Fig. 1
Aβ uptake in astrocytes occurs mainly through clathrin-dependent endocytosis. a Confocal fluorescence micrographs of WT (Mt3 +/+) astrocytes treated with 20 nM FITC-CtxB for 30 min at 37 °C. Cells were pretreated with vehicle, 1 μM CP, or 1 mM MβCD for 30 min before and during exposure to CtxB. After fixation, the cells were immunostained for the Golgi marker GM130 (red) and for the astrocyte marker GFAP (purple). Hoechst 33342 (blue) was used to counterstain the nuclei. CtxB uptake was disrupted by CP but not by MβCD, consistent with the involvement of the clathrin-dependent endocytotic pathway (n = 4 experiments). 3D images were added to help visualize the endocytosed CtxB (3D). All imaging experiments of the study were performed four times with different cultures. Scale bar, 20 μm. b, c Confocal fluorescence micrographs of cultured cortical astrocytes treated with FITC-Aβs. Cells treated with or without 1 mM MβCD or 1 μM CP were incubated with 1 μM FITC-Aβ1–42 (B) or FITC-Aβ1–40 (C) (green dots, arrowheads) for 1 h at 37 °C and then with Alexa Fluor 594-WGA (red) to stain the plasma membrane. The corresponding 3D images are also presented (bottom panels). Scale bar, 20 μm. d, e Bars indicate the percentage of FITC(+) cells. The values represent the mean changes in the percentage of FITC-Aβ1–42(+) (D) or FITC-Aβ1–40(+) (E) cells in the CP- or MβCD-treated groups relative to control (CTL), defined as 100 % (***P < 0.001; ns, not significant)