This study examined the hypothesis that activation of ATP-sensitive potassium channels produces vasodilation and contributes to dilator responses of the basilar artery to acetylcholine in vivo. Diameter of the basilar artery (baseline diam = 245 +/- 14 microns, means +/- SE) was measured through a cranial window in anesthetized rats. RP52891 (1 microM), a direct activator of ATP-sensitive potassium channels, increased the diameter of the basilar artery by 33 +/- 5%. Glibenclamide (1 microM), an inhibitor of ATP-sensitive potassium channels, did not alter baseline diameter but abolished responses of the basilar artery to RP52891. Topical application of acetylcholine (10 microM) for 3 min produced peak dilatation of 33 +/- 6% at 30 s and produced a sustained increase in diameter of 17 +/- 4%. Glibenclamide did not inhibit dilator responses of the basilar artery to acetylcholine. Nitro-L-arginine methyl ester (10 and 100 microM), which inhibits synthesis of endothelium-derived relaxing factor (EDRF), produced concentration-dependent inhibition of dilatation of the basilar artery in response to acetylcholine. Thus ATP-sensitive potassium channels are functional but do not appear to influence basal tone of the basilar artery. Dilator responses of the basilar artery to acetylcholine are dependent on formation of EDRF but not dependent on activity of glibenclamide-sensitive potassium channels.