This study aimed to investigate the effect of oral treatment with ketotifen, a mast cell (MC) stabilizer, in a rat model of surgically induced endometriosis. At 14 days after Sprague-Dawley rats had surgery, they were treated with ketotifen (1 or 10 mg/kg/day). Pain behaviors were evaluated 3 days prior to surgery and then at 7, 14, 21, and 28 days after surgery. At day 28, rats were sacrificed and all samples were then processed for biochemical studies. We found that ketotifen-treated rats showed significantly shorter duration of hyperalgesia (<0.05); smaller cyst diameter (<0.05) and lower histopathologic score (<0.001); significantly lower MC number and degranulation (<0.001), blood vessel number (<0.001), lower expression levels of nerve growth factor (<0.001), cyclooxygenase-2 (<0.001), intercellular cell adhesion molecule-1 (<0.001), and vascular endothelial growth factor (<0.05) in cysts, and nerve growth factor (<0.001) and transient receptor potential cation channel, subfamily V, member 1 (<0.001) in dorsal root ganglia; and lower histamine (<0.05) and tumor necrosis factor-alpha (<0.05) concentrations in serum compared with placebo-treated animal subjects. Oral treatment with ketotifen significantly suppressed the development of hyperalgesia, probably by modulating MC activity in cysts, thereby reducing peripheral sensitization due to noxious signals from endometriotic lesions. Our results suggest that ketotifen may inhibit the development of endometriotic lesions and hyperalgesia in rats.