Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Background: Hypothalamic AMPK acts as a cell energy sensor and can modulate food intake, glucose homeostasis, and fatty acid biosynthesis. Intrahypothalamic fatty acid injection is known to suppress liver glucose production, mainly by activation of hypothalamic ATP-sensitive potassium (K(ATP)) channels. Since all models employed seem to involve malonyl-CoA biosynthesis, we hypothesized that acetyl-CoA carboxylase can modulate the counter-regulatory response independent of nutrient availability. Methodology/Principal Findings: In this study employing immunoblot, real-time PCR, ELISA, and biochemical measurements, we showed that reduction of the hypothalamic expression of acetyl-CoA carboxylase by antisense oligonucleotide after intraventricular injection increased food intake and NPY mRNA, and diminished the expression of CART, CRH, and TRH mRNA. Additionally, as in fasted rats, in antisense oligonucleotide-treated rats, serum glucagon and ketone bodies increased, while the levels of serum insulin and hepatic glycogen diminished. The reduction of hypothalamic acetyl-CoA carboxylase also increased PEPCK expression, AMPK phosphorylation, and glucose production in the liver. Interestingly, these effects were observed without modification of hypothalamic AMPK phosphorylation. Conclusion/Significance: Hypothalamic ACC inhibition can activate hepatic counter-regulatory response independent of hypothalamic AMPK activation.84Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)FAPESP [2006/05660-5, 2009/12523-2]