The relaxant mechanisms of nicorandil were examined by comparing its effects with those of sodium nitroprusside and cromakalim in bovine tracheal smooth muscle. In preparations contracted with methacholine (0.3 µmol/l) or high K+ (40 mmol/l), nicorandil and sodium nitroprusside caused concentration-dependent relaxations. Their relaxant effects on high K+-contracted preparations were smaller than those on methacholine-contracted muscle. Cromakalim relaxed methacholine-contracted preparations, whereas it had no effect on high K+-contracted muscle. The inhibitor of soluble guanylyl cyclase 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ; 5 µmol/l) completely prevented the relaxation induced by lower concentrations (<30 µmol/l) of nicorandil, whereas it partially attenuated relaxation caused by higher concentrations. The ATP-sensitive K+ (KATP) channel blocker glibenclamide only partially attenuated the relaxant responses to nicorandil (at 100 and 300 µmol/l). Combination treatment with ODQ and glibenclamide almost completely prevented nicorandil-induced relaxations. The large-conductance Ca2+-activated K+ channel (Maxi K+ channel) inhibitor iberiotoxin significantly prevented the relaxations induced by lower concentrations (3 and 10 µmol/l) of nicorandil. The preventive effect of iberiotoxin was markedly enhanced under the blockade of KATP channels with glibenclamide. These results suggest that nicorandil relaxes bovine tracheal smooth muscle through 2 mechanisms: opening of KATP channels and activation of the nitric oxide (NO)-cyclic guanosine monophosphate (cGMP) pathway. Nicorandil may also activate Maxi K+ channels, possibly through the NO-cGMP pathway, and the interaction of KATP channels and Maxi K+ channels may affect the relaxant effect of nicorandil in bovine tracheal smooth muscle.