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

Publisher Website
Google Scholar
Abstract
Binding of 125I-.omega.-conotoxin GVIA and [3H]nitrendipine to membranes from bovine adrenal medulla was investigated to test for the presence of N- and L-type Ca2+ channels in adrenal chromaffin cells. Saturable, high-affinity binding sites for 125I-.omega.=-conotoxin and [3H]nitrendipine were detected in a membrane fraction from adrenal medulla. [3H]Nitrendipine binding sites were found to have a KD of 500 .+-. 170 pM and a Bmax of 26 .+-. 11 pmol/g of protein. 125I-.omega.-Conotoxin binding sites had a KD of 215 .+-. 56 pM and a Bmax of 105 .+-. 18 pmol/g of protein, about four times the number of sites found for [3H]nitrendipine. 125I-.omega.-Conotoxin binding was potently inhibited by unlabeled toxin and Ca2+ but was unaffected by dihydropyridines, verapamil, and diltiazem. [3H]Nitrendipine binding was not affected by .omega.-conotoxin, whereas it was inhibited by other dihydropyridines. Bay K 8644 potentiated K+-evoked cytosolic Ca2+ transients measured by fura-2 fluorescence, and this potentiation was completely blocked by nifedipine. In contrast, .omega.-conotoxin had no effect on Bay K 8644- evoked Ca2+ transients. Thus, the binding sites for .omega.-conotoxin and for nitrendipine appear to be different. The results confirm the presence of H-type Ca2+ channels and open the possibility of N-type Ca2+ channels as the .omega.-conotoxin binding sites in chromaffin cell membranes.