Although it is most well-known for its roles in central nervous system (CNS) function, the vast majority of serotonin, or 5-hydroxytryptamine (5-HT), is produced in the gastrointestinal (GI) tract. 5-HT is synthesized mostly by enterochromaffin (EC) cells of the GI epithelium and, in small part, by neurons of the enteric nervous system (ENS). The GI tract contains an array of broadly distributed 5-HT receptors, which participate in functions such as motility, sensation, inflammation, and neurogenesis. The roles of 5-HT in these functions are reviewed, as well as its role in the pathophysiology of disorders of gut-brain interaction (DGBIs) and inflammatory bowel diseases (IBD). © 2023 American Physiological Society. Compr Physiol 13:4851-4868, 2023.