Figure 4. Thin negatively regulates release-ready vesicle number.

(A) Representative excitatory postsynaptic currents (EPSCs) (individual sweeps and averages are shown in light colors and black, respectively), and mEPSCs (insets) of controls (elav^c155-Gal4>UAS-mCherry^RNAi, ‘Ctrl.’, gray) and presynaptic thin^RNAi (elav^c155-Gal4>UAS-thin^RNAi, ‘thin^RNAi (pre)’, red). Mean mEPSC amplitudes (B), EPSC amplitudes (C), and quantal content (D) of the indicated genotypes. (E) Representative EPSC train (60 Hz, 60 stimuli, top) and cumulative EPSC amplitudes (‘cum. EPSC’, bottom) of control and presynaptic thin^RNAi. The blue line is a line fit to the last 15 cum. EPSC amplitudes that was back-extrapolated to t = 0 (see Materials and methods). Mean readilyreleasable vesicle pool (RRP) size (cum. EPSC/mEPSC) (F), release probability (‘p_r’, EPSC1/cum. EPSC) (G), and paired-pulse ratio (‘PPR’, EPSC2/EPSC1) (H) of the indicated genotypes. Note the increase in EPSC amplitude and RRP size in presynaptic thin^RNAi. Mean ± standard error of the mean (SEM); Ctrl.: n = 16, thin^RNAi: n = 17; *p < 0.05; ***p < 0.001; n.s.: not significant; Student’s t-test.

Figure 4—figure supplement 1. Presynaptic thin^RNAi expression blocks presynaptic homeostatic plasticity (PHP) and induces a slight increase in AZ number.

(A) Maximum intensity projection of a control neuromuscular junction (NMJ) (elav^c155-Gal4>UAS-mCherry^RNAi, ‘Ctrl.’, left) and after presynaptic thin^RNAi expression (elav^c155-Gal4>UAS-thin^RNAi, ‘thin^RNAi’, right) (muscle 6) stained against the Drosophila neuronal membrane marker anti-HRP (‘HRP’) and the active-zone marker Bruchpilot (‘Brp’); scale bar, 10 µm. Mean HRP area per NMJ ‘HRP area’ (B), Brp puncta number per NMJ ‘Brp puncta #’ (C), Brp puncta number/HRP area per NMJ ‘Brp density’ (D), and Brp puncta fluorescence intensity (E) of the indicated genotypes. Elav^c155-Gal4>UAS-mCherry^RNAi: n = 12, elav^c155-Gal4>UAS-thin^RNAi: n = 16. (F) Representative mEPSCs (top) and excitatory postsynaptic currents (EPSCs) (bottom, individual sweeps and averages are shown in light colors and black, respectively) after presynaptic thin^RNAi expression (elav^c155-Gal4>UAS-thin^RNAi, ‘thin^RNAi’) in the absence (left, light red) and presence of the glutamate receptor (GluR) antagonist philanthotoxin-443 (PhTX) (right, dark red). Mean mEPSC amplitudes (G), EPSC amplitudes (H), and quantal content (I) in the absence (‘−’, light red) and presence (‘+’, dark red) of PhTX. Note that PhTX did not enhance quantal content after presynaptic thin^RNAi expression, indicating impaired PHP. Mean ± standard error of the mean (SEM); thin^RNAi − PhTX: n = 14, thin^RNAi + PhTX: n = 13; ***p < 0.001; n.s.: not significant; Student’s t-test.