Glutathione is a cofactor for H₂O₂-mediated stimulation of Ca²⁺- induced Ca²⁺ release in cardiac myocytes

Reactive oxygen species are known to cause attenuation of cardiac muscle contraction. This attenuation is usually preceded by transient augmentation of twitch amplitude as well as cytosolic Ca²⁺. The present study examines the role of an endogenous antioxidant, glutathione in the mechanism of H₂O₂-mediated augmentation of Ca²⁺ release from the sarcoplasmic reticulum. Whole-cell patch-clamped single rat ventricular myocytes were dialyzed with the Cs⁺-rich internal solution containing 200 μM fura-2 and 2 mM glutathione (reduced form). After equilibration of the myocyte with intracellular dialyzing solution, Ca²⁺ current-reduced Ca²⁺ release from the sarcoplasmic reticulum was monitored. Rapid perfusion with H₂O₂ (100 μM or 1 mM) for 20 s inhibited Ca²⁺ current, but enhanced the intracellular Ca²⁺ transients for 3-4 min. Thus, the efficacy of Ca²⁺- induced Ca²⁺ release mechanism was augmented in 71% of myocytes (n = 7). This enhancement ranged between 15- to threefold as the concentrations of H₂O₂ were raised from 100 μM to 1 mM. If glutathione were excluded from the patch pipette or replaced with glutathione disulfide, the enhancement of Ca²⁺-induced Ca²⁺ release was seen in only a minority (20%) of the myocytes H₂O₂ exposure did not increase the basal intracellular Ca²⁺ levels, suggesting that the mechanism of H₂O₂ action was not mediated by inhibition of the sarcoplasmic reticulum Ca²⁺ uptake or activation of passive Ca²⁺ leak pathway H₂O₂-mediated stimulation of Ca²⁺-induced Ca²⁺ release was also observed in myocytes dialyzed with dithiothreitol (0 5 mM). Therefore, reduced thiols support the action of H₂O₂ to enhance the efficacy of Ca²⁺-induced Ca²⁺ release, suggesting that redox reactions might regulate Ca²⁺ channel-gated Ca²⁺ release by the ryanodine receptor.