Using specific rabbit anti-GAP (gonadotropin-releasing hormone-associated peptide) serum, we have immunocytochemically localized GAP in the rat brain. Immunostaining of neuronal perikarya, fibers and terminals was demonstrated with GAP antiserum under conditions of tissue preparation which make immunostaining with LHRH anti-sera difficult or undectectable. GAP-immunoreactive perikarya were observed in sections of perfused or nonperfused brains without colchicine pretreatment. Using a double immunoperoxidase staining method, both GAP and LHRH immunoreactivities were shown to coexist in the same neurons. The common distributionof LHRH and GAP immunoreactivity in the rat brain is strongly supportive of GAP representing the non-LHRH portion of the LHRH precursor. The use of GAP antisera that can distinguish between LHRH and the remaining portion of its prohormone represents a valuable tool for studies of LHRH-prohormone processing and distribution.