It is known that the opioid peptide dynorphin A has a broad distribution throughout the neuraxis. Recent biochemical studies have extended the sequence of dynorphin A by 15 amino acids to include another [Leu]enkephalin-containing peptide known as dynorphin B. These sequence data have been validated by the elucidation of the structure of the hypothalamic mRNA coding for alpha- and beta-neo-endorphin, dynorphin A, and dynorphin B. Using specific antisera directed against each of the three opioid peptides, we have studied their cellular distribution in rat brain. Their distribution patterns are extremely similar, if not identical. Furthermore, all three peptide immunoreactivities can be localized to the same cells in five nuclear groups throughout the brainstem--the supraoptic nucleus, the paraventricular nucleus, a group of cells in the lateral hypothalamic area, the nucleus parabrachialis, and the nucleus tractus solitarius. The sequence of a common precursor for dynorphin A, B, and alpha- and beta-neo-endorphin was deduced from hypothalamic mRNA. The ability to localize all three peptides together within cells in widely placed nuclei strongly supports the use of the same biosynthetic precursor for the neo-endorphin and dynorphin peptides in other parts of the central nervous system as well.