Cotinine binding to nicotinic acetylcholine receptors in bovine chromaffin cell and rat brain membranes.

Abstract

Cotinine is the major metabolite of nicotine. It has nicotine-like biological activity, but its potency is low. We studied cotinine binding to nicotinic receptors labelled with [3H]epibatidine. In membranes from cultured bovine chromaffin cells [3H]epibatidine bound to two apparent sites with K(d) values of 93 and 1400 pM. The low-affinity binding represented two-thirds of the binding sites. In rat frontal cortex and hippocampus homogenate membranes, only one apparent binding site was detected. The Kd values were 40 and 62 pM, in frontal cortex and hippocampus, respectively. Nicotine displaced [3H]epibatidine 10 times more potently from the brain than from the chromaffin cell membranes, and cotinine had over two orders of magnitude lower affinity than nicotine. In addition, the competitive nicotinic receptor antagonists methyllycaconitine and dihydro beta-erythroidine displaced [3H]epibatidine (100 pM and 1 nM) from the chromaffin cell membranes. Alpha-bungarotoxin did not affect the binding of 100 pM [3H]epibatidine. However, upon labelling with 1 nM [3H]epibatidine alpha-bungarotoxin (10 nM to 10 microM) displaced one-sixth of the bound radioligand. Our results demonstrate that 100 pM to 1 nM [3H]epibatidine labels mostly neuronal heteropentameric nicotinic receptors in bovine chromaffin cell membranes, and that cotinine is a low-affinity nicotinic ligand both in the adrenal chromaffin cell and in the brain receptors.