Figure 7.

Hypothalamic AMPK knockdown attenuates autophagy and neuropeptide expression, and suppresses 2DG-induced hyperphagia. Cumulative food consumption after ip 2DG injection (500 mg/kg of body weight) in mice fed (A) a normal diet (ND) or (B) a high-fat diet (HFD) for the indicated time (**, p < 0.01; ***, p < 0.001; n.s. for saline vs. 2DG-injected group; n = 8). (C) Food consumption of mice that were sacrificed 1 h after 2DG injection. Mice were fed a HFD for 6 d after lentivirus injection, and 2DG was administered on d 6 (**, p < 0.01; ***, p < 0.001 for saline vs. 2DG in the control shRNA [shCont] group or Prkaa1+Prkaa2 shRNA [shAmpk] group; ^###, p < 0.001 for shCont vs. shAmpk in the 2DG-injected group; n = 8). (D) Relative levels of total AMPK in the ARC and lateral hypothalamus (LH) (**, p < 0.01 for shCont vs. shAmpk; ARC n = 8, LH n = 8). (E) Relative levels of p-AMPK (Thr172), p-ULK1 (Ser555), and p-RPTOR (Ser792) in the ARC from the shCont group; mice were sacrificed during the light or dark cycle (*, p < 0.05; **, p < 0.01 for light cycle vs. dark cycle; n = 3). (F) Relative levels of p-AMPK (Thr172), p-ULK1 (Ser555), p-RPTOR (Ser792), NBR1, and LC3-II in the ARC of shCont and shAmpk mice that were sacrificed 1 h after 2DG injection. p-ULK1 and p-RPTOR were normalized to corresponding total proteins. Other protein levels were normalized to GAPDH. Protein levels in the shCont group were arbitrarily set to 1.0 (*, p < 0.05; **, p < 0.01 for saline vs. 2DG in the shCont group; n = 4). (G) Npy and Pomc mRNA expression levels in the ARC from shCont and shAmpk mice that were sacrificed 1 h after 2DG injection. Mice were fed a HFD for 6 d after lentivirus injection, and 2DG was administered on d 6 (*, p < 0.05 for saline vs. 2DG in shCont; n = 6 or 7).