title: Glutathione is a cofactor for H2O2-mediated stimulation of Ca2+-induced Ca2+ release in cardiac myocytes.
abstract: 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 microM fura-2 and 2 mM glutathione (reduced form). After equilibration of the myocyte with intracellular dialyzing solution, Ca2+ current-induced Ca2+ release from the sarcoplasmic reticulum was monitored. Rapid perfusion with H2O2 (100 microM 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 1.5- to threefold as the concentrations of H2O2 were raised from 100 microM 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.
authors.affiliation: Department of Pharmacology, Georgetown University Medical Center, Washington, DC 20007, USA.
authors.forename: Y J
authors.initials: YJ
authors.lastname: Suzuki
authors.forename: L
authors.initials: L
authors.lastname: Cleemann
authors.forename: D R
authors.initials: DR
authors.lastname: Abernethy
authors.forename: M
authors.initials: M
authors.lastname: Morad
mesh_terms: D000818:Animals; D002118:Calcium / Q000378:metabolism* / Q000494:pharmacology*; D015220:Calcium Channels / Q000502:physiology; D004229:Dithiothreitol / Q000494:pharmacology; D004553:Electric Conductivity; D005978:Glutathione / Q000494:pharmacology*; D019803:Glutathione Disulfide / Q000494:pharmacology; D006861:Hydrogen Peroxide / Q000494:pharmacology*; D066298:In Vitro Techniques; D008297:Male; D009206:Myocardium / Q000166:cytology / Q000378:metabolism*; D010084:Oxidation-Reduction; D018408:Patch-Clamp Techniques; D051381:Rats; D017208:Rats, Wistar; D012519:Sarcoplasmic Reticulum / Q000187:drug effects / Q000378:metabolism; D013438:Sulfhydryl Compounds / Q000494:pharmacology
chemical_list: D015220:Calcium Channels; D013438:Sulfhydryl Compounds; D006861:Hydrogen Peroxide; D005978:Glutathione; D002118:Calcium; D004229:Dithiothreitol; D019803:Glutathione Disulfide