Vesicle-free transition zones, dense core vesicles, and vesicle pool redistribution contribute to synapse growth



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Long-term potentiation (LTP) is a widely studied cellular mechanism of learning and memory. LTP occurs at excitatory synapses on dendritic spines. Two hours after LTP induction in mature rat hippocampal slices, a reduction in spine number that is perfectly balanced by enlargement of the remaining synapses was previously observed. The sequence of events by which mature synapses enlarge is not well understood, but potential pre- and postsynaptic ultrastructural correlates of synapse growth have been identified. Vesicle-free transition zones (VFTZs) are postsynaptic thickenings contiguous with the PSD that have no apposing presynaptic vesicles perpendicular to the presynaptic membrane. VFTZs could be regions where synapses have expanded postsynaptically, but to which presynaptic vesicles have not yet been recruited. Presynaptic 80-nm dense core vesicles (DCVs) transport active zone proteins to the synapse during synaptogenesis, and may perform the same function during synaptic plasticity. 3-D reconstructions from ssTEM were used to investigate changes in VFTZs, DCVs, and presynaptic vesicles following LTP induction. By 30 minutes, VFTZ area and docked vesicle counts decreased, suggesting mobilization of additional vesicles to the synapse and enhanced release or delayed recycling. By two hours, VFTZs enlarged, suggesting VFTZ assembly contributes to synapse enlargement. DCV counts at 2 hours decreased relative to that at 30 minutes in both control and LTP conditions, suggesting DCVs were inserted at existing synapses to enlarge potentiated synapses in the LTP condition and to support ongoing spinogenesis in the control condition. The overall vesicle count in presynaptic boutons decreased at 2 hours following LTP induction, but docked vesicle count did not. Docked vesicle count was elevated at 2 hours relative to 30 minutes, suggesting that the depletion of docked vesicles observed at 30 minutes was followed by a replenishment and enhancement by 2 hours supplied by the non-docked vesicle pool. That the largest spines had more and larger VFTZs and recruited more DCVs and docked vesicles, and that the ratio of the sum of VFTZ area to the sum of PSD area is constant, provide further evidence that dendritic segments serve as functional units that manage resources in a coordinated and homeostatic way.