All-Optical Histology for 3-D Cortical Reconstructions

描述（由申请人提供）：超短激光脉冲作为实验室工具的出现为探测和操纵神经系统的解剖结构和功能开辟了新的机会。超短脉冲是驱动生物分子对光的非线性吸收的基本手段，这导致了激发的局部区域，并形成了双光子扫描显微镜的基础。最近，非线性光学吸收已被利用作为一种手段，以可靠和可重复地在脑组织中产生微米大小的消融，同时附带和热损伤最小。这些消融可以用作全光学组织学中的驱动技术，其允许在整个大脑中以微米分辨率对解剖结构进行成像。这些消融也可用于扰动新皮层血流，作为探测正常和患病组织的手段。然而，还需要更多的努力来使用和推进多光子消融和成像技术的混合物，作为一种手段，使神经元和血管结构的研究。 我们提出的研究涉及非线性光学和解剖学的融合。 .推进多光子消融和成像的混合，以建立全光学组织学作为标准解剖工具。这包括参数的优化和软件的进步，结合消融和成像。 .重建大鼠大脑皮层触须感觉区的细胞索马和血管系统。这些信息将用于评估基本的结构参数，包括横向和径向方向的细胞密度之间的相关性，以及基本的代谢参数，如脉管系统内的互连性和相对于毛细血管的胞体分布。 这两个目标，一个是技术性的，另一个是科学性的，有着内在的联系，并将平行进行。 拟议的进步将为组织学自动化提供一种新的工具，这是对大脑功能的理解的基础。我们将使这一工具随时提供给生物医学界。所提出的模型系统可能会大大提高我们的理解的大规模结构的大脑体系结构。

DESCRIPTION (provided by applicant): The advent of ultrashort laser light pulses as laboratory tool has opened up new opportunities to probe and manipulate anatomy and function in nervous systems. Ultrashort pulses are the essential means to drive the nonlinear absorption of light by biomolecules, which leads to a localized region of excitation and forms the basis of two-photon scanning microscopy. More recently, nonlinear optical absorption has been exploited as a means to reliably and reproducibly create micron-sized ablations in brain tissue with a minimum of collateral and thermal damage. These ablations can be use as the driving technology in an all-optical histology, which allows anatomy to be imaged with micrometer resolution throughout the entire brain. These ablations may also be used to perturb neocortical blood flow as a means to probe normal and diseased tissues. Yet much additional effort is required to use and advance the mixture of multi-photon ablation and imaging techniques as a means to enable studies of neuronal and vascular architectonics. Our proposed research concerns the confluence of nonlinear optics and anatomy. . Advance the mixture of multi-photon ablation and imaging to establish all-optical based histology as a standard anatomical tool. This includes the optimization of parameters and the advancement of software for combined ablation and imaging. . Reconstruct cell soma and vasculature positions throughout the vibrissa sensory areas in rat cortex. This information will be used to evaluate essential architectonic parameters, including correlations among cell densities in lateral as well as radial directions, as well as essential metabolic parameters, such as the interconnectivity within the vasculature and the distribution of somata relative to capillaries. These two goals, one technical and the other scientific, are intrinsically linked and will proceed in parallel. The proposed advancements will provide a novel tool for the automation of histology, which underlies an understanding of brain function. We will make this tool readily available to the biomedical community. The proposed model system may substantially improve upon our understanding of the large-scale structure of brain architectonics.