Adaptive optical microscopy for high-accuracy recording of neural activity in vivo

项目摘要 为了理解大脑中的计算，我们需要高精度地监测神经回路的活动， 这需要具有高空间和时间分辨率的方法。非侵入性，能够解决 亚细胞结构，光学显微镜已广泛应用于神经科学领域，具有多种 在高速、大深度和/或大空间尺度上对神经活动进行成像的方法。为 例如，自由空间角啁啾增强延迟双光子荧光显微镜被开发用于记录 在kHz帧速率下，在体内脑中的膜电压。三光子荧光显微镜，一种新兴的 使用比双光子荧光显微术更长波长的激发光的方法具有大的 穿透深度，并且能够对小鼠大脑中超过1 mm深的结构进行成像。的替代 点扫描多光子荧光显微镜以上，单光子宽场荧光显微镜 也被应用于大脑活动的体内监测。最常见的是，整个样品被照亮 并且发射的荧光由物镜透镜收集并用照相机成像，这使得能够快速活动 表面结构的成像，有时横向尺寸超过毫米。获得准确 然而，在活体神经活动的测量中，必须对抗神经元的分辨能力的退化。 这些显微镜方法应用于脑组织时。光学不均匀性的生物 组织本身会扭曲成像光，并阻止所有显微镜模式实现其设计 体内性能。当应用于活动成像时，这种退化可能导致错误的结论。在这里， 我们建议优化和应用Ji实验室开发的自适应光学方法，以选择尖端的高性能， 速度，大深度和大规模的活动记录模式，用于高精度测量神经功能 体内活性。

PROJECT SUMMARY To understand the computations in the brain, we need to monitor the activity of neural circuits at high accuracy, which requires methodologies with high spatial and temporal resolution. Non-invasive and capable of resolving subcellular structures, optical microscopy has been extensively applied in the field of neuroscience, with a variety of methods developed to image neural activity at high speed, large depths, and/or over large spatial scales. For example, free-space angular-chirp-enhanced delay two-photon fluorescence microscopy was developed to record membrane voltage at kHz frame rate in the brain in vivo. Three-photon fluorescence microscopy, an emerging method that uses excitation light of longer wavelengths than two-photon fluorescence microscopy, has large penetration depths and is capable of imaging structures over 1-mm deep in the mouse brain. An alternative to the point-scanning multiphoton fluorescence microscopy above, single-photon widefield fluorescence microscopy has also been applied to in vivo monitoring of brain activity. Most commonly, the entire sample is illuminated and the emitted fluorescence collected by an objective lens and imaged with a camera, which enables fast activity imaging of superficial structures, sometimes over millimeters in lateral dimension. To obtain accurate measurements of neural activity in vivo, however, one has to combat the degradation of the resolving power of these microscopy methods when they are applied to brain tissue. The optical inhomogeneity of the biological tissue itself distorts the image-forming light and prevents all microscopy modalities from achieving their designed performance in vivo. When applied to activity imaging, such degradation can lead to erroneous conclusions. Here, we propose to optimize and apply adaptive optics methods developed in the Ji lab to select cutting-edge high- speed, large-depth, and large-scale activity recording modalities for high-accuracy measurements of neural activity in vivo.