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. 2021 Nov 22;118(48):e2113859118. doi: 10.1073/pnas.2113859118

Fig. 4.

Fig. 4.

Lysine residues in the syt1 juxtamembrane linker regulate evoked neurotransmitter release. (A) Representative iGluSnFR traces from one field of view after a single field stimulus (indicated by a black arrow) imaged at 100 Hz. The plots show the iGluSnFR responses from multiple individual regions of interest (ROIs) (black) and the average responses (green). Note that the KO, KO + Juxta K, and KO + Juxta K, F349A conditions resulted in fewer responses after stimulation. (B) Histograms of iGluSnFR (ΔF/F0) peaks plotted using 10-ms bins. The timing of the stimulus is indicated with a black arrow. Peaks were binned over the entire 1.5 s of recording. Conditions were color-coded as follows: WT (black), syt1 KO (gray), KO + WT (green), KO + Juxta K (blue), KO + F349A (orange), and KO + Juxta K, F349A (magenta). The histograms include combined data from three independent trials. The y-axis zoom-in histograms of iGluSnFR (ΔF/F0) peaks are shown as Insets to emphasize the presence of asynchronous release. (C) Average traces of iGluSnFR ΔF/F0 from a single field stimulus, indicated by a black arrow, imaged at 100 Hz. Note that the 1-ms stimulus was applied in the last millisecond of the frame that is indicated by a black bracket; this causes a small portion of the rise in signal to be captured within that frame. Conditions are labeled in the figure, and the same color scheme is used throughout. Briefly, WT (black), syt1 KO (gray), WT syt1 rescue (green), Juxta K mutant rescue (blue), F349A mutant rescue (orange), and the double Juxta K + F349A mutant rescue (magenta). Values are means with 95% CI error bars; the number of ROIs analyzed were WT (1,155), syt1 KO (212), WT rescue (748), Juxta K rescue (547), F349A rescue (1,128), and double-mutant rescue (324), collected from three separate trials. (D) Average peak iGluSnFR responses 10-ms post-stimulus from each condition. Quantification of data in C. Values are mean ± SEM from three independent trials. These data were normally distributed, and only WT rescue and F349A mutant rescue were statistically similar (ns); all other groups are different using ordinary ANOVA with Holm–Sidak’s correction. All comparisons and statistical analysis are provided in the figure source data.