Temperature rise and copper exposure reduce heart mitochondrial reactive oxygen species scavenging capacity

Mitochondria produce and scavenge reactive oxygen species (ROS); however, whether oxidative distress due to exogenous stress arises from excessive production or impaired scavenging remains unclear. We assessed the effect of copper (Cu) and thermal stress on kinetics of ROS (H₂O₂) consumption in mitochondria isolated from fish heart. Mitochondria were energized with succinate, glutamate-malate or palmitoylcarnitine (PC) and incubated with 1-25 μM Cu at 11 (control) and 23 °C. We found that H₂O₂ consumption capacity of heart mitochondria varies with substrate and is additively reduced by temperature rise and Cu. While Cu is a potent inhibitor of H₂O₂ consumption in mitochondria oxidizing glutamate-malate and succinate, mitochondria oxidizing PC are resistant to the inhibitory effect of the metal. Moreover, the sensitivity of H₂O₂ consumption pathways to Cu depend on the substrate and are greatly impaired during oxidation of glutamate-malate. Pharmacological manipulation of mitochondrial antioxidant systems revealed that NADPH-dependent peroxidase systems are the centerpieces of ROS scavenging in heart mitochondria, with the glutathione-dependent pathway being the most prominent while catalase played a minimal role. Surprisingly, Cu is as efficacious in inhibiting thioredoxin-dependent peroxidase pathway as auranofin, a selective inhibitor of thioredoxin reductase. Taken together, our study uncovered unique mechanisms by which Cu alters mitochondrial H₂O₂ homeostasis including its ability to inhibit specific mitochondrial ROS scavenging pathways on a par with conventional inhibitors. Importantly, because of additive inhibitory effect on mitochondrial ROS removal mechanisms, hearts of organisms jointly exposed to Cu and thermal stress are likely at increased risk of oxidative distress.