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Diabetes Alters the Expression and Translocation of the Insulin-Sensitive Glucose Transporters 4 and 8 in the Atria.

Maria Z, Campolo AR, Lacombe VA - PLoS ONE (2015)

Bottom Line: In the healthy atria, insulin stimulation increased both GLUT-4 and -8 translocation to the cell surface (by 100% and 240%, respectively, P<0.05).Upon insulin stimulation, we reported an increase in Akt (Th308 and s473 sites) and AS160 phosphorylation, which was positively (P<0.05) correlated with GLUT4 protein content in the healthy atria.Akt and AS160 phosphorylation was not impaired in the diabetic atria, suggesting the presence of an intact insulin signaling pathway.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiological Sciences, Oklahoma State University, Stillwater, United States of America.

ABSTRACT
Although diabetes has been identified as a major risk factor for atrial fibrillation, little is known about glucose metabolism in the healthy and diabetic atria. Glucose transport into the cell, the rate-limiting step of glucose utilization, is regulated by the Glucose Transporters (GLUTs). Although GLUT4 is the major isoform in the heart, GLUT8 has recently emerged as a novel cardiac isoform. We hypothesized that GLUT-4 and -8 translocation to the atrial cell surface will be regulated by insulin and impaired during insulin-dependent diabetes. GLUT protein content was measured by Western blotting in healthy cardiac myocytes and type 1 (streptozotocin-induced, T1Dx) diabetic rodents. Active cell surface GLUT content was measured using a biotinylated photolabeled assay in the perfused heart. In the healthy atria, insulin stimulation increased both GLUT-4 and -8 translocation to the cell surface (by 100% and 240%, respectively, P<0.05). Upon insulin stimulation, we reported an increase in Akt (Th308 and s473 sites) and AS160 phosphorylation, which was positively (P<0.05) correlated with GLUT4 protein content in the healthy atria. During diabetes, active cell surface GLUT-4 and -8 content was downregulated in the atria (by 70% and 90%, respectively, P<0.05). Akt and AS160 phosphorylation was not impaired in the diabetic atria, suggesting the presence of an intact insulin signaling pathway. This was confirmed by the rescued translocation of GLUT-4 and -8 to the atrial cell surface upon insulin stimulation in the atria of type 1 diabetic subjects. In conclusion, our data suggest that: 1) both GLUT-4 and -8 are insulin-sensitive in the healthy atria through an Akt/AS160 dependent pathway; 2) GLUT-4 and -8 trafficking is impaired in the diabetic atria and rescued by insulin treatment. Alterations in atrial glucose transport may induce perturbations in energy production, which may provide a metabolic substrate for atrial fibrillation during diabetes.

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Related in: MedlinePlus

Regional heterogeneity of the insulin-sensitive GLUT4 and GLUT8 in the healthy myocardium.Higher total protein expression of A) GLUT4 and lower B) GLUT8 content in the healthy atrial compared to ventricular myocytes. Top panels: representative Western blot from total lysate of isolated rat myocytes; calsequestrin (CLSQ) was used as a loading control; representative bands were obtained from the same membrane. Bottom panels: Mean ± SE of total GLUT protein content (values expressed relative to atria; n = 6/group); * P<0.05 vs. atria. GLUT: glucose transporters. Insulin stimulation increases C) GLUT4 and D) GLUT8 protein content in the healthy atrial myocytes. Top panels: representative Western blot from total lysate of isolated rat myocytes, calsequestrin (CLSQ) was used as a loading control. Bottom panels: Mean ± SE of total GLUT protein content (values expressed relative to basal atria; n = 8-11/group); # P<0.05 vs. basal. Insulin stimulates E) GLUT4 and F) GLUT8 trafficking to the atrial and ventricular cell surface. Top panels: representative Western blot. Bottom panels: Mean ± SE of cell surface GLUT protein content (values expressed relative to labeled basal atria; n = 3-4/group); # P<0.05 vs. basal; *P<0.05 vs. atria. Methods: Cell surface GLUT measured using biotinylated photolabeling technique in the intact perfused mouse heart. L: Labeled (cell surface fraction); UL: Unlabeled (intracellular fraction).
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pone.0146033.g001: Regional heterogeneity of the insulin-sensitive GLUT4 and GLUT8 in the healthy myocardium.Higher total protein expression of A) GLUT4 and lower B) GLUT8 content in the healthy atrial compared to ventricular myocytes. Top panels: representative Western blot from total lysate of isolated rat myocytes; calsequestrin (CLSQ) was used as a loading control; representative bands were obtained from the same membrane. Bottom panels: Mean ± SE of total GLUT protein content (values expressed relative to atria; n = 6/group); * P<0.05 vs. atria. GLUT: glucose transporters. Insulin stimulation increases C) GLUT4 and D) GLUT8 protein content in the healthy atrial myocytes. Top panels: representative Western blot from total lysate of isolated rat myocytes, calsequestrin (CLSQ) was used as a loading control. Bottom panels: Mean ± SE of total GLUT protein content (values expressed relative to basal atria; n = 8-11/group); # P<0.05 vs. basal. Insulin stimulates E) GLUT4 and F) GLUT8 trafficking to the atrial and ventricular cell surface. Top panels: representative Western blot. Bottom panels: Mean ± SE of cell surface GLUT protein content (values expressed relative to labeled basal atria; n = 3-4/group); # P<0.05 vs. basal; *P<0.05 vs. atria. Methods: Cell surface GLUT measured using biotinylated photolabeling technique in the intact perfused mouse heart. L: Labeled (cell surface fraction); UL: Unlabeled (intracellular fraction).

