Distinguishing Neurotransmitter Carrying Pause Mechanics During Axonal Trafficking

Mature presynapses share neurotransmitter carrying vesicles with neighboring presynapses using a process called Inter-Synaptic Vesicle Exchange (ISVE). ISVE has been considered as a pathway to support synaptic transmission, by providing an extra pool of vesicles during activity. Understanding the mechanics of ISVE would thus allow us to better model the efficiency with which ISVE supports synaptic transmission. However, there are major gaps in our understanding about how ISVE vesicles transport along the axon in order to support transmission. ISVE vesicles utilize molecular motor transport, which is fast and efficient, but are predominantly rate-limited by vesicle pausing along the axon. We utilize high-resolution single vesicle fluorescence microscopy in live cells to measure ISVE transport at the nanometer and millisecond scales. We distinguish vesicles undergoing axonal transport, and use a computational approach to characterize motor-driven motion from pausing. We further developed a computational approach to quantify mechanics of vesicle pauses along the axonal cytoskeleton. We show that vesicles likely utilize multi-motor coordination in order to navigate the complex axonal cytoskeleton. We show that loss of actin-based transport results in altered pausing mechanics.