Effects of temperature and cadmium exposure on the mitochondria of oysters (Crassostrea virginica) exposed to hypoxia and subsequent reoxygenation
Authors: Ivanina Anna , Kurochkin Ilya , Leamy Larry , Sokolova Inna 
Published: June 2012

Summary
Marine organisms such as bivalves that live in estuarine and coastal zones are exposed to multiple stressors that include periodic anoxia, temperature fluctuations and pollution, all of which can strongly affect energy metabolism. In this study, we used top-down control and elasticity analyses to determine the interactive effects of these stressors on mitochondria in an intertidal bivalve, the eastern oyster Crassostrea virginica. Oysters were acclimated at 20°C for 30 days in the absence or presence of 50μg L-1 cadmium (Cd) in seawater (control and Cd-exposed oysters, respectively) and then subjected to a long-term anoxia (6 days at &lt;1% O2 in seawater) followed by the normoxic recovery. Respiration, membrane potential (Δψ) and Δψ-dependent kinetics of three major mitochondrial subsystems (substrate oxidation, proton leak and phosphorylation) were determined at 20 and 30°C in mitochondria from oysters exposed to normoxia, 6 days of anoxia and 1 hour of post-anoxic recovery. Mitochondria of control oysters endured prolonged anoxia and subsequent reoxygenation stress without deterioration in their ATP synthesizing capacity or without strong depolarization, reflecting resilience of the metabolic machinery of this intertidal organism. Despite the shut-down of anaerobic metabolism in anoxia, the Δψ-dependent flux capacity of the substrate oxidation and phosphorylation subsystems was significantly elevated in mitochondria from anoxic oysters, likely reflecting an anticipatory response to reoxygenation. During the post-anoxic recovery, oyster mitochondria maintained elevated flux capacity through the substrate oxidation subsystem, while the Δψ-dependent flux through the phosphorylation subsystem returned to the basal levels. Post-anoxic reoxygenation also led to a strong increase in proton conductance of oyster mitochondria. These changes in mitochondrial properties can confer resistance to anoxia-reoxygenation stress by maintaining high aerobic capacity and ATP synthesis rates and alleviating production of reactive oxygen species (ROS). Exposure to environmental stressors such as Cd and elevated temperatures abolished the putative adaptive responses of the substrate oxidation and phosphorylation subsystems and strongly enhanced proton leak in mitochondria of oysters subjected to anoxia/reoxygenation stress. Exposure to Cd or elevated temperatures also resulted in redistribution of control over mitochondrial respiration. This in turn led to an increased degree of control over state 3 (ADP-stimulated) and 4 (resting) respiration conferred by the substrate oxidation system, indicating that substrate oxidation may become a rate-limiting step in mitochondria under the stressful conditions. Our findings suggest that Cd and elevated temperature stress may lead to the loss of mitochondrial resistance to anoxia and reoxygenation and thus potentially affect the ability of oysters to survive periodic oxygen deprivation in the coastal and estuarine habitats.