Figure 4
SR58611A increases glutamate release probability and the number of readily releasable vesicles via activating the Ca2+/calmodulin/MLCK/myosin II pathway. (A) Depressions of EPSCs during trains (20 stimuli) of 50 Hz stimulation before and during application of SR58611A (15 µM). The amplitude of EPSCs by each stimulus was measured by resetting the base line each time at a point within 0.5 ms before the beginning of each stimulation artifact. (B) Cumulative amplitudes of EPSCs during the 50 Hz train before and during SR58611A (15 µM) application. Amplitudes of EPSCs from 16 to 20th were fitted with a linear regression line and extrapolated to time 0 for estimating the readily releasable pool size. (C) Application of SR58611A (15 µM) increased mean number of releasable vesicles (N) multiplied by mean quantal size (q), and SR58611A-enhanced Nq was abolished in the presence of BAPTA-AM (100 µM), W7 (50 µM), wortmannin (10 µM), or blebbistatin (1 µM), respectively. (D) Application of SR58611A (15 µM) increased mean release probability (Pr), and SR58611A-enhanced Pr was not observed in the presence of BAPTA-AM (100 µM), W7 (50 µM), wortmannin (10 µM), or blebbistatin (1 µM), respectively. (E) Upper: experimental protocol. A conditioning train (20 stimuli at 50 Hz) was followed by a test stimulus. The intervals between the end of the conditioning train and the beginning of the test stimulus were 0.1 s, 0.5 s, 1 s, 2 s, 5 s or 10 s. The interval between each sweep was 30 s to allow the refilling of the vesicles. Lower: EPSCs evoked by the test pulse from the same synapse at different intervals were aligned and superimposed before (left) and during (right) application of SR58611A (15 µM). (F) Time course of recovery from depletion before and during the application of SR58611A (15 µM) expressed as percentage recovery = (Itest − Is)/(I1st − Is) × 100, where Itest is the EPSCs evoked by the test pulse, Is is the steady-state current left after the conditioning train (the average of the last 5 EPSCs evoked by the conditioning train), I1st is the EPSCs evoked by the 1st stimulus of the conditioning train. Data before and during the application of SR58611A (15 µM) from 7 cells were fitted by a single exponential function. *P < 0.05 versus control.
