Stoichiometry of Na+-Ca2+ exchange in inside-out patches excised from guinea-pig ventricular myocytes

The stoichiometry (n(x)) of cardiac Na+-Ca2+ exchange was examined by measuring the reversal potential of the Na+-C2+ exchange current (INa-Ca) in large inside-out patches, 'macro patches', excised from intact guinea-pig ventricular cells. Cytoplasmic application of Na+ (Na-i(+)) or Ca2+(Ca-i(2+)) induced INa-Ca which showed properties similar to INa-Ca in the giant membrane patch. The outward INa-Ca was depressed by an exchanger inhibitory peptide, XIP. The reversal potential of the XIP-sensitive current indicated that n(x) was similar to 4 (3.6-4.2) at 9-40 mM Na-i(+), and n(x) tended to increase as Na-i(+) was increased. Proteolysis by trypsin did not significantly affect the stoichiometry Similar results were obtained from the reversal potential of INa-Ca that was induced by application of both Na-i(+) and Ca-i(2+). At 0.1 mu M Ca-i(2+), N-x was similar to 4 (3.7-4.4) at 6-25 mM Na-i(+) and tended to increase as Na-i(+) was increased. When Ca-i(2+) was changed from 0.1 to 1 and 1000 mu M at constant 50 mM Na-i(+), the value was similar to 4 (3.6-4.4). 5. When the extracellular Na+ (Na-o(2+)) and Ca2+ (Ca-o(2+)) concentrations were varied in the presence of 25 or 9 mM Na-i(+) and 1 mu M Ca-i(2+), n(x) was almost constant (similar to 4) over the range 0.3-20 mM Ca-o(2+) and 10-145 mM Na-o(+). These results indicated that the stoichiometry of Na+-Ca2+ exchange is different from generally accepted 3Na(+):1Ca(2+) and suggested that the stoichiometry is either 4Na(+):1Ca(2+) or variable depending on Na-i(+) and Ca-i(2+).