Zebrafish are becoming increasingly popular models for examining the mechanisms of and treatments for neurological diseases. The available methods and technology to examine disease processes in vivo are increasing, however, detailed observations of subcellular structures and processes are complex in whole organisms. To address this need, we developed a primary motor neuron (MN) culture technique for utilization with zebrafish neurological disease models. Our protocol enables the culturing of cells from embryos older than 24h post-fertilization, at points after MN axonal development and outgrowth begins, which enables MN axons to develop in vivo in the context of the normal endogenous cues of the model organism, while also providing the accessibility of an in vitro system. When utilized with the increasing number of genetically modified or transgenic models of neurological diseases, this approach provides a novel tool for the examination of cellular and subcellular disease mechanisms, and offers a new platform for therapeutic discoveries in zebrafish.
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