Article Figures & Data
Figures
- Figure 1.
Diagram of the experimental paradigm. A, Summary of proposed steps for our medium-throughput phenotypic assay on primary neurons. B–E, Representative image (B) undergoing postprocessing in CellProfiler (C–E) for automated measurement of morphologic properties.
- Figure 2.
RhebY35L neuron neurite outgrowth over time. A–C, RhebY35L primary mouse cortical neurons after 3 (A), 6 (B) and 9 (C) DIV. D, E, Automated measurement of total neurite length per cell (D) and terminal branch number per cell (E) using CellProfiler (DIV3: n = 116, DIV6: n = 81, DIV9: n = 198). F, G, Manual measurement of total length per cell (F) and terminal branch number per cell (G) using FIJI (DIV3: n = 74, DIV6: n = 72, DIV9: n = 126). One to eight individual points from DIV6 and DIV9 conditions were omitted from the graphs by y-axis truncation to better visualize the mean (bar) and the change between conditions. Scale bar = 50 μm. 1 μm = 1.5384 pixels. DIV = days in vitro. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001. Significant differences were determined by one-way ANOVA with Tukey’s post hoc test.
- Figure 3.
Torin 1-induced morphologic changes in RhebY35L neurons. A, B, DIV3 tdTomato-positive RhebY35L cortical neurons treated with 100 nm Torin 1 or vehicle. C, D, Automated measurement of total neurite length per cell (C) and terminal branch number per cell (D) using CellProfiler (DMSO: n = 116, Torin 1: n = 60). E, Automated measurement of soma size using CellProfiler (DMSO: n = 144, Torin 1: n = 73). F, G, Manual measurement of total neurite length per cell (F) and terminal branch number per cell (G) using FIJI (DMSO: n = 74, Torin 1: n = 32). H, Manual measurement of soma size using FIJI (soma size; DMSO: n = 78, Torin 1: n = 33). One to eight individual points were omitted from the DMSO condition by y-axis truncation to better visualize the mean (bar) and the change between conditions. Scale bar = 50 μm. 1 μm = 1.5384 pixels. DIV = days in vitro. **p < 0.01, ***p < 0.001, ****p < 0.0001. Significant differences were determined by an unpaired two-tailed Student’s t test.
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