Xiao Xian, Zhu Michael X, Xu Tian-Le
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.
Department of Integrative Biology and Pharmacology, The University of Texas Health Science Center at Houston, Houston, TX 77030, USA.
Neuropharmacology. 2013 Dec;75:126-37. doi: 10.1016/j.neuropharm.2013.07.018. Epub 2013 Aug 2.
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.
五聚体A型γ-氨基丁酸受体（GABAARs）是神经系统中主要的抑制性神经递质受体，长期以来一直被视为治疗多种神经或心理疾病的重要药物靶点。在此，我们表明，2-胍基-4-甲基喹唑啉（GMQ）是一种最近鉴定出的酸敏感离子通道（ASIC）调节剂，在培养的大鼠海马神经元的主要神经递质门控离子通道中，能强烈且优先抑制GABAAR。GMQ以竞争性方式抑制GABA（1μM）诱导的电流，其半数抑制浓度（IC50，0.39±0.05μM）与荷包牡丹碱相当。Schild分析显示，GMQ对在HEK293T细胞中表达的α1β2 GABAARs的斜率为1.04±0.06。单通道分析表明，GMQ降低了GABAARs的开放概率，而不影响电导。此外，GMQ抑制海马神经元中的GABA能神经传递，而对基础场兴奋性突触后电位（fEPSPs）和神经元的内在兴奋性没有显著影响。通过定点诱变，我们进一步证明，位于GABA结合口袋内的β2亚基的Glu155和α1亚基的Phe64处的突变，显著降低了受体对GABA和GMQ的敏感性。有趣的是，这些突变对氨氯地平的抑制作用没有显著影响，氨氯地平是一种结构与GMQ相似的利尿剂，也是一种已知的GABAAR抑制剂。我们得出结论，GMQ代表一种新的化学结构，其作用可能是通过与GABA竞争结合GABAARs。预计GMQ及其类似物将促进GABAARs新型化学探针的开发。
