The laboratory of Univ. Prof. Dr. Tibor Harkany at the Center of Brain Research in Vienna is interested in the diversification of neurons and their integration into neuronal networks during development. To add knowledge to the diversity of interneurons, Daniela Calvigioni and colleagues analyzed a subtype of GABAergic interneurons producing the neuropeptide cholecysteokinin (CCK). Due to difficulties in histochemical approaches, the migration of CCK interneurons and their population of the cerebral cortex at prenatal stages is poorly understood. Therefore, the researchers developed a novel transgenic mouse line marking CCK interneurons in vivo. This opened the possibility to localize this interneuron subtype for the first time within the intact brain structure and to analyze its migratory behavior. To do so, dissected brains of different embryonic time-points were fixed, cleared (CUBIC clear-ing) and imaged on a ZEISS Lightsheet Z.1 microscope using tile scanning to cover the size of the entire brain.
Tibor Harkany Lab Department of Molecular Neuroscience, Center for Brain Research, Medical University of Vienna - Daniela Calvigioni, Zoltán Máté, János Fuzik, Fatima Girach and colleague
Figure 1: Stitching of individual tiles using the Tile Sorter; shown are 35 individual z-stacks covering a dual labelled mouse brain (CCKBAC/DsRed::GAD67gfp/+) at embryonic day 16.5; GA-BAergic neurons are marked by GFP (green) due to expression of GAD67; CCK expression is marked by DsRed (red). Tile 4 is selected and highlighted by a white box. For aligning, the Tile sorter provides different methods including Grid, Manual and Alignments using advanced algorithms. In this case, the z-stack tiles were sorted using the Grid mode; Pixel overlap was set to 10% as during imaging. Several planes were checked to assure correct alignment. Due to stability during image acquisition no further adjustments as algorith-ms were used. Scale options and transparency settings (see selected Tile 4) help to assure correct alignment.
