Fig. 3. GTPase activities of Rac1 and Cdc42-GTPase are increased in neurons stimulated with A_1-42. Hippocampal neurons cultured for 4 days were treated with (A) 0.1, 1 and 10 µM A_1-42 for 24 hours and (B) with 10 µM of peptide for 30 min, 2, 4 and 24 hours. Active GTP-Rac1 was pulled down using the PAK-PBD conjugated with agarose and then tested by immunoblotting with anti-Rac1 monoclonal antibody. Values were normalized against total Rac1. Graphs show data from four independent experiments; ^*P<0.05, ^**P<0.01. (C,D) Hippocampal neurons stimulated 4 hours with 10 µM fibrilar A_1-42 were pre-treated with pertussis toxin and roscovitin. The GTP bound to Rac1 (C) and to Cdc42 (D) was determined using the Rac/Cdc42 Activation Assay Kit, according to manufacturer's recommendations. Values were normalized, with respect to total Rac1 and Cdc42; ^*P<0.05.