Activity-induced synaptic capture and exocytosis of the neuronal serine protease neurotrypsin

Extracellular proteolysis plays an essential role in synaptic remodeling that is indispensable for cognitive function. The extracellular serine protease neurotrypsin was implicated in cognitive function, because humans lacking a functional form of neurotrypsin suffer from severe mental retardation. By immunoelectron microscopy, neurotrypsin has been localized to presynaptic terminals, suggesting a local proteolytic function after its synaptic release. Here, we studied axonal trafficking and synaptic exocytosis of neurotrypsin by live imaging of hippocampal neurons expressing neurotrypsin fused with enhanced green fluorescent protein or its pH-sensitive variant, superecliptic pHluorin. In differentiated neurons, we identified neurotrypsin in mobile transport vesicles along axons and in both an intracellular and an extracellular pool at synapses. Short depolarization triggered rapid synaptic exocytosis of neurotrypsin. Once externalized, neurotrypsin lingered at its synaptic release site for several minutes before it disappeared. Cell depolarization also enhanced synaptic capture of intracellular neurotrypsin transport vesicles, and elevated synaptic activity increased both number and motility of mobile axonal neurotrypsin vesicles. We further observed trading of neurotrypsin vesicles between adjacent synapses. These activities may support the replenishment of neurotrypsin after activity-induced synaptic exocytosis. Together, the activity-dependent recruitment of neurotrypsin to synapses and its exocytosis and transient persistence at its synaptic release site argue for a spatially and temporally restricted proteolytic action at the synapse. Thereby, neurotrypsin may play a role in activity-dependent remodeling of the synaptic circuitry that is key to adaptive synaptic changes in the context of cognitive functions, such as learning and memory.