AMPK Activation through Mitochondrial Regulation Results in Increased Substrate Oxidation and Improved Metabolic Parameters in Models of Diabetes

Abstract

<div><p>Modulation of mitochondrial function through inhibiting respiratory complex I activates a key sensor of cellular energy status, the 5'-AMP-activated protein kinase (AMPK). Activation of AMPK results in the mobilization of nutrient uptake and catabolism for mitochondrial ATP generation to restore energy homeostasis. How these nutrient pathways are affected in the presence of a potent modulator of mitochondrial function and the role of AMPK activation in these effects remain unclear. We have identified a molecule, named R419, that activates AMPK <i>in vitro</i> via complex I inhibition at much lower concentrations than metformin (IC₅₀ 100 nM vs 27 mM, respectively). R419 potently increased myocyte glucose uptake that was dependent on AMPK activation, while its ability to suppress hepatic glucose production <i>in vitro</i> was not. In addition, R419 treatment of mouse primary hepatocytes increased fatty acid oxidation and inhibited lipogenesis in an AMPK-dependent fashion. We have performed an extensive metabolic characterization of its effects in the <i>db/db</i> mouse diabetes model. <i>In vivo</i> metabolite profiling of R419-treated <i>db/db</i> mice showed a clear upregulation of fatty acid oxidation and catabolism of branched chain amino acids. Additionally, analyses performed using both ¹³C-palmitate and ¹³C-glucose tracers revealed that R419 induces complete oxidation of both glucose and palmitate to CO₂ in skeletal muscle, liver, and adipose tissue, confirming that the compound increases mitochondrial function <i>in vivo</i>. Taken together, our results show that R419 is a potent inhibitor of complex I and modulates mitochondrial function <i>in vitro</i> and in diabetic animals <i>in vivo</i>. R419 may serve as a valuable molecular tool for investigating the impact of modulating mitochondrial function on nutrient metabolism in multiple tissues and on glucose and lipid homeostasis in diabetic animal models. </p> </div