Regulation of Na+/Ca2+ exchange activity by cytosolic Ca2+ in transfected Chinese hamster ovary cells

Transfected Chinese hamster ovary cells stably expressing the bovine cardiac Na+/Ca2+ exchanger (CK1.4 cells) were used to determine the range of cytosolic Ca2+ concentrations ([Ca2+](i)) that activate Na+/Ca2+ exchange activity. Ba2+ influx was measured in fura 2-loaded, ionomycin-treated cells under conditions in which the intracellular Naf concentration was clamped with gramicidin at similar to 20 mM. [Ca2+](i) was varied by preincubating ionomycin-treated cells with either the acetoxymethyl ester of EGTA or medium containing 0-1 mM added CaCl2. The rate of Ba2+ influx increased in a saturable manner with [Ca2+](i), with the half-maximal activation value of 44 nM and a Hill coefficient of 1.6. When identical experiments were carried out with cells expressing a Ca2+-insensitive mutant of the exchanger, Ba2+ influx did not vary with [Ca2+](i). The concentration for activation of exchange activity was similar to that reported for whole cardiac myocytes but approximately an order of magnitude lower than that reported for excised, giant patches. The reason for the difference in Ca2+ regulation between whole cells and membrane patches is unknown.