C75, a Fatty Acid Synthase Inhibitor, Reduces Food Intake via Hypothalamic AMP-activated Protein Kinase

Eun-Kyoung Kim,Ian Miller,Susan Aja,Leslie E Landree,Michael Pinn,Jill Mcfadden,Francis P Kuhajda,Timothy H Moran,Gabriele V Ronnett

doi:10.1074/jbc.m402165200

Abstract

Energy homeostasis and feeding are regulated by the central nervous system. C75, a fatty acid synthase (FAS) inhibitor, causes weight loss and anorexia, implying a novel central nervous system pathway(s) for sensing energy balance. AMP-activated protein kinase (AMPK), a sensor of peripheral energy balance, is phosphorylated and activated when energy sources are low. Here, we identify a role for hypothalamic AMPK in the regulation of feeding behavior and in mediating the anorexic effects of C75. 5-Aminoimidazole-4-carboxamide-1-beta-D-ribofuranoside (AICAR), an activator of AMPK, increased food intake, whereas compound C, an inhibitor of AMPK, decreased food intake. C75 rapidly reduced the level of the phosphorylated AMPK alpha subunit (pAMPKalpha) in the hypothalamus, even in fasted mice that had elevated hypothalamic pAMPKalpha levels. Furthermore, AICAR reversed both the C75-induced anorexia and the decrease in hypothalamic pAMPKalpha levels. C75 elevated hypothalamic neuronal ATP levels, which may contribute to the mechanism by which C75 decreased AMPK activity. C75 reduced the levels of pAMPKalpha and phosphorylated cAMP response element-binding protein (pCREB) in the arcuate nucleus neurons of the hypothalamus, suggesting a mechanism for the reduction in NPY expression seen with C75 treatment. These data indicate that modulation of FAS activity in the hypothalamus can alter energy perception via AMPK, which functions as a physiological energy sensor in the hypothalamus.

Highlights

Despite significant advances in the understanding of appetite and satiety at molecular levels [1,2,3], practical therapies for weight loss remain elusive
Since enzymes of the fatty acid metabolic pathways are highly expressed in hypothalamic neurons that regulate feeding behavior [9], we hypothesize that C75-induced alterations in fatty acid metabolism may affect neuronal energy flux, which could signal a change in energy status, leading to changes in feeding behavior
Feeding Behavior Is Changed by C75, AICAR, or Compound C Treatment—We investigated the relationships among C75induced alterations in fatty acid metabolism, AMPK activation or inhibition, and feeding behavior

Summary

EXPERIMENTAL PROCEDURES

C75 i.p./AICAR i.c.v. treatment groups were i.p. injected with 5 mg/kg of body weight C75 dissolved in 200 ␮l of glucose-free RPMI 1 h before the dark onset, followed by 3 ␮g/2.5 ␮l of saline i.c.v. AICAR immediately preceding the dark onset. Anti-AMPK␣ antibody (␣1 and ␣2, 1:1000, Cell Signaling) was used as a loading control. Hypothalamic neurons were lysed in TE (100 mM Tris-HCl, pH 7.4, 4 mM EDTA), and ATP levels were measured within the linear range using the ATP BioLuminescence Kit CLSII (Roche Applied Science) by following the manufacture’s recommendation. For double fluorescent in situ hybridization, digoxigenin-labeled riboprobe was generated from plasmid containing AMPK␣2 gene Sheep FITC-conjugated anti-digoxigenin antibody (1:50, Roche Applied Science) was incubated in TNB buffer (100 mM Tris-HCl, pH 7.5, 150 mM NaCl, and 0.5% blocking reagent) for FITC detection. Data were analyzed by one-way analysis of variance or t test

RESULTS

C75 Reduces Food Intake via Hypothalamic AMPK

DISCUSSION