Fast 3D multiphoton microscope

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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.