MECHANISM OF PERVANADATE STIMULATION AND POTENTIATION OF INSULIN-ACTIVATED GLUCOSE-TRANSPORT IN RAT ADIPOCYTES - DISSOCIATION FROM VANADATEEFFECT

Previous studies have shown that the combination of vanadate and H2O2 generates peroxide(s) of vanadate (pervanadate) that is able to mimic insulin in stimulating lipogenesis or protein synthesis and inhibiting lipolysis in rat adipocytes. Here we report that pervanadate is a pot ent trigger of 3-O-methylglucose transport in rat adipocytes, with an effective concentration of 5 muM and a maximum at 20 muM. Moreover, pe rvanadate produced an additional activation of approximately 60% on gl ucose influx in cells treated with maximally activating concentrations of insulin. Vanadate was ineffective in potentiating insulin-stimulat ed glucose uptake. Quercetin, a bioflavonoid that inhibits insulin rec eptor tyrosine kinase, blunted this effect of pervanadate. Treatment o f adipocytes with pervanadate inhibited protein phosphotyrosyl phospha tase activity of cell extracts in a dose-dependent manner, with an ID5 0 of 5 muM and complete inhibition at 80 muM. In contrast, vanadate (1 -800 muM) did not appreciably inhibit cell phosphotyrosyl phosphatases . The inhibitory effect of pervanadate correlated with the increase in protein phosphotyrosine accumulation, as determined by Western blotti ng with antiphosphotyrosine antibodies. The most prominent phosphotyro sine-containing band detected in pervanadate-treated adipocytes was th at of autophosphorylated insulin receptor, identified by immunoblottin g or immunoprecipitation with antiinsulin receptor antibodies. The add ition of insulin to pervanadate-treated adipocytes (20 muM) caused a f urther increase (approximately 70%) in receptor autophosphorylation. I n a cell-free system using partially purified insulin receptor devoid of tyrosine phosphatase activity, pervanadate did not stimulate the re ceptor autophosphorylation or interfere with the stimulating effect of insulin. These results suggest that 1) pervanadate triggers glucose u ptake by increasing autophosphorylation of insulin receptor, preventin g its dephosphorylation; 2) under physiological conditions, cellular p rotein phosphotyrosyl phosphatase activity is high, thereby significan tly opposing insulin-mediated hexose transport; and 3) pervanadate has the unique ability to markedly increase maximal cell responsiveness i n stimulating glucose transport achieved at a saturating insulin conce ntration. These findings suggest a possible clinical application in th e management of glucose uptake in pathological conditions of insulin r esistance and hyperinsulinemia.