Department of Neuroscience, Physiology and Pharmacology, University College London, Gower Street, London, WC1E 6BT, UK.
Department of Biomedicine, Neurophysiology Laboratory, Medical School, Institute of Neurosciences, University of Barcelona, Casanova 143, 08036, Barcelona, Spain.
Nat Commun. 2019 Sep 20;10(1):4312. doi: 10.1038/s41467-019-12280-9.
Desensitization is a canonical property of ligand-gated ion channels, causing progressive current decline in the continued presence of agonist. AMPA-type glutamate receptors (AMPARs), which mediate fast excitatory signaling throughout the brain, exhibit profound desensitization. Recent cryo-EM studies of AMPAR assemblies show their ion channels to be closed in the desensitized state. Here we present evidence that homomeric Q/R-edited AMPARs still allow ions to flow when the receptors are desensitized. GluA2(R) expressed alone, or with auxiliary subunits (γ-2, γ-8 or GSG1L), generates large fractional steady-state currents and anomalous current-variance relationships. Our results from fluctuation analysis, single-channel recording, and kinetic modeling, suggest that the steady-state current is mediated predominantly by conducting desensitized receptors. When combined with crystallography this unique functional readout of a hitherto silent state enabled us to examine cross-linked cysteine mutants to probe the conformation of the desensitized ligand binding domain of functioning AMPAR complexes.
脱敏是配体门控离子通道的一种典型特性，会导致在激动剂持续存在的情况下，电流逐渐下降。在大脑中起快速兴奋信号传递作用的 AMPA 型谷氨酸受体 (AMPAR) 会发生深度脱敏。最近关于 AMPAR 组装体的冷冻电镜研究表明，它们的离子通道在脱敏状态下是关闭的。在这里，我们提供的证据表明，当受体脱敏时，同型 Q/R 编辑的 AMPAR 仍然允许离子流动。单独表达的 GluA2(R)，或与辅助亚基 (γ-2、γ-8 或 GSG1L) 一起表达，会产生大的分数稳态电流和异常的电流方差关系。我们从波动分析、单通道记录和动力学建模中得到的结果表明，稳态电流主要由传导脱敏的受体介导。当与晶体学结合时，这种对沉默状态的独特功能读出使我们能够检查交联半胱氨酸突变体，以探测功能 AMPAR 复合物的脱敏配体结合域的构象。
