In this study the effects of neostigmine on metoclopramide-induced aldosterone secretion were examined in the presence of a relatively selective M1-antagonist, pirenzepine and of a non-selective muscarinic antagonist, atropine. Six normal male volunteers received metoclopramide, 10 mg i.v. on three different occasions, each study day being preceded by a day in which the intake of sodium and potassium was limited. The dosing was either metoclopramide alone or combined with either neostigmine and pirenzepine or with neostigmine and atropine. Serum aldosterone increased significantly with all three regimens. The highest levels were attained with the metoclopramide/neostigmine/pirenzepine regimen and those were significantly higher than those after metoclopramide alone and also, from 45 min onwards, from those after the metoclopramide/neostigmine/atropine regimen. The results of this investigation suggest that the metoclopramide-induced aldosterone secretion in humans is augmented by an accumulation of acetylcholine at the nerve-zona glomerulosa junctions and that the receptors mediating aldosterone secretion are of the M2-subclass of muscarinic receptors.