Necrotizing enterocolitis (NEC), a gastrointestinal inflammatory disease of unknown etiology, causes a great deal of morbidity and mortality in premature infants. The liver as well as the gut may be affected in NEC. We tested the hypothesis that signaling molecules, which are endogenous to the bowel, regulate the severity of experimental NEC in mice. Specifically, we postulated that mucosal serotonin (5-HT), which is proinflammatory, would increase the severity of NEC and that oxytocin (OT), which is present in enteric neurons and is anti-inflammatory, would oppose NEC. Genetic deletion of the 5-HT transporter (SERT), which increases and prolongs the effects of 5-HT, was found to increase the severity of the systemic manifestations of NEC as well as intestinal inflammation and associated hepatotoxicity. In contrast, genetic deletion of tryptophan hydroxylase 1 (TPH1), which is responsible for 5-HT biosynthesis in enterochromaffin (EC) cells of the intestinal mucosa, and TPH inhibition with LP-920540 both decrease the severity of NEC in intestine and liver. These observations suggest that 5-HT from EC cells helps to drive both the enteric and hepatic manifestations of NEC. Administration of OT decreased NEC-induced intestinal inflammation, while that of atosiban, an OT receptor antagonist, caused NEC-induced intestinal inflammation to become more severe. Data from the current investigation are consistent with the tested hypotheses, that the enteric signaling molecules, 5-HT (positively) and OT (negatively) regulate NEC severity; moreover, we suggest that mucosally restricted inhibition of 5-HT biosynthesis and/or administration of OT may be useful in the treatment of NEC.