Multi-Photon Laser-Scanning Mikroscope

维持低的细胞内钠浓度能够产生电信号，并为大脑中的多种运输过程提供能量。最近的研究，我们的实验室作出了重大贡献，表明生理活动引起短暂的增加钠浓度的神经元和星形胶质细胞。此外，我们和其他人已经证明，钠升高是兴奋性毒性损伤的核心，并在神经退行性疾病中严重参与细胞功能障碍和损伤。由于钠梯度变化的多方面后果，理解脑中细胞内钠稳态和信号传导的决定因素和特征是必不可少的。为了解决这些问题，多光子激光扫描显微镜已成为一种方法的选择，因为它使离子瞬态的测量在小隔间，如树突棘或精细星形胶质细胞的过程中的完整的，光散射组织深处。神经生物学研究所在2005/2006年安装了一个定制的多光子系统，基本上反映了2000年的技术状况。为了能够继续在竞争水平上进行研究，需要通过采用现代最先进的多光子成像来实现荧光信号的更好的时间和空间分辨率。因此，我们申请资金用于一种新的多光子激光扫描显微镜，该显微镜应配备动态荧光寿命成像组件，一个uncaging/光刺激单元和电生理学。商用主要模块将与定制设备和现有设备相结合，从而使拟议系统具有极佳的成本/性能比。新的多光子系统将成为我们大多数研究项目的核心，每天由神经生物学研究所运行约12小时。它将位于新的实验室设施中，这些设施已经配备了必要的基础设施，包括激光安全要求。该系统将由在多光子成像和荧光显微镜方面具有长期专业知识的终身学术人员建立和维护。数据的收集和管理将符合良好科学实践的标准，并坚持公平的原则。这包括使用电子实验室笔记本记录元数据，并在发表后立即通过开放获取文件，托管和特定神经科学数据库提供实验数据。新的多光子系统将为我们的方法库增加新的质量，并将大大加强我们未来的校外资助建议。此外，它将使我们能够在国际竞争力的水平上解决新的研究问题，从而也促进我们与我们的地方，国家和国际合作伙伴的合作。

The maintenance of a low intracellular sodium concentration enables the generation of electrical signaling and energizes a multitude of transport processes in the brain. Recent studies, to which our laboratory has contributed significantly, show that physiological activity evokes transient increases in the sodium concentration of both neurons and astrocytes. Moreover, we and others have demonstrated that sodium elevations are central to excitotoxic damage and are critically involved in cellular dysfunction and damage in neurodegenerative disease. Because of the manifold consequences of changes in the sodium gradient, understanding the determinants and characteristics of intracellular sodium homeostasis and signaling in the brain is indispensable. To address these questions, multi-photon laser scanning microscopy has become a method of choice since it enables the measurement of ion transients in small compartments such as dendritic spines or fine astrocytic processes deep in the intact, light-scattering tissue. The Institute of Neurobiology operates a custom-built multi-photon system installed in 2005/2006, which essentially reflects the technical status of the year 2000. To be able to continue performing research at a competitive level, a better temporal and spatial resolution of fluorescence signals by employing modern, state-of-the-art multi-photon imaging is required. We therefore apply for funds for a new multi-photon laser scanning microscope which shall be equipped with components for dynamic fluorescence lifetime imaging, an uncaging/opto-stimulation unit and electrophysiology. Commercially available major modules will be combined with customized and existing equipment, resulting in an excellent cost/performance ratio of the proposed system. The new multi-photon system will be central for the majority of our research projects and be operated daily for ~12 hours by the Institute of Neurobiology. It will be located in new laboratory facilities that are already equipped with the necessary infrastructure including laser safety requirements. The system will be set-up and maintained by tenured academic staff with long-standing expertise in multi-photon imaging and fluorescence microscopy. Data will be collected and managed complying the standards of good scientific practice and adhering the FAIR principles. This includes recording metadata using an electronic lab notebook, and making experimental data available through open access files, depositories, and specific neuroscience databases right after publication. The new multi-photon system will add a new quality to our methodological repertoire and will considerably strengthen our future proposals for extramural funding. It will moreover enable us to address new research questions at an internationally competitive level, thereby also fostering our collaborations with our local, national as well as international partners.