title: Cloning and characterization of a metabotropic glutamate receptor, mGluR4b.
abstract: An alternative spliced variant of metabotropic glutamate receptor subtype mGluR4a, termed mGluR4b was isolated from a rat cDNA library. Subtype mGluR4b was identical to the previously described mGluR4a, except for the last 64 amino acids in the C-terminal region in which were replaced by 135 new amino acids in mGluR4b. Recombinant baculoviruses coding for mGluR4a and mGluR4b were expressed in Spodoptera frugiperda, Sf-9, insect cells and characterized pharmacologically by measuring [3H]-L-2-amino-4-phosphonobutyrate ([3H]-L-AP4) binding and second messenger formation. [3H]-L-AP4 binding to membranes prepared from Sf-9 cells expressing mGluR4a and mGluR4b revealed respective affinities (Kd) of 480 and 360 nM and maximal binding densities (Bmax) of 4.2 and 0.8 pmol/mg protein. The ligand selectivity of mGluR4a and mGluR4b was similar: L-AP4 > L-serine-O-phosphate > L-glutamate > L-2-amino 2-methyl-4-phosphonobutyrate > (1S,3R)-1-aminocyclopentane-1,3-dicarboxylate > or = quisqualate. A decrease in the affinity of [3H]-L-AP4 was observed in the presence of 0.1 mM guanosine 5'-O-(3-thio)trisphosphate-gamma-S, indicating that mGluR4a and mGluR4b were functionally coupled to G-proteins in Sf-9 cells. Agonists of mGluR4 caused a minor decrease in forskolin-induced cAMP formation in Sf-9 cells expressing either mGluR4a or mGluR4b, suggesting that both receptors are coupled to adenylate cyclase in an inhibitory manner. Thus, mGluR4a and mGluR4b share a common signal transduction pathway and pharmacology when expressed in Sf-9 insect cells.
authors.affiliation: Department of Molecular Pharmacology, DK-2760 Målłv, Novo Nordisk A/S, Denmark. Cht@NOVO.DK
authors.forename: C
authors.initials: C
authors.lastname: Thomsen
authors.forename: R
authors.initials: R
authors.lastname: Pekhletski
authors.forename: B
authors.initials: B
authors.lastname: Haldeman
authors.forename: T A
authors.initials: TA
authors.lastname: Gilbert
authors.forename: P
authors.initials: P
authors.lastname: O'Hara
authors.forename: D R
authors.initials: DR
authors.lastname: Hampson
mesh_terms: D000595:Amino Acid Sequence; D000613:Aminobutyrates / Q000378:metabolism; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D003001:Cloning, Molecular; D000242:Cyclic AMP / Q000096:biosynthesis; D008969:Molecular Sequence Data; D051381:Rats; D018094:Receptors, Metabotropic Glutamate / Q000737:chemistry* / Q000187:drug effects / Q000378:metabolism
chemical_list: D000613:Aminobutyrates; D018094:Receptors, Metabotropic Glutamate; C107237:metabotropic glutamate receptor 4b; D000242:Cyclic AMP; C012729:2-amino-4-phosphonobutyric acid