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This study was undertaken to examine the effects of hydrogen peroxide on stimulatory guanine nucleotide-binding protein (Gs), and coupling in the beta-adrenergic receptor-Gs-adenylate cyclase system in rat heart, in vitro. Cardiac membranes were preincubated with various concentrations (0.1, 1, and 10 mM) of hydrogen peroxide at 30 degrees C for 5, 10, 30 and 60 min. Although the assay of beta-adrenergic receptors involving [3H]-dihydroalprenolol ([3H]-DHA) binding revealed that the maximal number of binding sites (Bmax) was not altered, the dissociation constant (Kd) for [3H]-DHA was increased in the presence of 1 mM and 10 mM hydrogen peroxide (control 0.68 +/- 0.16 nM, vs 1 mM H2O2 1.13 +/- 0.16, 10 mM H2O2 1.01 +/- 0.12). Conversely, no significant changes in Gs activities were observed in hydrogen peroxide-treated groups. Adenylate cyclase activity (stimulated by forskolin) was significantly reduced by 10 mM hydrogen peroxide after a 5 min preincubation period (control 277.1 +/- 19.2 pmol cAMP/mg protein/min, H2O2 230.3 +/- 14.9). The amounts of cyclic AMP produced by the stimulation of membranes with GTP, GTP+(l)-isoproterenol, guanylimidodiphosphate (Gpp(NH)p) or Gpp(NH)p+(l)-isoproterenol were significantly lower in 10 mM hydrogen peroxide-treated groups than those in controls (GTP: control 57.6 +/- 5.6 pmol cAMP/mg protein/min vs H2O2 46.4 +/- 6.9, GTP+(l)-isoproterenol: control 83.9 +/- 10.2 vs H2O2 67.7 +/- 10.3, Gpp(NH)p: control 77.5 +/- 8.8 vs H2O2 61.0 +/- 8.6, Gpp(NH)p+(l)-isoproterenol: control 105.0 +/- 13.1 vs H2O2 83.9 +/- 12.2, forskolin: control 223.2 +/- 13.8 vs H2O2 182.8 +/- 18.4).(ABSTRACT TRUNCATED AT 250 WORDS)
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