This study investigated the metabolic changes with age in the Fischer 344 × Brown Norway rat and its suitability as an animal model of postmaturational insulin resistance. Specifically, we determined whether an age-associated decrease in glucose disposal is associated with diminished whole body insulin responsiveness and/or a decrease in glucose transporter (GLUT-4) protein and mRNA content in medial gastrocnemius muscle of male Fischer 344 × Brown Norway rats of ages 8, 18, and 28 months. Fasting plasma glucose was unchanged with age. There was a significant age effect on visceral adiposity, fasting plasma insulin levels, insulin responsiveness, and GLUT-4 protein content. Insulin responsiveness and GLUT-4 protein were lower in the 18-month-old rats than in the 8-month-old rats. The findings of age-associated increases in visceral adiposity and insulin resistance, and decreases in GLUT-4 in the Fisher 344 × Brown Norway rat, suggest that this rat strain may be an appropriate model for studying the effects of aging on glucose homeostasis.
Although exercise training has well-known cardiorespiratory and metabolic benefits, low compliance with exercise training programs is a fact, and the harmful effects of physical detraining regarding these adaptations usually go unnoticed. We investigated the effects of exercise detraining on blood pressure, insulin sensitivity, and GLUT4 expression in spontaneously hypertensive rats (SHR) and normotensive Wistar Kyoto rats (WKY).
Studied animals were randomized into sedentary, trained (treadmill running/5 days a week, 60 min/day for 10 weeks), 1 week of detraining, and 2 weeks of detraining. Blood pressure (tail-cuff system), insulin sensitivity (kITT), and GLUT4 (Western blot) in heart, gastrocnemius and white fat tissue were measured.
Exercise training reduced blood pressure (19%), improved insulin sensitivity (24%), and increased GLUT4 in the heart (+34%); gastrocnemius (+36%) and fat (+22%) in SHR. In WKY no change in either blood pressure or insulin sensitivity were observed, but there was an increase in GLUT4 in the heart (+25%), gastrocnemius (+45%) and fat (+36%) induced by training. Both periods of detraining did not induce any change in neither blood pressure nor insulin sensitivity in SHR and WKY. One-week detraining reduced GLUT4 in SHR (heart: -28%; fat: -23%) and WKY (heart: -19%; fat: -22%); GLUT4 in the gastrocnemius was reduced after a 2-week detraining (SHR: -35%; WKY: -25%). There was a positive correlation between GLUT4 (gastrocnemius) and the maximal velocity in the exercise test (r = 0.60, p = 0.004).
The study findings show that in detraining, despite reversion of the enhanced GLUT4 expression, cardiorespiratory and metabolic beneficial effects of exercise are preserved.
In order to evaluate the role of visceral and subcutaneous fat tissue in insulin sensitivity and lipid metabolism, we measured the fasting levels of plasma free fatty acid (FFA) and insulin, glucose disappearance rate (Rd), and hepatic glucose production rate (HGP) after surgical removal of visceral (VF) or subcutaneous (SF) fat tissue in monosodium glutamate-obese (MSG-Ob) rats. Monosodium glutamate obesity was induced in rats by neonatal injection of MSG. Surgery to remove fat was done at 15 weeks of age. The experiments were done four weeks after the surgery. MSG-Ob rats showed increased levels of FFA, insulin, and HGP and decreased Rd compared to normal rats. In the VF group, the FFA level and HGP were decreased to normal values, Rd was partially normalized, but the level of insulin did not change significantly compared to MSG-Ob. In the SF group, FFA and Rd were partially normalized, but HGP was not suppressed significantly compared to MSG-Ob. These results suggest that visceral fat affects the insulin sensitivity of liver and FFA concentration more than subcutaneous fat; however, no significant difference was shown on whole body insulin sensitivity and fasting insulin concentration.
This study was designed to determine whether altered glucose transporter expression is essential for the in vivo insulin-resistant glucose uptake characteristic of streptozocin-induced diabetes. Immunofluorescence in rat skeletal muscle colocalizes GLUT4 with dystrophin, both intrinsic to muscle fibers. In contrast, GLUT1 is extrinsic to muscle fibers, probably in perineurial sheath. Immunoblotting shows that levels of GLUT1 and GLUT4 protein per DNA in hindlimb muscle are unaltered from control levels at 7 d of diabetes but decrease to approximately 20% of control at 14 d of diabetes. This decrease is prevented by insulin treatment. In adipose cells of 7 d diabetic rats, GLUT4 levels are depressed. Thus, GLUT4 undergoes tissue-specific regulation in response to diabetes. GLUT4 and GLUT1 mRNA levels in muscle are decreased 62-70% at both 7 and 14 d of diabetes and are restored by insulin treatment. At 7 d of diabetes, when GLUT4 protein levels in muscle are unaltered, in vivo insulin-stimulated glucose uptake measured by euglycemic clamp is 54% of control. This reflects impairment in both glycogen synthesis and glycolysis and the substrate common to these two pathways, glucose-6-phosphate, is decreased approximately 30% in muscle of diabetic rats. These findings suggest a defect early in the pathway of glucose utilization, probably at the step of glucose transport. Because GLUT1 and GLUT4 levels are unaltered at 7 d of diabetes, reduced glucose uptake in muscle probably reflects impaired glucose transporter translocation or intrinsic activity. Later, at 14 d of diabetes, GLUT1 and GLUT4 protein levels are reduced, suggesting that sequential defects may contribute to the insulin-resistant glucose transport characteristic of diabetes.
