title: Amiloride and GMQ Allosteric Modulation of the GABA-A ρ1 Receptor: Influences of the Intersubunit Site.
abstract: Amiloride, a diuretic used in the treatment of hypertension and congestive heart failure, and 2-guanidine-4-methylquinazoline (GMQ) are guanidine compounds that modulate acid-sensing ion channels. Both compounds have demonstrated affinity for a variety of membrane proteins, including members of the Cys-loop family of ligand-gated ion channels, such as the heteromeric GABA-A αβγ receptors. The actions of these guanidine compounds on the homomeric GABA-A ρ1 receptor remains unclear, especially in light of how many GABA-A αβγ receptor modulators have different effects in the GABA-A ρ1 receptors. We sought to characterize the influence of amiloride and GMQ on the human GABA-A ρ1 receptors using whole-cell patch-clamp electrophysiology. The diuretic amiloride potentiated the human GABA-A ρ1 GABA-mediated current, whereas GMQ antagonized the receptor. Furthermore, a GABA-A second transmembrane domain site, the intersubunit site, responsible for allosteric modulation in the heteromeric GABA-A receptors mediated amiloride's positive allosteric actions. In contrast, the mutation did not remove GMQ antagonism but only changed the guanidine compound's potency within the human GABA-A ρ1 receptor. Through modeling and introduction of point mutations, we propose that the GABA-A ρ1 intersubunit site plays a role in mediating the allosteric effects of amiloride and GMQ.
authors.affiliation: Department of Pharmacology and Neuroscience (H.D.S., E.B.G.), Institute for Aging and Alzheimer's Disease Research (E.B.G.), and Cardiovascular Research Institute (E.B.G.), University of North Texas Health Science Center, Fort Worth, Texas.
authors.forename: Heather D
authors.initials: HD
authors.lastname: Snell
authors.affiliation: Department of Pharmacology and Neuroscience (H.D.S., E.B.G.), Institute for Aging and Alzheimer's Disease Research (E.B.G.), and Cardiovascular Research Institute (E.B.G.), University of North Texas Health Science Center, Fort Worth, Texas eric.b.gonzales@unthsc.edu.
authors.forename: Eric B
authors.initials: EB
authors.lastname: Gonzales
mesh_terms: D000584:Amiloride / Q000494:pharmacology*; D000595:Amino Acid Sequence; D002478:Cells, Cultured; D004232:Diuretics / Q000494:pharmacology*; D006146:Guanidines / Q000494:pharmacology*; D006635:Histamine H2 Antagonists / Q000494:pharmacology*; D006801:Humans; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D018408:Patch-Clamp Techniques; D011799:Quinazolines / Q000494:pharmacology*; D011963:Receptors, GABA-A / Q000187:drug effects* / Q000235:genetics
chemical_list: C106067:2-guanidine-4-methylquinazoline; D004232:Diuretics; D006146:Guanidines; D006635:Histamine H2 Antagonists; D011799:Quinazolines; D011963:Receptors, GABA-A; D000584:Amiloride
references.citation: Mol Pharmacol. 1999 Mar;55(3):411-23
references.pmid: 10051524
references.citation: Proc Natl Acad Sci U S A. 1997 Sep 30;94(20):11031-6
references.pmid: 9380754
references.citation: Brain Res Dev Brain Res. 2005 Jan 1;154(1):15-23
references.pmid: 15617751
references.citation: Mol Pharmacol. 2005 Feb;67(2):470-9
references.pmid: 15509715
references.citation: Nucleic Acids Res. 2005 Jul 1;33(Web Server issue):W363-7
references.pmid: 15980490
references.citation: J Biol Chem. 2006 Aug 25;281(34):24455-61
references.pmid: 16754677
references.citation: Biochem Biophys Res Commun. 2006 Dec 1;350(4):900-4
references.pmid: 17034762
references.citation: J Pharmacol Exp Ther. 2007 Oct;323(1):236-47
references.pmid: 17636008
references.citation: Neurosci Lett. 2008 Jan 31;431(2):184-9
references.pmid: 18162311
references.citation: Pharmacol Rev. 2008 Sep;60(3):243-60
references.pmid: 18790874
references.citation: Neurochem Res. 2009 Oct;34(10):1704-11
references.pmid: 19387831
references.citation: Clin Exp Pharmacol Physiol. 2010 Apr;37(4):435-40
references.pmid: 19878215
references.citation: Nat Commun. 2013;4:1697
references.pmid: 23591864
references.citation: J Biol Chem. 2013 Jun 14;288(24):17420-31
references.pmid: 23640880
references.citation: Neuropharmacology. 2013 Dec;75:126-37
references.pmid: 23916476
references.citation: Nature. 2014 Aug 21;512(7514):333-7
references.pmid: 25143115
references.citation: PLoS One. 2014;9(10):e111470
references.pmid: 25347186
references.citation: Proc Natl Acad Sci U S A. 2000 Aug 1;97(16):9305-10
references.pmid: 10908659
references.citation: Eur J Neurosci. 2000 Oct;12(10):3575-82
references.pmid: 11029627
references.citation: J Biol Chem. 2002 Mar 15;277(11):9112-7
references.pmid: 11744711
references.citation: Mol Pharmacol. 2002 Jun;61(6):1322-8
references.pmid: 12021393
references.citation: Prog Neurobiol. 2002 Jun;67(2):113-59
references.pmid: 12126658
references.citation: Neurobiol Dis. 2002 Nov;11(2):298-307
references.pmid: 12505422
references.citation: Neurosci Lett. 2003 Jul 24;345(3):173-6
references.pmid: 12842284
references.citation: Future Med Chem. 2011 Feb;3(2):197-209
references.pmid: 21428815
references.citation: Nature. 2011 Jun 2;474(7349):54-60
references.pmid: 21572436
references.citation: PLoS One. 2011;6(9):e24892
references.pmid: 21949779
references.citation: Br J Pharmacol. 2004 Feb;141(4):717-27
references.pmid: 14732759
references.citation: Neurosci Lett. 2003 Sep 25;349(1):17-20
references.pmid: 12946576
references.citation: Vis Neurosci. 2012 May;29(3):157-68
references.pmid: 22564345
references.citation: J Pharmacol Exp Ther. 2004 Jun;309(3):1108-15
references.pmid: 14762104
references.citation: Acta Crystallogr D Biol Crystallogr. 2004 Aug;60(Pt 8):1355-63
references.pmid: 15272157
references.citation: Br J Pharmacol. 1988 Mar;93(3):679-83
references.pmid: 2453244
references.citation: Neuron. 1989 Mar;2(3):1195-205
references.pmid: 2483110
references.citation: EMBO J. 1992 Dec;11(12):4305-11
references.pmid: 1385113
references.citation: Br J Pharmacol. 1993 May;109(1):200-6
references.pmid: 8388298
references.citation: Receptors Channels. 1994;2(3):227-36
references.pmid: 7874449
references.citation: Proc Natl Acad Sci U S A. 1995 Dec 5;92(25):11756-60
references.pmid: 8524843
references.citation: Biophys J. 1995 Nov;69(5):1858-67
references.pmid: 8580329
references.citation: Receptors Channels. 1995;3(1):13-20
references.pmid: 8589989
references.citation: J Physiol. 1996 Dec 1;497 ( Pt 2):509-22
references.pmid: 8961191
references.citation: Vision Res. 1996 Dec;36(24):3997-4005
references.pmid: 9068852
references.citation: Nature. 1997 Sep 25;389(6649):385-9
references.pmid: 9311780
references.citation: Br J Pharmacol. 1999 Jun;127(3):601-4
references.pmid: 10401548