In various areas of rat and human brain and spinal cord the distributions of opioid peptides derived from the proenkephalin A precursor, the heptapeptide [Met]enkephalin-Arg6-Phe7 (MERF), the octapeptide [Met]enkephalin-Arg6-Gly7-Leu8 (MERGL), and bovine adrenal medulla dodecapeptide (BAM-12P), were determined by a combination of radioimmunoassay, gel filtration, and high-performance liquid chromatography. In the human central nervous system the highest concentrations were seen in the striatum (pallidum greater than caudate nucleus greater than putamen) and in substantia nigra, hypothalamus, and periaqueductal gray. Similarly, in rat brain high levels were found in striatum and hypothalamus. Bovine adrenal medulladocosa peptide (BAM-22P) only occurred in the rat brain, but could not be detected in human brain. No MERF, MERGL, BAM-12P, or BAM-22P could be found in either rat or human pituitary. In contrast to MERF, MERGL and BAM-12P, peptides derived from the proenkephalin B precursor, dynorphin1-8 and dynorphin B, showed high concentrations only in substantia nigra and pallidum, but quite low levels in the other regions of human brain and spinal cord. The present study provides evidence that the proenkephalin A precursor known from adrenal medulla also exists in the rat and human central nervous system. Moreover, the identification of BAM-12P in these tissues indicates that cleavage of the precursor molecule must also involve sites different from those with paired basic amino acids.