title: A novel assay of Gi/o-linked G protein-coupled receptor coupling to potassium channels provides new insights into the pharmacology of the group III metabotropic glutamate receptors.
abstract: The group III metabotropic glutamate receptors (mGluRs) represent a family of presynaptically expressed G-protein-coupled receptors (GPCRs) with enormous therapeutic potential; however, robust cellular assays to study their function have been difficult to develop. We present here a new assay, compatible with traditional high-throughput screening platforms, to detect activity of pharmacological ligands interacting with G(i/o)-coupled GPCRs, including the group III mGluRs 4, 7, and 8. The assay takes advantage of the ability of the Gbetagamma subunits of G(i) and G(o) heterotrimers to interact with G-protein regulated inwardly rectifying potassium channels (GIRKs), and we show here that we are able to detect the activity of multiple types of pharmacophores including agonists, antagonists, and allosteric modulators of several distinct GPCRs. Using GIRK-mediated thallium flux, we perform a side-by-side comparison of the activity of a number of commercially available compounds, some of which have not been extensively evaluated because of the previous lack of robust assays at each of the three major group III mGluRs. It is noteworthy that several compounds previously considered to be general group III mGluR antagonists have very weak activity using this assay, suggesting the possibility that these compounds may not effectively inhibit these receptors in native systems. We anticipate that the GIRK-mediated thallium flux strategy will provide a novel tool to advance the study of G(i/o)-coupled GPCR biology and promote ligand discovery and characterization.
authors.affiliation: 1215 MRB IV, Department of Pharmacology, Vanderbilt University, Nashville, TN 37232, USA. colleen.niswender@vanderbilt.edu
authors.forename: Colleen M
authors.initials: CM
authors.lastname: Niswender
authors.forename: Kari A
authors.initials: KA
authors.lastname: Johnson
authors.forename: Qingwei
authors.initials: Q
authors.lastname: Luo
authors.forename: Jennifer E
authors.initials: JE
authors.lastname: Ayala
authors.forename: Caroline
authors.initials: C
authors.lastname: Kim
authors.forename: P Jeffrey
authors.initials: PJ
authors.lastname: Conn
authors.forename: C David
authors.initials: CD
authors.lastname: Weaver
mesh_terms: D000322:Adrenergic Agonists / Q000494:pharmacology; D000494:Allosteric Regulation / Q000187:drug effects; D000596:Amino Acids / Q000494:pharmacology; D000818:Animals; D001681:Biological Assay / Q000379:methods*; D002217:Carbachol / Q000494:pharmacology; D002460:Cell Line; D004305:Dose-Response Relationship, Drug; D051676:G Protein-Coupled Inwardly-Rectifying Potassium Channels / Q000037:antagonists & inhibitors / Q000378:metabolism*; D019206:GTP-Binding Protein alpha Subunits, Gi-Go; D006801:Humans; D015640:Ion Channel Gating / Q000187:drug effects; D008024:Ligands; D018721:Muscarinic Agonists / Q000494:pharmacology; D037342:Pertussis Toxin / Q000494:pharmacology; D051381:Rats; D018094:Receptors, Metabotropic Glutamate / Q000819:agonists / Q000037:antagonists & inhibitors / Q000378:metabolism*; D011976:Receptors, Muscarinic / Q000378:metabolism; D015203:Reproducibility of Results; D013793:Thallium / Q000378:metabolism; D014966:Xanthenes / Q000494:pharmacology
chemical_list: D000322:Adrenergic Agonists; D000596:Amino Acids; D051676:G Protein-Coupled Inwardly-Rectifying Potassium Channels; C114624:LY 341495; D008024:Ligands; D018721:Muscarinic Agonists; D018094:Receptors, Metabotropic Glutamate; D011976:Receptors, Muscarinic; D014966:Xanthenes; C102718:metabotropic glutamate receptor 3; C111435:metabotropic glutamate receptor 8; D002217:Carbachol; D013793:Thallium; D037342:Pertussis Toxin; D019206:GTP-Binding Protein alpha Subunits, Gi-Go; C108823:metabotropic glutamate receptor 4