Figure 6.

Role of the Syt1 C2B domain in regulation of synaptic vesicle size and density. A, Electron micrographs are shown for wild-type, syt1^−/− (null), and syt1^−/− rescued with Ca²⁺-binding defective C2 domains (C2A*-C2B*) or with dual C2B domains (C2B-C2B). Boxed regions in each micrograph are magnified 2.5 × in insets. Synaptic vesicles with abnormally large diameters are indicated with arrowheads. Scale bar, 500 nm. B, Summary of the mean mEJP amplitude is shown for syt1^−/− as well as syt1^−/− rescued with wild-type Syt1 (C2A-C2B), Ca²⁺-binding defective C2A*-C2B*, or dual C2A/C2B domains. Data for syt1^−/− as well as syt1^−/−, C2A-C2B and syt1^−/−, C2A*-C2B* from Figure 5 are presented for comparison. Data are mean ± SEM. ***p < 0.001 and *p < 0.05, one-way ANOVA with multiple comparisons using the Fisher's LSD test between syt1^−/− rescued with the C2A-C2B and the indicated genotypes. The number of NMJs examined for each genotype is listed in Figure 5. C, Cumulative diameter distributions of synaptic vesicles residing within a 100 nm radius of active zones are shown for syt1^−/− as well as those rescued with wild-type, Ca²⁺-binding defective C2 domains, or dual C2B domains. A boxed inset is provided for a detailed comparison. D, E, The number of synaptic vesicles near or in the vicinity of active zones (D) and total synaptic vesicle density (E) are summarized for each genotype indicated. Data are mean ± SEM. ***p < 0.001 and **p < 0.01, one-way ANOVA with multiple comparisons using the Fisher's LSD test between syt1^−/− rescued with the C2A-C2B and the indicated genotypes. Number of active zones analyzed for D and E: syt^−/−, 24 and 14; syt^−/−, C2A-C2B, 17 and 14; syt^−/−, C2A*-C2B*, 13 and 5; and syt^−/−, C2B-C2B, 22 and 14.