ABSTRACT: ?-Synuclein is a presynaptic protein that regulates synaptic vesicle trafficking under physiological conditions. However, in several neurodegenerative diseases, including Parkinson's disease, dementia with Lewy bodies, and multiple system atrophy, ?-synuclein accumulates throughout the neuron, including at synapses, leading to altered synaptic function, neurotoxicity, and motor, cognitive, and autonomic dysfunction. Neurons typically contain both monomeric and multimeric forms of ?-synuclein, and it is generally accepted that disrupting the balance between them promotes aggregation and neurotoxicity. However, it remains unclear how distinct molecular species of ?-synuclein affect synapses where ?-synuclein is normally expressed. Using the lamprey reticulospinal synapse model, we previously showed that acute introduction of excess recombinant monomeric or dimeric ?-synuclein impaired distinct stages of clathrin-mediated synaptic vesicle endocytosis, leading to a loss of synaptic vesicles. Here, we expand this knowledge by investigating the effects of native, physiological ?-synuclein isolated from the brain of a neuropathologically normal human subject, which comprised predominantly helically folded multimeric ?-synuclein with a minor component of monomeric ?-synuclein. After acute introduction of excess brain-derived human ?-synuclein, there was a moderate reduction in the synaptic vesicle cluster and an increase in the number of large, atypical vesicles called "cisternae." In addition, brain-derived ?-synuclein increased synaptic vesicle and cisternae sizes and induced atypical fusion/fission events at the active zone. In contrast to monomeric or dimeric ?-synuclein, the brain-derived multimeric ?-synuclein did not appear to alter clathrin-mediated synaptic vesicle endocytosis. Taken together, these data suggest that excess brain-derived human ?-synuclein impairs intracellular vesicle trafficking and further corroborate the idea that different molecular species of ?-synuclein produce distinct trafficking defects at synapses. These findings provide insights into the mechanisms by which excess ?-synuclein contributes to synaptic deficits and disease phenotypes.
