Modulation of mitochondrial site-specific hydrogen peroxide efflux by exogenous stressors

Abstract

Oxygen (O₂) deprivation and metals are common environmental stressors and their exposure to aquatic organisms can induce oxidative stress by disrupting cellular reactive oxygen species (ROS) homeostasis. Mitochondria are a major source of ROS in the cell wherein a dozen sites located on enzymes of the electron transport system (ETS) and substrate oxidation produce superoxide anion radicals (O₂˙‾) or hydrogen peroxide (H₂O₂). Sites located on ETS enzymes can generate ROS by forward electron transfer (FET) and reverse electron transfer (RET) reactions; however, knowledge of how exogenous stressors modulate site-specific ROS production is limited. We investigated the effects of anoxia-reoxygenation and cadmium (Cd) on H₂O₂ emission in fish liver mitochondria oxidizing glutamate-malate, succinate or palmitoylcarnitine-malate. We find that anoxia-reoxygenation attenuates H₂O₂ emission while the effect of Cd depends on the substrate, with monotonic responses for glutamate-malate and palmitoylcarnitine-malate, and a biphasic response for succinate. Anoxia-reoxygenation exerts a substrate-dependent inhibition of mitochondrial respiration which is more severe with palmitoylcarnitine-malate compared with succinate or glutamate-malate. Additionally, specific mitochondrial ROS-emitting sites were sequestered using blockers of electron transfer and the effects of anoxia-reoxygenation and Cd on H₂O₂ emission were evaluated. Here, we find that site-specific H₂O₂ emission capacities depend on the substrate and the direction of electron flow. Moreover, anoxia-reoxygenation alters site-specific H₂O₂ emission rates during succinate and glutamate-malate oxidation whereas Cd imposes monotonic or biphasic H₂O₂ emission responses depending on the substrate and site. Contrary to our expectation, anoxia-reoxygenation blunts the effect of Cd. These results suggest that the effect of exogenous stressors on mitochondrial oxidant production is governed by their impact on energy conversion reactions and mitochondrial redox poise. Moreover, direct increased ROS production seemingly does not explain the increased adverse effects associated with combined exposure of aquatic organisms to Cd and low dissolved oxygen levels.

Keywords: Anoxia-reoxygenation; Cadmium; Mitochondria; Reactive oxygen species; Respiration.