Adipose tissue-derived inflammation may contribute to metabolic alterations and eventually to the metabolic syndrome (MetS). The purpose of this study was to: 1) examine the role of adipocytokines in the association between obesity and the MetS; and 2) to determine whether the association is different in obese and non-obese persons.
Cross-sectional population-based InCHIANTI study.
944 community-dwelling adults aged 65 years and older living in Tuscany, Italy.
Obesity was defined as body mass index ≥ 30 kg/m2 and MetS as ≥ 3 of the ATP-III criteria. Circulating levels of CRP, IL-6, IL-1ra, IL-18, TNF-α R1, adiponectin, resistin, and leptin were measured. Additionally, insulin resistance was determined using the homeostasis model assessment (HOMA-IR).
The prevalence of the MetS was 32%. Both overall and abdominal obesity were significantly associated with the MetS after adjusting for inflammatory cytokines, adipokines and lifestyle factors. After adjusting for multiple confounders and HOMA-IR, IL-1ra, TNF-α R1 and adiponectin (p < 0.05) remained significantly associated with the MetS. Having multiple cytokines in the highest tertile increased the likelihood of having the MetS in both obese (p for trend 0.002) and non-obese persons (p for trend 0.001) independent of insulin resistance.
Non-obese and obese individuals who develop an intense pro-inflammatory state may be more prone to develop the MetS than those with lower levels of inflammation.
adipocytokines; adiponectin; cytokines; inflammation; metabolic syndrome; obesity
Obesity is associated with the rise of noncommunicable diseases worldwide. The pathophysiology behind this disease involves the increase of adipose tissue, being inversely related to adiponectin, but directly related to insulin resistance and metabolic syndrome (MetS). Therefore, this study aimed to determine the relationship between adiponectin levels with each component of MetS in eutrophic and obese Mexican children.
A cross sectional study was conducted in 190 school-age children classified as obese and 196 classified as eutrophic. Adiponectin, glucose, insulin, high density lipoprotein cholesterol (HDL-C) and triglycerides were determined from a fasting blood sample. Height, weight, waist circumference, systolic and diastolic blood pressures (BP) were measured; MetS was evaluated with the IDF definition. The study groups were divided according to tertiles of adiponectin, using the higher concentration as a reference. Linear regression analysis was used to assess the association between adiponectin and components of the MetS. Finally, stepwise forward multiple logistic regression analysis controlling for age, gender, basal HOMA-IR values and BMI was performed to determine the odds ratio of developing MetS according to adiponectin tertiles.
Anthropometric and metabolic measurements were statistically different between eutrophic and obese children with and without MetS (P <0.001). The prevalence of MetS in obese populations was 13%. Adiponectin concentrations were 15.5 ± 6.1, 12.0 ± 4.8, 12.4 ± 4.9 and 9.4 ± 2.8 μg/mL for eutrophic and obese subjects, obese without MetS, and obese with MetS, respectively (P <0.001). Obese children with low values of adiponectin exhibited a higher frequency of MetS components: abdominal obesity, 49%; high systolic BP, 3%; high diastolic BP, 2%; impaired fasting glucose, 17%; hypertriglyceridemia, 31%; and low HDL-C values, 42%. Adjusted odds ratio of presenting MetS according to adiponectin categories was 10.9 (95% CI 2.05; 48.16) when the first tertile was compared with the third.
In this sample of eutrophic and obese Mexican children we found that adiponectin concentrations and MetS components have an inversely proportional relationship, which supports the idea that this hormone could be a biomarker for identifying individuals with risk of developing MetS.
Obesity; Adiponectin; Child; Insulin resistance; Metabolic syndrome; Biomarker
Obesity is commonly associated with diabetes, cardiovascular diseases and cancer. The purpose of this study was to determinate the effect of a lower dose of fish oil supplementation on insulin sensitivity, lipid profile, and muscle metabolism in obese rats.
