title: Glucocorticoid regulation of insulin receptor and substrate IRS-1 tyrosine phosphorylation in rat skeletal muscle in vivo.
abstract: To test the hypothesis that glucocorticoid-induced insulin resistance might originate from abnormalities in insulin receptor signaling, we investigated the effects of glucocorticoids on in vivo tyrosine phosphorylation of the insulin receptor and the insulin receptor substrate IRS-1 in rat skeletal muscle. Male Sprague-Dawley rats were treated with cortisone (100 mg/kg for 5 d) and compared to pair-fed controls. Cortisone treatment of rats resulted in both hyperglycemia and hyperinsulinemia. Anesthetized animals were injected with 10 U/kg insulin via cardiac puncture and, after 2 min, hindlimb muscles were removed, snap-frozen, and homogenized in SDS. Protein tyrosine phosphorylation was studied by immunoblotting with phosphotyrosine antibody. Insulin receptors and substrate IRS-1 were identified and quantified with specific antibodies. Cortisone treatment increased the amount of insulin receptor protein by 36%, but decreased the total level of receptor tyrosine phosphorylation (69 +/- 4% of control, P < 0.05). The decreased level of receptor phosphorylation was explained by a reduced number of receptors containing phosphorylated tyrosine residues (64.6 +/- 5% of control, P < 0.05). Glucocorticoid excess decreased skeletal muscle IRS-1 content by 50%, but did not significantly alter the total level of IRS-1 tyrosine phosphorylation. The apparent M(r) of IRS-1 was reduced by approximately 10 kD. Treatment with protein phosphatase-2A reduced IRS-1 M(r) in control but not in glucocorticoid-treated muscle indicating that the lower M(r) likely results from lower phosphoserine and/or phosphothreonine content. To investigate the role of hyperinsulinemia in the glucocorticoid response, rats were made insulin-deficient with streptozotocin (100 mg/kg, i.p.). Subsequent treatment with cortisone for 5 d had no effects on insulin levels, tyrosine phosphorylation of insulin receptors or IRS-1, or the M(r) of IRS-1. In conclusion, glucocorticoid-treated skeletal muscle is characterized by: (a) decreased total tyrosine phosphorylation of insulin receptors as a result of a reduction in the pool of receptors undergoing tyrosine phosphorylation; (b) decreased IRS-1 content and reduced serine and/or threonine phosphorylation of IRS-1. Glucocorticoid-induced hyperinsulinemia appears to be essential for the development of these alterations.
authors.affiliation: Research Division, Joslin Diabetes Center, Boston, Massachusetts 02215.
authors.forename: F
authors.initials: F
authors.lastname: Giorgino
authors.forename: A
authors.initials: A
authors.lastname: Almahfouz
authors.forename: L J
authors.initials: LJ
authors.lastname: Goodyear
authors.forename: R J
authors.initials: RJ
authors.lastname: Smith
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies / Q000302:isolation & purification; D001786:Blood Glucose / Q000378:metabolism; D003348:Cortisone / Q000494:pharmacology*; D003921:Diabetes Mellitus, Experimental / Q000378:metabolism; D015151:Immunoblotting; D007328:Insulin / Q000494:pharmacology*; D055504:Insulin Receptor Substrate Proteins; D046911:Macromolecular Substances; D008297:Male; D008969:Molecular Sequence Data; D009132:Muscles / Q000187:drug effects / Q000378:metabolism*; D010455:Peptides / Q000138:chemical synthesis / Q000276:immunology; D010750:Phosphoproteins / Q000302:isolation & purification / Q000378:metabolism*; D010766:Phosphorylation; D019000:Phosphotyrosine; D051381:Rats; D017207:Rats, Sprague-Dawley; D011972:Receptor, Insulin / Q000187:drug effects / Q000302:isolation & purification / Q000378:metabolism*; D013311:Streptozocin / Q000494:pharmacology; D014443:Tyrosine / Q000031:analogs & derivatives* / Q000378:metabolism
chemical_list: D000906:Antibodies; D001786:Blood Glucose; D007328:Insulin; D055504:Insulin Receptor Substrate Proteins; C526296:Irs1 protein, rat; D046911:Macromolecular Substances; D010455:Peptides; D010750:Phosphoproteins; D019000:Phosphotyrosine; D014443:Tyrosine; D013311:Streptozocin; D011972:Receptor, Insulin; D003348:Cortisone