Figure 9.

BFA Markedly Reduced ASPD Accumulation

Primary rat hippocampal neuronal cultures, with or without AAV-APPswe transduction, were treated with 75 nM MG132 for 24 h in the absence or presence of 0.6 μg/mL BFA. We determined this concentration by examining BFA concentrations from 0.6 to 10 μg/mL in our system and found that 0.6 μg/mL BFA treatment was enough to destroy the Golgi structure without affecting the cell viability. The cultures were triple-stained with rpASD1, anti-TGN38 antibody (TGN marker), and DAPI. Representative images are shown. High-power images on the right were obtained with a highly sensitive, direct photon-counting system with a 100X oil objective lens. Solid and hatched scale bars, 10 and 1 μm, respectively. See also representative images of control cultures without AAV transduction in Figure S7. The total amount of rpASD1-immunostained ASPD detected in MAP2-positive neurons was determined by using an IN Cell Analyzer as in Figure 4B, right, and is shown as “ASPD level” below (mean ± SD, n = 3; Scheffé post hoc test, ***p < 0.0001, **p = 0.001).

See Figures S7 and S8.