Monosodium glutamate (MSG) (4 mg/g body weight) was injected in neonatal Wistar male rats. Three-month-old rats were divided in normal-weight control group (C), coconut fat-treated normal weight group (CO), fish oil-treated normal weight group (FO), obese control group (Ob), coconut fat-treated obese group (ObCO) and fish oil-treated obese group (ObFO). Obese insulin-resistant rats were supplemented with fish oil or coconut fat (1 g/kg/day) for 4 weeks. Insulin sensitivity, fasting blood biochemicals parameters, and skeletal muscle glucose metabolism were analyzed.
Obese animals (Ob) presented higher Index Lee and 2.5 fold epididymal and retroperitoneal adipose tissue than C. Insulin sensitivity test (Kitt) showed that fish oil supplementation was able to maintain insulin sensitivity of obese rats (ObFO) similar to C. There were no changes in glucose and HDL-cholesterol levels amongst groups. Yet, ObFO revealed lower levels of total cholesterol (TC; 30%) and triacylglycerol (TG; 33%) compared to Ob. Finally, since exposed to insulin, ObFO skeletal muscle revealed an increase of 10% in lactate production, 38% in glycogen synthesis and 39% in oxidation of glucose compared to Ob.
Low dose of fish oil supplementation (1 g/kg/day) was able to reduce TC and TG levels, in addition to improved systemic and muscle insulin sensitivity. These results lend credence to the benefits of n-3 fatty acids upon the deleterious effects of insulin resistance mechanisms.
Evidence suggests that insulin-sensitive glucose transporters (GLUTs) other than GLUT4 may exist. To investigate whether GLUT12 may represent another insulin-sensitive GLUT, transgenic (TG) mice that overexpress GLUT12 were characterized.
RESEARCH DESIGN AND METHODS
TG mice that overexpressed GLUT12 under a β-actin promoter were generated. Glucose metabolism in TG and wild-type control mice was compared using glucose and insulin tolerance tests and hyperinsulinemic-euglycemic clamps. In addition, basal and insulin-stimulated glucose clearance rates into insulin-sensitive peripheral tissues were measured using [3H]-2-deoxy-d-glucose.
GLUT12 was overexpressed by 40–75% in TG compared with wild-type mice in insulin-sensitive tissues with no change in GLUT4 content. Body weight and fasting blood glucose did not differ between wild-type and TG mice; however, insulin concentrations were reduced in TG mice. Enhanced oral glucose tolerance was noted in TG mice by a reduced blood glucose excursion compared with wild-type mice (P < 0.05). Enhanced insulin sensitivity was noted by a greater decrease in blood glucose in TG mice during insulin tolerance testing. Hyperinsulinemic-euglycemic clamps confirmed enhanced insulin sensitivity in GLUT12-overexpressing mice (P < 0.01). Tissues of TG mice exhibited normal basal glucose clearance rates; however, under insulin-stimulated conditions, glucose clearance was significantly increased (P < 0.01) in tissues of TG mice.
Increased expression of GLUT12 results in improved whole-body insulin sensitivity mediated by an increased glucose clearance rate in insulin-responsive tissues under insulin-stimulated, but not basal, conditions. These findings provide evidence that GLUT12 represents a novel, second insulin-sensitive GLUT.
Purpose of review
Low-grade inflammation is characteristic of the metabolic syndrome (MetS). C-reactive protein (CRP), the best characterized biomarker of inflammation, is also an independent predictor of future cardiovascular events. The purpose of this review is to outline the role of inflammation and high sensitivity CRP in the MetS.
Emerging laboratory and epidemiological data now link inflammation and high sensitivity CRP to insulin resistance and adiposity and other features of MetS. Furthermore, in large prospective studies, increased high sensitivity CRP levels in MetS confer greater cardiovascular risk. CRP has been shown to impair insulin signaling and contributes to atherothrombosis.
Thus, although a high CRP level predisposes to increased cardiovascular risk in MetS, future investigation is warranted on the in-vivo role of CRP in mediating vascular effects and resulting in increased cardiovascular events in MetS patients.
