Ultra-large field two-photon microscope to image transcortical brain dynamics

超大视场双光子显微镜对经皮层脑动力学进行成像

• 批准号: 8893665
• 负责人: David Kleinfeld
• 金额: $ 21.76万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN DIEGO
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-05-01 至 2017-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8893665
• 关键词: Achievement; Address; Animals; Area; Astigmatism; Attention; Back; Beds; Biological Assay; Blood Vessels; Blood flow; Brain; Calcium; Caliber; Cells; Cephalic; Chronic; Color; Complex; Cone; Coupling; Craniotomy; Data; Decision Making; Development; Diagnosis; Distant; Dyes; Equilibrium; Functional Imaging; Glass; Goals; Health; Image; Imaging Techniques; Individual; Investigation; Label; Laser Scanning Microscopy; Lasers; Light; Literature; Magnetism; Measurement; Measures; Membrane; Metabolic; Microscope; Microscopy; Movement; Mus; Natural regeneration; Nature; Neocortex; Neoplasm Metastasis; Nervous System Physiology; Neurologic; Neurons; Optics; Pathway interactions; Pattern; Physiologic pulse; Physiology; Process; Publishing; Reporter; Resolution; Rest; Sampling; Scanning; Schools; Secondary to; Services; Signal Transduction; Speed; Staining method; Stains; Surface; System; Techniques; Testing; Tissues; Touch sensation; Transgenic Animals; Vasomotor; Vibrissae; Work; abstracting; arteriole; base; cell motility; cell type; cranium; data acquisition; design; imaging system; in vivo; in vivo imaging; instrument; instrumentation; interest; meetings; millimeter; multisensory; neocortical; neuronal cell body; novel; recombinase; sensory cortex; tool; tumor; two-photon; vasomotion; voltage

DESCRIPTION (provided by applicant): The PI introduces novel instrumentation to image and then analyze and quantify the whole- cortex electrical dynamics of neurons and the analogous vasomotor dynamics of brain vasculature. The spatial-scales encompassed by this instrument span from cell resolution - at the one-micrometer scale - to all of mouse cortex - at the then-millimeter scale. Large-scale brain activity encompasses the neurovascular issue of the redistribution of blood flow to areas of heightened metabolic need as well as patterns of blood flow secondary to vasomotor activity. It further concerns neurological issues of correlated neuronal activity in attention and decision making, distant signaling in multisensory processing and, at a more abstract level, issues such as the patterning of electrical waves in the brain. Further, large- scale Thus the proposed instrument gains broad intellectual merit as a means to diagnose neuronal and neurovascular processing that extends across different cortical areas. Our approach involves the design and realization of a novel, large-field in vivo two-photon microscope with an unprecedented ten-millimeter field of view with one micrometer resolution across the field. This is sufficient to image the entire cortical mantle in mouse with subcellular resolution. Our design and realization must meet the complex yet not insurmountable challenges of maintaining an aberration-free beam across large scan-angles, which are traditionally associated with chromatic and spatial aberrations. While there is limited guidance in the published literature on correcting such problems, we are able to successfully address these issues in a systematic way. Our novel microscopy will be combined with the use of transgenic animals in which individual specific cell types express a functional reporter. This will allow us t record from specific cell types - deep to the pial surface - and thus provide an anatomical basis for neuronal and neurovascular dynamics. Our approach provides a qualitative improvement of past, whole-field imaging techniques in which uniformly stained neuronal tissue was studied with neither depth nor cell-specific information. Beyond our scientific interests in neuronal processing and neurovascular coupling, our unique instrument will be of service to investigations of many other topics in the physiology of whole systems, including for example cell migration in development and regeneration as well as the flow of cells and tissue in tumor genesis and metastasis. Technical aspects of work will be broadly disseminated through the involvement of the PI and colleagues in graduate and post-graduate summer schools.

描述（由申请人提供）：PI 引入了新颖的仪器来成像，然后分析和量化神经元的整个皮质电动力学和脑脉管系统的类似血管舒缩动力学。该仪器所涵盖的空间尺度涵盖从细胞分辨率（一微米尺度）到整个小鼠皮层（当时的毫米尺度）。大规模的大脑活动包括血流重新分配到代谢需求增加的区域的神经血管问题以及继发于血管舒缩活动的血流模式。它还涉及注意力和决策中相关神经元活动的神经学问题、多感觉处理中的远程信号传递，以及在更抽象的层面上，诸如大脑中电波模式等问题。此外，大规模因此，所提出的仪器作为诊断跨不同皮质区域的神经元和神经血管处理的手段获得了广泛的智力价值。我们的方法涉及设计和实现一种新颖的大视场活体双光子显微镜，具有前所未有的十毫米视场和一微米的视场分辨率。这足以以亚细胞分辨率对小鼠的整个皮质套进行成像。我们的设计和实现必须满足复杂但并非不可克服的挑战，即在大扫描角度上保持无像差光束，这些挑战传统上与色差和空间像差相关。尽管已发表的文献中关于纠正此类问题的指导有限，但我们能够以系统的方式成功解决这些问题。我们的新型显微镜将与转基因动物的使用相结合，其中个体特定细胞类型表达功能性报告基因。这将使我们能够记录特定细胞类型（深入到软脑膜表面），从而为神经元和神经血管动力学提供解剖学基础。我们的方法对过去的全视野成像技术进行了质的改进，在这种技术中，既没有深度信息也没有细胞特异性信息来研究均匀染色的神经元组织。超越我们对神经元处理的科学兴趣 和神经血管耦合，我们独特的仪器将有助于研究整个系统生理学中的许多其他主题，包括例如发育和再生中的细胞迁移以及肿瘤发生和转移中细胞和组织的流动。技术方面的工作将通过 PI 及其同事在研究生和研究生暑期学校的参与得到广泛传播。