The synaptic vesicle cycle: a single vesicle budding step involving clathrin and dynamin

Area of Science:

• Neuroscience
• Cell Biology
• Molecular Biology

Background:

• Synaptic vesicle reformation after exocytosis is crucial for neuronal function.
• Clathrin-coated vesicles and endosome-like vacuoles are implicated in this process.
• The precise mechanism of vesicle budding from internal intermediates remained unclear.

Purpose of the Study:

• To investigate the mechanisms of synaptic vesicle budding from endosome-like intermediates.
• To clarify the role of clathrin and dynamin in synaptic vesicle recycling.

Main Methods:

• Electron microscopy (EM) of lysed nerve terminals and subfractions incubated with GTPγS.
• Immunoreactivity for AP2 and AP180 in coated budding intermediates.
• Observation of coated vesicle buds in intact cultured hippocampal neurons stimulated with high K+.

Main Results:

• Numerous clathrin-coated budding intermediates, positive for AP2/AP180 and dynamin, were observed on both plasma membrane and internal vacuoles.
• Internal vacuoles were functionally segregated but sometimes connected to the plasma membrane.
• Coated vesicle buds resembled synaptic vesicles in size and contained the synaptic vesicle protein synaptotagmin.

Conclusions:

• Endosome-like intermediates likely originate from bulk plasma membrane uptake.
• Clathrin- and dynamin-mediated budding occurs in parallel from both plasma membrane and internal membranes.
• A unified synaptic vesicle recycling model involving a single clathrin/dynamin-dependent budding step is proposed.