Abstract 174: Biophysical Basis of Protein Kinase C Inhibition of the Novel alpha1D Calcium Channel
The recently reported α1D calcium channel in the heart is known to be regulated by protein kinase C (PKC) at the whole cell level and has been implicated in atrial fibrillation. The biophysical basis of this regulation at the single channel level is not known. Therefore, the effect of PKC activation was studied on α1D calcium channel expressed in tsA201 cells using cell-attached method. Unitary currents were recorded in the presence of 70 mM Ba2+ as the charge carrier. Unitary currents were evoked by 500 ms depolarizing pulses from a holding potential of −80 mV every 0.5 Hz. Under basal condition, channel activity was rare and infrequent, however Bay K 8644 (1 μM) induced channel openings with a conductance of 22.3 pS. Single channel analysis of open and closed time distributions were best fitted with a single exponential. PKC activation by PMA (10 nM), a phorbol ester derivative, resulted in a decrease in open probability and increase in closed-time without any significant effect on the conductance of the α1D calcium channel. This is consistent with a decreased entry of α1D Ca channel into open states in the presence of PMA. These data show, for the fist time, 1) the α1D calcium channel activity at the single channel level and 2) the biophysical basis of by which PKC activation inhibits the α1D calcium channel. The shortening of the open-time and the lengthening of the closed-time constants and the increase in blank sweeps may explain the inhibition of the α1D Ca-channel activity and the reduction in whole-cell α1D Ca current previously reported. Altogether, these data are relevant to the understanding of the patho-physiology of α1D calcium channel and its regulation by the autonomics.