ANTISERA were generated against nociceptin/orphanin FQ, the putative ligand of the opioid receptor-like ORL1 receptor. Dot blot analysis showed that the antibodies selectively detect nociceptin but not dynorphin or other opioid peptides. Immunofluorescent staining of tissue sections revealed dense plexus of nociceptin-immunoreactive nerve fibres and terminals within the spinal cord dorsal horn, sensory trigeminal complex, raphe nuclei, locus coeruleus, periaqueductal grey, amygdala, habenula, hypothalamic region and septal area in mice and rats. When adjacent sections were stained either with the nociceptin antibody or the pan-opioid 3-E7 mouse monoclonal antibody, an overlapping distribution was observed in many nociceptive centres including the superficial dorsal horn, sensory trigeminal complex and periaqueductal grey. However, confocal microscopic examination of dual-labelled spinal cord and brain stem sections showed no instances of co-localization of nociceptin and opioid peptides in these regions. Intra- cerebroventricular administration of nociceptin has been shown to induce hyperalgesia. Thus, the present results suggest that nociceptin and opioids are released from different terminals thereby modulating pain signals in opposite ways.