Pharmacological Characterization and Identification of Amino Acids Involved in the Positive Modulation of Metabotropic Glutamate Receptor Subtype 2

Abstract

In the present study, we describe the characterization of a positive allosteric modulator at metabotropic glutamate subtype 2 receptors (mGluR2). 35 S]GTP␥S stimulation. This effect of LY487479 was also observed to a greater extent on the concentration-response curves to selective hmGluR2/3 agonists. In radioligand binding studies to rat cortical membranes, LY487379 increased the affinity of the radiolabeled agonist, [ 3 H]DCG-IV, without affecting the binding affinity of the radiolabeled antagonist, [ N-(4-(2-Methoxyphenoxy)-phenyl-N-(2,2,2-trifluoroethylsulfonyl)-pyrid-3-ylmethylamine3 H]LY341495. In rat hippocampal slices, coapplication of LY487379 potentiated synaptically evoked mGluR2 responses. Finally, to elucidate the site of action, we systematically exchanged segments and single amino acids between hmGluR2 and hmGluR3. Substitution of Ser688 and/or Gly689 in transmembrane IV along with Asn735 located in transmembrane segment V, with the homologous amino acids of hmGluR3, completely eliminated LY487379 allosteric modulation of hmGluR2. We propose that this allosteric binding site defines a pocket that is different from the orthosteric site located in the amino terminal domain.G-protein-coupled receptors (GPCRs) are a family of membrane bound proteins that play a central role in the recognition and signal transduction of neurotransmitters. It is generally believed that binding of the agonist induces a conformational change in the intracellular domain responsible for G-protein activation, thereby initiating a cascade of signaling events in the cell. In contrast, competitive antagonist binding will stabilize the inactive conformation of the receptor and block agonist-induced conformational changes and signal transduction (Gether and Kobilka, 1998). Although the exact mechanism of receptor/G-protein interaction is still unclear, GPCRs share a common motif of seven transmembrane helices connected by intra-and extracellular loops, an extracellular amino terminus, and a cytoplasmic carboxyl terminus. Based on their sequence homology, GPCRs have been subdivided into five families (Pin et al., 1994). Family 3 comprises the calcium-sensing receptors, pheromones, GABA B , and metabotropic glutamate (mGlu) receptors. To date, eight mGlu receptor subtypes have been cloned and classified into three groups based on their primary sequence, second messenger coupling, and pharmacology. Signal transduction from group I receptor subtypes B.A.R. and S.M. participated equally in this work.

Interdomain movements in metabotropic glutamate receptor activation

“…In addition, the mutations did not prevent the 7TM domain from activating G proteins. The positive allosteric modulator (PAM), LY487379, which binds to the 7TM (26,27), activated all mutants except the C500A mutant (Fig. 1C); this is in agreement with our previous reports that mGluR PAMs become full agonists when the 7TM is disconnected from the VFT (24,28).…”

Pharmacological Characterization of a New, Orally Active and Potent Allosteric Metabotropic Glutamate Receptor 1 Antagonist, 4-[1-(2-Fluoropyridin-3-yl)-5-methyl-1H-1,2,3-triazol-4-yl]-N-isopropyl-N-methyl-3,6-dihydropyridine-1(2H)-carboxamide (FTIDC)

“…FTIDC (Litschig et al, 1999;Pagano et al, 2000;Maj et al, 2003;Malherbe et al, 2003;Schaffhauser et al, 2003). The activity of FTIDC toward chimeric mGluR5(680)1a was similar to that toward wildtype mGluR1a.…”

Evaluation of Four Pyridine Analogs to Characterize 6-OHDA-Induced Modulation of mGluR5 Function in Rat Brain Using microPET Studies

“…These receptors are affected on several intracellular signal transduction mechanisms through G-proteins, having various physiological functions in the mammalian central nervous system (Bruno et al, 2001). According to their signal transduction mechanism, amino-acid sequence homologies, and pharmacological properties, metabotropic glutamate receptors have been identified as having eight subtypes, divided into three groups (Conn and Pin, 1997;Bhave et al, 2003;Schaffhauser et al, 2003). The metabotropic glutamate subtype-5 receptors (mGluR5s) are included in the first group.…”