Beating oxygen: chronic anoxia exposure reduces mitochondrial F1FO-ATPase activity in turtle (Trachemys scripta) heart

Abstract

SUMMARYThe freshwater turtle Trachemys scripta can survive in the complete absence of O 2 (anoxia) for periods lasting several months. In mammals, anoxia leads to mitochondrial dysfunction, which culminates in cellular necrosis and apoptosis. Despite the obvious clinical benefits of understanding anoxia tolerance, little is known about the effects of chronic oxygen deprivation on the function of turtle mitochondria. In this study, we compared mitochondrial function in hearts of T. scripta exposed to either normoxia or 2weeks of complete anoxia at 5°C and during simulated acute anoxia/reoxygenation. Mitochondrial respiration, electron transport chain activities, enzyme activities, proton conductance and membrane potential were measured in permeabilised cardiac fibres and isolated mitochondria. Two weeks of anoxia exposure at 5°C resulted in an increase in lactate, and decreases in ATP, glycogen, pH and phosphocreatine in the heart. Mitochondrial proton conductance and membrane potential were similar between experimental groups, while aerobic capacity was dramatically reduced. The reduced aerobic capacity was the result of a severe downregulation of the F 1 F O -ATPase (Complex V), which we assessed as a decrease in enzyme activity. Furthermore, in stark contrast to mammalian paradigms, isolated turtle heart mitochondria endured 20min of anoxia followed by reoxygenation without any impact on subsequent ADP-stimulated O 2 consumption (State III respiration) or State IV respiration. Results from this study demonstrate that turtle mitochondria remodel in response to chronic anoxia exposure and a reduction in Complex V activity is a fundamental component of mitochondrial and cellular anoxia survival.

Mitochondrial responses to prolonged anoxia in brain of red-eared slider turtles

“…The mechanism by which CS activity is decreased in anoxia may involve post-translational modifications but further experiments are required to evaluate this possibility. In addition, F 1 F O -ATPase activity decreased by approximately 80% in anoxic samples (figure 1d; t 10 ¼ 0.04, p ¼ 0.97), which is similar to the 85% decrease previously reported in anoxic turtle heart mitochondria [5]. (b) Electron transport chain flux and C m are reduced by chronic anoxia…”

“…In addition, a two-way repeated measures ANOVA revealed a significant treatment effect between normoxia and anoxia on C m (treatment: F 1,14 ¼ 10.88, p , 0.0001; interaction: F 1,14 ¼ 0.92, p ¼ 0.52), matching the treatment effect observed for ETC flux; however, this trend did not reach significance with any individual treatment (electronic supplementary material, figure S1). Overall, the respiratory flux pattern observed in figure 1 is divergent from anoxic turtle heart mitochondria [5]: anoxia-mediated changes in turtle brain result in a more robust downregulation of the ETC as a whole, whereas in turtle heart, reductions in the respiration rates of individual ETC complexes are observed. Conversely, the decrease in F 1 F O -ATPase activity is similar between turtle brain and heart, but reduced relative to skeletal muscle from hypoxia- rsbl.royalsocietypublishing.org Biol.…”

“…This suggests an overall reduction in the aerobic capacity of brain in response to two weeks of anoxia. This change is tissue-specific to turtle brain because CS activity does not change in turtle heart following acclimatization to anoxia [5]. The anoxic decrease in CS activity is likely not indicative of a change in mitochondrial volume density, however, because total CS protein expression was not different between treatments (figure 1b,c; t 10 ¼ 0.78, p ¼ 0.45).…”

“…Lett. 12: 20150797 acclimated frogs [4,5] For example, in vitro, turtle brain mitochondria become depolarized when exposed to acute anoxia due to the activation of mitochondrial ATP-sensitive K þ (mK ATP ) channels [12]. mK ATP channel activation increases mitochondrial membrane permeability to K þ , and the resulting K þ flux through these channels must be opposed by the activity of H þ -fuelled antiporters.…”

“…Not surprisingly, studies of these species have revealed important adaptations at the mitochondrial level that limit the deleterious effects of hypoxia. For example, F 1 F O -ATPase activity is reduced by approximately 95% in skeletal muscle of the anoxia-tolerant frog Rana temporaria [4] and by approximately 85% in the heart of red-eared slider turtles (Trachemys scripta elegans) [5]. This adaptation is thought to prolong cellular viability by limiting ATP consumed by reversed activity of the F 1 F O -ATPase in anoxia.…”