C-reactive protein; inflammation; metabolic syndrome; vascular cells
We have examined the regulation of GLUT-4 phosphorylation in adipocytes isolated from diabetic rats. Despite progressive (40-70%) reductions in GLUT-4 protein contents on the 2nd, 7th, and 14th day of diabetes, the phosphorylation of GLUT-4 was increased two- to fourfold. These alterations were accompanied by concomitant reductions (40-66%) in the insulin-stimulated 2-deoxyglucose transport. Insulin treatment of diabetic animals for 5 d restored glucose transport activity, GLUT-4 protein, and GLUT-4 phosphorylation to control levels whereas vanadate and phlorizin were ineffective. In control adipocytes, insulin promoted GLUT-4 translocation from the low density microsomal (LDM) pool to the plasma membranes (PM) and decreased the state of GLUT-4 phosphorylation. In adipocytes isolated from the diabetic rats, insulin failed to stimulate GLUT-4 translocation and to decrease GLUT-4 phosphorylation. To explore the mechanism of the diabetes-induced increases in the GLUT-4 phosphorylation, we investigated phosphoserine phosphatase (PSPase) activities using 32P-labeled GLUT-4 and phosphorylase "a" as substrates. Diabetes resulted in 50-60% increase in the particulate PSPase activity and concomitant reductions in cytosolic PSPase activities. Although reduced cytosolic PSPase activity correlated with an inadequate dephosphorylation of LDM GLUT-4, the existence of highly phosphorylated PM GLUT-4 in the presence of increased particulate PSPase activity required additional explanation. To address this problem, we used PM GLUT-4 from diabetic rats as a substrate of particulate PSPase. Highly active diabetic particulate PSPase, which dephosphorylated control GLUT-4 and phosphorylase a, failed to dephosphorylate PM GLUT-4 from diabetic rats. These data suggest that PM GLUT-4 from diabetic rats is unable to interact with PSPase or that its phosphorylation sites are not accessible to PSPase action. In summary, an induction of diabetes with streptozotocin resulted in significant increases in GLUT-4 phosphorylation. In contrast to normal cells, insulin failed to promote GLUT-4 recruitment to the plasma membranes and its dephosphorylation in diabetic adipocytes. At the same time, diabetes appears to induce redistribution of PSPases, resulting in lower cytosolic activity and higher particulate activity. It also appears that the existence of highly phosphorylated GLUT-4 in the plasma membranes of diabetic adipocytes resulted from its inability to interact with particulate PSPases.
Epidemiology and animal models suggest that dietary monosodium glutamate (MSG) may contribute to the onset of obesity and the metabolic syndrome.
Families (n = 324) from a rural area of Thailand were selected and provided MSG as the sole source for the use in meal preparation for 10 days. Three hundred forty-nine subjects aged 35–55 years completed the study and were evaluated for energy and nutrient intake, physical activity, and tobacco smoking. The prevalence of overweight and obesity (BMI ≥ 25 kg/m2), insulin resistance (HOMA-IR >3), and the metabolic syndrome (ATP III criteria) were evaluated according to the daily MSG intake.
The prevalence of the metabolic syndrome was significantly higher in the tertile with the highest MSG intake. Further, every 1 g increase in MSG intake significantly increased the risk of having the metabolic syndrome (odds ratio 1.14, 95% confidence interval-CI- 1.12 - 1.28) or being overweight (odds ratio 1.16, 95% CI 1.04 - 1.29), independent of the total energy intake and the level of physical activity.
Higher amounts of individual MSG consumption are associated with the risk of having the metabolic syndrome and being overweight independent of other major determinants.
Functional food; Obesity; Metabolic syndrome; Thailand
Glucose enters the heart via GLUT1 and GLUT4 glucose transporters. GLUT4-deficient mice develop striking cardiac hypertrophy and die prematurely. Whether their cardiac changes are caused primarily by GLUT4 deficiency in cardiomyocytes or by metabolic changes resulting from the absence of GLUT4 in skeletal muscle and adipose tissue is unclear. To determine the role of GLUT4 in the heart we used cre-loxP recombination to generate G4H–/– mice in which GLUT4 expression is abolished in the heart but is present in skeletal muscle and adipose tissue. Life span and serum concentrations of insulin, glucose, FFAs, lactate, and β-hydroxybutyrate were normal. Basal cardiac glucose transport and GLUT1 expression were both increased approximately 3-fold in G4H–/– mice, but insulin-stimulated glucose uptake was abolished. G4H–/– mice develop modest cardiac hypertrophy associated with increased myocyte size and induction of atrial natriuretic and brain natriuretic peptide gene expression in the ventricles. Myocardial fibrosis did not occur. Basal and isoproterenol-stimulated isovolumic contractile performance was preserved. Thus, selective ablation of GLUT4 in the heart initiates a series of events that results in compensated cardiac hypertrophy.
