Fig. 5.

Rac inhibition affected CDK5 shRNA-miR-induced spine morphogenesis in mature neurons. a Primary hippocampal neurons were treated with 0.01-, 0.05-, 0.1-, 1-, 2.5-, 5-, 10-, 25-, 50-, and 100-μM NSC23766 for 24 h. After treatment, the LDH released from medium was measured to determine cytotoxicity. b Then, neurons were transduced with Scr or CDK5 shRNA-miR and DIV19 treated with 1- or 5-μM NSC23766 for 30 min to analyze spine morphogenesis. Rac and RhoA activities were quantified for the 30-min NSC23766 treatment as the amount of Rac-GTP and RhoA-GTP corresponding to the optical density observed using ELISA (λ = 490 nm). The data are presented as the mean ± SEM from n = 4 per duplicate. c Morphological characters show: AAV vector viral eGFP-tagged Scr and CDK5 shRNA-miR (green), nuclei visualized with Hoechst staining (blue), and F-actin cytoskeleton visualized with Alexa 594 Phalloidin dye (red). Magnification ×60. Scale bar 20 µm, n = 4. Insets are showing the proximal dendritic shaft for each treatment. Scale bars 2 μm. *p < 0.05; **p < 0.01; ANOVA with Tukey’s test for b