We investigated the effect of 24 h exposure to 100 nmol/l glibenclamide on insulin secretion in isolated rat pancreatic islets. The insulin content was similar in control islets and in islets preincubated with 100 nmol/l glibenclamide for 24 h. In islets preexposed to glibenclamide: 1) the subsequent response to a maximal glibenclamide stimulatory concentration (10 mumol/l, 1 h at 37 C) was greatly reduced in comparison to control islets (0.69 +/- 0.20% vs 2.16 +/- 0.41%; mean +/- SE; n = 14; p less than 0.001); 2) the response to 100 mumol/l tolbutamide stimulation was also reduced (0.55 +/- 0.15% vs 2.38 +/- 0.44%; n = 8; p less than 0.001); 3) the response to 16.7 mmo/l glucose, both in the presence or in the absence of 1 mmol/l IBMX, a phosphodiesterase inhibitor, was also diminished by about 50% (1.79 +/- 0.39% vs. 3.22 +/- 0.42%; n = 14, p less than 0.001). In glibenclamide pretreated islets, blunted responses to stimuli were confirmed also by dynamic studies using a perifusion system. The effect of glibenclamide preincubation was fully reversible: when islets cultured in the presence of glibenclamide were transferred to a glibenclamide-free medium for further 24 h, insulin release in response to glibenclamide stimulation returned to control values. We conclude that prolonged exposure of rat pancreatic islets to glibenclamide induces a reversible desensitization to a variety of metabolic stimuli. The inhibition by prolonged glibenclamide exposure of a common pathway in the mechanism of insulin release is one possible explanation for these results.