Glutathione oxidation as a trigger of mitochondrial depolarization and oscillation in intact hearts

Depolarization of the mitochondrial inner membrane potential (Delta psi(m).) associated with oxidative stress is thought to be a critical factor in cardiac dysfunction and cell injury following ischemia-reperfusion or exposure to cardiotoxic agents. In isolated cardiomyocytes, mitochondrially-generated reactive oxygen species (ROS) can readily trigger cell-wide collapse or oscillations of Delta psi(m) but it is not known whether these phenomena scale to the level of the whole heart. Here we utilize two-photon laser scanning fluorescence microscopy to track Delta psi(m), ROS, and reduced glutathione (GSH) levels in intact perused guinea-pig hearts subjected to simulated ischemia reperfusion or GSH depletion with the thiol oxidizing agent diamide. Exposure to oxidative stress by either method provoked heterogeneous AT, depolarization and occasional oscillation in clusters of myocytes in the epicardium in association with increased mitochondrial ROS production. Furthermore, the whole-heart oxidative stress dramatically increased the sensitivity of seemingly quiescent cells to Delta psi(m) depolarization induced by a localized laser flash. These effects were directly correlated with depletion of the intracellular GSH pool. Unexpectedly, hearts perfused with nominally Ca2+ -free solution or those switched from 0.5 mM Ca2+ to nominally Ca2+ -free solution also displayed heterogeneous AT. depolarization and oscillation, in parallel with net oxidation of the GSH pool. The findings demonstrate that metabolic heterogeneity initiated by mitochondrial ROS-induced ROS release is present in the intact heart, and that the redox state of the glutathione pool is a key determinant of loss of Delta psi(m). (C) 2008 Elsevier Inc. All rights reserved.