Figure 9.
PI(3)P synthesis is necessary for the recycling of SNX17 and SNX27 cargoes upon cLTP. (A) Representative confocal images of surface β1-integrin levels of DIV17 hippocampal neurons that were infected at DIV11 with lentiviruses carrying either ctrl- or SNX27-shRNAs. Neurons were treated in the presence or absence of cLTP and live labeled with an anti-surface β1-integrin antibody for 15 min, followed by fixation and immunostaining for MAP2. Scale bar, 5 µm. (B) The mean intensity of β1-integrin in the first 50 µm of secondary dendrites was quantified, and values were normalized to crtl-shRNA. ctrl-shRNA: 1,000 ± 0.031, N = 40 neurons; ctrl-shRNA cLTP: 1.179 ± 0.038, N = 40 neurons; VPS34-shRNA: 0.770 ± 0.029, N = 40 neurons; VPS34-shRNA cLTP: 0.779 ± 0.031, N = 40 neurons. Three independent experiments. Data were analyzed by one-way ANOVA with Tukey’s post hoc test, ***P < 0.005, ****P < 0.001. Error bars are SEM. (C) Representative confocal images of surface GluA1 levels of DIV17 hippocampal neurons that were infected at DIV11 with lentiviruses carrying either ctrl- or VPS34-shRNAs. Neurons were treated in the presence or absence of cLTP and live labeled with an anti-surface GluA1 antibody for 15 min, followed by fixation and immunostaining for MAP2. Scale bar, 5 µm. (D) The mean intensity of surface GluA1 in the first 50 µm of secondary dendrites was quantified, and values were normalized to crtl-shRNA. ctrl-shRNA: 1,000 ± 0.033, N = 40 neurons; ctrl-shRNA cLTP: 1.171 ± 0.052, N = 40 neurons; VPS34-shRNA: 0.653 ± 0.027, N = 40 neurons; VPS34-shRNA cLTP: 0.688 ± 0.031, N = 40 neurons. Three independent experiments. Data were analyzed by one-way ANOVA with Tukey’s post hoc test, **P < 0.01, ****P < 0.001. Error bars are SEM. DIV, days in vitro.

PI(3)P synthesis is necessary for the recycling of SNX17 and SNX27 cargoes upon cLTP. (A) Representative confocal images of surface β1-integrin levels of DIV17 hippocampal neurons that were infected at DIV11 with lentiviruses carrying either ctrl- or SNX27-shRNAs. Neurons were treated in the presence or absence of cLTP and live labeled with an anti-surface β1-integrin antibody for 15 min, followed by fixation and immunostaining for MAP2. Scale bar, 5 µm. (B) The mean intensity of β1-integrin in the first 50 µm of secondary dendrites was quantified, and values were normalized to crtl-shRNA. ctrl-shRNA: 1,000 ± 0.031, N = 40 neurons; ctrl-shRNA cLTP: 1.179 ± 0.038, N = 40 neurons; VPS34-shRNA: 0.770 ± 0.029, N = 40 neurons; VPS34-shRNA cLTP: 0.779 ± 0.031, N = 40 neurons. Three independent experiments. Data were analyzed by one-way ANOVA with Tukey’s post hoc test, ***P < 0.005, ****P < 0.001. Error bars are SEM. (C) Representative confocal images of surface GluA1 levels of DIV17 hippocampal neurons that were infected at DIV11 with lentiviruses carrying either ctrl- or VPS34-shRNAs. Neurons were treated in the presence or absence of cLTP and live labeled with an anti-surface GluA1 antibody for 15 min, followed by fixation and immunostaining for MAP2. Scale bar, 5 µm. (D) The mean intensity of surface GluA1 in the first 50 µm of secondary dendrites was quantified, and values were normalized to crtl-shRNA. ctrl-shRNA: 1,000 ± 0.033, N = 40 neurons; ctrl-shRNA cLTP: 1.171 ± 0.052, N = 40 neurons; VPS34-shRNA: 0.653 ± 0.027, N = 40 neurons; VPS34-shRNA cLTP: 0.688 ± 0.031, N = 40 neurons. Three independent experiments. Data were analyzed by one-way ANOVA with Tukey’s post hoc test, **P < 0.01, ****P < 0.001. Error bars are SEM. DIV, days in vitro.

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