The structurally related orphan G-protein-coupled receptors GPR7 and GPR8 are expressed in the central nervous system, and their ligands have not been identified. Here, we report the identification of the endogenous ligand for both of these receptors. We purified the peptide ligand from porcine hypothalamus using stable Chinese hamster ovary cell lines expressing human GPR8 and cloned the cDNA encoding its precursor protein. The cDNA encodes two forms of the peptide ligand with lengths of 23 and 30 amino acid residues as mature peptides. We designated the two ligands neuropeptide W-23 (NPW23) and neuropeptide W-30 (NPW30). The amino acid sequence of NPW23 is completely identical to that of the N-terminal 23 residues of NPW30. Synthetic NPW23 and NPW30 activated and bound to both GPR7 and GPR8 at similar effective doses. Intracerebroventricular administration of NPW23 in rats increased food intake and stimulated prolactin release. These findings indicate that neuropeptide W is the endogenous ligand for both GPR7 and GPR8 and acts as a mediator of the central control of feeding and the neuroendocrine system. Searches for ligands for orphan G-protein-coupled receptors (GPCRs)1 have discovered many novel peptides and have identified the previously unknown receptors of bioactive substances (1-9). Studies on the newly identified ligands and their receptors have given us a more precise understanding of the physiological processes involved in the endocrine, cardiovascular, reproductive, immune, inflammatory, digestive, metabolic, and central nervous systems (1-11). In addition, these studies have provided opportunities to discover innovative drugs that exert their pharmacological effects by interacting with an identified receptor as an agonist or antagonist (12). GPR7 and GPR8, for which the ligands have not been identified, are structurally related orphan GPCRs. Two genes for GPR7 and GPR8 were originally isolated from human genomic DNA by O'Dowd et al. (13). Human GPR7 highly resembles human GPR8, with an amino acid identity of 64%. Among various families of GPCRs, GPR7 and GPR8 share high similarity to the opioid and somatostatin receptor families. In mammalian brain, gene expression of GPR7 and GPR8 was detected by Northern blot and in situ hybridization analyses (13). Especially in rat brain, GPR7 mRNA was detected in regions of the cortex, hippocampus, and hypothalamus (14). Profiles of GPR7 and GPR8 expressed mainly in brain suggest that the endogenous ligands for the two receptors have several functions in the central nervous system.In this study, we report the purification, cloning, and characterization of neuropeptide W (NPW). We attempted to purify the agonist peptide for GPR8. The cDNA encoding the agonist peptide for GPR8 demonstrates the existence of neuropeptide W-23 (NPW23) and neuropeptide W-30 (NPW30), which exhibit no meaningful similarity to any known peptides. With the functional and binding characterization of NPW for GPR7 and GPR8, we show that NPW is the endogenous ligand for both of these recept...

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