Fig. 5

Vesicle movements and exocytosis. (A) Cross-section of prolate-ellipsoidal “cell” with long axis 1.8 µm and short axis 1 µm, membrane thickness 3 voxels containing vesicles of diameter = 100 nm. Red spheres represent single molecules in the cytoplasm, green spheres represent molecules in the vesicle membrane. (B) Cross-section of a synthetic membrane bound volume showing vesicles fused to the plasma membrane via a 50 nm diameter tube (left), a vesicle-width (200 nm diameter) tube (center) and a vesicle that fuses via direct contact with the plasma membrane (right). (C) Main graph shows fluorescence time-course of modelled exocytosis region averaged over ten simulated fusion events, synchronized to docking time (defined as t = −1 s). Note decay rate increases when fusion tube diameter was increased from 50 to 200 nm (green vs blue lines). Fluorescence decay was significantly faster when molecules were instantaneously transferred from the vesicle to the cell membrane (red line). Furthermore, a fluorescence spike was observed at the time of fusion because molecules in the vesicle membrane (located ~ 100 nm above the coverslip surface) moved rapidly to the basal cell membrane, where TIR illumination intensity is higher. A similar spike in fluorescence was observed in experimental data obtained using TIRFM illumination (insert). The model indicates that observation of a fluorescence spike and fast decay rate is consistent with rapid vesicle fusion. The finite delay between vesicle “docking” and fusion requires a rate-limiting step occurring before fusion pore formation.