Methodological comparison of different approaches to cultivate postnatal organotypic hippocampal cultures

Abstract

ABSTRACTTo study molecular and cellular processes of the brainin vitro, the cultivation of postnatal organotypic hippocampal slices should be established. The methodological approach had to be optimized several times because the result of the cultivation was not optimal. Initially, cultivation was performed by roller tube culture, which is the first developed cultivation method for these cultures. As the cultures detached or even dissolved, further cultivation was carried out using inserts, which represent a modification of the roller tube cultures. In the further course it came also to the use of different preparation methods and to the exchange of the medium and the incubator, in order to obtain a result of the organ cultivation corresponding to the expectations. The results show that cultivation on semi-permeable membranes has an advantage over roller tube cultures. On the one hand, there is no detachment and dissolution of hippocampal slices due to the mechanical influence and on the other hand, the tissue shows an improvement in general condition compared to roller tube cultures. Furthermore, it was found that an acceleration of the preparation time has positive effects on the neurons. In addition, it could be shown by the recorded time series that a cultivation over a longer period of time stabilizes the tissue. If the hippocampi remain connected to the entorhinal cortex, wallerian degeneration does not occur, resulting in longer survival of the cells in the postnatal cultures and thus strengthening the tissue.