FIGURE 3.

Aβ₄₂ oligomer-induced PAK activation and translocation in primary neurons. A, immunofluorescent staining of PAK activation and translocation induced by Aβ₄₂ oligomer in primary hippocampal neurons. Primary hippocampal neurons 7 DIV were treated with 100 nm Aβ oligomers for 30 min and then stained with anti-pPAK (red), anti-Aβ (6E10, green), and the purple nuclear stain, 4′,6-diamidino-2-phenylindole; scale bar, 25 μm. CTRL, control. B-G, Western immunoblot analysis of PAK activation and translocation induced by Aβ₄₂ oligomer in primary hippocampal neurons. Hippocampal neurons 7 DIV were treated with 250 nm of Aβ oligomers and Western-blotted with anti-pPAK antibody. pPAK levels were significantly increased in membrane and cytosol (M/C) fractions after 30 min in Aβ₄₂ oligomer-treated neurons when compared with controls (B,^*, p < 0.05; C, ^**, p < 0.01). The pPAK ratio of membrane to cytosol had a strong trend to increase by 30 min in Aβ₄₂ oligomer-treated neurons when compared with controls (D, p = 0.0630). In contrast pPAK levels were no longer increased in membrane fractions (E) and were significantly reduced in cytosol fraction by 9.5 h (F,^*, p < 0.05). As a result, the membrane/cytosol ratio was no longer significantly altered (G). H, Aβ oligomer preparations detected by 6E10 antibody.