Mentions: Since glucose transport has not been well characterized in the atria, we first quantified total GLUT-4 and -8 protein content in the healthy myocardium. Our results indicated a significant regional heterogeneity of both the GLUTs in the atria vs. the ventricle. Total GLUT4 protein content was significantly greater in the isolated rat atrial myocytes compared to the ventricular myocytes (Fig 1A). In contrast, there was significantly greater GLUT8 content in the healthy ventricle compared to the atria (Fig 1B). In order to investigate the insulin sensitivity of the GLUTs in the healthy atria, we then measured protein content of GLUT-4 and -8 upon insulin stimulation in isolated rat atrial myocytes. Our results indicated an increase in total GLUT-4 and -8 total protein content (by 44% and 60% respectively, P<0.05) upon insulin stimulation (Fig 1C and 1D). In order to measure GLUT translocation to the cell surface, the rate limiting step in glucose uptake, we used the biotinylation photolabeled assay in the intact perfused mouse heart to reliably quantify the proportion of active cell surface GLUTs at the atrial and ventricular cell surface (Fig 1E and 1F). Our results confirmed the regional heterogeneity of GLUT4 and GLUT8 in the atrial and ventricular cell surface, showing significantly greater cell surface GLUT4 content in the atria and greater cell surface GLUT8 content in the ventricle respectively (Fig 1E and 1F). To further determine whether GLUT-4 and -8 are insulin-sensitive, the biotinylation photolabeled assay was performed in the heart perfused with or without (basal) physiological concentration of insulin. Upon insulin stimulation, there was significant increase in cell surface GLUT-4 and -8 content in both the atrial (by 100% and 240% vs. basal condition, respectively, P<0.05) and ventricular cell surface (by 97% and 40% vs. basal condition, respectively, P<0.05), indicating that both GLUT-4 and -8 are insulin-sensitive in the atria and ventricle. Interestingly, the response to insulin was greater in the atria compared to the ventricle, with an increase in GLUT8 trafficking by 240% vs. 40% (compared to basal condition), respectively (P<0.05).


Diabetes Alters the Expression and Translocation of the Insulin-Sensitive Glucose Transporters 4 and 8 in the Atria.

Maria Z, Campolo AR, Lacombe VA - PLoS ONE (2015)

