Deep and fast imaging using adaptive excitation sources

摘要 活动的光学记录对于探测神经系统至关重要，因为它们提供了高分辨率， 非侵入性测量，范围从单个神经元到完整神经系统中的整个群体，以及 很容易与遗传学方法结合，以提供细胞类型特异性记录。然而，有限的 穿透深度、空间尺度和时间分辨率仍然是光学成像的主要挑战。细胞- 散射脑中的分辨率成像通常用多光子显微镜（MPM）实现。因为 非线性激发过程，多光子成像的发展关键取决于超快技术， 特别是飞秒光源。从二次谐波产生（SHG）和2- 光子荧光（红宝石激光），第一个2光子成像（锁模飞秒激光），最深的3- 光子成像到目前为止（长波长光学参量放大器），多光子成像的进展已经 在很大程度上受到激光技术创新的推动。这项研究提案旨在继续这一点 趋势我们将开发和推广新一代超快激光器和多光子成像工具， 将能够以细胞和亚细胞分辨率对结构和功能进行深度、快速和大规模成像。 为了接近由“光子预算”定义的基本限制，我们将开发自适应激发源 (AES)在1300 nm处进行深层组织3光子显微镜检查（3 PM）。通过输入样本的结构信息 对于激光源，AES仅在感兴趣区域（ROI）内生成按需脉冲，并将其转换为 将传统的多光子显微镜转变为用于ROI的“随机访问”显微镜。我们将整合 AES与高速扫描仪和优化光子预算和扫描系统。我们将进一步测试， 在动物模型的三个概念验证实验中验证新成像技术的性能。 本研究涉及PI（Chris Xu）和合作研究者（Alex Kwan，Frank Wise， Nilay Yapici和Rafael Yuste）。此外，我们将与行业合作伙伴共同探索商业化 这将为广泛传播提供一条直接途径。1300 nm AES和 3 PM将改变我们深入和快速成像的能力，并将对神经科学产生广泛的影响， 需要在完整的大脑深处进行高分辨率、高速的成像。团队成员是积极的支持者 多样性，公平和包容性（DEI）在他们的机构，并将整合本研究计划的目标 与先进的DEI。

Abstract Optical recordings of activity are critical to probe neural systems because they provide high-resolution, non-invasive measurements, ranging from single neurons to entire populations in intact nervous systems, and are readily combined with genetic methods to provide cell type-specific recordings. Nevertheless, the limited penetration depth, spatial scale and temporal resolution remain major challenges for optical imaging. Cellular- resolution imaging in scattering brains is typically achieved with multiphoton microscopy (MPM). Because of the nonlinear excitation process, the development of multiphoton imaging depends critically on ultrafast technologies, particularly femtosecond sources. From the first demonstrations of second harmonic generation (SHG) and 2- photon fluorescence (ruby laser), the first 2-photon imaging (mode-locked femtosecond laser), to the deepest 3- photon imaging so far (long wavelength optical parametric amplifiers), advances in multiphoton imaging have been largely propelled by the innovations in laser technologies. This research proposal aims to continue this trend. We will develop and disseminate a new generation of ultrafast lasers and multiphoton imaging tools that will enable deep, fast, and large-scale imaging of structure and function with cellular and subcellular resolution. To approach the fundamental limits defined by the “photon budget”, we will develop an adaptive excitation source (AES) at 1300 nm for deep tissue 3-photon microscopy (3PM). By feeding the structural information of the sample to the laser source, the AES generates on-demand pulses only within regions of interest (ROIs) and transforms a conventional multiphoton microscope into a “random-access” microscope for the ROIs. We will integrate the AES with high speed scanners and optimize the photon budget and scanning systems. We will further test and validate the performance of the new imaging technology in three proof-of-concept experiments in animal models. The research involves close interactions between the PI (Chris Xu) and Co-investigators (Alex Kwan, Frank Wise, Nilay Yapici, and Rafael Yuste). Furthermore, we will work with industry partners to explore commercialization of the technology, which will provide a direct path to broad dissemination. The combination of 1300 nm AES and 3PM will transform our ability to image deep and fast and will have a broad impact on neuroscience where high- resolution, high speed imaging deep within an intact brain is required. The team members are active proponents of diversity, equity and inclusion (DEI) in their institutions, and will integrate the goals of this research program with advancing DEI.