Clarity histology and planar fluorescence microscopy implementation for central nervous system study

• Grant number: 14/21984-1
• Support Opportunities: Regular Research Grants
• Start date: March 01, 2015
• End date: August 31, 2017
• Field of knowledge: Biological Sciences - Physiology - Physiology of Organs and Systems
• Principal Investigator: Gabriel Maisonnave Arisi
• Grantee: Gabriel Maisonnave Arisi
• Host Institution: Escola Paulista de Medicina (EPM). Universidade Federal de São Paulo (UNIFESP). Campus São Paulo. São Paulo , SP, Brazil

Abstract

Microscopy and histological techniques had greatly advanced our understanding of the fine cellular structure of nervous tissue. Nonetheless, the complex tridimensional organization of neurons, glial cells and vasculature is lost in sectioning the brain in order to analyze it under the microscope. Laborious and time consuming methods are employed to reconstruct the 3D organization of cells.Advanced histological techniques like CLARITY, that preserves the cellular protein structure in situ and renders the nervous tissue transparent to light by removing the cellular lipid layers, will allow studies encompassing large volumes of different brain regions. This histology is performed by perfusion of hydrogel monomers that after polymerization creates a protein-monomer hybrid that allows electrophoretic removal of the lipids.Combining this histology to Selective Plane Illumination Microscopy (SPIM), or planar fluorescence microscopy, makes possible a digital scanning of different proteins of interest in huge cellular populations. The planar fluorescence microscopy is implemented by the use of orthogonal lenses, one for specimen illumination and the other for image capture. At low cost and in short time frame is possible to map complete neural circuits, neurotransmitter systems and glia-neuron cellular structure in the brain. Open platforms using common off-the-shelf parts will create local trained personnel able to build and maintain the microscopy setups. CLARITY histology and SPIM could be implemented as routine histology and microscopy for multiple neuroscience research labs in the state of São Paulo. (AU)