Figure 6. Regulation of mTOR pathway on hypothalamic HIF.

(A&B) 24-h fasted adult C57BL/6 mice received 5-h third-ventricle infusion of glucose (Glu) versus vehicle (Veh) in the presence or absence a prior third-ventricle injection of rapamycin (Rap). Hypothalami were harvested and analyzed by Western blotting for phosphorylated S6K (pS6K) and HIF2α. β-act, β-actin. Bar graphs: Western blots were quantitated and analyzed statistically. * p<0.05, ** p<0.01; n = 4 per group. Error bars reflect mean ± SEM. AU, arbitrary unit. (C–F) POMC/HIFβ^lox/lox mice (P/HIFβ^l/l) and control littermate HIFβ^lox/lox mice (HIFβ^l/l) received MBH injection of lentiviruses expressing the constitutive-active form of Rheb tagged with Flag (Lenti-^CARheb/Flag) or matched GFP-expressing control lentiviruses (Lenti-GFP). Site-specific gene delivery was verified by immunostaining (C) and Western blots of Rheb/mTOR signaling proteins (D) as well as the quantitation analysis of Western blots (E). Food intake of these mice was monitored on a daily basis for 2 wk (F). * p<0.05, ** p<0.01; n = 5–6 per group. Error bars reflect mean ± SEM. AU, arbitrary unit. (A–F) All mice were adult males and normal chow-fed.