Dissection and Direct Imaging of Axonal Transport in Drosophila Segmental Nerves

For neurons, especially those with long axons, the forceful transport of mitochondria, vesicles, and other cytoplasmic components by cytoskeletal motors is vital. Defects in cytoplasmic transport machinery cause a degradation of signaling capacity that is most severe for neurons with the longest axons. In humans, with motor axons up to a meter long, even a mild mutation in one copy of the gene that codes for kinesin-1, the primary anterograde axonal transport motor, can cause spastic paraplegia and other distal neuropathies.To address questions about the molecular mechanisms of organelle movement, we turned to Drosophila as a model system, because it offered rigorous genetic and molecular approaches to the identification and inhibition of specific elements of transport machinery. However, methods for direct observation of organelle transport were largely lacking. We describe here an approach that we developed for imaging the transport behaviors of specific organelles in the long motor axons of larvae. It is straightforward, the equipment is commonly available, and it provides a powerful tool for studying the contributions of specific proteins to organelle transport mechanisms.