The localization of Barrington's nucleus in the dorsolateral pons of the rabbit and its projections to the sacral spinal cord were examined by using retrograde and anterograde labeling methods combined with immunohistochemistry. After injection of wheat germ agglutinin-horseradish peroxidase (WGA-HRP) or a fluorescence tracer, tetramethylrhodamine-dextran amine (TMR), into the sacral spinal cord segments, a cluster of neurons labeled with WGA-HRP or TMR were seen in the pontine dorsolateral tegmentum. To identify whether the retrogradely labeled neurons were situated within the locus coeruleus, the sections containing TMR-labeled neurons through the pons were incubated with anti-tyrosine hydroxylase (TH) antibody and observed under epifluorescence microscope. It was shown that the cluster of TMR-labeled neurons in the dorsolateral tegmentum were surrounded by TH-positive neurons, but they were negatively immunostained with TH-like immunoreactivity. In anterograde experiment, injection of WGA-HRP into the dorsolateral tegmentum resulted in many anterogradely labeled nerve fibers and terminals in the sacral spinal cord, including the sacral parasympathetic nucleus. The present results suggest that the cluster of neurons in the dorsolateral tegmentum of the rabbit may correspond to Barrington's nucleus revealed in the rat and cat, and thus may be involved in micturtion reflex of the rabbit.