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. 2009 Sep 17;284(47):32803–32812. doi: 10.1074/jbc.M109.039149

FIGURE 3.

FIGURE 3.

The rate of exocytosis and the size of recycling vesicle pool in μ2 mutant-expressing neurons. A, representative trace of exocytosis of vG-pH in μ2 mutant-expressing neurons. Neurons were triple-transfected with vG-pH, shRNA μ2, and rescue of each mutant and stimulated (1200 action potentials at 10 Hz) to trigger exocytosis of the entire recycling vesicle in the presence of bafilomycin (1 μm). YK, Y334A,K335A; KKKEEE, K341E/K343E/K345E. Error bars indicate S.E. B, the rate of exocytosis was determined from a single exponential fit. Time constants for exocytosis are not significantly different across the different mutants (WT, 42.3 ± 4.5 s, n = 12; YXXϕ, 45.5 ± 7.5 s, n = 14; T156A, 37.6 ± 5.0 s, n = 6; Y334A,K335A, 43.4 ± 7.2 s, n = 13; K341E/K343E/K345E, 45.3 ± 6.9 s, n = 12; 3M, 45.4 ± 11.2 s, n = 7; All-in-one, 42.0 ± 7.0 s, n = 5). C, the relative size of the recycling pool in transfected neurons was determined by measuring the maximum intensity of the change in vG-pH fluorescence during prolonged stimulation in the presence of bafilomycin. All cells were triple-transfected with same DNA mixture condition. Relative pool size to WT was determined as follows: YXXϕ, 92.2 ± 9.7%, n = 12; T156A, 94.1 ± 16.4 s, n = 12; Y334A,K335A, 90.2 ± 8.2 s, n = 12; K341E/K343E/K345E, 94.7 ± 9.9 s, n = 11; 3M, 81.9 ± 15.5 s, n = 6; All-in-one, 71.9 ± 11.1 s, n = 8) *, p <0.05. The solid line indicates the SV recycling pool of AP-2KD (12).