Non-invasive Chemical Genetic Control of Neuronal Activity

描述（由申请人提供）：绘制功能电路是细胞和系统神经科学的主要目标。由于缺乏唤起细胞特异性神经活动的技术，目前绘制神经回路的方法受到限制。现有的神经刺激方法依赖于局部应用无区别的电流场、谷氨酸释放或人工感官刺激的呈现。尽管最近使用光门控离子通道提供了在快速时间尺度上对神经元活动的光学控制，但这种方法受到直接光学访问感兴趣的神经元群的要求的限制，并且目前不适合激活大的大脑区域或分散神经元群。神经科学的一项变革性技术将是对哺乳动物大脑中由基因决定的神经元群体的神经活动进行非侵入性控制。这样的目标需要结合神经元亚群的遗传敏感性和一种方法来远程操纵它们的电活动，而不需要手术或颅内植入。为了创造这样的技术，我的实验室已经启动了一个体内化学遗传和生理研究项目，以设计一个小鼠模型，适合对体内遗传定义的神经元群体的神经活动进行精确的非侵入性操作。我们开发了一种条件小鼠模型，通过异源受体（TRPV1）的细胞类型特异性表达，使遗传定义的神经元对人工配体（辣椒素）敏感。我们发现辣椒素对表达TRPV1的神经元的作用诱导了强大的内向电流，触发了强大的动作电位，并激活了刻板行为。利用这些初步数据，以及TRPV1广泛的药理学和生物物理特性，我们建议扩展和修改该模型，使外周给药激动剂能够激活指定的神经元亚群。此外，由于大TRPV1通道孔可渗透小分子，包括膜外钠通道阻滞剂QX-314，我们建议测试这种新型小鼠模型，以激活和抑制神经元活动。这项工作将允许开发一种新的体内技术，用于神经元活动的化学遗传调控，该技术：(1)与光学和光遗传学策略正交，(2)基于当前唯一的基于Cre/lox的神经元激活模型，(3)可以通过药物注射实现完全无创的中枢神经系统激活，(4)可以实现靶向小分子递送到定义的神经元亚群。公共卫生相关性：拟议的研究将开发一种新技术，用于非侵入性控制基因定义的脑细胞群体的电活动。大脑中异常的电活动会导致癫痫、记忆力下降、抑郁、自闭症、精神分裂症和成瘾。通过开发一种关键的新技术来有针对性地操纵脑细胞活动和代谢，拟议的研究将定义新的脑回路和治疗策略，以治疗这些破坏性的神经和精神疾病，这些疾病目前对公众健康产生了深远的负面影响。

DESCRIPTION (provided by applicant): Mapping functional circuits is a major goal for both cellular and systems neuroscience. Current approaches for mapping neural circuits are limited by the lack of technologies for evoking cell-specific neural activity. Available methods of neural stimulation rely on either local application of undiscriminating fields of electrical currents, glutamate uncaging, or the presentation of artificial sensory stimuli. Although recent use of light- gated ion channels has provided optical control of neuronal activity on rapid time scales, such approaches are limited by the requirement for direct optical access to neuronal populations of interest, and are not currently suitable for activating large brain areas or disperse neuronal populations. A transformative technology for neuroscience would be non-invasive control over neural activity in genetically defined populations of neurons in the mammalian brain. Such a goal requires combining genetic sensitization of neuronal subsets with a means to manipulate their electrical activity remotely without surgery or intracranial implants. To create such a technology, my laboratory has initiated a program of in vivo chemical genetic and physiological studies to engineer a mouse model suitable for precise non-invasive manipulation of neural activity in genetically defined populations of neurons in vivo. We have developed a conditional mouse model that sensitizes genetically defined neurons to an artificial ligand (capsaicin) by cell type-specific expression of a heterologous receptor (TRPV1). We have found that application of capsaicin to neurons expressing TRPV1 induces strong inward currents, triggers robust firing of action potentials, and activates stereotyped behaviors. Taking advantage of these preliminary data, and the extensive pharmacological and biophysical characterization of TRPV1, we propose to extend and modify this model to enable peripheral administration of agonists for central activation of defined neuronal subsets. Moreover, because the large TRPV1 channel pore is permeable to small molecules, including the membrane-impermeant sodium channel blocker QX-314, we propose to test this novel mouse model to enable both activation and inhibition of neuronal activity. This work will allow for the development of a novel in vivo technology for chemical genetic regulation of neuronal activity that is (1) orthogonal to optical and optogenetic strategies, (2) based on the only current Cre/lox-based model for neuronal activation, (3) may allow for fully non-invasive CNS activation by drug injection, and (4) may enable targeted small molecule delivery to defined neuronal subsets. PUBLIC HEALTH RELEVANCE: The proposed research will develop a novel technology for non-invasive control over electrical activity in genetically defined populations of brain cells. Abnormal electrical activity in the brain contributes to epilepsy, memory decline, depression, autism, schizophrenia, and addiction. By developing a crucial new technology for targeted manipulation of brain cell activity and metabolism, the proposed research will define novel brain circuits and therapeutic strategies for treating these devastating neurological and psychiatric disorders, which currently have a profound negative impact on public health.