The mouse GalR2 galanin receptor: genomic organization, cDNA cloning, and functional characterization.

The diverse physiological actions of galanin are thought to be mediated through activation of galanin receptors (GalRs). We report the genomic and cDNA cloning of a mouse GalR that possesses a genomic structure distinct from that of GalR1 and encodes a functional galanin receptor. The mouse GalR gene consists of two exons separated by a single intron within the protein-coding region. The splicing site for the intron is located at the junction between the third transmembrane domain and the second intracellular loop. The cDNA encodes a 370-amino acid putative G protein-coupled receptor that is markedly different from human GalR1 and rat GalR3 (38 and 57%) but shares high homology with rat GalR2 (94%). In binding studies utilizing membranes from COS-7 cells transfected with mouse GalR2 cDNA, the receptor displayed high affinity (K(D) = 0.47 nM) and saturable binding with 125I-galanin (Bmax = 670 fmol/mg). The radioligand binding can be displaced by galanin and its analogues in a rank order: galanin approximately = M40 approximately = M15 approximately = M35 approximately = C7 approximately = galanin(2-29) approximately = galanin(1-16) >> galanin(10-29) approximately = galanin(3-29), which resembles the pharmacological profile of the rat GalR2. Receptor activation by galanin in COS-7 cells stimulated phosphoinositide metabolism, which was not reversed by pertussis toxin. Thus, the galanin receptor encoded in the cloned mouse GalR gene is the type 2 galanin receptor and is active in both ligand binding and signaling assays.