Regional heterogeneity of the insulin-sensitive GLUT4 and GLUT8 in the healthy myocardium.Higher total protein expression of A) GLUT4 and lower B) GLUT8 content in the healthy atrial compared to ventricular myocytes. Top panels: representative Western blot from total lysate of isolated rat myocytes; calsequestrin (CLSQ) was used as a loading control; representative bands were obtained from the same membrane. Bottom panels: Mean ± SE of total GLUT protein content (values expressed relative to atria; n = 6/group); * P<0.05 vs. atria. GLUT: glucose transporters. Insulin stimulation increases C) GLUT4 and D) GLUT8 protein content in the healthy atrial myocytes. Top panels: representative Western blot from total lysate of isolated rat myocytes, calsequestrin (CLSQ) was used as a loading control. Bottom panels: Mean ± SE of total GLUT protein content (values expressed relative to basal atria; n = 8-11/group); # P<0.05 vs. basal. Insulin stimulates E) GLUT4 and F) GLUT8 trafficking to the atrial and ventricular cell surface. Top panels: representative Western blot. Bottom panels: Mean ± SE of cell surface GLUT protein content (values expressed relative to labeled basal atria; n = 3-4/group); # P<0.05 vs. basal; *P<0.05 vs. atria. Methods: Cell surface GLUT measured using biotinylated photolabeling technique in the intact perfused mouse heart. L: Labeled (cell surface fraction); UL: Unlabeled (intracellular fraction).
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
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pone.0146033.g001: Regional heterogeneity of the insulin-sensitive GLUT4 and GLUT8 in the healthy myocardium.Higher total protein expression of A) GLUT4 and lower B) GLUT8 content in the healthy atrial compared to ventricular myocytes. Top panels: representative Western blot from total lysate of isolated rat myocytes; calsequestrin (CLSQ) was used as a loading control; representative bands were obtained from the same membrane. Bottom panels: Mean ± SE of total GLUT protein content (values expressed relative to atria; n = 6/group); * P<0.05 vs. atria. GLUT: glucose transporters. Insulin stimulation increases C) GLUT4 and D) GLUT8 protein content in the healthy atrial myocytes. Top panels: representative Western blot from total lysate of isolated rat myocytes, calsequestrin (CLSQ) was used as a loading control. Bottom panels: Mean ± SE of total GLUT protein content (values expressed relative to basal atria; n = 8-11/group); # P<0.05 vs. basal. Insulin stimulates E) GLUT4 and F) GLUT8 trafficking to the atrial and ventricular cell surface. Top panels: representative Western blot. Bottom panels: Mean ± SE of cell surface GLUT protein content (values expressed relative to labeled basal atria; n = 3-4/group); # P<0.05 vs. basal; *P<0.05 vs. atria. Methods: Cell surface GLUT measured using biotinylated photolabeling technique in the intact perfused mouse heart. L: Labeled (cell surface fraction); UL: Unlabeled (intracellular fraction).
Mentions: Since glucose transport has not been well characterized in the atria, we first quantified total GLUT-4 and -8 protein content in the healthy myocardium. Our results indicated a significant regional heterogeneity of both the GLUTs in the atria vs. the ventricle. Total GLUT4 protein content was significantly greater in the isolated rat atrial myocytes compared to the ventricular myocytes (Fig 1A). In contrast, there was significantly greater GLUT8 content in the healthy ventricle compared to the atria (Fig 1B). In order to investigate the insulin sensitivity of the GLUTs in the healthy atria, we then measured protein content of GLUT-4 and -8 upon insulin stimulation in isolated rat atrial myocytes. Our results indicated an increase in total GLUT-4 and -8 total protein content (by 44% and 60% respectively, P<0.05) upon insulin stimulation (Fig 1C and 1D). In order to measure GLUT translocation to the cell surface, the rate limiting step in glucose uptake, we used the biotinylation photolabeled assay in the intact perfused mouse heart to reliably quantify the proportion of active cell surface GLUTs at the atrial and ventricular cell surface (Fig 1E and 1F). Our results confirmed the regional heterogeneity of GLUT4 and GLUT8 in the atrial and ventricular cell surface, showing significantly greater cell surface GLUT4 content in the atria and greater cell surface GLUT8 content in the ventricle respectively (Fig 1E and 1F). To further determine whether GLUT-4 and -8 are insulin-sensitive, the biotinylation photolabeled assay was performed in the heart perfused with or without (basal) physiological concentration of insulin. Upon insulin stimulation, there was significant increase in cell surface GLUT-4 and -8 content in both the atrial (by 100% and 240% vs. basal condition, respectively, P<0.05) and ventricular cell surface (by 97% and 40% vs. basal condition, respectively, P<0.05), indicating that both GLUT-4 and -8 are insulin-sensitive in the atria and ventricle. Interestingly, the response to insulin was greater in the atria compared to the ventricle, with an increase in GLUT8 trafficking by 240% vs. 40% (compared to basal condition), respectively (P<0.05).

Bottom Line: In the healthy atria, insulin stimulation increased both GLUT-4 and -8 translocation to the cell surface (by 100% and 240%, respectively, P<0.05).Upon insulin stimulation, we reported an increase in Akt (Th308 and s473 sites) and AS160 phosphorylation, which was positively (P<0.05) correlated with GLUT4 protein content in the healthy atria.Akt and AS160 phosphorylation was not impaired in the diabetic atria, suggesting the presence of an intact insulin signaling pathway.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiological Sciences, Oklahoma State University, Stillwater, United States of America.

ABSTRACT
Although diabetes has been identified as a major risk factor for atrial fibrillation, little is known about glucose metabolism in the healthy and diabetic atria. Glucose transport into the cell, the rate-limiting step of glucose utilization, is regulated by the Glucose Transporters (GLUTs). Although GLUT4 is the major isoform in the heart, GLUT8 has recently emerged as a novel cardiac isoform. We hypothesized that GLUT-4 and -8 translocation to the atrial cell surface will be regulated by insulin and impaired during insulin-dependent diabetes. GLUT protein content was measured by Western blotting in healthy cardiac myocytes and type 1 (streptozotocin-induced, T1Dx) diabetic rodents. Active cell surface GLUT content was measured using a biotinylated photolabeled assay in the perfused heart. In the healthy atria, insulin stimulation increased both GLUT-4 and -8 translocation to the cell surface (by 100% and 240%, respectively, P<0.05). Upon insulin stimulation, we reported an increase in Akt (Th308 and s473 sites) and AS160 phosphorylation, which was positively (P<0.05) correlated with GLUT4 protein content in the healthy atria. During diabetes, active cell surface GLUT-4 and -8 content was downregulated in the atria (by 70% and 90%, respectively, P<0.05). Akt and AS160 phosphorylation was not impaired in the diabetic atria, suggesting the presence of an intact insulin signaling pathway. This was confirmed by the rescued translocation of GLUT-4 and -8 to the atrial cell surface upon insulin stimulation in the atria of type 1 diabetic subjects. In conclusion, our data suggest that: 1) both GLUT-4 and -8 are insulin-sensitive in the healthy atria through an Akt/AS160 dependent pathway; 2) GLUT-4 and -8 trafficking is impaired in the diabetic atria and rescued by insulin treatment. Alterations in atrial glucose transport may induce perturbations in energy production, which may provide a metabolic substrate for atrial fibrillation during diabetes.

Show MeSH
Related in: MedlinePlus