Alignment of Synaptic Vesicle Macromolecules with the Macromolecules in Active Zone Material that Direct Vesicle Docking (original) (raw)
Figure 2
The lumen of synaptic vesicles after staining under different conditions.
Each panel shows a virtual slice, 1–2 nm thick, through vesicles at or near an active zone (PM, presynaptic membrane; asterisk, main body of AZM). (A) The NMJ was fixed and stained with glutaraldehyde, osmium tetroxide and uranyl acetate at room temperature. (B) The NMJ was fixed with glutaraldehyde at room temperature and, after rapid freezing, fixed further and stained with osmium tetroxide and uranyl acetate in acetone by freeze-substitution. (C–E) The NMJ’s were fixed by rapid freezing, stained with osmium tetroxide and uranyl acetate in acetone by freeze-substitution. While the lumen of synaptic vesicles at the NMJ fixed and stained with glutaraldehyde, osmium tetroxide and uranyl acetate at room temperature appears empty (A), the lumen of synaptic vesicles at the NMJ’s, stained with osmium tetroxide and uranyl acetate in acetone by freeze-substitution, regardless of whether they were fixed by rapid freezing or with glutaraldehyde at room temperature, contains an assembly of macromolecules (B–D; arrows). The staining of the vesicle membrane and luminal assembly in all cases is particulate. The particles in the luminal assemblies have greater electron density than those in the membrane. In (E), the vesicle membrane and luminal assemblies shown in (D) are overlaid with the portion of the 3D surface models of the entire vesicle membrane (blue) and the luminal assembly (orange) that were generated from this virtual slice, which establishes the limits of the membrane width throughout the membrane’s circumference, the edges of the luminal assemblies, and the sites of connection of the luminal assemblies to the membrane. Scale bar (A–C) = 50 nm, (D–E) = 25 nm.