Fernández-Busnadiego et al. use cryo-electron tomography to investigate the structure of the presynapse, revealing important functions for short protein tethers in organizing synaptic vesicles at the nerve terminal.

The technique allowed the researchers to capture accurate, three-dimensional views of the presynaptic cytoplasm, and quantify the effects of different pharmacological treatments. Actin is thought to be involved in organizing synaptic vesicle release, but Fernández-Busnadiego et al. saw relatively few actin filaments in the nerve terminals. Instead, many short filaments of unknown composition linked vesicles to each other and to the active zone of the presynaptic plasma membrane, where vesicles are exocytosed.

Vesicles were clustered together through links that the researchers termed “connectors.” These connections were rearranged when the synapses were strongly stimulated or treated with the phosphatase inhibitor okadaic acid. The researchers think the connectors regulate vesicle mobility and release in resting and active synapses.

In conventional electron microscopy studies, vesicles were proposed to stably contact the plasma membrane, but the team only saw direct contact when the vesicles were being exocytosed. Instead, vesicles near the active zone were docked by links that the authors named “tethers.” The tethers existed in long and short versions, the latter of which were absent in samples treated with tetanus toxin, indicating that they consist of SNARE proteins involved in membrane fusion. Vesicles with longer tethers were retained after mild synaptic stimulation, suggesting that they are at an early stage of docking and haven't yet formed a readily releasable pool of SNARE-tethered vesicles.

Author Vladan Lučić now plans to examine more specific drug treatments and the effects of genetic mutants on synaptic vesicle organization.

et al
J. Cell Biol.