Hyperglycemia inhibits insulin activation of Akt protein kinase B but not phosphatidylinositol 3-kinase in rat skeletal muscle

Sustained hyperglycemia impairs insulin-stimulated glucose utilization in t he skeletal muscle of both humans and experimental animals-a phenomenon ref erred to clinically as glucose toxicity. To study how this occurs, a model was developed in which hyperglycemia produces insulin resistance in vitro. Rat extensor digitorum longus muscles were preincubated for 4 h in Krebs-He nseleit solution containing glucose or glucose + insulin at various concent rations, after which insulin action was studied. Preincubation with 25 mmol /l glucose + insulin (10 mU/ml) led to a 70% decrease in the ability of ins ulin (10 mU/ml) to stimulate glucose incorporation into glycogen and a 30% decrease in 2-deoxyglucose (2-DG) uptake, compared with muscles incubated w ith 0 mmol/l glucose. Glucose incorporation into lipid and its oxidation to CO, were marginally diminished, if at all. The alterations of glycogen syn thesis and 2-DG uptake were first evident after 1 h and were maximal after 2 h of preincubation; they were not observed in muscles preincubated with 2 5 mmol/l glucose + insulin for 5 min. Preincubation for 4 h with 25 mmol/l glucose in the absence of insulin produced a similar although somewhat smal ler decrease in insulin-stimulated glycogen synthesis; however, it did not alter 2-DG uptake, glucose oxidation to CO,, or incorporation into Lipids. Studies of insulin signaling in the latter muscles revealed that: activatio n of Akt/ protein kinase B (PKB) was diminished by 60%, compared with that of muscles preincubated in a glucose-free medium; whereas activation of pho sphatidylinositol (PI) 3-kinase, an upstream regulator of Akt/PKB in the in sulin-signaling cascade, and of mitogen-activated protein (MAP) kinase, a p arallel signal, was unaffected. Immunoblots demonstrated that this mas not due to a change in Akt/PKB abundance. The results indicate that hyperglycem ia-induced insulin resistance can be studied in rat skeletal muscle in vitr o. They suggest that impairment of insulin action in these muscles is relat ed to inhibition of Akt/PKB by events that do not affect PI 3-kinase.