J. Clin. Invest. 104:1703–1714 (1999).
Stearoyl-CoA desaturase 1 (SCD1) deficiency protects mice from diet-induced obesity and insulin resistance. To understand the tissue-specific role of SCD1 in energy homeostasis, we have generated mice with an adipose-specific knockout of Scd1 (AKO), and report here that SCD1 deficiency increases GLUT1 expression in adipose tissue of AKO mice, but not global SCD1 knockout (GKO) mice. In 3T3-L1 adipocytes treated with a SCD inhibitor, basal glucose uptake and the cellular expression of GLUT1 were significantly increased while GLUT4 expression remained unchanged. Consistently, adipose-specific SCD1 knockout (AKO) mice had significantly elevated GLUT1 expression, but not GLUT4, in white adipose tissue compared to Lox counterparts. Concurrently, adiponectin expression was significantly diminished, whereas TNF-α expression was elevated. In contrast, in adipose tissue of GKO mice, GLUT4 and adiponectin expression were significantly elevated with lowered TNF-α expression and little change in GLUT1 expression, suggesting a differential responsiveness of adipose tissue to global- or adipose-specific SCD1 deletion. Taken together, these results indicate that adipose-specific deletion of SCD1 induces GLUT1 up-regulation in adipose tissue, associated with decreased adiponectin and increased TNF-α production, and suggest that GLUT1 may play a critical role in controlling glucose homeostasis of adipose tissue in adipose-specific SCD1-deficient conditions.
stearoyl-coenzyme A desaturase; adipose tissue; GLUT1; GLUT4; adiponectin
We have taken the approach of introducing the muscle-specific myosin light chain (MLC)-GLUT4 transgene into the GLUT4-null background to assess the relative role of muscle and adipose tissue GLUT4 in the etiology of the GLUT4-null phenotype. The resulting MLC-GLUT4-null mice express GLUT4 predominantly in the fast-twitch extensor digitorum longus (EDL) muscle. GLUT4 is nearly absent in female white adipose tissue (WAT) and slow-twitch soleus muscle of both sexes of MLC-GLUT4-null mice. GLUT4 content in male MLC-GLUT4-null WAT is 20% of that in control mice. In transgenically complemented EDL muscle, 2-deoxyglucose (2-DOG) uptake was restored to normal (male) or above normal (female) levels. In contrast, 2-DOG uptake in slow-twitch soleus muscle of MLC-GLUT4-null mice was not normalized. With the normalization of glucose uptake in fast-twitch skeletal muscle, whole body insulin action was restored in MLC-GLUT4-null mice, as shown by the results of the insulin tolerance test. These results demonstrate that skeletal muscle GLUT4 is a major regulator of skeletal muscle and whole body glucose metabolism. Despite normal skeletal muscle glucose uptake and insulin action, the MLC-GLUT4-null mice exhibited decreased adipose tissue deposits, adipocyte size, and fed plasma FFA levels that are characteristic of GLUT4-null mice. Together these results indicate that the defects in skeletal muscle and whole body glucose metabolism and adipose tissue in GLUT4-null mice arise independently.
Inflammation is associated with obesity and insulin resistance. Proinflammatory cytokines produced by adipose tissue in obesity could alter insulin signaling and action. Recent studies have shown a relationship between interleukin (IL)-1β amount and metabolic syndrome or type 2 diabetes. However, the ability of IL-1β to alter insulin signaling and action remains to be explored. We demonstrated that IL-1β sligthly increased Glut 1 translocation and basal glucose uptake in 3T3-L1 adipocytes. Importantly, we found that prolonged-IL-1β treatment reduced the insulin-induced glucose uptake whereas an acute treatment had no effect. Chronic treatment with IL-1β slightly decreased the expression of Glut 4 and markedly inhibited its translocation to the plasma membrane in response to insulin. This inhibitory effect was due to a decrease in the amount of IRS-1 but not IRS-2 expression both in 3T3-L1 and human adipocytes. The decrease in IRS-1 amount resulted in a reduction in its tyrosine phosphorylation and in the alteration of insulin-induced PKB activation and AS160 phosphorylation. Pharmacological inhibition of ERK totally inhibited IL-1β–induced down regulation of IRS-1 mRNA. Moreover, IRS-1 protein expression and insulin-induced PKB activation, AS160 phosphorylation and Glut 4 translocation were partially recovered following treatment with the ERK inhibitor. These results demonstrate that IL-1β reduces IRS-1 expression at a transcriptional level through a mechanism that is ERK dependent and at a posttranscriptional level independently of ERK activation. By targeting IRS-1, IL-1β is capable of impairing insulin signaling and action, and could thus participate in concert with other cytokines, in the development of insulin resistance in adipocytes.
