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

It has been reported previously that the beta subunit increases both the ionic current and the gating charge movement of the human cardiac L-type Ca2+ channel alpha1 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 (Ig), and determines the relationship of these components to the activation of the Ca channel ionic current (ICa). Cloned human cardiac L-type alpha1+alpha2+beta3 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 (QON-Vm) were fit by a sum of two Boltzmann components: QON1, which ranged over more negative potentials, and QON2, 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 (QONfast and QONslow) yielded distributions that were similar in their voltage dependence and relative proportion to those measured directly by steady-state integration of QON1 and QON2. Changes in the initial conditions were found to affect QON1 and QON2 differently. The time constants of the ON gating current decays were similar to those of the activation of ICa. 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 (approximately 1/4 of the total charge; QON1 or QONfast) precedes a slower charge movement (approximately 3/4 of the total charge; QON2 or QONslow); and (3) channel opening is associated with the conformational change(s) producing QONslow.