Dendritic spine elongation and new filopodia formation are induced by
RGD-containing peptide in 14 DIV hippocampal neurons.
A–C, Confocal images of GFP-labeled 14 DIV
mouse hippocampal neurons untreated (A), treated with
500 μm control RAD-containing peptide
(B), or treated with 500 μm
RGD-containing peptide (C). GFP fluorescence was used
to visualize dendritic morphology. RGD-treated neurons showed increased number of
filopodia-like protrusions and increased length of dendritic spines, whereas neurons
treated with control RAD-containing peptide showed no changes in dendritic spine
morphology or length. Scale bars: top panel, 10 μm; bottom panel, 5 μm.
D–F, Live images of GFP-labeled dendritic
spines in 14 DIV hippocampal neurons before (0 min) and after (60 min) treatment
with blank (D), control RAD
(E), or RGD (F).
RGD-treated neurons showed formation of new filopodia (arrowheads) and elongation of
existing dendritic spines (arrows). G,
H, Quantification of dendritic protrusion length
in 14 DIV GFP-labeled hippocampal neurons before and after treatment with blank
(control), control RAD, or RGD. G, Dendritic
protrusions were significantly longer in RGD-treated neurons. Data represent the
mean protrusion length. Error bars indicate SEM (n = 15 neurons per
group). H, The time course of the elongation of
dendritic protrusions showed a significant increase in mean protrusion length
starting ∼30 min after RGD treatment. Live images were taken at 3 min
intervals for 1 h. The arrow indicates RGD/RAD application. Data represent the mean
protrusion length. Error bars indicate SEM (n = 5 neurons per
group). *p < 0.05; **p < 0.01;
***p < 0.001 with one-way ANOVA.
I, J, Quantification
of dendritic protrusion density in 14 DIV GFP-labeled hippocampal neurons before and
after treatment with blank (control), control RAD, or RGD.
I, There were more dendritic protrusions in
RGD-treated neurons. Data represent the average number of protrusions per 10
μm of dendrite. Error bars indicate SEM (n = 15 neurons per
group). J, Time-lapse imaging has revealed a
significant increase in the number of dendritic protrusion ∼30 min after RGD
application. Live images were taken at 3 min interval for 1 h. The arrow indicates
RGD/RAD application. Data represent the average number of protrusions per 10
μm of dendrite. Error bars indicate SEM (n = 5 neurons per
group). *p < 0.05; **p < 0.01;
***p < 0.001 with one-way ANOVA.