In the present study, direct projections from the lumbosacral cord to Barrington's nucleus in the rat were investigated by using retrograde and anterograde tracing techniques. After injection of cholera toxin B subunit (CTb) into Barrington's nucleus, a number of moderately CTb-labeled neurons were observed in the lumbosacral cord, with a slight ipsilateral dominance; most were located in the spinal parasympathetic and dorsal commissural nuclei of the lumbosacral cord. In addition, some retrogradely labeled neurons were found in the periaqueductal gray (PAG). These findings were confirmed by an anterograde labeling experiment. After biotinylated dextran amine (BDA) was injected into the lumbosacral cord, dense BDA-labeled axon terminals were found in Barrington's nucleus as well as in the PAG. Injection of BDA into the PAG resulted in many BDA-labeled terminals in Barrington's nucleus. The present results provided clear evidence for a direct projection from the spinal parasympathetic and dorsal commissural nuclei to Barrington's nucleus that could subserve conveying bladder-filling information from the lumbosacral cord to Barrington's nucleus in the micturition reflex of the rat.