Fig. 4

Nlg1 mutants differently affect excitatory synaptic transmission and spine density. CA1 neurons of organotypic hippocampal slices from Nlg1 KO mice were single-cell electroporated at DIV 3–5 with GFP plus EV, Nlg1-WT, Nlg1-Y782A, or Nlg1-Y782F, then processed for electrophysiology or immunostaining and confocal imaging at DIV 13–15. a Dual whole-cell recording configuration (left) and corresponding image from an experiment (right). Scale bar, 10 µm. b Representative traces of evoked AMPAR- and NMDAR-mediated EPSCs recorded at −70 mV and +40 mV, respectively. Color sample traces correspond to electroporated neurons in the different conditions, and black traces correspond to control, unelectroporated neurons. c, d Average AMPAR- and NMDAR-mediated EPSC amplitudes, respectively, normalized to the control condition (the dashed line indicates 100%, dot plots corresponds to different pairs, from 4 independent experiments). e Average ratio between paired NMDAR- and AMPAR-mediated EPSCs. f Representative traces of spontaneous EPSCs recorded at −70 mV from electroporated and non-electroporated neurons. Black dots indicate synchronous events. g Average of paired spontaneous EPSC amplitudes recorded in electroporated neurons, normalized to the control condition (dashed line at 100%, dot plots corresponds to different pairs, from 3 independent experiments). h Representative confocal images of secondary apical dendritic segments from CA1 pyramidal neurons co-expressing GFP along with EV, Nlg1-WT, Nlg1-Y782A, or Nlg1-Y782F. Scale bar, 5 µm. i Average spine density for the same conditions (dot plots corresponds to different cells, from 4 independent experiments). Data from graphs c, d, and g were compared to the control condition (non-electroporated neurons) by a Wilcoxon matched-pairs signed rank test. Data in graph e were compared by a Kruskal–Wallis test followed by Dunn’s multiple comparison test. Data in graphs g and i were compared by one-way ANOVA followed by Tukey’s multiple comparison test (ns: not significant, *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001). Data represent mean ± SEM