Kinetic components of the gating currents of human cardiac L-type Ca²⁺ channels

Abstract

 It has been reported previously that the β subunit increases both the ionic current and the gating charge movement of the human cardiac L-type Ca²⁺ channel α₁ subunit, and that steady-state measurements reveal the presence of two distinct components of the charge movement [Josephson IR, Varadi G (1996) Biophys J 70:1285–1293]. The present work identifies and characterizes the kinetic properties of the components of the human cardiac L-type Ca channel gating currents (I _g), and determines the relationship of these components to the activation of the Ca channel ionic current (I _Ca). Cloned human cardiac L-type α₁+α₂+β₃ subunits were transiently expressed in HEK293 cells and calcium channel gating currents were recorded following the addition of 5 mM Co²⁺. The steady-state charge integrals of the gating currents (Q _ON-V _m) were fit by a sum of two Boltzmann components: Q _ON1, which ranged over more negative potentials, and Q _ON2, which ranged over more positive potentials. The kinetic components of the ON and OFF gating currents were identified using bi-exponential curve fitting. Reconstruction of the two kinetic components of charge (Q _ONfast and Q _ONslow) yielded distributions that were similar in their voltage dependence and relative proportion to those measured directly by steady-state integration of Q _ON1 and Q _ON2. Changes in the initial conditions were found to affect Q _ON1 and Q _ON2 differently. The time constants of the ON gating current decays were similar to those of the activation of I _Ca. The results suggest that: (1) the activation of the human cardiac L-type Ca channel involves the movements of at least two, functionally distinct gating structures; (2) a fast charge movement (≈1/4 of the total charge; Q _ON1 or Q _ONfast) precedes a slower charge movement (≈3/4 of the total charge; Q _ON2 or Q _ONslow); and (3) channel opening is associated with the conformational change(s) producing Q _ONslow.