Neuropeptides form a part of the brain-gut axis which may regulate gastrointestinal functions, including immune regulation. Various changes in the neuropeptides--most important, vasoactive intestinal peptide and substances P (VIP and SP)--have been described in inflammatory bowel disease. We employed a sensitive immunoperoxidase (avidin-biotin-peroxidase complex) technique, using anti-VIP and anti-SP antibodies to localize and compare the distribution of VIP and SP in the colon. Colon specimens from 19 normal subjects, eight patients with ulcerative colitis (UC), and eight with Crohn's disease (CD) were used. In the normal colon, VIP and SP immunoreactivity (IR) were localized in the muscularis mucosa, circular muscles, walls of blood vessels, nerve fibers, and some distinct cells, probably enterochromaffin cells. SP-IR was also present in the epithelial cells, mainly along the basolateral domain. VIP-IR was considerably diminished at all locations in patients with UC and CD. However, the SP-IR was increased in UC in the colonic epithelial cells along the basolateral areas. The SP-IR was intense in patients with CD, in the epithelium, the granulomas, cells lining the mucosal fissure, and in the muscle layers. In contrast to normals, SP-IR in patients with CD was observed both in the longitudinal and circular muscles. We conclude that VIP-IR and SP-IR are distributed widely in the mucosa, submucosa, and in the circular muscle in normal colon. VIP-IR is decreased in UC and CD, whereas SP-IR is increased in both, but more so in CD.