Regulation and deregulation of cardiac Na⁺-Ca²⁺ exchange in giant excised sarcolemmal membrane patches

A plasmalemmal Na⁺-Ca²⁺ exchange mechanism^1,2 is an important electrogenic determinant of contractility in cardiac cells^3-5. As in other cell types^6-8, calcium influx by Na⁺-Ca²⁺" exchange is secondarily activated by cytoplasmic calcium⁴ and probably ATP⁹, but these modulatory mechanisms are either absent or altered in isolated cardiac sarcolemmal vesicles^5,12. Involvement of a calcium-dependent protein kinase in exchange regulation has been suggested^7,10 but not verified^5,11. Here I describe measurements of outward Na⁺-Ca²⁺ exchange current, corresponding to calcium influx, in giant excised sarcolemmal Patches¹¹ from guinea pig myocytes. The exchange current is stimulated by both calcium and Mg-ATP from the cytoplasmic face, evidently through separate mechanisms. Activation by cytoplasmic calcium takes place within seconds, is reversible, and does not require ATP. Stimulation by Mg-ATP reverses only slowly over > 10 min, or not at all. Unexpectedly, a substantial decrease in exchange current occurs during activation by cytoplasmic sodium, which seems to reflect an inactivation process rather than ion concentration changes or a 'first pass' exchange cycle. This apparent inactivation, and the modulations by cytoplasmic calcium and Mg-ATP, are all abolished by brief treatment of the cytoplasmic surface with chymotrypsin, leaving the exchanger in a maintained state of high activity. Therefore, limited proteolysis deregulates Na⁺-Ca²⁺ exchange and could contribute to the loss of secondary regulation of the exchange in isolated sarcolemmal vesicles.