(125I-His1, D-Ala2, Nleu27)-growth hormone-releasing factor (GRF) (l-29)-NH2, initially developed as a possible radioligand for identifying GRF receptors in the anterior pituitary, was found to bind to rat hepatic membranes. The tracer was stable, bound rapidly and reversibly, and its dissociation was accelerated by GTP. Radioligand binding was enhanced by the divalent cations Mg2+, Ca2+ and Mn2+ and inhibited by the chelating agent EDTA. Vasoactive intestinal peptide (VIP), PHI, secretin, GRF(l-29)-NH2, (His1, D-Ala2, Nleu27)-GRF(l-29)-NH2, and (D-Ala2, Nleu27)-GRF(l-29)-NH2 dose-dependently inhibited tracer binding. The order of potency of the unlabelled peptides tested suggested that (123I-His1, D-Ala2, Nleu27)-GRF(l-29)-NH2 specifically identified a high-affinity subclass of VIP receptors in rat liver membranes.