A widespread distribution of immunoreactive dynorphin (ir-Dyn) in rat brain and spinal cord was demonstrated by means of a highly specific radioimmunoassay. The highest concentrations of ir-Dyn (greater than 399 pg/mg protein) were found in hypothalamic nuclei, i.e. the premamillary, anterior hypothalamic and dorsomedial nuclei and median eminence. Relatively high concentrations of ir-Dyn (between 320 and 399 pg/mg protein) were found in other hypothalamic nuclei such as the medial and lateral preoptic, perifornical, suprachiasmatic, ventromedial nuclei and in the medulla oblongata in the area postrema and in the nucleus of the solitary tract (commissural part). Moderate levels of ir-Dyn (between 140 and 320 pg/mg protein) were found in most diencephalic areas other than the hypothalamic nuclei and further nuclei in the medulla oblongata, in the mesencephalon, pons and spinal cord. Low to moderate levels of ir-Dyn were found in the telencephalon, with lowest levels (less than 140 pg/mg protein) found in the cerebral cortex, olfactory bulb, dorsal septal nucleus, medial amygdaloid nucleus, caudate-putamen, superior collicle, cerebellum and certain areas of the reticular formation.