Figure 4.
EVL is required for DF morphogenesis and influences dendritic spine plasticity. (A) Live primary cortical neurons derived from wild-type (WT) or EVL knockout (KO) mice at indicated days in vitro expressing mRuby2-LifeAct. Top: Full cell image. Middle: Indicated segment of dendrite presented with an intensity-coded LUT. Bottom: Maximum intensity projection of temporally color-coded binary mask outline, illustrating DF dynamics during imaging (5 s interval, 5 min duration). Scale bars = 10 µm. (B) Scatterplot of average length reached during the duration of imaging for indicated conditions. Median ± interquartile range (IQR). Mixed-effects model; n = 280–641 total DF from 4–6 neurons per biological replicate, N = 3 biological replicates. (C) Scatterplot of median protrusion rates of DFs (the median of values when instantaneous change in length was greater than +0.0128 µm/s (motile), protruding). Median ± IQR. Mixed-effects model; n = 280–641 total DF from 4–6 neurons per biological replicate, N = 3 biological replicates. (D) High-density cultures of wild-type, EVL knockout, or wild-type overexpressing EVL neurons at day in vitro 14 (D14) expressing EGFP-LifeAct and synapse reporter PSD95-FingR-mRuby2. Left: Full cell image. Inset: EVL expression. Right: Indicated segment of dendrite. Scale bars = 10 µm. (E) Scatterplot of average dendritic spine density per neuron of indicated conditions. Median ± IQR. One-way ANOVA corrected with Holm-Sidak multiple comparisons; n = 38 total neurons for each condition, N = 3 biological replicates. (F) Filmstrip of segment of dendrite from live cortical neurons derived from wild-type or EVL knockout mice at D14 expressing EGFP-LifeAct and PSD95-FingR-mRuby2 showing synapse formation and dynamics. Inset: Illustration of region indicated by asterisks. Scale bars = 10 µm. (G) Scatterplot of average duration of synapse persistence per neuron as quantified by PSD95-positive foci tracked in time. Median ± IQR. Unpaired t test; n = 12 (wild-type) and 13 (knockout) total neurons; N = 3 biological replicates. (H) Scatterplot of average speed per neuron of PSD95-positive foci tracked in time. Median ± IQR. Unpaired t test; n = 12 (wild-type) and 13 (knockout) total neurons; N = 3 biological replicates. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001, n.s. is not significant. See also Fig. S5, Video 3, and Video 4.

EVL is required for DF morphogenesis and influences dendritic spine plasticity. (A) Live primary cortical neurons derived from wild-type (WT) or EVL knockout (KO) mice at indicated days in vitro expressing mRuby2-LifeAct. Top: Full cell image. Middle: Indicated segment of dendrite presented with an intensity-coded LUT. Bottom: Maximum intensity projection of temporally color-coded binary mask outline, illustrating DF dynamics during imaging (5 s interval, 5 min duration). Scale bars = 10 µm. (B) Scatterplot of average length reached during the duration of imaging for indicated conditions. Median ± interquartile range (IQR). Mixed-effects model; n = 280–641 total DF from 4–6 neurons per biological replicate, N = 3 biological replicates. (C) Scatterplot of median protrusion rates of DFs (the median of values when instantaneous change in length was greater than +0.0128 µm/s (motile), protruding). Median ± IQR. Mixed-effects model; n = 280–641 total DF from 4–6 neurons per biological replicate, N = 3 biological replicates. (D) High-density cultures of wild-type, EVL knockout, or wild-type overexpressing EVL neurons at day in vitro 14 (D14) expressing EGFP-LifeAct and synapse reporter PSD95-FingR-mRuby2. Left: Full cell image. Inset: EVL expression. Right: Indicated segment of dendrite. Scale bars = 10 µm. (E) Scatterplot of average dendritic spine density per neuron of indicated conditions. Median ± IQR. One-way ANOVA corrected with Holm-Sidak multiple comparisons; n = 38 total neurons for each condition, N = 3 biological replicates. (F) Filmstrip of segment of dendrite from live cortical neurons derived from wild-type or EVL knockout mice at D14 expressing EGFP-LifeAct and PSD95-FingR-mRuby2 showing synapse formation and dynamics. Inset: Illustration of region indicated by asterisks. Scale bars = 10 µm. (G) Scatterplot of average duration of synapse persistence per neuron as quantified by PSD95-positive foci tracked in time. Median ± IQR. Unpaired t test; n = 12 (wild-type) and 13 (knockout) total neurons; N = 3 biological replicates. (H) Scatterplot of average speed per neuron of PSD95-positive foci tracked in time. Median ± IQR. Unpaired t test; n = 12 (wild-type) and 13 (knockout) total neurons; N = 3 biological replicates. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001, n.s. is not significant. See also Fig. S5, Video 3, and Video 4.

This Feature Is Available To Subscribers Only

or Create an Account

Close Modal
Close Modal