Queensland Brain Institute, The University of Queensland , Brisbane, QLD 4072, Australia.
School of Biomedical Sciences, The University of Queensland , Brisbane, QLD 4072, Australia.
ACS Chem Neurosci. 2016 Dec 21;7(12):1647-1657. doi: 10.1021/acschemneuro.6b00168. Epub 2016 Sep 27.
The ability to control neuronal activation is rapidly advancing our understanding of brain function and is widely viewed as having eventual therapeutic application. Although several highly effective optogenetic, optochemical genetic, and chemogenetic techniques have been developed for this purpose, new approaches may provide better solutions for addressing particular questions and would increase the number of neuronal populations that can be controlled independently. An early chemogenetic neuronal silencing method employed a glutamate receptor Cl channel engineered for activation by 1-3 nM ivermectin. This construct has been validated in vivo. Here, we sought to develop cation-permeable ivermectin-gated receptors that were either maximally Ca-permeable so as to induce neuro-excitotoxic cell death or minimally Ca-permeable so as to depolarize neurons with minimal excitotoxic risk. Our constructs were based on the human α1 glycine receptor Cl channel due to its high conductance, human origin, high ivermectin sensitivity (once mutated), and because pore mutations that render it permeable to Na alone or Na plus Ca are well characterized. We developed a Ca-impermeable excitatory receptor by introducing the F207A/P-2'Δ/A-1'E/T13'V/A288G mutations and a Ca-permeable excitatory receptor by introducing the F207A/A-1'E/A288G mutations. The latter receptor efficiently induces cell death and strongly depolarizes neurons at nanomolar ivermectin concentrations.
控制神经元激活的能力正在迅速提高我们对大脑功能的理解，并且被广泛认为具有最终的治疗应用前景。尽管已经开发出几种非常有效的光遗传学、光化学遗传和化学遗传技术来实现这一目标，但新的方法可能会为解决特定问题提供更好的解决方案，并增加可以独立控制的神经元群体的数量。一种早期的化学遗传神经元沉默方法使用了一种谷氨酸受体 Cl 通道，该通道经过工程设计可被 1-3 nM 伊维菌素激活。该构建体已在体内得到验证。在这里，我们试图开发阳离子渗透性伊维菌素门控受体，要么最大程度地允许 Ca 渗透，以诱导神经兴奋性细胞死亡，要么最小程度地允许 Ca 渗透，以最小的兴奋性风险去极化神经元。我们的构建体基于人α1 甘氨酸受体 Cl 通道，因为它具有高电导率、人类起源、高伊维菌素敏感性（一旦突变），并且可以很好地描述使其对 Na 或 Na 加 Ca 具有渗透性的孔突变。我们通过引入 F207A/P-2'Δ/A-1'E/T13'V/A288G 突变开发了一种非钙渗透性兴奋性受体，通过引入 F207A/A-1'E/A288G 突变开发了一种钙渗透性兴奋性受体。后者受体在纳摩尔伊维菌素浓度下有效诱导细胞死亡并强烈去极化神经元。
