Next-gen Opto-GPCRs: spatiotemporal simulation of neuormodulator signaling

下一代 Opto-GPCR：神经调节信号传导的时空模拟

• 批准号: 9815886
• 负责人: Michael R. Bruchas
• 金额: $ 103.87万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-11-01 至 2020-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9815886
• 关键词: Next; gen; Opto; GPCRs; spatiotemporal

Project Summary/Abstract: The emerging field of optogenetics — using light to engage biological systems — holds tremendous promise for dissection of neural circuits, cellular signaling and manipulating neurophysiological systems in awake, behaving animals. However, the technological limits for implementing optogenetics in dissecting neuromodulators in awake, freely-moving behavior is clear while working with paradigms that require discrete spatiotemporal control of receptor signaling and when investigating neural circuits that have very small diverse, “hard to reach” architecture, such as heterogeneous brain nuclei. To engage neuropharmacological receptor substrates, neuroscientists in nearly every field use cannulas (simple metal tubes) and have more recently adopted tethered fiber optics for in vivo optogenetics to control local release of neuromodulator monoamine or neuropeptides. Unfortunately, these current methods are rather limited and difficult to implement because they severely limit the spatiotemporal control over receptor signaling pathways in discrete cell types. Moreover, current technology lacks a full tool box for multiplexed, subcellular, spatiotemporal control of G protein coupled receptor signaling, the predominant means for neuromodulator communication in the brain. For these reasons, an innovative effort combining neuroscience with biochemistry and pharmacology was necessary in order to bring spatial-temporal in vitro and in vivo control over GPCR- neuromodulator signaling. Therefore, here we directly address the central goals of this RFA-NS-16-775 in the following manner. The central goal of this proposal is to develop a cutting-edge v2.0 Opto-XR receptors that spatially and temporally control neuromodulator signaling in vitro and in freely moving animals. We have proposed an uniquely integrated approach to achieve this goal that brings pharmacologists, physiologists, biochemists, and neuroscientists together in a unique parallel manner. In the two specific aims we will develop and test these novel tools in vitro and in vivo: 1) To develop mutant Gi and Gs, Opto-XR v2.0 receptors with greater signaling dynamics and altered color spectra and sensitivity using structure-function analyses and thorough in vitro characterization; and 2) To develop utility and characterize Gi and Gs versions of Opto-XR v2.0 constructs in vivo and in models of freely-moving behavior using both traditional and wireless optogenetic approaches. Successful completion of the proposal will provide the wider community of neuroscience with a long awaited spatiotemporal manipulation of GPCRs – neuromodulator signaling within neural circuits in awake freely behaving animals. This new technology will also further widen the field for approaches that are capable of discrete control and optodynamic simulation of neuromodulator function in brain tissue.

项目摘要/摘要：光遗传学的新兴领域——利用光参与生物系统 - 为神经回路的解剖、细胞信号传导和操纵带来了巨大的希望 清醒、行为动物的神经生理系统。然而，实施的技术限制 光遗传学在清醒、自由移动行为中解剖神经调节剂的作用是清楚的，同时与 需要对受体信号进行离散时空控制以及研究神经系统时的范例 具有非常小、多样化、“难以到达”架构的电路，例如异构脑核。到 涉及神经药理学受体底物，几乎每个领域的神经科学家都使用插管（简单 金属管），最近采用系留光纤进行体内光遗传学来控制局部 释放神经调节剂单胺或神经肽。不幸的是，这些当前的方法相当 有限且难以实施，因为它们严重限制了对受体信号传导的时空控制 离散细胞类型中的途径。此外，当前的技术缺乏用于多重、亚细胞、 G 蛋白偶联受体信号传导的时空控制，神经调节剂的主要手段 大脑中的沟通。由于这些原因，将神经科学与生物化学相结合的创新努力 为了在体外和体内对 GPCR 进行时空控制，药理学是必要的 神经调节信号传导。因此，在这里我们直接解决 RFA-NS-16-775 的中心目标 以下方式。该提案的中心目标是开发一种尖端的 v2.0 Opto-XR 受体 在体外和自由活动的动物中空间和时间控制神经调节信号。我们有 提出了一种独特的综合方法来实现这一目标，该方法使药理学家、生理学家、 生物化学家和神经科学家以独特的并行方式聚集在一起。我们将制定两个具体目标 并在体外和体内测试这些新工具：1) 开发突变型 Gi 和 Gs、Opto-XR v2.0 受体 使用结构功能分析增强信号动态并改变色谱和灵敏度 彻底的体外表征； 2) 开发实用程序并表征 Opto-XR 的 Gi 和 Gs 版本 v2.0 使用传统和无线光遗传学构建体内和自由移动行为模型 接近。该提案的成功完成将为更广泛的神经科学界提供 期待已久的 GPCR 时空操纵——清醒状态下神经回路内的神经调节信号传导 行为自由的动物。这项新技术还将进一步拓宽能够实现的方法的领域 脑组织神经调节功能的离散控制和光动力学模拟。

项目成果

Optogenetic approaches for dissecting neuromodulation and GPCR signaling in neural circuits.

• DOI: 10.1016/j.coph.2016.11.001
• 发表时间: 2017-02
• 期刊: Current opinion in pharmacology
• 影响因子: 4
• 作者: Spangler SM;Bruchas MR
• 通讯作者: Bruchas MR