All optical control and monitoring of neural activity

项目总结：光遗传神经活动干预的成功需要优化 将编码光敏蛋白(Opsins)的基因传递到特定的细胞，并记录 光发生刺激过程中细胞和组织的变化。最常用的方法是 传递视蛋白(S)是使用病毒载体，容易引起意想不到的炎症 反应、免疫反应和不适当的基因整合。此外，病毒方法 限制可包装和递送的质粒的大小，因此不能携带多个 视蛋白编码基因或大启动子。此外，在一些人类疾病的案例中，例如 视网膜色素变性(RP)周围视网膜发生进行性的光感受器丧失，它 将有助于定位opsins的表达，不仅在特定的细胞类型中，而且在 受限制的空间区域(在RP中为周边)。该提案的第一个目标是优化一种非病毒 减轻病毒传播带来的挑战的传播方法。我们最近使用了Focus 近红外超快激光束方法将视黄素(ChR2)输送到空间图案区 神经组织(视网膜)。但是，该技术需要进行优化，以最大限度地减少 有害的影响。此外，开发无标记光学技术将被证明是有用的(相比之下 到电生理学)无创地评估光遗传敏化的功能激活 具有高空间分辨率和大吞吐量的神经元。最近，我们演示了如何使用 相敏频域光学相干层析成像(PSFD-OCT) 光基因刺激细胞的波动。PSFD-OCT是一种基于 可探测几十个数量级位移的低相干干涉测量原理 皮克计。由于光的低相干长度，检测到的信号必须在 光源的相干长度(~10？m)。这使PSFD-OCT能够调查 纳米在组织体积的非常小的区域内变化，并且适合于高度 局部化检测。本研究的总体目标是优化基因编码的光学传递 OPTINS，并开发了基于PSFD-OCT的无标签非侵入性光学读出方法 监测激活引起的大脑皮层神经元和组织的变化。 1

Project Summary: Success of optogenetic intervention of neural activity requires optimization of delivery of genes encoding light sensitive proteins (opsins) to specific cells, and to record the changes in cells and tissue during optogenetic stimulation. The most-commonly used method for delivering opsin(s) is use of viral vector, which is prone to cause unexpected inflammatory responses, immunological reactions, and improper gene integration. Further, the viral methods limit the size of plasmid that can be packaged and delivered and therefore cannot carry multiple opsin-encoding genes or large promoters. Further, in several cases of human diseases such as retinitis pigmentosa (RP) where progressive loss of photoreceptors happens in peripheral retina, it will be useful to localize the expression of the opsins not only in specific cell types, but in a restricted spatial region (peripheral in RP). The first aim of the proposal is to optimize a non-viral delivery method to mitigate the challenges posed by viral delivery. We have recently used focused near-IR ultrafast laser beam method to deliver opsins (ChR2) into spatially-patterned regions of neural tissue (retina). However, this technique needs to be optimized so as to minimize the deleterious effects. Further, it will prove useful to develop a label-free optical technique (in contrast to electrophysiology) to non-invasively evaluate functional activation of optogenetically-sensitized neurons with high spatial resolution and large throughput. Recently, we demonstrated use of Phase-Sensitive Frequency Domain Optical Coherence Tomography (PSFD-OCT) for detection of fluctuations in optogenetically-stimulated cells. PSFD-OCT is a novel technique based on the principles of low-coherence interferometry that can detect displacements of the order of tens of picometers. Because of the low-coherence length of the light, the detected signal has to be within the coherence length of the light source (~10¿m). This enables PSFD-OCT to investigate sub- nanometer changes within a very small region of the tissue volume and is suitable for highly localized detection. The overall aim of this study is to optimize optical delivery of gene encoding opsins, and develop label-free non-invasive optical readout method based on PSFD-OCT to monitor the changes in cortical neurons and tissue resulting from the activation. 1