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

 描述（由申请人提供）：检查神经回路关键依赖于激活或沉默各个电路组件的能力，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。最近对基因传递病毒工具的改进使得光遗传学方法能够根据特定的细胞类型、定位和连接性来靶向细胞。目标电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应用仍然有限。我们计划开发和测试新一代病毒工具，使我们能够激活和 抑制非人类灵长类动物模型中的不同细胞类型。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，由生物化学家和光生物学家 (John Spudich)、分子神经科学家 (Roger Janz) 以及系统和计算神经科学家 (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（ACR）的基础上，它具有完美的阴离子选择性，光敏性比当前的光遗传学视紫红质高出几个数量级，并且能够实现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）阳离子传导视紫红质通道 (CCR)，其单位电导比常用的视紫红质通道高出约 3 倍，从激发中恢复更快，并且钠选择性更高。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，在大量人群中实现有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的原位小鼠神经元以及行为猴子的初级视觉皮层（V1）中进行测试。开发这些强大的工具对于探测非人类灵长类动物模型中的神经回路将具有无价的价值，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.