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和功能开辟了新的机会。超短脉冲是驱动生物分子非线性吸收光的重要手段，它导致局部激发区域，是双光子扫描显微镜的基础。最近，非线性光学吸收被用作一种可靠、可重复地在脑组织中产生微米级烧蚀的手段，同时最小化了附带损伤和热损伤。这些消融可以用作全光学组织学的驱动技术，这使得整个大脑的解剖结构可以以微米分辨率成像。这些消融也可以用来干扰新皮层血流，作为探测正常和病变组织的一种手段。然而，使用和推进多光子消融和成像技术的混合，作为一种使神经元和血管结构研究成为可能的手段，还需要付出更多的努力。

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.