Pre-Steady-State Kinetics of Ba-Ca Exchange Reveals a Second Electrogenic Step Involved in Ca2+ Translocation by the Na-Ca Exchanger

AbstractKinetic 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 Ca2+ concentration jumps with either 5 mM extracellular Sr2+ or Ba2+. After a Ca2+ 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 Ca2+ concentration jump is ∼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 Ca2+ concentration on the cytosolic side before the Ca2+ concentration jump. At low initial Ca2+ levels (∼0.5 μM), charge translocation is voltage independent. At a higher initial concentration (1 μM Ca2+), 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 Ca2+ concentration was reduced to ≤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