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)和具体的挑战主题，06-MH-103神经科学研究的新技术。该项目开发工具来解决两个假设：第一，局部脑区或亚区域中的特定类型的神经元可以被改变，以便它们容易受到系统输送的低毒化学品或药物的可逆失活或激活；第二，神经元的发育、分化和迁移可以通过操纵离子通量来研究，包括钙离子通量，这将导致不同的信号转导事件。将产生可调节的、可逆的、细胞特异性的、电生理上可测量的、对神经元兴奋性和神经元内钙激活的信号转导进行非抗原操纵的试剂。阿维菌素受体(AVMRs)是一种配体激活的小通道，可以在靶神经元中表达，这方面的开发将继续进行。这些通道将通过系统地突变现有的五聚体Cys-loop受体的亚单位来开发：线虫GluClα和β亚单位，以及人类甘氨酸受体。这些通道对内源性配体(如神经递质)不敏感，但将被伊维菌素及其类似物剂量依赖地激活，伊维菌素及其类似物是广泛使用的药物，可以口服或外周注射。渗透氯的AVMR-Cl将被用作构建这些通道的AVMR-Na和AVMR-Ca、钠和钙渗透版本的起点。AVMR系统的“治疗或研究指标”将得到改善，方法是找到一种中枢神经系统副作用较少的AVM类似物，提高现有优化的GluCl亚单位的AVM敏感性，或增加现有GluCl亚单位的通道开放时间或电导，同时保持可逆性。该项目还将通过产生一种基于人类甘氨酸受体的版本来避免对AVMR蛋白的免疫反应。AVMR将用于许多脊椎动物物种的研究：将对电路分析、控制分化、神经发生和迁移、兴奋性毒性模型和神经胶质激活的研究产生相当大的影响。治疗效果包括“药理脑深部刺激”；神经保护；以及超越中枢神经科学的其他用途。 与公共健康相关：该项目继续使用受体离子通道，以获得对神经元如何在回路中连接以及这些回路如何控制行为的新见解。我们将设计新的受体通道，这种通道只对阿维菌素类药物起反应，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，这种既涉及基因疗法，又涉及FDA批准的一套药物的程序，也可能有助于使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.