The distribution of Met-enkephalin- and substance P-immunoreactive neurons was studied by indirect immunofluorescence in some areas related to pain and analgesia. Met-enkephalin- and substance P-positive cell bodies and nerve terminals were observed in the periaqueductal central gray, the nucleus raphe magnus, the marginal layers and substantia gelatinosa of the spinal trigeminal nucleus, and the dorsal horn of the spinal cord. Lesion experiments suggest that Met-enkephalin neurons in the dorsal horn and possibly in the spinal trigeminal nucleus are interneurons or propriospinal neurons with nerve terminals in the laminae I and II of the cord and in the superficial layers of the spinal trigeminal nucleus, respectively. These areas are also very rich in substance P-positive nerve terminals, mainly representing central branches of primary afferent neurons. The present immunohistochemical-anatomical findings support the hypothesis that stimulation-produced analgesia is related to activation of spinal and spinal trigeminal enkephalin interneurons forming axo-axonic synapses with (substance P?) pain afferents in the superficial laminae of the dorsal horn and the spinal trigeminal nucleus. These interneurons may be activated by sensory fibers and by descending fibers from medullary stimulation sites. Transmitter substances in these descending fibers may be 5-hydroxytryptamine and substance P.