Substance P-immunoreactive neurons projecting from the midbrain to the spinal cord of the chicken were examined by the use of the retrograde tract-tracing method combined with immunohistochemical techniques. Many small neurons were densely clustered in the rostral midline area of the midbrain (RMA), and showed substance P-immunoreactivity. These substance P-immunoreactive neurons sent axons to the intermediomedial cell column (avian autonomic preganglion) and its vicinity in the lumbar spinal segments. On the basis of the strong neuroanatomical analogy in the cytoarchitectural features, immunoreactivity, and fiber connections, the RMA was assumed to be the avian homologue of the anteromedian nucleus in the mammalian midbrain.