The Journal of Neuroscience, March 28, 2007, ():
Phospholipase C Is Required for Changes in Postsynaptic Structure and Function Associated with NMDA Receptor-Dependent Long-Term Depression
J. Neurosci. Horne and Dell'Acqua
27: 3523
Supplemental Data
Files in this Data Supplement:
- supplemental material
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Supplemental Figure 1: CaN activation is not required for NMDA receptor stimulation of PLC. A) Hippocampal neurons cotransfected with PLCδPH-YFP (green) and PSD95-CFP (blue) show colocalization in dendritic spines of untreated neurons. Treatment with NMDA (50μM) showed a loss in PLCδPH-YFP colocalization with PSD95-CFP in dendritic spines 30 minutes after NMDA addition. Neurons treated with the CaN inhibitor ascomycin (Asc) (2μM for 1hour) did not prevent the translocation of PLCδPH-YFP seen 30 minutes after NMDA addition. B) Quantification of PLCδPH-YFP colocalization with PSD95-CFP showed that addition of NMDA caused a decrease in PSD95 colocalization that was not blocked by ascomycin (control 1.00±0.02 n=6; NMDA 0.31±0.1 ***p<0.001, n=3; Asc and NMDA 0.26±0.067 ***p<0.001, n=3).
- supplemental material
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Supplemental Figure 2: Calcium release from internal stores through IP3 receptors and PKC activation are not necessary for the loss of AKAP150 and F-actin from dendritic spines in cLTD. A) Hippocampal neurons untreated or treated with cLTD in the presence of bis-indolmaleamide (BIM, 1 μM), calphostin C (CalC, 1 μM), or xestospongin (XeD, 1μM) added 20 minutes prior to cLTD and stained with antibodies to label AKAP150 and PSD95 and phalloidin to visualize F-actin. Small panels show magnifications of dendrites. B) Quantification of F-actin and AKAP150 colocalization in dendrites 15 minutes after cLTD stimulation as measured by a correlation value (r). Induction of cLTD in the presence of BIM, CalC, or XeD shows a reduction in the amount of F-actin colocalized with AKAP150 compared to control (0.16±0.03, 0.08±0.02, 0.04±.06 , 0.58±0.03. respectively, n=10). C) Quantification of F-actin colocalization with PSD95 in dendritic spines 15 minutes after cLTD stimulation normalized to controls shows a loss of F-actin from spines in neurons treated with cLTD in the presence of BIM, CalC, or XeD (0.16±0.03, 0.39±0.07, 0.17±0.02, respectively; n=10). D) Quantification of AKAP150 colocalization with PSD95 in dendritic spines 15 minutes after cLTD stimulation normalized to control shows a loss of AKAP150 from spines in neurons treated with cLTD in the presence of BIM, CalC, or XeD (0.19±0.04, 0.23±0.04, 0.20±0.04, respectively; n=10). **p<0.001. All data analyzed using one way One-way ANOVA with Tukey’s post hoc test.
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