Depolarization-dependent binding of the calcium channel antagonist, (+)-[3H]PN200-110, to intact cultured PC12 cells.

Voltage-dependent Ca++ channels of excitable cell membranes are coupled to drug recognition sites that influence Ca++ channel gating behavior. In cardiac tissue, these sites are themselves regulated by membrane potential, which may explain the apparent dependence of drug effects on Ca++ channel conductance state. Whether a similar relationship pertains in other (e.g., neural) cells is unknown. To examine this issue, we investigated the effect of K+-depolarization on binding of the dihydropyridine Ca++ channel antagonist, (+)-[3H]PN200-110, to intact PC12 cells in culture. Specific (nifedipine-sensitive) binding of 50 pM (+)-[3H]PN200-110 to intact PC12 cells was increased approximately 3-fold by K+-depolarization. Binding was also increased when membrane potential was abolished by treatment with digitonin; elevated [K+] had no additional effect under these conditions. Enhancement of binding by K+-depolarization was reversible upon repolarization and resulted from an increase in binding affinity (decrease in KD from 274 to 55 pM in equilibrium saturation experiments and from 625 to 44 pM in kinetic studies), without an increase in binding site number. These findings are in accord with a modulated receptor model of Ca++ channel function in which affinity for dihydropyridine Ca++ channel antagonists is enhanced by depolarization, and provide evidence that this form of Ca++ channel regulation occurs in neural, as well as muscle, cells.