MULTI-PHOTON/CONFOCAL MICROSCOPE FOR BRAIN INJURY STUDY

我们申请资金，在一个共享的成像设施的多光子/共聚焦激光扫描共聚焦显微镜的研究脑和脊髓损伤。 该设施将由华盛顿大学神经系统损伤研究中心内的四名研究人员组成的核心小组操作和指导。我们利用光学和电生理学方法，以及离子或氧化敏感的荧光探针，以了解缺氧缺血性损伤，癫痫，阿尔茨海默氏病和其他神经系统疾病的机制。以前我们的研究主要集中在原代细胞培养模型上，但我们已经扩展了NIH资助的项目，包括更完整的模型系统，包括急性脑切片制备和器官型切片培养。这些模型保留了细胞关系和大脑代谢相互作用，并提供了检查活检获得的人体组织的可能性。 最近获得的具有细胞特异性表达绿色荧光蛋白的转基因小鼠为动态观察活组织内的神经元和神经胶质细胞提供了特别令人兴奋的机会。然而，在传统的光学显微镜的限制，排除在厚标本的高分辨率成像。 双光子显微镜最大限度地减少光损伤和光散射，并允许持续的高分辨率动态成像深入脑切片。 为了最大限度地利用这些方法的发展，我们寻求全职访问一个专用设施内的激光扫描显微镜，允许修改组织灌注，温度和大气控制，和电生理学。 我们提出了一个商业化的激光扫描共聚焦显微镜与多光子能力，锁模亚皮秒钛：蓝宝石激光器，和直立的生理显微镜。 该系统将安装在神经系统损伤研究中心现有的共享显微镜设施中。 它将得到神经病学系的支持，并可供核心用户和大学社区的科学家使用。多光子和共聚焦显微镜的可用性将促进新的进展，了解细胞间的相互作用，导致神经系统损伤。

We request funds for a multi-photon/confocal laser scanning confocal microscope in a shared imaging facility for the study of brain ans spinal cord injury. The facility will be operated and directed by a core group of four investigators within the Center for the Study of Nervous System Injury at Washington University. We utilize optical and electrophysiological approaches, together with ion- or oxidation- sensitive fluorescent probes, to understand mechanisms of hypoxic- ischemic injury, epilepsy, Alzheimer's disease, and other nervous system conditions. Previously our research focused on primary cell culture models, but we have extended our NIH-funded projects to include more intact model systems, including acute brain slice preparations and organotypic slice cultures. These models preserve cellular relationships and brain metabolic interactions, and offer the possibility of examining human tissue obtained at biopsy. The recent availability of transgenic mice with cell-specific expression of green fluorescent protein offers particularly exciting opportunities for dynamic observation of neurons and glia within living tissue. However, limitations in conventional optical microscopy preclude high resolution imaging in thick specimens. Two photon microscopy minimizes photodamage and light scattering, and allows sustained high-resolution dynamic imaging deep in brain slices. To take maximal advantage of these methodological developments, we seek full-time access to a laser scanning microscope within a dedicated facility which allows modifications for tissue perfusion, temperature and atmospheric control, and electrophysiology. We propose to acquire a commercial laser scanning confocal microscope with multiphoton capability, mode- locked sub-picosecond ti:sapphire laser, and upright physiology microscope. The system will be installed in an existing shared microscopy facility within the Center for the Study of Nervous System Injury. It will be supported by the Department of Neurology, and be accessible to core users and to scientists in the university community. Availability of multiphoton and confocal microscopy will promote new advances in understanding cell-cell interactions which lead to nervous system injury.