Division of Preclinical Research, Egis Pharmaceuticals PLC, Budapest, Hungary.
Division of Preclinical Research, Egis Pharmaceuticals PLC, Budapest, Hungary; Centre of Integrative Physiology, University of Edinburgh, Edinburgh, Scotland, UK.
Neuropharmacology. 2018 Jan;128:408-415. doi: 10.1016/j.neuropharm.2017.10.042. Epub 2017 Nov 3.
Gamma-amino butyric acid (GABA) is an abundant neurotransmitter in the CNS. GABAergic interneurons orchestrate pyramidal neurons in the cerebral cortex, and thus control learning and memory. Ionotropic receptors for GABA (GABAR) are heteropentameric complexes of α, β and γ integral membrane-protein subunits forming Cl -channels operated by GABA, which are vital for brain function and are important drug targets. However, knowledge on how GABAR bind GABA is controversial. Structural biology versus functional modelling combined with site-directed mutagenesis suggest markedly different roles for loop F of the extracellular domain of the α-subunit when complexed with GABA. Here, we report that contrary to the results of structural studies, loop F of the α-subunit controls the potency of GABA on GABAR. We examined the effect of replacing a short, variable segment of loop F of the GABA α5-subunit with the corresponding segment of the α2-subunit (GABAA5_LF2) and vice versa (GABAA2-LF5). When compared with their respective wild-type counterparts, GABAA5_LF2 receptors displayed enhanced sensitivity towards GABA, whilst in GABAA2-LF5 sensitivity was diminished. Mice homozygous for the genetic knock-in of the GABAA5_LF2 subunit showed a marked deficit in long- but not short-term object recognition memory. Working memory in place learning, spontaneous alternation and the rewarded T-maze were all normal. The deficit in long-term recognition memory was reversed by an α5-GABA negative allosteric modulator compound. The data show that loop F governs GABA potency in a receptor isoform-specific manner in vitro. Moreover, this mechanism of ligand recognition appears to be operative in vivo and impacts cognitive performance.
γ-氨基丁酸（GABA）是中枢神经系统中丰富的神经递质。GABA 能中间神经元协调大脑皮层中的锥体神经元，从而控制学习和记忆。GABA 的离子型受体（GABAR）是由α、β和γ整合膜蛋白亚基组成的异五聚体复合物，形成由 GABA 控制的 Cl-通道，对大脑功能至关重要，是重要的药物靶点。然而，关于 GABAR 如何结合 GABA 的知识存在争议。结构生物学与功能建模相结合，再加上定点突变，表明当与 GABA 结合时，α 亚基细胞外结构域的环 F 具有明显不同的作用。在这里，我们报告与结构研究的结果相反，α 亚基环 F 控制 GABA 对 GABAR 的效力。我们研究了用 GABA α5 亚基的环 F 的短可变片段替代相应的 α2 亚基（GABAA5_LF2）和反之亦然（GABAA2-LF5）的影响。与各自的野生型相比，GABAA5_LF2 受体对 GABA 的敏感性增强，而 GABAA2-LF5 的敏感性降低。GABAA5_LF2 亚基的基因敲入纯合子小鼠在长但不在短时间物体识别记忆中表现出明显缺陷。位置学习中的工作记忆、自发交替和奖励 T 迷宫都是正常的。长效识别记忆缺陷可通过 α5-GABA 负变构调节剂化合物逆转。数据表明，环 F 以受体同工型特异性的方式在体外控制 GABA 的效力。此外，这种配体识别机制似乎在体内起作用，并影响认知表现。
