Cardiac ischemia oxidizes regulatory thiols on ryanodine receptors: Captopril acts as a reducing agent to improve Ca2+ uptake by ischemic sarcoplasmic reticulum

We tested the hypothesis that ischemia alters sarcoplasmic reticulum (SR) C a2+ transport by oxidizing regulatory thiols on ryanodine receptors (RyRs), and that membrane-permeable sulfhydryl-containing angiotensin-converting e nzyme (ACE) inhibitors protect against ischemia-induced oxidation and expla in in part, the therapeutic actions of captopril. Ca2+ uptake and adenosine triphosphatase (ATPase) activity was measured from SR vesicles isolated fr om control or ischemic dog and human ventricles and compared with or withou t sulfhydryl reductants. The rate and amount of Ca2+ uptake was lower for c anine ischemic SR compared with control (6.5 +/- 0.2 --> 18.5 +/- 1.1 nmol Ca2+/mg/min and 123.1 +/- 4.7 --> 235.0 +/- 17.3 nmol Ca2+/mg; n = g each). Captopril, dithiothreitol (DTT), glutathione (GSH), and L-cysteine increas ed the rate and amount of Ca2+ uptake by canine and human ischemic SR vesic les by similar to 50%. Reducing agents had no effect on Ca2+-ATPase activit y in either canine control or ischemic (similar to 40% less than control) S R. Captopril was as potent as DTT at reversing the oxidation of skeletal an d cardiac RyRs induced by reactive disulfides (RDSs) or nitric oxide (NO). In neonatal rat myocytes, RDSs or NO triggered SR Ca2+ release and increase d cytosolic Ca2+, an effect reversed by captopril and DTT but not GSH or cy steine. Pretreatment of myocytes with captopril (exposure and then wash) in hibited Ca2+ elevation elicited by RDSs or NO, indicating that captopril is an effective, membrane-permeable intracellular reducing agent. Thus, net S R Ca2+ accumulation is reduced by ischemia in part due to the oxidation of thiols that gate RyRs, an effect reversed by captopril.