title: 2-Guanidine-4-methylquinazoline acts as a novel competitive antagonist of A type γ-aminobutyric acid receptors.
abstract: The pentameric A type γ-aminobutyric acid receptors (GABAARs) are the major inhibitory neurotransmitter receptors in the nervous system and have long been considered as important pharmaceutical targets for the treatment of multiple neurological or psychological disorders. Here, we show that 2-guanidine-4-methylquinazoline (GMQ), a recently identified acid-sensing ion channel (ASIC) modulator, strongly and preferentially inhibits GABAAR among the major neurotransmitter-gated ion channels in cultured rat hippocampal neurons. GMQ inhibited GABA (1 μM)-induced currents in a competitive manner, with an IC50 (0.39±0.05 μM) comparable to that of bicuculline. Schild analysis revealed a slope of 1.04±0.06 for GMQ on α1β2 GABAARs expressed in HEK293T cells. Single-channel analysis showed that GMQ decreased open probability of GABAARs without affecting conductance. Moreover, GMQ inhibited GABAergic neurotransmission in hippocampal neurons, while having no significant effect on the basal field excitatory postsynaptic potentials (fEPSPs) and the intrinsic excitability of neurons. Using site-directed mutagenesis, we further demonstrated that mutations at Glu155 of β2 subunit and Phe64 of α1 subunit, both located inside the GABA binding pocket, profoundly decreased the sensitivity of the receptor to both GABA and GMQ. Interestingly, these mutations did not significantly affect the inhibition by amiloride, a diuretic structurally similar to GMQ and a known GABAAR inhibitor. We conclude that GMQ represents a novel chemical structure that acts, possibly, by competing with GABA binding to GABAARs. It is anticipated that GMQ and its analogs will facilitate the development of new chemical probes for GABAARs.
authors.affiliation: Institute of Neuroscience, State Key Laboratory of Neuroscience, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031, China; Department of Anatomy and Embryology, Shanghai Key Laboratory for Reproductive Medicine, Institute of Medical Sciences, Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China; Department of Biochemistry and Molecular Cell Biology, Shanghai Key Laboratory for Tumor Microenvironment and Inflammation, Institute of Medical Sciences, Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China.
authors.forename: Xian
authors.initials: X
authors.lastname: Xiao
authors.affiliation: Department of Integrative Biology and Pharmacology, The University of Texas Health Science Center at Houston, Houston, TX 77030, USA.
authors.forename: Michael X
authors.initials: MX
authors.lastname: Zhu
authors.affiliation: Department of Anatomy and Embryology, Shanghai Key Laboratory for Reproductive Medicine, Institute of Medical Sciences, Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China; Department of Biochemistry and Molecular Cell Biology, Shanghai Key Laboratory for Tumor Microenvironment and Inflammation, Institute of Medical Sciences, Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China. Electronic address: xu-happiness@shsmu.edu.cn.
authors.forename: Tian-Le
authors.initials: TL
authors.lastname: Xu
mesh_terms: D000109:Acetylcholine / Q000494:pharmacology; D000200:Action Potentials / Q000187:drug effects / Q000235:genetics; D000818:Animals; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D004622:Embryo, Mammalian; D018683:Excitatory Amino Acid Agents / Q000494:pharmacology; D018682:GABA Agents / Q000494:pharmacology*; D005998:Glycine / Q000494:pharmacology; D006146:Guanidines / Q000494:pharmacology*; D006624:Hippocampus / Q000166:cytology; D006801:Humans; D066298:In Vitro Techniques; D009154:Mutation / Q000235:genetics; D009474:Neurons / Q000187:drug effects*; D021122:Protein Subunits / Q000235:genetics / Q000378:metabolism; D011799:Quinazolines / Q000494:pharmacology*; D051381:Rats; D017207:Rats, Sprague-Dawley; D011963:Receptors, GABA-A / Q000235:genetics / Q000378:metabolism*; D005680:gamma-Aminobutyric Acid / Q000494:pharmacology
chemical_list: C106067:2-guanidine-4-methylquinazoline; D018683:Excitatory Amino Acid Agents; D018682:GABA Agents; D006146:Guanidines; D021122:Protein Subunits; D011799:Quinazolines; D011963:Receptors, GABA-A; D005680:gamma-Aminobutyric Acid; D000109:Acetylcholine; D005998:Glycine
keywords: 2-guanidine-4-methylquinazoline; 6-cyano-7-nitroquinoxaline-2,3-dione; A type γ-aminobutyric acid receptor; ACSF; ACh; AMPAR; ASIC; Bic; CNQX; Competitive antagonist; D(−)-2-amino-5-phosphonopentanoic acid; D-AP5; DMEM; DR; Dulbecco's Modified Eagle's Medium; GABA; GABA(A) receptor; GABA(A)R; GLIC; GMQ; Gloeobacter ligand-gated ion channel; HBSS; HEPES; Hank's Buffered Salt Solution; I-V; N-hydroxyethylpiperazine-N-2-ethanesulphonic acid; N-methyl-D-aspartic acid receptor; NMDAR; PS; TTX; acetylcholine; acid-sensing ion channel; artificial cerebrospinal fluid; bicuculline-methobromide; current-voltage; dose ratio; fEPSPs; field excitatory postsynaptic potentials; mIPSC; miniature inhibitory postsynaptic current; nAChR; nicotinic acetylcholine receptor; population spike; tetrodotoxin; α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor; γ-aminobutyric acid