Identification of omega-conotoxin binding sites on adrenal medullary membranes: possibility of multiple calcium channels in chromaffin cells.
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 L-type Ca2+ channels and open the possibility of N-type Ca2+ channels as the omega-conotoxin binding sites in chromaffin cell membranes.
Jan
C R
CR
Department of Pharmacology and Toxicology, Albany Medical College, New York 12208.
Titeler
M
M
Schneider
A S
AS
D000313:Adrenal Medulla / Q000378:metabolism*; D000818:Animals; D001572:Benzofurans; D001665:Binding Sites; D015220:Calcium Channels / Q000378:metabolism*; D002417:Cattle; D002462:Cell Membrane / Q000378:metabolism; D005456:Fluorescent Dyes; D016257:Fura-2; D007700:Kinetics; D008978:Mollusk Venoms / Q000378:metabolism; D009568:Nitrendipine / Q000378:metabolism; D011869:Radioligand Assay; D013050:Spectrometry, Fluorescence; D020866:omega-Conotoxin GVIA
D001572:Benzofurans; D015220:Calcium Channels; D005456:Fluorescent Dyes; D008978:Mollusk Venoms; D020866:omega-Conotoxin GVIA; D009568:Nitrendipine; D016257:Fura-2