A newly developed, primary culture of human antral epithelial cells has been utilized to examine the effect of parasympathomimetics on somatostatin release. The cholinergic agonists, carbachol and methacholine, stimulated somatostatin secretion in a concentration-dependent manner. Maximal release in response to carbachol was observed at 0.1 mmol/l. Methacholine was 10 times more potent with a significant release being observed at 1 mumol/l, maximal secretion was observed at 10 mumol/l. Somatostatin release, stimulated by the mixed nicotinic and muscarinic agonist, carbachol, was attenuated by the addition of atropine at 0.1 mumol/l but was unaffected by the same concentration of pirenzepine. Methacholine-stimulated release was attenuated by addition of 0.1 mumol/l atropine and unaffected by the same concentration of pirenzepine. The response to methacholine was reversed by the addition of 0.1 mumol/l 4-diphenylacetoxy-n-methylpiperidine methiodide (4-DAMP) and attenuated by 1 nmol/l 4-DAMP indicating that the effect was mediated by an M3 receptor. In conclusion, human antral D cells are stimulated by parasympathomimetics acting at an M3 receptor.