Multiphoton laser scanning microscope

描述(申请人提供)：请提供资金购买集成多光子激光扫描显微镜系统，用于活体脑组织成像。该非线性光学(NLO)系统将基于蔡司LSM 510扫描系统和Axioskop 2Plus FS电动直立显微镜。主要的荧光激发源将是一台5瓦的相干钛蓝宝石飞秒激光器。该系统将在活体脑片、线虫和半完整的脊髓标本中进行优化，以同时进行NLO成像和膜片钳电生理学。拟议的仪器将由神经科学成像中心(NIC)容纳和管理。NIC目前维护着另外两个主要的显微镜系统，并有一名全职技术人员负责用户培训和少量维护。描述了来自10个用户(4个大用户和6个小用户)的项目。所有拟议的项目都利用了这种直立式NLO系统的独特优势，包括降低光毒性、通过厚实的活体制剂进行深度光学切片以及同时进行全细胞膜片钳电生理。本·斯特罗布里奇博士将使用NLO成像技术记录嗅球和齿状回脑片中单一的突触前钙瞬变。林恩·兰德梅瑟博士将使用NLO成像技术研究鸡胚胎和转基因小鼠脊髓发育过程中的局部回路。斯特凡·赫利策博士计划使用这种多光子显微镜来研究活着的线虫的钙通道功能。David Friel博士将使用NLO成像技术研究正常和突变小鼠小脑切片中浦肯野神经元的亚细胞钙处理。

DESCRIPTION (provided by applicant): Funds are requested to purchase integrated multiphoton laser scanning microscope system for imaging living brain tissue. This non-linear optical (NLO) system will be based on the Zeiss LSM 510 scan system and an Axioskop 2Plus FS motorized upright microscope. The primary fluorescent excitation source will be a 5 watt Coherent titanium-sapphire femtosecond laser. This system will be optimized for simultaneous NLO imaging and patch clamp electrophysiology in living brain slices, nematodes and semi-intact spinal cord preparations. The proposed instrument will be housed and administered by the Neuroscience Imaging Center (NIC). The NIC currently maintains two other major microscope systems and has a full-time technician available for user training and minor maintenance. Projects from ten users (four major and six minor) are described. All proposed projects exploit the unique advantages of this upright NLO system including reduced phototoxicity, deep optical sectioning through thick living preparations, and simultaneous whole cell patch clamp electrophysiology. Dr. Ben Strowbridge will use NLO imaging to record unitary presynaptic calcium transients in olfactory bulb and dentate gyrus brain slices. Dr. Lynn Landmesser will use NLO imaging to study local circuits in the developing spinal cord in chick embryos and transgenetic mice. Dr. Stefan Herlitze plans to use this multiphoton microscope to study calcium channel function in living C. elegans. Dr. David Friel will use NLO imaging to study subcellular calcium handling in Purkinje neurons in cerebellar brain slices from normal and mutant mice.