Leu-enkephalin is an opioid peptide that has been found to modulate nociception in the spinal cord. Both pre- and postsynaptic interactions by enkephalins have been proposed. By the peroxidase . anti-peroxidase immunocytochemical method, we studied the distribution and ultrastructure of neuronal elements in the monkey dorsal horn to elucidate possible morphological substrates for postulated opioid actions. Biochemical analysis of immunoreactive Leu-enkephalin-like peptides in the cord was performed to characterize the forms present in labeled neurons and terminals. At the light microscopic level, fiber immunostaining was found in most areas of gray matter, especially in laminae I to V, and in the dorsolateral funiculus. Cell bodies were located in laminae I, II, III, and V. At the ultrastructural level, in the superficial dorsal horn, we found that neurons with Leu-enkephalin receive numerous types of axon inputs, some of which have been identified previously as originating from the dorsal root. Leu- enkephalin terminals formed primarily axosomatic and axodendritic synapses and less frequently synapsed with other axons. With the same Leu-enkephalin antiserum as used in the immunocytochemistry, a peptide physicochemically similar to intact Leu-enkephalin and two larger Leu- enkephalin-like peptides were identified in monkey spinal cord extracts. It is likely that a family of Leu-enkephalin-like peptides is present in monkey spinal cord and that the labeled elements may contain any or all of these substances. It is concluded that both pre- and postsynaptic physiologic effects of Leu-enkephalin are possible, although the preponderance of axodendritic synapses favors a principal postsynaptic site of action. The anatomical results suggest that neurons containing immunoreactive Leu-enkephalin in the dorsal horn, some of which may receive input from primary afferents, modulate nociception by directly synapsing with cells of origin of the spinothalamic tract and also by interacting with primary afferent terminals.