Molecular mechanisms of contraction-induced translocation of GLUT4 in isolated cardiomyocytes.
Isolated adult rat ventricular cardiomyocytes were used to investigate the effects of contractile activity on 3-O-methylglucose transport on the translocation of the insulin-responsive glucose transporter GLUT4, and the possible activation of intermediates of the insulin signaling cascade. When elicited by field stimulation, contraction at 1 Hz did not significantly affect the adenosine triphosphate (ATP) content of cardiac cells, even after 60 min. At 5 Hz, a stable ATP level was observed until 15 minutes with a rapid decline at later time points. Stimulation of cardiomyocytes at 5 Hz for 5 minutes induced a 2-3 fold increase of 3-O-methylglucose transport with no additional stimulation in the presence of insulin (10[-7] M). Subcellular fractionation and immunoblotting analysis of GLUT4 distribution indicated that both contraction and insulin induced an identical increase (8-9-fold) of GLUT4 in the plasma membrane with a concomitant decrease (one third) in the microsomal fraction. Treatment of cardiomyocytes with wortmannin produced a complete inhibition of insulin- and contraction-induced glucose uptake. However, immunoprecipitation of insulin receptor substrate-1 (IRS-1) showed that the p85 regulatory subunit of phosphatidylinositol-3 kinase did not associate with IRS-1 upon contraction but with a marked stimulated association in response to insulin. These data suggest the existence of identical insulin- and contraction-recruitable GLUT4 pool. Contraction-induced signaling may use a limited part of the insulin-signaling cascade, possibly involving IRS-2. We further suggest that insulin resistance at the level of IRS-1 will not affect contraction-regulated glucose uptake by the heart.
Till
M
M
Molecular Cardiology, Diabetes Research Institute, Düsseldorf, Germany.
Kolter
T
T
Eckel
J
J
D019325:3-O-Methylglucose / Q000378:metabolism; D000730:Androstadienes / Q000494:pharmacology; D000818:Animals; D001693:Biological Transport, Active; D051275:Glucose Transporter Type 4; D066298:In Vitro Techniques; D007328:Insulin / Q000502:physiology*; D007329:Insulin Antagonists / Q000494:pharmacology; D008297:Male; D009004:Monosaccharide Transport Proteins / Q000378:metabolism*; D009124:Muscle Proteins; D009200:Myocardial Contraction / Q000502:physiology*; D009206:Myocardium / Q000166:cytology / Q000378:metabolism*; D011233:Precipitin Tests; D051381:Rats; D017208:Rats, Wistar; D015398:Signal Transduction; D000077191:Wortmannin
D000730:Androstadienes; D051275:Glucose Transporter Type 4; D007328:Insulin; D007329:Insulin Antagonists; D009004:Monosaccharide Transport Proteins; D009124:Muscle Proteins; C495694:Slc2a4 protein, rat; D019325:3-O-Methylglucose; D000077191:Wortmannin