Voltage and Ca 2+ Dependence of Pre–Steady-State Currents of the Na-Ca Exchanger Generated by Ca 2+ Concentration Jumps

Abstract

The Ca 2+ concentration and voltage dependence of the relaxation kinetics of the Na-Ca exchanger after a Ca 2+ concentration jump was measured in excised giant membrane patches from guinea pig heart. Ca 2+ concentration jumps on the cytoplasmic side were achieved by laser flash-induced photolysis of DM-nitrophen. In the Ca-Ca exchange mode a transient inward current is generated. The amplitude and the decay rate of the current saturate at concentrations >10 μM. The integrated current signal, i.e., the charge moved is fairly independent of the amount of Ca 2+ released. The amount of charge translocated increases at negative membrane potentials, whereas the decay rate constant shows no voltage dependence. It is suggested that Ca 2+ translocation occurs in at least four steps: intra- and extracellular Ca 2+ binding and two intramolecular transport steps. Saturation of the amplitude and of the relaxation of the currrent can be explained if the charge translocating reaction step is preceded by two nonelectrogenic steps: Ca 2+ binding and one conformational transition. Charge translocation in this mode is assigned to one additional conformational change which determines the equilibrium distribution of states. In the Na-Ca exchange mode, the stationary inward current depends on the cytoplasmic Ca 2+ concentration and voltage. The K m for Ca 2+ is 4 μM for guinea pig and 10 μM for rat myocytes. The amplitude of the pre–steady-state current and its relaxation saturate with increasing Ca 2+ concentrations. In this mode the relaxation is voltage dependent.