3T3-L1 Cells; Adipocytes; drug effects; immunology; metabolism; Animals; Down-Regulation; drug effects; immunology; Extracellular Signal-Regulated MAP Kinases; metabolism; GTPase-Activating Proteins; metabolism; Gene Expression; drug effects; immunology; Glucose; metabolism; Glucose Transporter Type 1; genetics; metabolism; Glucose Transporter Type 4; genetics; metabolism; Inflammation; immunology; metabolism; Insulin Resistance; immunology; Interleukin-1beta; immunology; pharmacology; Interleukin-6; pharmacology; MAP Kinase Signaling System; drug effects; immunology; Mice; Mice, Obese; Phosphoproteins; genetics; Phosphorylation; drug effects; Proto-Oncogene Proteins c-akt; metabolism; Tyrosine; metabolism
Antibodies specific for the insulin-regulatable glucose transporter (GLUT 4) were used to immunolocalize this protein in brown adipose tissue from basal- and insulin-treated rats. Cryosections of fixed tissue were incubated with antibodies, which were subsequently labeled with Protein A/gold and examined by EM. Antibodies against albumin and cathepsin D were also used with gold particles of different sizes to identify early and late endosomes, respectively. Under basal conditions 99% of the GLUT 4 labeling was located within the cell. Labeling was predominantly in the trans-Golgi reticulum and tubulo-vesicular structures elsewhere in the cytoplasm. In insulin-stimulated cells approximately 40% of the GLUT 4 labeling was at the cell surface, where it was randomly distributed, except for occasional clustering in coated pits. Moreover, after insulin treatment, GLUT 4 was also enriched in early endosomes. We conclude that translocation of GLUT 4 to the cell surface is the major mechanism by which insulin increases glucose transport. In addition, these results suggest that in the presence of insulin GLUT 4 recycles from the cell surface, probably via the coated pit-endosome pathway that has been characterized for cell surface receptors, and also that insulin causes the redistribution of GLUT 4 by stimulating exocytosis from GLUT 4-containing tubulo-vesicular structures, rather than by slowing endocytosis of GLUT 4.
In the central nervous system (CNS) insulin mediates a variety of effects including feeding, metabolism and cognition. The cognitive enhancing effects of insulin are proposed to be mediated through activation of insulin receptors in the hippocampus, an important integration center for learning and memory in the mammalian brain. Since less is known regarding insulin signaling events in the hippocampus, the aim of the current study was to determine whether insulin stimulates similar signaling cascades and GLUT4 translocation in the rat hippocampus as has been described in peripheral tissues. Intracerebroventricular administration of insulin increases hippocampal insulin levels and also stimulates the phosphorylation of Akt in a time-dependent manner. Insulin also stimulates the translocation of GLUT4 to hippocampal plasma membranes in a time course that mirrors the increases in glucose uptake observed during the performance of hippocampal-dependent tasks. Insulin stimulated phosphorylation of Akt and translocation of GLUT4 were blocked by pre-treatment with the PI3-kinase inhibitor LY294002. Confocal immunofluorescence determined that insulin stimulated phosphorylation of Akt was localized to neurons and colocalized with the insulin receptor and GLUT4 in the rat hippocampus, thereby identifying the functional anatomical substrates of insulin signaling in the hippocampus. These results demonstrate that insulin-stimulated translocation of GLUT4 to the plasma membrane in the rat hippocampus occurs via similar mechanisms as described in peripheral tissues and suggests that insulin mediated translocation of GLUT4 may provide a mechanism through which hippocampal neurons rapidly increase glucose utilization during increases in neuronal activity associated with hippocampal-dependent learning.
glucose; Akt; cognition; diabetes; confocal microscopy
Insulin-mediated glucose uptake is highly sensitive to the levels of the facilitative GLUT protein GLUT4. Transcription of the GLUT4 gene is repressed in states of insulin deficiency and insulin resistance and can be induced by states of enhanced energy output, such as exercise. The cellular signals that regulate GLUT4 transcription are not well understood. We hypothesized that changes in energy substrate flux regulate GLUT4 transcription.
RESEARCH DESIGN AND METHODS
To test this hypothesis, we used transgenic mice in which expression of the chloramphenicol acetyltransferase (CAT) gene is driven by a functional 895-bp fragment of the human GLUT4 promoter, thereby acting as a reporter for transcriptional activity. Mice were treated with a single dose of etomoxir, which inhibits the transport of long-chain fatty acids into mitochondria and increases basal, but not insulin-mediated, glucose flux. GLUT4 and transgenic CAT mRNA were measured.
Etomoxir treatment significantly reduced CAT and GLUT4 mRNA transcription in adipose tissue, but did not change transcription in heart and skeletal muscle. Downregulation of GLUT4 transcription was cell autonomous, since etomoxir treatment of 3T3-L1 adipocytes resulted in a similar downregulation of GLUT4 mRNA. GLUT4 transcriptional downregulation required the putative liver X receptor (LXR) binding site in the human GLUT4 gene promoter in adipose tissue and 3T3-L1 adipocytes. Treatment of 3T3-L1 adipocytes with the LXR agonist, TO901317, partially restored GLUT4 expression in etomoxir-treated cells.
Our data suggest that long-chain fatty acid import into mitochondria in adipose tissue may produce ligands that regulate expression of metabolic genes.
