Calcium transport mechanisms in the cardiac sarcolemma and sarcoplasmic reticulum

CALCIUM TRANSPORT MECHANISMS IN THE CARDIAC SARCOLEMMA AND SARCOPLASMIC RETICULUM. M. Kadoma and M. Tada The First Department of Medicine, Osaka University School of Medicine, Osaka, Japan Ca plays a central role in the excitation-contraction in that most important is Ca coupling of the myocardium, movement across the two membrane systems: sarcolemma (SL) and sarcoplasmic reticulum (SR). A considerable Ca-gradient exists across the SL and SR because of the low sarcoplasmic Ca concentrations. Special Ca transport mechanisms maintain the Ca-gradients across these membrane systems, among which Na-Ca exchange mechanism in SL and Ca2+-ATPase in SR are extensively investigated. The original idea of Na-Ca exchange across SL was first proposed to explain the effect of the levels of extracellular Na+ and Ca2+ on the contractility of the heart. The existence of this mechanism has been shown in the isolated SL vesicles, in which the transmembrane movement of Ca2+ in one direction is coupled to the movement of Na+ in the opposite direction. Employing stopped-flow spectrophotometry and the Ca2+ sensitive dye arsenazo III, we examined the initial time course of Na+-stimulated Ca2+ efflux in SL vesicles. SL vesicles were loaded with Ca2+ in the presence of KC1 then mixed with variable amount of NaCl. The initial velocity of the Na+ stimulated Ca2+effl,lx increased sigmoidally with [Na+] reaching half-saturation at 30 mM with Hill coefficient 2.53, consistent with a stoichiometry of 3 Na+ per Ca2+. The electrogenic nature of Na-Ca exchange mechanism was also examined employing ionophore and less permeant ion. In SR, Ca-ATPase (E; Mr=lOO,OOO) serves as an energy transducer and a translocator of Ca across the membrane. By binding of Ca and ATP, the ATPase forms a phosphoprotein intermediate (EP) and undergoes a conformational change, thus translocating Ca from outside to inside the membrane. Cardiac SR forms another phosphoprotein (Mr=22,000), termed phospholamban (PN). CAMP-dependent phosphorylation resulted in a marked increase in turnover of Ca pump by enhancing the rate of formation and decomposition of EP, indicating that PN could function as a regulator of ATPase. Transient state analyses of EP and Pi indicated that PN phosphorylation enhances the steps at which ATPase greatly alters its affinity for Ca (E2-+El; ElP-+ E2P). These biochemical analyses would provide a more rational basis for the understanding of the transmembrane control of Ca ions in the myocardial cells.