beta-Endorphin-containing neurons in the rat central nervous system were localized using three improvements of the unlabelled antibody-enzyme bridge immmunocytochemical technique. These improvements were (1) the use of brains from colchicine-treated rats; (2) the proteolytic pretreatment of sections with pronase, and (3) a 'double-bridge' staining procedure. In addition to the known localization of beta-endorphin-like immunoreactivity in perikarya in the medial basal hypothalamus, we have observed nerve fibers and terminals with beta-endorphin-like immunoreactivity to be more widely distributed than reported in previous studies. This includes discrete areas of the septal, preoptic, hypothalamic, thalamic and subthalamic regions, the amygdala, the periaqueductal gray, the inferior colliculus, the nucleus tegmenti pontis, the nucleus raphe dorsalis, several regions of the reticular formation, the locus ceruleus, the parabrachial nuclei, the mesencephalic trigeminal nucleus, the nucleus raphe magnus, the solitary tract and the nucleus of the solitary tract. The distribution of beta-endorphin-like immunoreactivity is in good agreement with many of the physiological, neuroendocrine and behavioral effects attributed to this peptide such as analgesia, the regulation of the release of pituitary hormones, thermoregulation and feeding behavior. This implicates beta-endorphin as an important neurotransmitter or modulator with specific functions within the central nervous system.