Figure 2.
Overexpression of wild-type synaptotagmin 1 results in a larger fast burst and a smaller slow burst of secretion without changing fusion rates. A, Mean calcium concentration (top; error bars represent SEM), capacitance (middle), and amperometric (bottom) responses after a step-like elevation of [Ca2+]i induced by flash photolysis (flash at arrow). Both the capacitance and amperometric traces display a burst-like increase within the first 0.5 s after the flash in the control cells (+/+ and +/-, black; n = 43 cells, N = 8 animals), followed by a slower sustained phase of secretion representing vesicle recruitment (priming) and consecutive fusion. Syt1 null cells lack the fast burst component either measured as cell capacitance or by amperometry (red; n = 41, N = 8), which is reconstituted when cells overexpress wild-type Syt1 (Syt1-WT, blue; n = 42, N = 8). The inset shows the first half-second of the capacitance traces scaled to control level (black). Whereas secretion from the null cells is slowed down (red), overexpression of Syt1 (blue) appears to be faster than control. B, C, The mean ± SEM of kinetic parameters. There was no significant difference between the time constants of both the fast or the slow burst components (B), whereas overexpression of Syt1 in null cells significantly increased the size of the fast burst component and decreased the size of the slow burst component compared with control cells (C). Note that knock-out (KO) cells completely missed the fast burst component, without significant changes in the other kinetic parameters (for color coding, see A). D-F, Overexpression of Syt1-WT in control cells (Syt1-WT, gray; n = 28) also increased fast burst without changing total burst size compared with non-infected control cells (black; n = 28), as seen in the middle capacitance trace (D). Triple-exponential fit revealed that, like in the knock-out cells, overexpression of Syt1 in control cells did not change time constants (E) but led to an increased size of the fast burst accompanied with a decreased slow burst size (F). **p < 0.01; ***p < 0.001.