Insulin Resistance due to Nutrient Excess: Mechanisms of AMPK Downregulation

Abstract

Excesses of glucose and the branched chain amino acids lead to insulin resistance in skeletal muscle by activating mTOR/p70S6 kinase (p70S6K). In a recent study we demonstrated that glucose and leucine may produce these effects by downregulating AMP‐activated protein kinase (AMPK) which, when activated, diminishes mTOR/p70S6 K signaling. How these excesses of glucose and leucine decrease AMPK activity is unclear, since they do not alter the cell's energy state (AMP/ATP or CrP). We explored whether the decrease in AMPK activity might be due to decreases in the activity of SIRT1, a histone protein deacetylase that can activate AMPK or by changes in the activity of an unidentified kinase or phosphatase that results in the phosphorylation of Ser485/491 on the catalytic subunit of AMPK leading to its inhibition. Incubation of extensor digitorum longus muscle, with high glucose (25 mM) or 5 mM glucose and leucine (100 μM) persistently diminished AMPK activity within 30 min an effect that lasted for 2 h. Likewise both high glucose and leucine decreased the activity of SIRT1as reflected by increases in the deacetylation of histone H3 and a decrease in the activity of the rate‐limiting enzyme for SIRT1 activation, nicotinamide phosphoribosyltransferase (NAMPT). In addition, SIRT1 abundance was decreased. These changes only become evident after 2 h, however. In contrast, excess glucose and leucine caused increased phosphorylation of AMPK at Ser485/491 after 1 h. In studies with intact rats, we found that prolonged (3–8 h) hyperglycemia, produced by a glucose infusion, downregulates AMPK Thr172 phosphorylation and causes insulin resistance as early as 3 h. However, SIRT1, AMPK (Ser485/491) and NAMPT activity only decreased between 5–8 h. The results indicate that both leucine and high glucose downregulate AMPK by at least two mechanisms. This could account for sustained decreases in AMPK activity; however, temporally they do not account for the initial decrease in AMPK activity caused by these fuels.