Effect of NMDA and IL-1β on IL-1RI subcellular localisation. A: Western blot analysis of the TIF fraction obtained from control, IL-1β-treated (0.05 ng/ml) and NMDA-treated hippocampal cultures (50 μM). The same amount of proteins was loaded in each lane. IL-1β increases IL-1RI localization in the Triton-insoluble fraction (TIF) (*p <0.05) leaving unaffected IL-1RAcP and MyD88 levels. Values are means ± S.E of 4 independent experiments. B: Western blot of IL-1RI from control, IL-1β-treated (0.05 ng/ml) and NMDA-treated (50 μM) hippocampal cultures exposed (+BS3 lanes) or not (-BS3 lanes) to the cross-linking agent BS3. IL-1RI high-molecular-weight complexes that didn't enter the gel are not shown. C: Hippocampal neurons were either left untreated (control) or treated with IL-1β (0.05 ng/ml, 30 minutes) or NMDA (50 μM) fixed, and immunolabeled for IL-1RI (green) and PSD-95 (red) as a postsynaptic marker. Data are expressed as percentage of IL-1RI colocalization with PSD-95 (AIM4.2 software, Zeiss). White arrows indicate PSD-95 positive clusters in the merge panel. Scale bar: 5 μM.