Characterization of the functional subunit combination of nicotinic acetylcholine receptors in bovine adrenal chromaffin cells.

The combination of nicotinic acetylcholine receptors (nAChRs) subunits connecting with the secretion of catecholamines in bovine adrenal chromaffin cells was pharmacologically investigated using selective agonists and antagonists for their nAChRs. The EC(50) values (microM) for the agonists that stimulate the catecholamine secretion and the rank order were as follows: nicotine (3.3)> or =1,1-dimethyl-4-phenylpiperazinium (3.5) > (E)-N-methyl-4-(3-pyridinyl)-3-butene-1-amine (14) > cytisine (23) > or =acetylcholine (25). However, because both the rank order and the EC(50) values differed considerably from those in the various subunits' combinations expressed in Xenopus oocytes or mammalian cells (e.g. alpha2beta2, alpha3beta4, alpha4beta4, etc.), we could not compare them. On the other hand, the IC(50) values (microM) for the antagonists that inhibit the secretion and the rank order were mecamylamine (0.08) > alpha-conotoxin-MII (alpha-CTX-MII) (0.71) > dihydro-beta-erythroidine (DHbetaE) (48) > alpha-bungarotoxin (alpha-BTX) (no effect). Mecamylamine is a highly selective antagonist for alpha3beta4 nAChRs, and alpha-CTX-MII and alpha-BTX are specific antagonists for alpha3beta2 and alpha7 nAChRs, respectively. DHbetaE is a selective antagonist for the alpha4beta2. It has already been shown that the mRNAs for alpha3, alpha5, alpha7 and beta4 subunits are expressed in the chromaffin cells. Therefore, the subunit combination of nAChRs associated with the catecholamine secretion from bovine adrenal chromaffin cells is suggested to be at least alpha3beta4 or alpha3beta4alpha5. Further, the results indicate that the utilization of the nicotinic agonists as selective ligands for the subunit combination of nAChRs may be not suitable for the characterization of nAChRs.