Figure 5From: Automated 4D analysis of dendritic spine morphology: applications to stimulus-induced spine remodeling and pharmacological rescue in a disease modelA PI3 kinase inhibitor rescues spine morphology in neurons from Fmr1 knockout mice. (a) Hippocampal neurons cultured from wild type (WT) or Fmr1 knockout (KO) mice were treated with vehicle or a PI3 kinase inhibitor (LY294, 10 μM) for 72 hours starting at 15 DIV. Neurons were transfected with a plasmid encoding Lifeact-ruby at 17 DIV and fixed 24 hours later. The images depict representative dendritic regions from deconvolved z-series images (scale bar is 5 μm). (b) Spine density was measured using our automated approach for WT and KO neurons treated with either vehicle or LY294 (n = 55-60 neurons; ANOVA [F = 3.996, P = 0.009]; post-hoc Fisher's LSD: *P = 0.017, #P = 0.028). Cumulative distributions of (c) spine head width, (d) length, and (e) volume were plotted for each group (Kolmogorov-Smirnov test: head width [WT vs. KO: P = 0.002, WT vs. WT LY294: P = 0.235, KO vs. KO LY294: P = 0.009], spine length [WT vs. KO: P = 0.009, WT vs. WT LY294: P = 0.537, KO vs. KO LY294: P = 0.014], spine volume [WT vs. KO: P < 0.001, WT vs. WT LY294: P = 0.158, KO vs. KO LY294: P < 0.001]). (f) Spines were classified as stubby, mushroom, and thin based on the automated geometric measurement, and the values were plotted as percentage of total spines per treatment group (n = 55 - 60 neurons; ANOVA with post-hoc Fisher's LSD: *P = 0.043, #P = 0.006, ‡P = 0.043).Back to article page