Effect of hydrogen peroxide on reoxygenation-induced Ca2+ accumulation in rat cardiomyocytes

Reactive oxygen species (ROS) contribute to cell damage during reperfusion of the heart. ROS may exert their effects partly by interfering with Ca2+ homeostasis of the myocardium. The purpose of this study was to investigate the effects of hydrogen peroxide (H2O2) on Ca2+ accumulation during reoxygenation of isolated adult rat cardiomyocytes exposed to 1 h of hypoxia and to relate the effects to possible changes in release of lactate dehydrogenase (LDH), free intracellular Ca2+ ([Ca2+]i) and Mg2+([Mg2+]i), and mitochondrial membrane potential (Δψm). Cell Ca2+ was determined by 45Ca2+ uptake. Free [Mg2+]i and [Ca2+]i and Δψm were measured by flow cytometry. Reoxygenation-induced Ca2+ accumulation was attenuated by 23 and 34% by 10 and 25 μM H2O2, respectively, added at reoxygenation. H2O2 at 100 and 250 μM increased cell Ca2+ by 50 and 83%, respectively, whereas 500 μM H2O2 decreased cell Ca2+ by 20%. H2O2 at (25 μM) reduced LDH release and [Mg2+]i and increased Δψm, indicating cell protection, whereas 250 μM H2O2 increased LDH release and [Mg2+]i and decreased Δψm, indicating cell damage. Clonazepam (100 μM) attenuated the increase in Ca2+ accumulation, the elevation of [Ca2+]i, and the decrease in Δψm induced by 100 and 250 μM H2O2 during reoxygenation. We report for the first time that 25 μM H2O2 attenuates Ca2+ accumulation, LDH release, and dissipation of Δψm during reoxygenation of hypoxic cardiomyocytes, indicating cell protection.