Fig. 6: The second phase of LTD-associated PIP2 dynamics is dependent on PTEN in the spine head whereas the early phase of LTD induction is not.
a Scheme visualizing suggested routes towards PIP2 during LTD and the inhibition by bpV(HOpic). b–g Representative TEM images of cryo-preserved, freeze-fractured and anti-PIP2 immunogold-labeled spines of DIV14-16 hippocampal neurons pretreated for 60 min with ddH2O as a vehicle control (b–d) or with 15 nM bpV(HOpic) (e–g) and subjected to NMDA-induced LTD for 2 min (c, f) and for 3 min NMDA followed by 7 min postincubation time (d, g) in comparison to non-NMDA-treated controls (0 min) (b, e). Bars, 200 nm. h Quantitative analyses of the anti-PIP2 labeling density in neurons pretreated with bpV(HOpic) compared to the merely ddH2O-treated neurons (−bpV(HOpic)) in the same assays not revealing any statistically significant effect of PTEN inhibition on basal PIP2 levels. The labeling densities were normalized to the respective - bpV(HOpic) control. i, j Quantitative analyses of PIP2 signaling during the first and second phase of LTD induction in the total spine (i) and in the spine head (j), respectively, (both normalized to the respective 0 min data for the total spine labeling densities of each of the two conditions, i.e., with and without inhibitor). Data, mean ± SEM. −bpV(HOpic), 0 min, n = 39; 2 min, n = 29; 10 (3 + 7) min, n = 33 ROIs each (total spines; spine heads) and +bpV(HOpic), 0 min, n = 33; 2 min, n = 30; 10 (3 + 7) min, n = 27 ROIs each (total spines; spine heads) of primary neurons from 3 independent assays. Mann–Whitney (h); Two-way ANOVA/Bonferroni’s Multiple Comparison between ±bpV(HOpic) conditions (i, j). *P < 0.05. Additional Kruskal–Wallis/Dunn’s Multiple Comparisons were conducted for comparison of + and −bpV(HOpic) data at 2 min and at 3 + 7 min to 0 min control data (i, j) #P < 0.05; ##P < 0.01. P-values are reported directly in the figure. For numerical source data, see Supplementary Data 1.
