Galanin receptor and its ligands in the rat hippocampus

Receptors for the 29-amino-acid peptide, galanin, in membranes from the rat ventral hippocampus were examined using chloramine-T-iodinated porcine galanin as ligand. The equilibrium binding of 125I-galanin showed the presence of a high-affinity binding site (Kd = 1.91 +/- 0.40 nM). The concentration of the high-affinity-binding sites was 107 +/- 15 fmol/mg membrane protein. The on rate constant was estimated to be 2.6 +/- 0.1 M-1 min-1 at 37 degrees C. The affinity of rat galanin (differing in three amino acid residues from the porcine protein) was equal to that of porcine galanin. The 125I--galanin-binding site is a trypsin-sensitive membrane protein, which is heat-denaturated at 60 degrees C within 5 min. The effect of GTP and its analogs and of pertussis-toxin-catalyzed ADP-ribosylation on the binding of 125I-galanin suggest that the galanin receptor is coupled to an inhibitory G protein (Gi protein). 127I-galanin was shown to be a ligand with affinity equal to that of galanin in displacing 125I-galanin. The 125I-galanin-binding site in the ventral hippocampus recognizes as a ligand the tryptic fragments 1-20 and 21-29 of rat galanin and the synthetic fragments 12-29, 18-29 and 21-29 of porcine galanin. None of these afforded full inhibition of the binding of fragment 1-29 of 125I-galanin at a concentration of 1 microM.