A custom-built lightsheet microscope that can image an entire intact mouse brain at half micron in-plane resolution is now up and running in the microscopy facility at Campus Biotech. Development of the new microscope was made possible thanks to the collaboration of the world leading neuroscience groups of Professor Lüscher and Professor Holtmaat from the University of Geneva, and Professor Deisseroth from Stanford University.
One of only three in the world, this state-of-the-art microscope, which was built by Dr Stéphane Pagès, can image at the level of dendritic spines, the micron sized protrusions that emerge from the dendrites of a neuron.
Video: Mouse brain injected with a fluorescent anterograde virus in the brain stem shows wide labelling of glutamatergic projection. The video zooms into the cortical area. Credit: Courtine Lab (Leonie Asboth and Elodie Rey), EPFL
Video: Mouse brain injected with a fluorescent anterograde virus in ventral tegmental area shows dopaminergic projections, widely implicated in the drug and natural reward circuitry. Credit: Christian Lüscher Lab, UNIGE
The microscope offers neuroscientists a unique opportunity to capture high resolution, whole brain images without perturbing neuronal architecture.
The combination of viral neuronal tracing, with optical clearing methods and lightsheet technologies enables the distribution of neuronal pathways of an entire mouse central nervous system (CNS) to be pin pointed at submicronic resolution. The lightsheet microscope can also be used to visualize and confirm the location of novel neural implants.
How it works
The Clarity Optimized Light-sheet Microscope (COLM) was developed by Dr. Raju Tomer at Professor Deisseroth’s laboratory. The microscope uses a digitally scanned light-sheet to illuminate a transparent brain sample and create images in a single plane. Axial movement of the sample generates multiple optical slices, that are then combined to create a 3D reconstruction. The fusion of images from multiple positions into a single mosaic allows visualization of the entire brain at cellular or sub-cellular resolution.
The microscope is optimized for large scale clarified tissues, such as a whole mouse CNS or brains from transgenic mice that express endogenous florescent proteins.
Clarification using the diffusion based ‘clarity’ technique generates a completely transparent brain by removal of the opaque lipids present in brain tissue.
High resolution imaging over 1-2 days can generate up to 20TB of data, creating data management and analysis challenges, unprecedented in optical imaging.
How to use it
New virtual reality tools are being developed that allow the user to visualize the huge data volumes, and navigate through the image to explore the 3D brain from all angles.
The light sheet microscope is part of the larger microscopy facility at Campus Biotech which offers both technical support and innovation in imaging technology to the scientific community in academia and in industry.
The facility includes microscopy instrumentation, ranging from a slide scanner and confocal laser scanning to advanced software for image analysis.
Video: Mouse brain injected with a fluorescent anterograde virus in the brain stem shows wide labelling of glutamatergic projection. 2D navigation through the several z-optical slices. Credit: Courtine Lab (Leonie Asboth and Elodie Rey)