For each KO line, the average intracellular [Ca2+] ± SEM and the average △Cm are shown in panels (A–C, E). Single gene deletions of (A) Unc13a, (B) Unc13c, or (C) Baiap3 did not impair LDCV exocytosis. (D) Summary of burst sizes, sustained release rates, and time constants. (E) LDCV exocytosis is dramatically reduced in Unc13aKOUnc13bKO cells, Unc13aWTUnc13bKO cells, and Unc13aHetUnc13bKO cells. This reduction is primarily due to the absence of ubMunc13-2. (F) Fast burst, slow burst and the rate of sustained release are reduced in the absence of Munc13-1 and ubMunc13-2, as well as in the absence of ubMunc13-2 alone (ANOVA with post-hoc Tukey’s test). (G) Compared to Unc13aWTUnc13bKO cells, the deletion of Munc13-1 causes significant reductions in the slow burst and the rate of sustained release (Student’s t-test, two-tailed). (H) Delay of the onset of exocytosis after the flash stimulus (ANOVA with post-hoc Tukey’s test). (I) Normalized traces show identical release kinetics of the exocytotic burst. (J) Summary of the release components shown in panels (E, F, G). Time constants are not significantly different (ANOVA with post-hoc Tukey’s test). (*p < 0.05, **p < 0.01, ***p < 0.001). See also Figure 2—figure supplement 1.