Glucose transporter type 4 (GLUT4) is insulin responsive and is expressed in striated muscle and adipose tissue. To investigate the impact of a partial deficiency in the level of GLUT4 on in vivo insulin action, we examined glucose disposal and hepatic glucose production (HGP) during hyperinsulinemic clamp studies in 4-5-mo-old conscious mice with one disrupted GLUT4 allele [GLUT4 (+/-)], compared with wild-type control mice [WT (+/+)]. GLUT4 (+/-) mice were studied before the onset of hyperglycemia and had normal plasma glucose levels and a 50% increase in the fasting (6 h) plasma insulin concentrations. GLUT4 protein in muscle was approximately 45% less in GLUT4 (+/-) than in WT (+/+). Euglycemic hyperinsulinemic clamp studies were performed in combination with [3-3H]glucose to measure the rate of appearance of glucose and HGP, with [U-14C]-2-deoxyglucose to estimate muscle glucose transport in vivo, and with [U-14C]lactate to assess hepatic glucose fluxes. During the clamp studies, the rates of glucose infusion, glucose disappearance, glycolysis, glycogen synthesis, and muscle glucose uptake were approximately 55% decreased in GLUT4 (+/-), compared with WT (+/+) mice. The decreased rate of in vivo glycogen synthesis was due to decreased stimulation of glucose transport since insulin's activation of muscle glycogen synthase was similar in GLUT4 (+/-) and in WT (+/+) mice. By contrast, the ability of hyperinsulinemia to inhibit HGP was unaffected in GLUT4 (+/-). The normal regulation of hepatic glucose metabolism in GLUT4 (+/-) mice was further supported by the similar intrahepatic distribution of liver glucose fluxes through glucose cycling, gluconeogenesis, and glycogenolysis. We conclude that the disruption of one allele of the GLUT4 gene leads to severe peripheral but not hepatic insulin resistance. Thus, varying levels of GLUT4 protein in striated muscle and adipose tissue can markedly alter whole body glucose disposal. These differences most likely account for the interindividual variations in peripheral insulin action.
Metabolic syndrome (MetS) is associated with an increased risk of major cardiovascular events. Increased high-sensitivity C-reactive protein (hsCRP) levels are associated with MetS and its components. Changes in gamma-glutamyl transferase (GGT) levels in response to oxidative stress are also associated with MetS, and the levels could be modulated by hsCRP.
From a single community, we recruited 822 men (mean age, 61 ± 14 years) and 1,097 women (63 ± 12 years) during their annual health examination. We investigated whether increased hsCRP and GGT levels are synergistically associated with MetS and insulin resistance evaluated by Homeostasis of model assessment of insulin resistance (HOMA-IR).
Of these subjects, 141 men (17.2%) and 170 women (15.5%) had MetS. Participants with MetS had a higher hsCRP and GGT level than those without MetS in both genders, and the HOMA-IR increased significantly in correlation with an increase in hsCRP and GGT. In men, the adjusted odds ratios (95% confidence interval) for MetS across tertiles of hsCRP and GGT were 1.00, 1.69 (1.01-2.80), and 2.13 (1.29-3.52), and 1.00, 3.26 (1.84-5.78) and 6.11 (3.30-11.3), respectively. In women, the respective corresponding values were 1.00, 1.54 (0.92-2.60), and 3.08 (1.88-5.06), and 1.00, 1.70 (1.04-2.79) and 2.67 (1.66-4.30). The interaction between increased hsCRP and GGT was a significant and independent determinant for MetS and insulin resistance in both genders.
These results suggested that higher CRP and GGT levels were synergistically associated with MetS and insulin resistance, independently of other confounding factor in the general population.
The effect of insulinopenic diabetes on the expression of glucose transporters in the small intestine was investigated. Enterocytes were sequentially isolated from jejunum and ileum of normal fed rats, streptozotocin-diabetic rats, and diabetic rats treated with insulin. Facilitative glucose transporter (GLUT) 2, GLUT5, and sodium-dependent glucose transporter 1 protein content was increased from 1.5- to 6-fold in enterocytes isolated from diabetic animals in both jejunum and ileum. Insulin was able to reverse the increase in transporter protein expression seen after induction of diabetes. There was a four- to eightfold increase in the amount of enterocyte glucose transporter mRNA after diabetes with greater changes in sodium-dependent glucose transporter 1 and GLUT2 than in GLUT5 levels. In situ hybridization showed that after the induction of diabetes there was new hybridization in lower villus and crypt enterocytes that was reversed by insulin treatment. Thus, the increase in total hexose transport caused by diabetes is due to a premature expression of hexose transporters by enterocytes along the crypt-villus axis, causing a cumulative increase in enterocyte transporter protein during maturation. These changes are likely to represent an adaptive response by the organism to increase nutrient absorption in a perceived state of tissue starvation. These adaptive changes may lead to exacerbation of hyperglycemia in uncontrolled diabetes.
