Fig. 6

Inhibition of OGA by thiamet G (TG) decreased autophagic flux even in the presence of rapamycin. (a) DIV7 primary rat cortical neurons were exposed to rapamycin (1 μM) and/or TG (0.25 μM) for 7 d. Western blot analyses of LC3 were performed. The increase of LC3-II by thiamet G was attenuated by rapamycin. (b) Western blot analyses of LC3-II were performed in the presence and absence of chloroquine (CQ) as described in Fig. 2. (c) Quantification of western blot band intensities was performed using NIH image J. (d) Calculated values of LC3-II with CQ minus LC3-II without CQ (Autophagic Flux) from b-c and normalized to 0 μM TG control. (e) DIV7 primary rat cortical neurons were exposed to rapamycin (1 μM) and/or TG (0.25 μM) for 7 d. Western blot analysis of p62 was performed. Inhibition of mTOR decreased p62. For all panels, β-actin was used as a loading control, Data = mean ± SEM (n = 3), *p < 0.05 compared to no TG, #p < 0.05 compared to –rapamycin. Results were analyzed by ANOVA followed by Bonferroni’s Multiple Comparison post-hoc test. *** p = 0.0001