Open-source miniaturized two-photon microscopes for large field-of-view and volumetric imaging

用于大视场和体积成像的开源小型双光子显微镜

• 批准号: 10516900
• 负责人: Daniel Aharoni
• 金额: $ 110.93万
• 依托单位: UNIVERSITY OF CALIFORNIA LOS ANGELES
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-08-15 至 2026-07-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10516900
• 关键词: Animals; Anterior; Axon; Behavior; Brain; Bypass; Calcium; Cells; Collaborations; Communities; Coupled; Crystallization; Custom; Cytoplasmic Granules; Decision Making; Dendrites; Educational workshop; Elements; Ensure; Excision; Fiber; Glutamates; Hippocampus (Brain); Image; Lateral; Letters; Link; Memory; Methods; Microscope; Microscopy; Mus; Neurons; Neurosciences; Operative Surgical Procedures; Optics; Parietal; Parietal Lobe; Pattern; Persons; Photons; Population; Production; Publishing; Rattus; Recording of previous events; Research; Research Personnel; Resolution; Role; Scanning; Social Interaction; Structure; System; Techniques; Technology; Testing; Thalamic structure; Tissue imaging; Tissues; Work; cingulate cortex; cost; data acquisition; design; design-build-test; equipment acquisition; imaging capabilities; insight; lens; memory encoding; miniaturize; neural network; neuronal cell body; nonhuman primate; novel; open source; open source tool; photonics; sensor; spatial memory; theories; tool; two photon microscopy; two-photon; wiki

Abstract: Single-photon (1P) epifluorescence miniaturized microscopy coupled with genetically encoded calcium sensors has allowed investigators to record the activity of large populations of identified neurons over days to weeks in freely behaving animals, answering fundamental questions in neuroscience. Our group's efforts with the UCLA Miniscope Project have allowed over 600 labs to build and use over 2500 open-source miniaturized microscopes with expanded capabilities at a small fraction of the cost of those offered by commercial versions, thus democratizing access. Yet, 1P miniscopes lack the lateral and axial resolution to image activity in fine structures such as dendrites and axons. In addition, 1P imaging is limited to superficial structures or requires removal of overlying tissue for imaging of deeper neurons. Two-photon (2P) microscopy has exquisite lateral and axial resolution and bypasses all of these obstacles. Recent advances in technology have made the construction of two-photon miniaturized microscopes for mice possible. However, the field of view (FOV) is still limited, and these microscopes require custom-built optics and cost several hundred thousand dollars to acquire commercially. We have designed and built a two-photon miniaturized microscope for mice, including a custom- made objective lens, that allows 2P imaging of an 800 micrometer FOV nearly quadrupling the FOV from the latest published 2P miniaturized microscope (Mini2P-V1). In this proposal, we will optimize this microscope and test it in freely behaving mice for axonal, dendritic and deep somatic imaging. This microscope will be tested in three labs. The Golshani Lab will test the scope with calcium imaging of thalamic axons in anterior cingulate cortex during social interaction. The Silva Lab will test the scope by performing dendritic calcium and glutamate imaging in retrosplenial cortex during memory linking. The Shtrahman Lab will test deep imaging capability by imaging dentate granule neurons through an intact CA1. We will also build a larger miniaturized microscope suitable for rats and non-human primates with expanded capabilities, including a higher numerical aperture (NA), large FOV and temporal multiplexing capability to allow volumetric imaging at high frame rates (MiniMux2P). This microscope will be tested by the Blair Lab to dissect the role of superficial and deep CA1 neurons of rats in navigation. It will also be tested in the Churchland Lab to image rat posterior parietal cortical neurons during decision-making tasks. Finally, we will disseminate the technology using our open-source wiki that has already disseminated miniscope technology to thousands of users. We will provide parts-lists, optical designs and methods for obtaining custom lens elements. As we have done before, we will educate users through online videos and hands-on workshops where imaging basics, surgical techniques and analysis tools are demonstrated. We hope these cutting edge, novel and open-source tools will allow investigators to extend their research beyond that of what is possible with currently available technology.

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