Pre-steady-state kinetics of Ba-Ca exchange reveals a second electrogenic step involved in Ca2+ translocation by the Na-Ca exchanger.

Kinetic properties of the Na-Ca exchanger (guinea pig NCX1) expressed in Xenopus oocytes were investigated with excised membrane patches in the inside-out configuration and photolytic Ca(2+) concentration jumps with either 5 mM extracellular Sr(2+) or Ba(2+). After a Ca(2+) concentration jump on the cytoplasmic side, the exchanger performed Sr-Ca or Ba-Ca exchange. In the Sr-Ca mode, currents are transient and decay in a monoexponential manner similar to that of currents in the Ca-Ca exchange mode described before. Currents recorded in the Ba-Ca mode are also transient, but the decay is biphasic. In the Sr-Ca mode the amount of charge translocated increases at negative potentials in agreement with experiments performed in the Ca-Ca mode. In the Ba-Ca mode the total amount of charge translocated after a Ca(2+) concentration jump is approximately 4 to 5 times that in Ca-Ca or Sr-Ca mode. In the Ba-Ca mode the voltage dependence of charge translocation depends on the Ca(2+) concentration on the cytosolic side before the Ca(2+) concentration jump. At low initial Ca(2+) levels (approximately 0.5 microM), charge translocation is voltage independent. At a higher initial concentration (1 microM Ca(2+)), the amount of charge translocated increases at positive potentials. Biphasic relaxation of the current was also observed in the Ca-Ca mode if the external Ca(2+) concentration was reduced to < or =0.5 mM. The results reported here and in previous publications can be described by using a 6-state model with two voltage-dependent conformational transitions.