A novel neurotensin-like peptide, designated neuromedin N, has been isolated from porcine spinal cord by using a bioassay for a stimulant effect on guinea pig ileum. By microsequencing, the amino acid sequence of the peptide has been determined to be Lys-Ile-Pro-Tyr-Ile-Leu, which is found to be quite homologous to the COOH-terminal sequence of neurotensin. This structure has been confirmed by synthesis. Neuromedin N exhibits a contractile activity on guinea pig ileum and induces a hypotensive response in the rat similar to that with neurotensin. These findings suggest that neuromedin N may be a new neuromediator or hormone with a specific spectrum of biological activity.