Background The 5-HT4 receptor agonist, mosapride citrate, accelerates gastric emptying. However, the effect of mosapride on colonic function has not been well investigated. We examined whether mosapride changes rectosigmoid motility and perception in patients with irritable bowel syndrome (IBS). Methods Thirty-seven patients with IBS and 18 healthy subjects were studied. All subjects underwent a rectosigmoid barostat test to measure pain perception to intraluminal distention and resting smooth muscle motility for 20 min in the fasting state. Irritable bowel syndrome patients were then randomly assigned to receive either mosapride 15 mg (n = 19) or placebo (n = 18) orally with 200 mL water. Rectosigmoid motility and perception were measured again for 60 min following dosing. Rectosigmoid tone and contractility were evaluated in each 10-min period. Key Results The pain threshold in the patients was significantly lower than that in controls (P < 0.01). There were no differences between mosapride and placebo groups in pain threshold, barostat bag volume, or number of contractions at baseline. Mosapride significantly decreased the mean bag volume (P < 0.01; group x period interaction by two-way ANOVA) and increased the mean number of contractions (P < 0.05) compared with placebo, but did not affect the perception. In IBS patients with constipation (i.e., excluding diarrhea-predominant subjects), mosapride (n = 13) increased rectosigmoid tone (P < 0.01) and contractions (P < 0.05) more than placebo (n = 14). Conclusions & Inferences Mosapride stimulates colonic motility without any adverse effect. These findings suggest that mosapride may have the potential to treat IBS patients with constipation and/or functional constipation. Further clinical trials are warranted to confirm the efficacy of this agent.