Direct wavefront sensing and adaptive optics to enable two-photon imaging axons and spines throughout all of cortex

直接波前传感和自适应光学器件可实现整个皮层的双光子成像轴突和脊柱

基本信息

批准号：
10640249
负责人：
David Kleinfeld
金额：
$ 34.03万
依托单位：
UNIVERSITY OF CALIFORNIA, SAN DIEGO
依托单位国家：
美国
项目类别：
财政年份：
2019
资助国家：
美国
起止时间：
2019-09-30 至 2025-03-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10640249
关键词：
Anatomy Area Astigmatism Award Axon Blood Brain Brain Stem Calcium Cells Cephalic Chronic Communities Compensation Dendrites Dextrans Dyes Educational workshop Elements Engineering Feedback Glutamates Grant Hemoglobin Image Imaging problem Individual Label Laboratories Laser Scanning Microscopy Methods Microscope Mus Nature Neocortex Neurons Neurosciences Noise Optics Output Photobleaching Photons Phototoxicity Plasma Preparation Principal Investigator Process Records Research Design Research Personnel Resolution Scanning Side Signal Transduction Structure Surface Synapses System Tissues Update Vertebral column Water Work Writing absorption adaptive optics awake base design handbook imaging study imaging system improved in vivo instrument instrumentation meter multiphoton imaging multiphoton microscopy neocortical neuroimaging neurophysiology novel optical imaging programs quantum red fluorescent protein segregation success two-photon usability

项目摘要

Principal Investigator (Last, first, middle):KLEINFELD, DAVID Project summary Two-photon laser scanning microscopy is indispensable for imaging the structure and function of the mammalian brain with subcellular resolution. However, the resolution and efficiency decreases with tissue depth as a result of scattering and optical aberrations. Adaptive optics can improve multi-photon imaging by synthesizing a distortion to the wavefront of the excitatory beam that compensates for aberrations in the wavefront that are created by the tissue. Our system utilized adaptive optics to enable investigators to probe subcellular dynamics in individual synapses along the full depth of cortex. This is a crucial advance, particularly as layer 5 and 6 cortical output neurons lie deep to the surface of the brain. Many contemporary studies within the neuroimaging community are limited by the current inability to record synaptic dynamics within output regions of cortex, e.g., layer 5, as opposed to within the dominant input layer, i.e., layer 4, and the intermediate levels, e.g., layers 2/3. We will remove this limit and thus open up a new subfield of in vivo studies on subcellular determinates of cortical output. Our proposed work incorporates "good engineering practice" in the design of our current adaptive optics two-photon microscope design. We will disseminate accurate plans and construction details to enable other laboratories to duplicate this system. We will further educate the neuroimaging community on the principles of adaptive optics and the design and utility of adaptive optics-based two-photon microscopes. This effort includes workshops at UC San Diego. Throughout the period of the proposed grant, we will continue to advance the adaptive optics two-photon system and update and expand our user base. Proposed new directions include a rapid shift in focus together with aberration correction for diffraction limited focus over planes separated by as much as 300 µm and the incorporation of a resonant scanner for fast cell-based imaging. Lastly, we will form a team effort among users and incorporate feedback from the team to extend adaptive optics into new areas of inquiry in neuroscience as they arise.

首席研究员（最后、第一位、中间）：KLEINFELD、DAVID 项目概要双光子激光扫描显微镜对于对生物体的结构和功能进行成像是必不可少的。具有亚细胞分辨率的哺乳动物大脑然而，分辨率和效率随着组织的增加而降低。散射和光学像差导致的深度变化可以通过以下方式改善多光子成像。合成激励光束波前的畸变，以补偿像差我们的系统利用自适应光学技术使研究人员能够探测组织产生的波前。整个皮层深度的单个突触的亚细胞动力学这是一个至关重要的进步，尤其如此。因为第 5 层和第 6 层皮质输出神经元位于大脑表面深处。神经影像学界受到当前无法记录输出中突触动态的限制皮层区域，例如第 5 层，而不是主要输入层（即第 4 层）和中间层我们将消除这个限制，从而开辟一个新的体内研究子领域。皮质输出的亚细胞决定因素。我们提出的工作将“良好的工程实践”融入到我们当前的自适应光学器件的设计中我们将传播准确的计划和施工细节，以使其他人能够使用双光子显微镜设计。实验室复制该系统，我们将进一步教育神经影像学界的原理自适应光学以及基于自适应光学的双光子显微镜的设计和实用性。在提议的拨款期间，我们将继续推进在加州大学圣地亚哥分校举办的研讨会。自适应光学双光子系统以及更新和扩大我们的用户群提出的新方向包括：焦点快速移动以及像差校正，用于衍射限制焦点在由 as 分隔的平面上高达 300 µm 并结合共振扫描仪进行快速细胞成像。用户之间的团队努力，并结合团队的反馈，将自适应光学扩展到新的领域当神经科学出现时进行探究。