文摘
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<sup>–sup> 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<sup>2+sup>-permeable so as to induce neuro-excitotoxic cell death or minimally Ca<sup>2+sup>-permeable so as to depolarize neurons with minimal excitotoxic risk. Our constructs were based on the human α1 glycine receptor Cl<sup>–sup> channel due to its high conductance, human origin, high ivermectin sensitivity (once mutated), and because pore mutations that render it permeable to Na<sup>+sup> alone or Na<sup>+sup> plus Ca<sup>2+sup> are well characterized. We developed a Ca<sup>2+sup>-impermeable excitatory receptor by introducing the F207A/P-2′Δ/A-1′E/T13′V/A288G mutations and a Ca<sup>2+sup>-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.
