Identification of ω‐Conotoxin Binding Sites on Adrenal Medullary Membranes: Possibility of Multiple Calcium Channels in Chromaffin Cells

Binding of 125 I‐ω‐conotoxin GVIA and [ 3 H]nitrendipine to membranes from bovine adrenal medulla was investigated to test for the presence of N‐ and L‐type Ca 2+ channels in adrenal chromaffin cells. Saturable, high‐affinity binding sites for 125 I‐ω‐conotoxin and [ 3 H]nitrendipine were detected in a membrane fraction from adrenal medulla. [ 3 H]Nitrendipine binding sites were found to have a K D of 500 ± 170 p M and a B max of 26 ± 11 pmol/g of protein. 125 I‐ω‐Conotoxin binding sites had a K D of 215 ± 56 p M and a B max of 105 ± 18 pmol/g of protein, about four times the number of sites found for [ 3 H]nitrendipine. 125 I‐ω‐Conotoxin binding was potently inhibited by unlabeled toxin and Ca 2+ but was unaffected by dihydropyridines, verapamil, and diltiazem. [ 3 H]Nitrendipine binding was not affected by ω‐conotoxin, whereas it was inhibited by other dihydropyridines. Bay K 8644 potentiated K + ‐evoked cytosolic Ca 2+ transients measured by fura‐2 fluorescence, and this potentiation was completely blocked by nifedipine. In contrast, ω‐conotoxin had no effect on Bay K 8644‐evoked Ca 2+ transients. Thus, the binding sites for ω‐conotoxin and for nitrendipine appear to be different. The results confirm the presence of L‐type Ca 2+ channels and open the possibility of N‐type Ca 2+ channels as the ω‐conotoxin binding sites in chromaffin cell membranes.