Short-term regulation of [3H]methylscopolamine binding to muscarinic receptors and acetylcholineinduced stimulation of insulin release was investigated in pancreatic islets of the rat. Binding of methylscopolamine was reversible; 47% of label was displaced 10 min and 70% 30 min after addition of unlabelled substance. 0.1 mM chloromercuribensoic acid, when present during binding incubations, inhibited binding by 54%, whereas acetylcholine-induced insulin release was unaffected by the presence of the thiol reactant. Pre-incubation for 60 min in a calcium-deprived medium or in the presence of 50 \g=m\mtrifluoroperazine likewise inhibited binding. Pre-incubation with 1.0 mm 3-isobutyl-l-methylxanthine or 16.7 m glucose failed to influence subsequent binding although acetylcholine-induced insulin release was 4-fold enhanced by priming with glucose. We conclude that 1) binding to muscarinic receptors is influenced by thiol interaction, 2) short-term alterations in calcium fluxes influence binding, whereas short-term changes in cyclic AMP (cAMP) or glucose metabolism do not, 3) a priming effect of glucose on insulin secretion is not mediated by changes in receptor binding. The parasympathetic nervous system exerts an important influence on insulin secretion. Cephalic phase insulin secretion may thus be mediated to a major part by vagai stimulation ; its absence leads to delayed insulin secretion out of phase with mealinduced hyperglycaemia (Trimble et al. 1981). The receptors mediating the cholinergic effect are muscarinic in nature since acetylcholine-induced insu¬ lin secretion is readily blocked by atropine (Froh¬ man et al. 1967).To gain further insight into mechanisms behind acetylcholine-induced insulin release, we previous¬ ly studied binding of [3H]methylscopolamine to rat pancreatic islets. A high-affinity, rapid and specific binding of the muscarinic antagonist could be demonstrated (Grill & Östenson 1983). Half-maxi¬ mal inhibition of acethylcholine-induced release by methylscopolamine occurred at concentrations which were not lower than those causing halfmaximal displacement of label, suggesting that the binding parameter measured reflected an initial step in acetylcholine-induced insulin secretion.The aim of the present study was to gain further insight into islet regulation of muscarinic receptor binding and its role for insulin release. To this end we have investigated the effects on binding of factors of importance for insulin release such as the functional state of membrane thiol groups, per¬ turbations in cyclic AMP and calcium levels as well as changes in glucose metabolism. Additionally the dissociation of [3H]methylscopolamine was studied in order to further validate our procedure for measuring binding. Material and Methods Material and animah[3H]methylscopolamine chloride (S.A. 85 Ci/mmol) and [14C]sucrose (S.A. 673 mCi/mmol) was obtained from New England Nuclear (Dreieichenhain, FRG). Unlabel¬ led methylscopolamine nitrate was from Pharmacia, Uppsala, Sweden. Acetylcholine chloride, p-chloromerc...