Integrated compressive sensing microscope for high-speed functional biological imaging

从遗传指定的神经元直接读出尖峰和亚阈值电压活动 人口将促进我们对神经计算的理解的重大进展， 网络级。出于这个原因，开发一种遗传编码电压指示器（GEVI）， 足够的速度、膜定位和亮度来报告哺乳动物的动作电位 在过去的二十年里，细胞一直是神经科学的主要目标。最近，我们开发了一种 基于视蛋白的GEVI，称为执政官，它在多个物种的神经元中表现出良好的定位， 亮度比先前基于视蛋白的报告子提高数倍，数量级 电压敏感性和光漂白优于GFP样报告基因，以及 与光遗传学控制的相容性。然而，动作电位成像，即使在一个单一的 Archon表达神经元，需要最先进的sCMOS相机，以满足采样率和 信噪比（SNR）要求。这些设备体积大、耗电量大、带宽密集， 并支持最大帧速率下的有限视场。这使他们无法融入 并且极大地限制了它们对大量细胞成像的能力。 该提案的目标是创建一个头戴式微型显微镜（“微型镜”）， 能够对神经群进行高速（>1000帧/秒; FPS）电压成像， 移动的动物为此，我们将调整我们最近开发的集成压缩传感 CMOS图像传感器，具有逐像素曝光（“CS-PCE相机”），用于GEVI成像。CS- PCE相机的关键创新是允许传感器阵列上的每个像素独立曝光 而不是用帧时钟以锁步方式曝光整个阵列。这使得CS重建 高速视频的采样速度比香农/奈奎斯特速率慢得多。在 这个建议，我们证明了1000 FPS的执政官读出神经元的行动视频， 可以从以100 FPS操作的CS-PCE相机准确地重建电势。我们 证明使用长像素曝光和慢读出节省了功率，减少了系统 大小，并增加SNR，同时保持动作电位检测能力相比， 现有技术的sCMOS成像器件。 总之，我们的目标是提供一个遗传靶向替代微电极为基础的 记录，这一直是系统神经科学研究中长期追求的目标。评价 设备性能，我们将使用CS-PCE微型记录仪记录海马位置细胞序列 回放，其超过了基于钙的功能成像技术的时间带宽 并且目前只能使用电子记录来访问。

Direct readout of spiking and subthreshold voltage activity from genetically-specified neural populations will facilitate major advances in our understanding of neural computation at the network level. For this reason, developing a genetically encoded voltage indicator (GEVI) with adequate speed, membrane localization, and brightness to report action potentials in mammalian cells has been a major goal in neuroscience for the past two decades. Recently, we developed an opsin-based GEVI, called Archon, which exhibits good localization in neurons of multiple species, several fold improved brightness over previous opsin-based reporters, order-of-magnitude improvements in voltage sensitivity and photo bleaching over GFP-like reporters, and compatibility with optogenetic control. However, action potential imaging, even in a single Archon-expressing neuron, requires a state of the art sCMOS camera to meet sample-rate and signal-to-noise (SNR) requirements. These devices are large, power-hungry, bandwidth intensive, and support a limited field of view at maximal frame rate. This precludes their integration into head-mountable devices and greatly limits their ability to image large numbers of cells. The goal of this proposal is to create a head mountable miniature microscope (“miniscope”) that enables high speed (>1000 frames per second; FPS) voltage imaging of neural populations in freely moving animals. To do this, we will adapt our recently developed integrated compressed sensing CMOS image sensor with pixel-wise exposure (“CS-PCE Camera”) for GEVI imaging. The CS- PCE camera's key innovation is to permit independent exposure of each pixel on the sensor array instead of exposing the entire array in lock-step with a frame clock. This enables CS reconstruction of high speed video from samples acquired much more slowly than the Shannon/Nyquist rate. In this proposal, we demonstrate that a 1000 FPS video of Archon readout of neuronal action potentials can be accurately reconstructed from a CS-PCE camera operated at 100 FPS. We demonstrate that using long pixel-wise exposure and slow readout saves power, reduces system size, and increases the SNR while maintaining action potential detectability compared to state of the art sCMOS imaging devices. Taken together, we aim to provide a genetically-targetable replacement for microelectrode-based recordings, which has been a long sought after goal in systems neuroscience research. To evaluate device performance, we will use the CS-PCE miniscope to record hippocampal place-cell sequence replay, which exceeds the temporal bandwidth of calcium-based functional imaging techniques and is currently only accessible using electrical recordings.