The effect of nebivolol (NBV), a third-generation selective β1- antagonist, on indomethacin (IND) - induced gastric ulcer in rats was investigated. Thirty two male rats were used in this study, divided into four groups as follows: control group, IND ulcer-induced group, IND + NBV (1mg/kg/day, p.o.) group, and IND + NBV (5mg/kg/day, p.o.) group. Gastric ulcer was induced by a single injection of IND (30 mg/Kg, i.p.); NBV was administered for seven days prior to ulcer induction. At the end of the experiment, the stomach of each rat was removed for macroscopic examination and gastric ulcer scoring. The gastric mucosa was prepared for quantitative real-time polymerase chain reaction (qRT-PCR) and tissue homogenate preparation for biochemical study. Then, the remaining part was used for histopathological and immunohistochemical examination. Results showed that NBV significantly protected rats from indomethacin-induced gastric ulceration. There is a significant decreased in the mean ulcer number, score and index along with increased preventive index. Also, NBV significantly decreased the elevations of nitrite⁄nitrate, malondialdehyde, tumor necrosis factor-α, and interleukin-1beta in IND-treated rats. In addition, NBV significantly ameliorated the IND-induced reductions of glutathione level, glutathione peroxidase, superoxide dismutase, and catalase activities, and prostaglandin E2. Moreover, histopathological, immunohistochemical analysis of inducible nitric oxide synthase (iNOS) and qRT-PCR for mRNA expression of iNOS gene:confirmed the biochemical results. It was concluded that NBV renders protection in IND-induced gastric ulceration in rats via maintenance of mucosal nitric oxide and prostaglandin E2, reducing oxidative stress and inflammatory responses.