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. 2019 May 31;20(11):2692. doi: 10.3390/ijms20112692

Figure 2.

Figure 2

Vesicle membrane–presynaptic membrane (VM–PM) contact sites of two docked synaptic vesicles on the presynaptic membrane, at two different active zones and measured membrane thicknesses. (A,E) Virtual slices (0.5 nm thick) of the two docked synaptic vesicles on their presynaptic membranes at active zones of the frog NMJs. Scale bar = 25 nm. (B,F) Surface models of the two docked synaptic vesicles (light blue) and the stretch of presynaptic membranes (light gray) are superimposed on the two virtual slices. (C,G) Portions of the models (10 nm thick) of the vesicles and presynaptic membranes are visualized. The vertices used to generate the models provide continuous surfaces for the vesicle membrane and presynaptic membrane, while accounting for irregularities in the membrane staining. (D,H) Frequency distributions of thousands of vertex-to-vertex thickness measurements made across the entire membranes of the two docked synaptic vesicles and the presynaptic membranes, from their 3D surface models. For the docked synaptic vesicle in A–C, the membrane thicknesses of the individual membranes away from their contact sites are 8.3 nm ± 1.5 nm (mean ± SD) for the VM and 8.1 nm ± 1.1 nm (mean ± SD) for the PM, and the combined thickness of the vesicle membrane and presynaptic membrane at the contact site (16.4 nm ± 1.5 nm; mean ± SD) is not different from the expected combined thickness (16.5 nm ± 1.9 nm; mean ± SD) obtained by summing randomly the vesicle membrane and presynaptic membrane thicknesses away from the contact site (Bootstrap test, p = 0.97; See Materials and Methods). For the docked synaptic vesicle in E–G, the VM and the PM are 7.5 nm ± 1.0 nm and 7.8 nm ± 1.5 nm, respectively. The combined thickness of the vesicle membrane and presynaptic membrane at the contact site (11.6 nm ± 2.4 nm; mean ± SD) is less than its expected combined thickness (Bootstrap test, p < 0.05), showing that the docked vesicle is merged with the presynaptic membrane. Note that each membrane varies in thickness from one vertex to another, mainly due to irregular staining of the membrane.