Fig. 5

L100P hippocampal neurons exhibit normal expression of excitatory synaptic markers and dendrites morphology. a Immunostaining for PSD-95 (red) and synapsin-1(SYN, blue) in primary neuronal cultures from WT (a-c) and L100P (d-f) hippocampus. Scale bar figures, 50 μm; scale bar blow-out, 10 μm. b-d Summary of immunostaining density for PSD-95 (b), synapsin-1 (c) and excitatory synapse formation (d) in primary cultured L100P and WT hippocampal neurons. Spine density was calculated on secondary and tertiary dendrites. Unpaired t-test, n = 21 neurons for WT group and n = 24 neurons for L100P group. All data are shown as means ± SEM. e-f Summary histogram showing that L100P and WT neurons have similar total dendrites length (e) and dendrites number (f) in primary, secondary and tertiary dendritic segments. Two-way ANOVA and Bonferroni post-tests. WT group, n = 18 neurons; L100P group, n = 21 neurons. Data were collected from three to four independent experiments. All data are shown as means ± SEM. g-h Immunoblots showing synaptic proteins expression in the forebrain of WT mice and L100P mutants. Immunoblots of synaptosome (SS) fraction (g) and PSD-fraction (h) with antibodies against synapsin-1, synaptophysin, PSD-95, GluA1, αCaMKII, GluN2B and β-tubulin. Upper, representative sample immunoblot. β-tubulin is used to control for protein loading. Lower, summary of synaptic proteins expression in SS and PSD fractions. For quantification, the amount of each protein is first standardized as a ratio of protein/tubulin and then normalized by average protein/tubulin ratio of WT group before statistical analysis. Quantifications are based on the average of 6–12 independent experiments. *P < 0.05, unpaired t-test. All data are shown as means ± SEM.