By using a combined technique of immunocytochemistry and [3H]thymidine autoradiography, we have determined the "birth date" of opioid peptide-containing neurons in several hypothalamic nuclei and regions. These include proopiomelanocortin (POMC) neurons (represented by ACTH immunoreactivity) in the arcuate nucleus; dynorphin A neurons in the supraoptic and paraventricular nuclei and the lateral hypothalamic area; and leu-enkephalin neurons in the periventricular, ventromedial, and medial mammillary nuclei, as well as in preoptic and perifornical areas. Arcuate POMC neurons were born very early in embryonic development, with peak heavy [3H]thymidine nuclear labelling occurring on embryonic day E12. Supraoptic and paraventricular dynorphin A neurons were also labelled relatively early (peak at E13). The lateral hypothalamic dynorphin A neurons showed peak heavy labelling also on day E12. By contrast, leu-enkephalin neurons in the periventricular nucleus and medial preoptic area exhibited peak heavy nuclear labelling on day E14. Furthermore, perifornical and ventromedial leu-enkephalin neurons were also born relatively early (peak on days E12 and E13, respectively). However, the leu-enkephalin neurons in the medial mammillary nucleus were born the latest of all cell groups studied (i.e., peak at E15). The results indicate a differential genesis of these opioid peptide-containing neuronal groups in different hypothalamic nuclei and regions.