Figure 5.

Ultrastructural analysis reveals reduced synaptic vesicle (SV) complement at PicMut AZ. (A,B) Representative electron micrographs of PicWT (A) and PicMut (B) endbulb synapse active zones (AZs) demonstrating reduced SVs in the mutant AZ. Red arrows demarcate the boundaries of the AZ. Yellow arrow heads mark a region of the PicMut AZ almost devoid of SVs. (C) Unaltered postsynaptic density (PSD) length in PicMut endbulb AZs. Box and whisker plots present grand median, lower/upper quartiles, 10–90th percentiles. n.s.—p-value ≥ 0.05, Wilcoxon’s rank sum test. Each data point represents the PSD length of individual endbulb AZs. (D) Mean SV number per 100 nm of the PSD length significantly reduced at the PicMut endbulb AZs. Data represented as mean ± SEM (**p-value < 0.01, Student’s t-test). Each data point represents the mean SV number per 100 nm of PSD for each endbulb AZ imaged. (E) Mean SV number within 0–10, 10–20, 20–30, 40–50, till 90–100 nm of the AZ perpendicular to the AZ membrane into the presynaptic cytosol normalized to PSD length. Normality of distribution was tested with Jarque-Bera test and variances were compared with F-test. Non-normally distributed data are presented as box and whisker plots, lower/upper quartiles, 10–90th percentile; *p-value < 0.05, **p-value < 0.01, ***p-value < 0.001 and were tested by Wilcoxon rank sum test. Normally distributed data were tested by Student’s t-test and are presented as bar plot and error bars represent SEM. (F) Mean SV number within 40 nm of the AZ membrane (perpendicular to the AZ membrane into the presynaptic cytosol) normalized to the length of the PSD reduced in PicMut endbulb AZs. Box and whisker plots present grand median, lower/upper quartiles, 10–90th percentiles. ***p-value < 0.001, Wilcoxon rank sum test. Each data point represents the SV number per 100 nm of PSD within 40 nm for each endbulb AZ imaged. PicWT (N = 3; n = 32) in black and PicMut (N = 2; n = 33) in red (N, number of animals; n, number of AZs).