Adrenal medullary cells contain opiate recognition sites that cannot be classified with any of the accepted conventional criteria. In primary cultures of bovine adrenal chromaffin cells, stimulation of nicotinic receptors by acetylcholine causes an increase in the release of catecholamines. When the action of acetylcholine is studied in the presence of opiate receptor agonists, the acetylcholine secretory action is curtailed. The action of the opiates is stereoselective and is blocked by naloxone and diprenorphine. The blocking activity of each opiate correlates with its Ki for the displacing of [3H]etorphine bound to specific recognition sites of adrenal medulla. Enkephalin-like opiate peptides are stored in the splanchnic nerves; they appear to act as a cotransmitter because they decrease the gain at which nicotinic receptors operate. This regulation appears to involve a down-regulation of the nicotinic receptor recognition sites because these opiate peptides elicit a decrease in the Bmax of the specific binding to adrenal medullary membranes of a radioactive fraction of alpha-bungarotoxin, a compound that inhibits the action of acetylcholine in releasing catecholamines from chromaffin cells.