Chemical Targeting of Sensors and Pharmacological Probes in the Brain

摘要 大脑中传感器和药物探针的化学靶向 没有通用的工具能够监测和/或调节哺乳动物的特定电路和细胞 用纯化学的方法。我们最近披露了一种用于化学输送电压的聚合物平台 首次发现对大脑中特定细胞类型(单胺能神经元及其延伸)敏感的染料(VSD) 一个非遗传系统的例子，能够在脑组织中定向输送高度亲脂的分子。我们有 研究表明，葡聚糖是一种细菌多糖，可以作为一种极性聚合物载体，动态地 将亲脂性VSD包裹起来，通过脑组织携带，并通过以下方式将其部署到特定类型的感兴趣细胞 寻的配基的手段。拟议的研究旨在建立在此概念验证示例的基础上并开发 将亲脂性传感器和致动器/药物输送到大脑中特定细胞类型的通用平台。在AIM 我们将开发基于配体导向的酰基咪唑化学的无痕迹共价标记技术。 使细胞类型的特异性靶向，而不会长期扰动系统的药理作用 配基，以及b)共价固定离子转运蛋白(AMPA受体)附近的电压传感器 和多巴胺转运体)，以测量局部膜电位的变化。在目标1B中，我们将利用 通过将平台扩展到双药物靶向来实现配基的药理作用 多巴胺受体D2激动剂对D2表达的中棘神经元(D2-MSN)的特异性，但不是 纹状体中的多巴胺能轴突，通过腺苷受体A2a拮抗剂(D2-MSN标记物)。 这将为细胞选择性药理学提供治疗神经退行性变(例如 帕金森氏症)和精神障碍。在目标2中，我们集中于a)优化用于高密度的荧光VSD 使用葡聚糖平台的灵敏度和目标性；以及b)将目标平台扩展到电压传感器 支持包括短波红外荧光显微镜和光声在内的新型成像方式 成像。在目标3中，我们将通过调节聚合物分子来系统地优化葡聚糖给药平台 重量和代用模式。我们还将探索其他以糖为基础的聚合物作为输送系统 大脑中的亲脂性物质，包括环糊精和聚苯乙烯-乙醚-海藻糖。这五年的结果是 该计划承诺提供一个通用的和优化的平台，将亲脂性传感器和执行器输送到 大脑中的特定细胞类型。化学靶向方法消除了对基因操作的需要 大脑，因此，从长远来看，将适用于人类和其他不容易的生物体 适应于基因操作。

SUMMARY Chemical Targeting of Sensors and Pharmacological Probes in the Brain There are no general tools that enable monitoring and/or modulation of specific circuits and cells in the mammalian brain by purely chemical means. We have recently disclosed a polymer platform for chemical delivery of voltage sensitive dyes (VSDs) to specific cell types in the brain (monoaminergic neurons and their extensions), as a first example of a non-genetic system enabling targeted delivery of highly lipophilic molecules in brain tissue. We have shown that dextran, a bacterial polysaccharide, can function as a polar polymer carrier which dynamically encapsulates the lipophilic VSD to carry it through brain tissue and deploy it at a specific cell type of interest by the means of a homing ligand. The proposed research aims to build on this proof-of-concept example and develop a general platform for delivery of lipophilic sensors and actuators/drugs to specific cell types in the brain. In Aim 1A we will develop traceless covalent labeling techniques based on ligand-directed acyl imidazole chemistry to a) enable cell-type specific targeting without long-term perturbation of the system by pharmacological effects of the ligand, and b) covalently immobilize voltage sensors in the vicinity of ion transporting proteins (AMPA receptor and dopamine transporter) to measure local changes of membrane potential. In Aim 1B, we will leverage the pharmacological effects of the ligands by extending the platform to dual-drug targeting which will enable delivery of dopamine receptor D2 agonists specifically to D2-expressing medium spiny neurons (D2-MSNs), but not dopaminergic axons in the striatum, by the means of an adenosine receptor A2A antagonist (a D2-MSN marker). This will provide a platform for cell selective pharmacology with therapeutic potential for neurodegenerative (e.g. Parkinson’s disease) and psychiatric disorders. In Aim 2 we focus on a) optimization of fluorescent VSDs for high sensitivity and targetability using the dextran platform; and b) extension of the targeting platform to voltage sensors enabling novel imaging modalities including short wave infrared fluorescence microscopy and photoacoustic imaging. In Aim 3 we will systematically optimize the dextran delivery platform by tuning the polymer molecular weight and substitution pattern. We will also explore other saccharide-based polymers as delivery systems for lipophilic cargo in the brain, including cyclodextrins and poly(styrene-ether-trehalose). The outcome of this 5-year program promises to provide a general and optimized platform for delivery of lipophilic sensor and actuators to specific cell types in the brain. The chemical targeting approach eliminates the need for genetic manipulation of the brain and thus will be, in a long-term perspective, applicable to humans and other organisms not readily adaptable for genetic manipulations.