Figure 1.

CpxII controls magnitude and time course of synchronous exocytosis. (A) Mean flash-induced [Ca²⁺]i levels (top) and corresponding CM responses (bottom) of wt (n = 26), CpxII ko (n = 33), and wt cells expressing either GFP (wt + gfp, n = 16) or CpxII (wt + CpxII, n = 23). Flash is at t = 0.5 s. (B) RRP and SRP size are reduced in the absence and increased with overexpression of CpxII. (C) CpxII loss slows the time constants of RRP and SRP exocytosis (τ_RRP and τ_SRP). (D) Normalized CM (as shown in A) scaled to the wt response 1 s after the flash. (inset) Extended scaling of normalized CM during the first 50 ms after flash (arrow) depicting the delayed onset of CpxII secretion. (E) Mean exocytotic delay determined from fitting individual cellular responses. (F–J) CpxII-dependent control of tonic exocytosis determines the EB size. (F) Exemplary recording of CM, series conductance (Gs), membrane conductance (Gm), and [Ca²⁺]i during infusion of submicromolar [Ca²⁺]i. A sustained capacitance increase (ΔCM) over 120 s precedes the rapid CM rise in response to the flash-evoked Ca²⁺ increase (arrow). (G) Mean tonic exocytosis determined from the cells shown in A. (H) Rate of tonic exocytosis. (I) Correlation between tonic secretion rate and EB size for the indicated groups. The regression line (r² = 0.94) intercepts the y axis (b, y intercept) at 35 fF/10 s (representing ∼4 vesicles/s), a value that comes close to the vesicle priming rate at 500 nM [Ca²⁺]i (rate_(500nM) = 5.35 × 10⁻³/s × 780 unprimed vesicles = 4 vesicles/s; Sørensen, 2004), suggesting that a balance between Ca²⁺-dependent priming and CpxII-dependent clamping of tonic secretion determines the EB size. (J) The mean preflash [Ca]i values for the tonic CM responses shown in G are nearly identical. *, P < 0.05; **, P < 0.01; ***, P < 0.001, Student’s t test versus the corresponding control. Error bars indicate means ± SEM.