FIGURE 8.

Inhibition of PI3K reduced CHT1 induction by ChAT. A, real time RT-PCR analysis of hChAT in wt-hChAT-GFP (wt-hChAT) and GFP alone cells treated with either the PI3K inhibitor LY294002 in DMSO or with DMSO alone for 48 h. Total RNA was then extracted from the cells and the relative level of hChAT mRNA evaluated by a real time PCR assay. Values are expressed as the mean ± S.E. A post hoc Tukey-Kramer test and the non-parametric Mann-Whitney test were used for statistical analysis. ns represents no significant difference. B, real time RT-PCR analysis of CHT1 in wt-hChAT-GFP and GFP alone cells treated with either LY294002 in DMSO or with DMSO alone for 48 h. Total RNA was then extracted from the cells and the relative level of CHT1 mRNA was evaluated by a real time PCR assay. Values are expressed as the mean ± S.E. A post hoc Tukey-Kramer test and the non-parametric Mann-Whitney test were used for statistical analysis. ns represents no significant difference. *, p < 0.05 by the non-parametric Mann-Whitney test. wt-hChAT-GFP cells treated with LY294002 continued to express CHT1 mRNA at a level that was ∼8-fold higher than that of GFP alone cells. Activation of CHT1 transcription was partially reduced in LY294002-treated wt-hChAT-GFP cells compared with DMSO alone-treated wt-hChAT-GFP cells. C, dose-response curves obtained by using real time RT-PCR analysis of CHT1 in wt-hChAT-GFP and GFP alone cells with 0 nm to 50 μm LY294002 in DMSO for 24 h. Squares indicate values of wt-hChAT cells, and triangles represent values of GFP alone cells. There is an approximate straight line relationship between the reduction of CHT1 induction in wt-hChAT-GFP and the dose of LY294002. y = −0.0536x + 2.7265, R² = 0.8573.