Na+/Ca2+ exchange of isolated sarcolemmal membrane: effects of insulin, oxidants and insulin deficiency

Abstract

In this study we prepared sarcolemmal fractions from bovine and rat hearts; their Na+K+ ATPase activities, measured in the presence of saponin to unmask latent Na+K+ ATPase, were 59.4 and 48.8 mu mol Pi/mg protein.h, respectively. The rate of Na+ dependent Ca2+ uptake was linear for the first 10 s and a plateau was reached in 3 min. Oxidation by free radical generation either with H2O2, FeSO4 plus DTT or xanthine oxidase plus hypoxanthine stimulated Na+/Ca2+ exchange in a time-dependent manner. The stimulation was abolished by deferoxamine or o-phenanthroline. By contrast, oxidation by HOCl inhibited Na+/Ca2+ exchange in proportion to its concentration, and this inhibition was antagonized by DTT. DTT alone had no effect on the exchange. Insulin stimulated Na+/Ca2+ exchange, its maximal effect was attained after 30 min incubation with 100 mu units/ml. N-ethylmaleimide inhibited the exchange both in the presence and in the absence of insulin. Sarcolemmal fractions prepared from hearts of alloxan-treated, acutely diabetic rats showed a significant decrease in Na+/Ca2+ exchange. Addition of insulin in vitro significantly stimulated Na+/Ca2+ exchange of both diabetic and control groups. The results indicate that sarcolemmal Na+/Ca2+ exchange function is modulated by oxidation-reduction states and by the presence of insulin.