Cardiac arrhythmias induced by glutathione oxidation can be inhibited by preventing mitochondrial depolarization

We have previously proposed that the heterogeneous collapse of mitochondrial inner membrane potential (ΔΨ_m) during ischemia and reperfusion contributes to arrhythmogenesis through the formation of metabolic sinks in the myocardium, wherein clusters of myocytes with uncoupled mitochondria and high K_ATP current levels alter electrical propagation to promote reentry. Single myocyte studies have also shown that cell-wide ΔΨ_m depolarization, through a reactive oxygen species (ROS)-induced ROS release mechanism, can be triggered by global depletion of the antioxidant pool with diamide, a glutathione oxidant. Here we examine whether diamide causes mitochondrial depolarization and promotes arrhythmias in normoxic isolated perfused guinea pig hearts. We also investigate whether stabilization of ΔΨ_m with a ligand of the mitochondrial benzodiazepine receptor (4′-chlorodiazepam; 4-ClDzp) prevents the formation of metabolic sinks and, consequently, precludes arrhythmias. Oxidation of the GSH pool was initiated by treatment with 200 μM diamide for 35 min, followed by washout. This treatment increased GSSG and decreased both total GSH and the GSH/GSSG ratio. All hearts receiving diamide transitioned from sinus rhythm into ventricular tachycardia and/or ventricular fibrillation during the diamide exposure: arrhythmia scores were 5.5 ± 0.5; n = 6 hearts. These arrhythmias and impaired LV function were significantly inhibited by co-administration of 4-ClDzp (64 μM): arrhythmia scores with diamide + 4-ClDzp were 0.4 ± 0.2 (n = 5; P < 0.05 vs. diamide alone). Imaging ΔΨ_m in intact hearts revealed the heterogeneous collapse of ΔΨ_m beginning 20 min into diamide, paralleling the timeframe for the onset of arrhythmias. Loss of ΔΨ_m was prevented by 4-ClDzp treatment, as was the increase in myocardial GSSG. These findings show that oxidative stress induced by oxidation of GSH with diamide can cause electromechanical dysfunction under normoxic conditions. Analogous to ischemia-reperfusion injury, the dysfunction depends on the mitochondrial energy state. Targeting the mitochondrial benzodiazepine receptor can prevent electrical and mechanical dysfunction in both models of oxidative stress.