Abstract The relationship between the central processes of classical conditioning and conditioned responses of the gastrointestinal function is incompletely understood in humans. We tested the hypothesis that the rectosigmoid motility becomes conditioned with anticipatory painful somatosensory stimulus and that characteristic brain areas become activated during anticipation. In nine right-handed healthy male subjects, a loud buzzer (CS, conditional stimulus) was paired with painful transcutaneus electrical nerve stimulation to the right hand (unconditional stimulus). Rectosigmoid muscle tone measured by the barostat as the intrabag volume, phasic contractions of the bowel measured as the number of phasic volume events (PVEs), and regional cerebral blood flow assessed by positron emission tomography (PET), were measured before and after conditioning. Following conditional trials, the bag volume after CS alone did not show significant changes between before and after the stimulus, but the number of PVEs after 2-minute interval of the CS alone was significantly greater than that before the stimulus (P < 0.05). The PET data showed the conditioning elicited significant cerebral activation of the prefrontal, anterior cingulate, parietal and insula cortices (P ≤ 0.001, uncorrected). Rectosigmoid motility can be conditioned with increase in phasic contractions in humans.