The effect of diazoxide on the secretion of glucagon and insulin was studied using the isolated perfused rat pancreas. The perfusate concentration of D-glucose was kept constant at 5.6 mM. Five secretagogues of both glucagon and insulin--10 mM L-arginine, 5 mM L-leucine, 1.4 muM prostaglandin F2alpha, 100 nM bovine growth hormone, and 10 nM theophylline--were administered individually in the presence or absence of 325 muM diazoxide. Basal secretion of glucagon or insulin was not discernibly affected by diazoxide. With diazoxide the secretion of glucagon was (a) abolished completely in response to L-arginine or L-leucine; (b) inhibited partially in response to prostaglandin F2alpha; (c) unaltered in response to growth hormone; and (d) unchanged or, at times, enhanced in response to theophylline. On the other hand, the secretion of insulin induced by each of these agents was inhibited effectively by diazoxide. CONCLUSIONS (a) Diazoxide inhibits the secretion of glucagon as well as insulin in response to certain secretagogues independent of any changes in prevailing levels of glucose. (b) At the concentration tested, diazoxide is a more potent and consistent inhibitor of the release of insulin than of glucagon.