysis.

No oxygen? No problem! Intrinsic brain tolerance to hypoxia in vertebrates

“…Unlike the mammalian brain, which shows an overproduction of reactive oxygen species (ROS) following hypoxia or ischemia/reperfusion (Hashimoto et al, 2003), the turtle brain appears to suppress ROS production upon re-oxygenation (Milton et al, 2007;Pamenter et al, 2007). As with other protective mechanisms, adenosine also impacts the production of ROS upon re-oxygenation (Fig.…”

δ-Opioid receptor antagonism induces NMDA receptor-dependent excitotoxicity in anoxic turtle cortex

“…There is some indirect evidence to support a role for G protein-mediated responses in the turtle's anoxia tolerance. Whole-brain cAMP concentration decreases significantly in the anoxic cortex, and since cAMP is directly mediated by G i activity these data suggest anoxia-induced changes in G i signaling occur in turtle brain (Pamenter et al, 2007). Furthermore, we have recently shown that the anoxic depression in turtle NMDAR activity is blocked by pertussis toxin, a G i inhibitor (Pamenter et al, 2008b).…”

Scavenging ROS dramatically increases NMDA receptor whole cell currents in painted turtle cortical neurons

“…The effect of changing [ROS] on NMDA/AMPA receptor activity or [Ca 2+ ] i in turtle cortical pyramidal neurons has not been explored, although studies in other vertebrate species demonstrate that NMDA receptor activity is significantly increased by a decrease in ROS levels (Aizenman et al, 1989;Bodhinathan et al, 2010;Choi and Lipton, 2000). However, because ROS levels naturally decrease in the anoxic turtle brain, we propose that this will trigger an increase in [Ca 2+ ] i and a decrease in NMDA and AMPA receptor whole-cell currents (Pamenter et al, 2007). The aims of this study were to determine: (1) whether decreasing [ROS] i decrease whole-cell evoked NMDA and AMPA receptor currents in turtle pyramidal neurons, (2) whether mitochondrial Ca 2+ release is induced by ROS scavenging, and (3) using a complex I inhibitor, whether these responses are dependent on mitochondrial ROS production.…”

“…Furthermore, diazoxide application to turtle cortical neurons results in a depolarization of Ψ m , release of mitochondrial [Ca 2+ ], and a decrease in NMDA/AMPA receptor currents to levels comparable to those seen during anoxia (Hawrysh and Buck, 2013;Pamenter et al, 2008a;Zivkovic and Buck, 2010). Concentrations of rotenone and diazoxide were based on previous experiments on turtle cortical tissue (Pamenter et al, 2007). Diazoxide was initially solubilized in DMSO (1% in final solution) and rotenone was solubilized in chloroform before being diluted further in aCSF (0.05% in final solution).…”