The endoplasmic reticulum (ER) extends throughout neurons and regulates many neuronal functions, including neurite outgrowth, neurotransmission, and synaptic plasticity. Mutations in proteins that control ER shape are linked to the neurodegenerative disorder Hereditary Spastic Paraplegia (HSP), yet the ultrastructure and dynamics of neuronal ER remain largely unexplored, especially at presynaptic terminals. Using super-resolution and live imaging in Drosophila melanogaster larval motor neurons, we investigated ER structure at presynaptic terminals of wild-type animals and null mutants of the HSP-linked gene, Atlastin, which encodes an ER-shaping protein. Previous studies reported diffuse localization of an ER luminal marker at Atlastin null mutant presynaptic terminals, which was attributed to ER fragmentation. However, using an ER membrane marker, we found that Atlastin mutant ER forms robust ER networks with only mild defects in structure and dynamics, indicating that the primary defect is functional rather than architectural. We demonstrate that Atlastin mutants progressively displace a luminal ER protein reporter to the cytosol during larval development, specifically at synapses, while this reporter remains correctly localized in cell bodies, axons, and muscles. This synapse-specific displacement phenotype, previously unreported in non-neuronal cells, emphasizes the importance of studying neurons to understand HSP pathogenesis.
