Carbenoxolone, a semi-synthetic triterpenoid, exhibits gastroprotective activity related to the participation of nitric oxide (NO); however, the complete NO/cGMP/KATP channels pathway for carbenoxolone is unknown. Therefore the aim of this study was to examine the NO/cGMP/KATP channels pathway as the gastroprotective mechanism of carbenoxolone in the ethanol-induced gastric injury model in the rat. Oral administration of carbenoxolone (30 mg/kg, p.o.) exhibited gastroprotective effect against ethanol-induced gastric injury in rats. Pretreatment with NG-nitro-L-arginine methyl ester (L-NAME, 70 mg/kg, i.p.); 1H-[1,2,4]oxadiazolo[4,3-a]quinoxaline-1-one (ODQ, guanylate cyclase inhibitor, 10 mg/kg, i.p.); or glibenclamide (KATP channels inhibitor, 1 mg/kg, i.p.) reversed the gastroprotective effect of carbenoxolone for ethanol-induced gastric injury. Furthermore, gastric prostaglandins and NO levels increased after carbenoxolone administration in ethanol-induced gastric injury in rats. In conclusion, our results suggest that the increase of NO levels in gastric tissue after pretreatment with carbenoxolone activates the NO/cGMP/KATP channels pathway, the principal gastroprotective mechanism of carbenoxolone.