Fast 3D multiphoton microscope

该应用程序是一个超快多光子成像平台，融合三维（3D）荧光采集与（有针对性的）3D光刺激。我们研究的小鼠脑或人类诱导多能干细胞（iPSC）衍生的脑类器官包含通过精细过程连接的神经元和神经胶质细胞的3D网络，以了解神经退行性疾病的基本机制。可视化这种复杂的结构，特别是检测在3D中的分散位置同时发生的快速时间事件（例如，跨越神经元树突树上的棘）需要非常快速的扫描。这可以通过使用具有允许随机访问点扫描的kHz采样率的声光（AO）技术来利用。与荧光钙指示剂结合，可以在大体积内研究数百个互连神经元的活性，包括相对稀疏的神经元群体如抑制性中间神经元的成像。这对于同时分析多种不同神经元亚型中的活性变化的结果或细胞亚群中的细胞内蛋白质聚集体的结果尤其重要。由于其扫描速率限制，这不能用常规的基于振镜/共振的多光子逐面成像来实现。

This application is for an ultrafast multiphoton imaging platform that fuses three-dimensional (3D) fluorescence acquisition with (targeted) 3D photostimulation. The mouse brain or human induced pluripotent stem cells (iPSCs) derived brain organoids which we study to understand basic mechanisms of neurodegenerative diseases contain 3D networks of neurons and glia cells connected via fine processes. Visualizing such elaborate structures in action, in particular detecting fast temporal events occurring simultaneously at dispersed locations in 3D (e.g., across spines on a neuronal dendritic tree) need very fast scanning. This can be utilized by the use of acousto-optic (AO) technology with kHz sampling rates which allow random-access point scanning. In combination with fluorescent calcium indicators, the activity of hundreds of interconnected neurons can be studied within large volumes including the imaging of relatively sparse neuronal populations such as inhibitory interneurons. This is especially important in order to simultaneously analyse the consequence of activity changes in a multitude of different neuronal subtypes or the consequence of intracellular protein aggregates in a subset of cells. Due to its scanning rate limitations, this cannot be achieved with conventional galvo /resonant based multiphoton plane-by-plane imaging.