Anion channelrhodopsin-based viral tools to manipulate brain networks in behaving animals

 描述(由申请人提供)：检查神经电路关键取决于激活或静音单个电路组件的能力，以随后评估它们对电路其他部分的影响及其对行为的影响。最近对病毒基因传递工具的改进使光遗传学方法能够根据特定的细胞类型、定位和连接来靶向细胞。靶向回路的生理解剖在小鼠大脑中非常成功，但在非人类灵长类动物大脑中的使用仍然有限。我们计划开发和测试新一代病毒工具，它将允许我们激活和 在非人类灵长类动物模型中抑制不同类型的细胞。为了实现我们的目标，我们组建了一个具有互补专业知识的专家团队，由一名生物化学家兼光生物学家(John Spudich)、一名分子神经学家(Roger Janz)以及一名系统和计算神经学家(Valentin Dragoi)组成。我们的方法建立在最近发现的阴离子传导通道视紫红质(ACRs)的基础上，ACR具有完美的阴离子选择性，其光敏性比目前的光遗传视紫红质大几个数量级，并使高效的神经元超极化。我们相信，我们的ACR结构将开启靶向神经抑制的新篇章。此外，我们将使用新的神经元激活(去极化)阳离子传导通道视紫红质(CCR)，它比常用的通道视紫红质-2具有~3倍的单位电导，从兴奋中恢复更快，以及更高的钠选择性。我们将构建编码ACR-CCR对的病毒载体，并使用光谱不同的ACR、ACR-ACR对，在大量人群中实现高效的波长选择神经元激活或抑制。这些病毒载体的有效性将在培养和原位培养的小鼠神经元以及行为正常的猴子的初级视皮层(V1)中进行测试。开发这些强大的工具对于探索非人类灵长类动物模型中的神经电路将是非常宝贵的，最终允许询问灵长类动物认知功能的微电路。

 DESCRIPTION (provided by applicant): Examining neural circuits crucially relies on the ability to activate or silence individual circuit components to subsequently assess their impact on other parts of the circuit and their influence on behavior. Recent refinements of viral tools for gene delivery have allowed optogenetic methods to target cells based on specific cell types, localization, and connectivity. The physiological dissection of targeted circuits has been extremely successful in the mouse brain, but remains of limited use in non-human primate brain. We plan to develop and test a new generation of viral tools that will allow us to both activate and suppress different cell types in non-human primate models. To accomplish our aims we have assembled an expert team with complementary expertise composed of a biochemist and photobiologist (John Spudich), a molecular neuroscientist (Roger Janz), and a systems and computational neuroscientist (Valentin Dragoi). Our approach builds upon recently discovered anion-conducting channelrhodopsins (ACRs), which perform with perfect anion selectivity, photosensitivity orders of magnitude greater than current optogenetic rhodopsins, and enable highly efficient neuron hyperpolarization. We believe that our ACR constructs will open a new chapter in targeted neuro-suppression. In addition, we will use new neuron-activating (depolarizing) cation-conducting channelrhodopsins (CCRs) that have ~3-fold greater unitary conductance, faster recovery from excitation, and higher sodium selectivity than the commonly used channelrhodopsin-2. We will construct viral vectors encoding ACR-CCR pairs and, using spectrally different ACRs, ACR-ACR pairs, enabling efficient wavelength-selected neuron activation or suppression in large populations. The effectiveness of these viral vectors will be tested in cultured and in situ mouse neurons and in the primary visual cortex (V1) of behaving monkeys. Developing these powerful tools will be invaluable for probing neural circuits in non-human primate models, finally allowing the interrogation of microcircuits underlying primate cognitive function.