Figure 5

Nedd4-2 knockdown in MPTP-treated mice increases the expression and function of glutamate transporters. (a) Western blotting showing efficient lentiviral knockdown of Nedd4-2 in the midbrain of mice. 2.0, 3.0 or 4.0 μl of virus were stereotaxically injected into the right side of the SN, and 3, 5 or 7 days after injection, the mice were killed for dissection of the midbrain. Nedd4-2 levels were measured by western blotting. Based on the results of this experiment, 3.0 μl virus was injected into the SN for 5 days in subsequent experiments. (b and c) Western blotting showing that Nedd4-2 knockdown increases GLAST total protein expression (b) and GLT-1 and GLAST membrane protein expression (c). (d and e) Immunohistochemistry staining showing that Nedd4-2 knockdown increases GLT-1 and GLAST expression in the SN of MPTP-treated mice. Scale bar: 40 μm. (f) Nedd4-2 knockdown increased glutamate uptake in the synaptosome in the midbrain of MPTP-treated mice. Results are expressed as the mean±S.E. One-way ANOVA. n=3 per group in (a–c). n=6 per group in (d–f). **P<0.01, *P<0.05, ^##P<0.01, ^#P<0.05. *Represents a significant difference between other group and ctrl shRNA group; while ^#represents the significant difference between the indicated group and MPTP+ctrl shRNA group