Afferents to the interpeduncular nucleus (IPN) of the rat were studied with the horseradish peroxidase (HRP) retrograde transport method. HRP was deposited microelectrophoretically in the IPN of adult rats. Major projections to the IPN originate in the medial habenular nucleus, the region surrounding the dorsal tegmental nucleus (accessory dorsal tegmental nucleus and the so-called dorsal tegmental nucleus pars lateralis), and the midbrain raphe (nucleus centralis superior and nucleus raphe dorsalis). Also, minor projections originate in the central gray and nucleus locus coeruleus. Our results indicate that the habenulointerpeduncular projection originates solely from the medial habenular nuclei and is topographically organized; medial regions of the medial habenular nuclei project to ventral portions of IPN and lateral regions project to the dorsal IPN.