Journal: iScience
Article Title: TAOK2 controls synaptic plasticity and anxiety via ERK and calcium signaling
doi: 10.1016/j.isci.2025.113712
Figure Lengend Snippet: MAPK and calcium signaling is reduced in Taok2 cKO neurons (A–D) Luciferase assay using EGR1p (A), SARE- (B), FOSBp- (C), and CRE- (D) sensor-detected MAPK (A–C) and calcium (B, D)-dependent signaling activity in primary cortical neurons. Neurons were treated with AMPA (1 μM), BDNF (10 ng/mL), or bicuculline (BIC, 1 μM) on day in vitro 12 (DIV12) for 4 h. ∗ p < 0.05; ∗∗ p < 0.01; ∗∗∗ p < 0.001; ∗∗∗∗ p < 0.0001; n.s., not significant; Student’s t test (two-sided). (E) Imaging of control and Taok2 cKO primary cortical neurons infected with AVVs expressing the synapsin1-driven GCaMP6f calcium sensor. Images were taken before and after stimulating cells with 25 mM KCl. (F) Quantification of cellular GCaMP6f responses to 25 mM KCl. Intensities from responding cells were normalized to the background and pooled for analysis. Gray arrowheads on the x axis represent the time points at which images were taken before and after stimulation as shown in (E). The line represents the mean, and the shaded area represents the SEM; n = 30 responding cells from two independent cultures. ∗∗∗ p < 0.001 and mixed ANOVA. (G) Western blots indicate reduced phosphorylated Mek1/2 (p-Mek1/2) and Erk1/2 (p-Erk1/2) in primary mouse cortical neurons. Neurons were treated with AMPA (1 μM) on DIV12 for 4 h. (H and I) Quantification of western blot data shown in (G). Data of p-Erk1/2 are relative to total Erk1/2 (H), and data of p-Mek1/2 are relative to total Mek1/2 (I). Values are presented as mean ± SD. ∗ p < 0.05; ∗∗ p < 0.01; ∗∗∗ p < 0.001; ∗∗∗∗ p < 0.0001; Wilcoxon rank-sum test (two-sided).
Article Snippet: On DIV6, half the medium was changed and the neurons were infected with AAVs for expression of mRuby2-P2A-GCaMP6f under the control of the human synapsin1 promoter (hSyn1p) (Addgene #50943) at an MOI of 1000.
Techniques: Luciferase, Activity Assay, In Vitro, Imaging, Control, Infection, Expressing, Western Blot
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