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引入了新颖的仪器以图像，然后分析和量化神经元的全皮质电气动力学以及脑脉管系统的类似血管舒张动力学。该仪器跨度从细胞分辨率（以单微晶计尺度）到所有小鼠皮层的空间尺度以当时的毫米尺度为单位。大规模的大脑活动涵盖了血液流动重新分布到代谢需求增强区域的神经血管问题，以及继发于血管舒缩活性的血液流动模式。它进一步涉及关注和决策中神经元活性相关的神经系统问题，多感官处理中的遥远信号传导，在更抽象的层面上，诸如大脑中电波的模式等问题。此外，大规模的仪器获得了广泛的智力优点，作为诊断神经元和神经血管加工的一种手段，该方法遍及不同的皮质区域。我们的方法涉及设计和实现一种新型的大型体内两光子显微镜，并具有前所未有的十毫米视野，整个场均分辨率有一个千分尺。这足以使小鼠中细胞分辨率的整个皮质地幔成像。我们的设计和认识必须应对复杂但无法克服的挑战，即在大型扫描角度保持无像差的光束，这些扫描角传统上与色差和空间畸变有关。尽管已发表的有关纠正此类问题的文献的指导有限，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使用转基因动物的使用相结合，其中各个特定细胞类型表达功能报告基因。这将使我们从特定的细胞类型（深到丘脑表面深处）记录记录，从而为神经元和神经血管动力学提供了解剖基础。我们的方法对过去的全场成像技术进行了定性改进，其中研究了均匀染色的神经元组织，既没有深度也不是细胞特异性信息。除了我们在神经元处理中的科学利益之外 和神经血管耦合，我们独特的仪器将为整个系统生理学中许多其他主题的研究提供服务，包括例如开发和再生中的细胞迁移以及肿瘤起源和转移中细胞和组织的流动。通过PI及其同事在研究生和研究生暑期学校的参与中，工作的技术方面将广泛传播。