Fig. 2
Synaptic density, mEPSC properties and pre-synaptic dysfunction. a For each line, [control, C9ORF72RE- (C9) and C9ORF72RE gene-edited- (C9-Δ) cortical neurons], low magnification images (scale bar, 20 μm) show immunostaining for βIII-tubulin (yellow, top left), synapsin-1 (red), PSD-95 (green) and a PSD-95/synapsin-1 composite view (lower) from which the region of interest is highlighted. The region of interest is shown with greater higher magnification image below (scale bar, 10 μm). b Mean ± s.e.m. co-localised PSD-95/synapsin-1 puncta per 20 μm of neurite length for each line type (Control, N = 3; C9, N = 12; C9-Δ, N = 12). Data for C9 and C9-Δ are pooled from three respective C9ORF72RE- and gene-edited pairs – individual pair comparisons are presented in Supplementary Figure 6A. Data from each de novo preparation represents a mean obtained from four coverlips each consisting of five randomly selected image fields. c Sample traces from recordings of mEPSC events from control, C9ORF72RE- and C9ORF72RE gene-edited-cortical neurons. Recordings were made at a holding potential of − 84 mV. Scale bar; 20 pA, 10 s. mEPSCs in all lines are mediated by Ca2+-impermeable (NASPM-insensitive) AMPA receptors (CNQX-sensitive). d Mean ± s.e.m. mEPSC frequency and amplitude for each line (Control, n = 14, N = 3; C9, n = 52, N = 12; C9-Δ, n = 42, N = 11). Data are pooled for C9 and C9-Δ – individual pair comparisons are presented in Supplementary Figure 7. e Sample traces from recordings of mEPSC events before and in the presence of sucrose from control (Con), C9ORF72RE-(C9) and C9ORF72RE gene-edited (C9-Δ)-cortical neurons. The transient and steady-state phases evoked by sucrose addition are highlighted. Note the difference in mEPSC frequency in these phases versus baseline. Scale bar, 50 pA, 5 s. f Mean ± s.e.m. fold change in mEPSC frequency for each line for the transient (Control, n = 14, N = 3; C9, n = 28, N = 10; C9-Δ, n = 30, N = 9) and steady-state phases (Control, n = 8, N = 2; C9, n = 23, N = 8; C9-Δ, n = 23, N = 8). Data are pooled for C9 and C9-Δ – individual pair comparisons are presented in Supplementary Figure 8. Significance of data in the figure determined by one-way ANOVA with Bonferroni’s post hoc test