MATERIALS AND METHODS

the anatomical changes of all these brain neuropils throughout metamorphic development is missing. Thus, the aim of the current study was to visualize the anatomical changes of discernable brain areas in parallel and thus to provide a tool which allows a compari-son of these changes along the developmental time line. To obtain the desired datasets, we labeled whole brains of dif-ferent metamorphic stages with an antiserum against the synap-tic vesicle protein synapsin to visualize neuropil areas, analyzed the staining using confocal laser scanning microscopy, 3D recon-structed the selected brain neuropils using the software AMIRA (Visage Imaging, Fürth, Germany), and subsequently transformed the data sets into the 4D visualization software Cinema 4D. The used technique can in principal be adapted to any brain area which can be visualized and 3D reconstructed at different times of development