Avermectin Receptors for Neuronal Engineering

描述（由申请人提供）：本申请涉及广泛的挑战领域（06）和特定的挑战主题，06-MH-103神经科学研究的新技术。该项目开发了解决两个假设的工具：第一，局部脑区域或次区域中的特定类型的神经元可以改变，以便它们对系统传递的低毒性化学品或药物的可逆失活或激活敏感;第二，神经元的发育、分化和迁移可以通过操纵离子流，包括钙流，然后导致不同的信号转导事件。将产生可以实现神经元兴奋性和神经元内Ca激活信号转导的可调节、可逆、细胞特异性、电生理学可测量和非抗原性操纵的试剂。将继续开发一个小的配体激活通道，阿维菌素受体（AVMR），可以在靶神经元中表达。这些通道将通过系统地突变现有五聚体Cys环受体的亚基来开发：C。elegans GluCl α和β亚基，以及人甘氨酸受体。这些通道对内源性配体（如神经递质）不敏感，但会被伊维菌素及其类似物剂量依赖性地激活，伊维菌素及其类似物是广泛使用的药物，可以口服或注射到外周。氯离子渗透性AVMR-Cl将用作构建AVMR-Na和AVMR-Ca、钠和钙渗透性通道的起点。AVMR系统的“治疗或研究指数”将通过发现具有更少CNS副作用的AVM类似物、通过增加现有优化GluCl亚基的AVM敏感性、或通过增加现有GluCl亚基的通道开放时间或电导、同时保持可逆性来改善。该项目还将通过产生基于人类甘氨酸受体的版本来避免对AVMR蛋白的免疫反应。AVMRs将用于许多脊椎动物物种的研究：对电路分析的研究将产生相当大的影响;控制分化，神经发生和迁移;兴奋性毒性模型;和胶质细胞活化。治疗影响包括“药理学脑深部刺激”;神经保护;以及其他超出CNS神经科学的用途。 公共卫生相关性：这个项目继续使用受体离子通道来获得神经元如何在电路内连接以及这些电路如何控制行为的新见解。我们将设计新的受体通道，只对药物，阿维菌素，可以在动物的饮食中传递。一旦这些受体被开发出来，就有可能研究激活或抑制选定的神经元如何影响行为。最终，这些涉及基因治疗和FDA批准的一系列药物的程序也可以帮助精神疾病中过于活跃或不够活跃的神经元正常化。

DESCRIPTION (provided by applicant): This application addresses broad Challenge Area (06) and specific Challenge Topic, 06-MH-103 New technologies for neuroscience research. This project develops tools to address two hypotheses: first that specific types of neurons in localized brain regions or subregions can be altered so that they are susceptible to reversible inactivation or activation in response to systemically delivered low-toxicity chemicals or drugs; and second, that neuronal development, differentiation, and migration can be studied via manipulating ion fluxes, including calcium fluxes, which then lead to diverse signal transduction events. Reagents will be generated that can achieve adjustable, reversible, cell-specific, electrophysiologically measurable, and non-antigenic manipulation of neuronal excitability, and of Ca-activated signal transduction within neurons. Development will continue on a small repertoire of ligand-activated channels, avermectin receptors (AVMRs), that can be expressed in target neurons. These channels will be developed by systematically mutating subunits of existing pentameric Cys-loop receptors: the C. elegans GluCl alpha and beta subunits, and the human glycine receptor. The channels will be insensitive to endogenous ligands (such as neurotransmitters), but will be activated dose-dependently by ivermectin and its analogs, widely used drugs that can be given orally or by injection into the periphery. The chloride-permeable AVMR-Cl will be used as a starting point for constructing AVMR-Na and AVMR-Ca, sodium- and calcium-permeable versions of these channels. The "therapeutic or research index" of the AVMR system will be improved by finding an AVM analog with fewer CNS side effects, by increasing the AVM sensitivity of the existing optimized GluCl subunits, or by increasing the channel open time or conductance of the existing GluCl subunits, while maintaining reversibility. The project will also avoid immune reactions to AVMR proteins by generating a human glycine receptor-based version. The AVMRs will be useful for research in many vertebrate species: there will be considerable impact for research on circuit analysis; controlling differentiation, neurogenesis, and migration; models for excitotoxicity; and glial activation. Therapeutic impacts include "pharmacological deep-brain stimulation"; neuroprotection; and other uses that extend beyond CNS neuroscience. PUBLIC HEALTH RELEVANCE: This project continues to use receptor ion channels to gain new insight into how neurons are connected within circuits and how such circuits control behavior. We will engineer new receptor channels that respond only to drugs, avermectins, that can be delivered in an animal's diet. Once these receptors are developed, it will be possible to study how activating or inhibiting selected neurons influences behavior. Ultimately, such procedures, involving both gene therapy and an FDA-approved set of drugs, could also help normalize neurons that are either too active, or not active enough, in psychiatric diseases.