2 FUNCTIONAL NA+/K+-ATPASE ISOFORMS IN THE LEFT-VENTRICLE OF GUINEA-PIG HEART

Guinea pig left ventricular muscle contains two distinct molecular forms of the Na+/K+-ATPase catalytic alpha-subunit. Sarcolemmal vesicles highly enriched in Na+/K+-ATPase were isolated by a new procedure that yielded specific activities of 60-100-mu-mol P(i).h-1.mg-1. SDS/PAGE of isolated sarcolemma after reduction and alkylation of the sulfhydryl groups and identification on immunoblots with specific anti-(alpha-subunit) antibodies indicated the presence of two major polypeptides of 100 kDa and 103 kDa, respectively. The two alpha-subunits were functional: the dose/response curves of Na+/K+-ATPase activity with ouabain, dihydroouabain and digitoxigenin were biphasic, revealing the presence of high-affinity [concentration of drug causing 50% inhibition (IC50) = 10 nM] and low-affinity (IC50 = 2-mu-M) forms with proportional contributions of 55% and 45%, respectively. The involvement of the high-affinity form in the positive inotropic effect of digitalis and of the low-affinity sites in both inotropy and toxicity are consistent with the literature data on rodents.