ZeBraInspector, a whole organism screening platform enabling volumetric analysis of zebrafish brain white matter

In recent years, zebrafish have become well established for the study of fundamental biology, toxicology and drug discovery. Here we propose fast, standardized protocols for high-throughput and high-content 3D imaging of 5dpf zebrafish EE using a confocal laser scanning microscope. Using a lipophilic fluorescent dye, we generated a reference labeling in cleared fish. We developed a mounting method based on 3D-printed stamps used to create a grid of wells in an agarose cast. We demonstrate that the quality and homogeneity of the fluorescence signal allows an unequivocal segmentation of the whole specimen and the white matter. From there, we built an automated image registration and analysis pipeline for the volumetric analysis of morphological defects. The platform, termed ZeBraInspector, is focused on the analysis of the shape of the whole EE or the white-matter of the central nervous system. A user-friendly software to visualize at large scale registered and segmented 3D images can be downloaded from https://tefor.net/portfolio/zebrainspector. We finally provide two examples of application: the characterization of a phenotype associated with a neuro-developmental mutation, and the defects caused by treatments with a toxic anti-cancer compound. ZeBraInspector can be adapted to a wide variety of high-content analyses.