Nigrostriatal organotypic cultures to study neuromelanin accumulation in dopaminergic circuits

In this protocol we describe the preparation and maintenance of rat organotypic cultures from parasagittal brain slices. We use a 13º slicing angle and a customized cocktail of small molecules and growth factors, to maximize the integrity of the nigrostriatal pathway and ensure the survival of dopaminergic neurons, respectively. The slices maintain the basic cytoarchitecture of the brain, including glia and the extracellular matrix (not including vessels nor immune system). The cultures can be used to study dopaminergic degeneration, cell-cell and cell-matrix interactions and are particularly suitable for AAV-mediated overexpression of transgenes, time-lapse live imaging and longitudinal studies.