The aim of this study was to evaluate cardiovascular autonomic function in a rodent obesity model induced by monosodium glutamate injections during the first seven days of life.
The animals were assigned to control (control, n = 10) and monosodium glutamate (monosodium glutamate, n = 13) groups. Thirty-three weeks after birth, arterial and venous catheters were implanted for arterial pressure measurements, drug administration, and blood sampling. Baroreflex sensitivity was evaluated according to the tachycardic and bradycardic responses induced by sodium nitroprusside and phenylephrine infusion, respectively. Sympathetic and vagal effects were determined by administering methylatropine and propranolol.
Body weight, Lee index, and epididymal white adipose tissue values were higher in the monosodium glutamate group in comparison to the control group. The monosodium glutamate-treated rats displayed insulin resistance, as shown by a reduced glucose/insulin index (-62.5%), an increased area under the curve of total insulin secretion during glucose overload (39.3%), and basal hyperinsulinemia. The mean arterial pressure values were higher in the monosodium glutamate rats, whereas heart rate variability (>7 times), bradycardic responses (>4 times), and vagal (∼38%) and sympathetic effects (∼36%) were reduced as compared to the control group.
Our results suggest that obesity induced by neonatal monosodium glutamate treatment impairs cardiac autonomic function and most likely contributes to increased arterial pressure and insulin resistance.
Monosodium Glutamate; Obesity; Insulin Resistance; Arterial Pressure; Autonomic Function
Given the established fact that obesity interferes with male reproductive functions, the present study aimed to evaluate sperm production in the testis and storage in the epididymis in a glutamate-induced model of obesity.
Male rats were treated neonatally with monosodium glutamate (MSG) at doses of 4 mg/kg subcutaneously, or with saline solution (control group), on postnatal days 2, 4, 6, 8 and 10. On day 120, obesity was confirmed by the Lee index in all MSG-treated rats. After this, all animals from the two experimental groups were anesthetized and killed to evaluate body and reproductive organ weights, sperm parameters, plasma hormone levels (FSH, LH and testosterone), testicular and epididymal histo-morphometry and histopathology.
Significant reductions in absolute and relative weights of testis, epididymis, prostate and seminal vesicle were noted in MSG-treated animals. In these same animals plasma testosterone and follicle-stimulating hormone (FSH) concentrations were decreased, as well as sperm counts in the testis and epididymis and seminiferous epithelium height and tubular diameter. The sperm transit time was accelerated in obese rats. However, the number of Sertoli cells per seminiferous tubule and stereological findings on the epididymis were not markedly changed by obesity.
Neonatal MSG-administered model of obesity lowers sperm production and leads to a reduction in sperm storage in the epididymis of adult male rats. The acceleration of sperm transit time can have implications for the sperm quality of these rats.
Obesity; Monosodium glutamate; Epididymis; Testosterone; Sperm; Rat
BACKGROUND: Adipose and muscle tissues express an insulin-sensitive glucose transporter (GLUT4). This transporter has been shown to translocate from intracellular stores to the plasma membrane following insulin stimulation. The molecular mechanisms signalling this event and the details of the translocation pathway remain unknown. In type II diabetes, the cellular transport of glucose in response to insulin is impaired, partly explaining why blood-glucose levels in patients are not lowered by insulin as in normal individuals. MATERIALS AND METHODS: Isolated rat epididymal adipocytes were stimulated with insulin and subjected to subcellular fractionation and to measurement of glucose uptake. A caveolae-rich fraction was isolated from the plasma membranes after detergent solubilization and ultracentrifugal floatation in a sucrose gradient. Presence of GLUT4 and caveolin was determined by immunoblotting after SDS-PAGE. RESULTS: In freshly isolated adipocytes, insulin induced a rapid translocation of GLUT4 to the plasma membrane fraction, which was followed by a slower transition of the transporter into a detergent resistant caveolae-rich region of the plasma membrane. The insulin-stimulated appearance of transporters in the caveolae-rich fraction occurred in parallel with enhanced glucose uptake by cells. Treatment with isoproterenol plus adenosine deaminase rapidly inhibited insulin-stimulated glucose transport by 40%, and at the same time GLUT4 disappeared from the caveolae-rich fraction and from plasma membranes as a whole. CONCLUSIONS: Insulin stimulates glucose uptake in adipocytes by rapidly translocating GLUT4 from intracellular stores to the plasma membrane. This is followed by a slower transition of GLUT4 to the caveolae-rich regions of the plasma membrane, where glucose transport appears to take place. These results have implications for an understanding of the defect in glucose transport involved in type II diabetes.
The metabolic syndrome (MetS) confers an increased risk for diabetes and cardiovascular disease. Although high-sensitive C-reactive protein (hsCRP) concentrations are higher and adiponectin concentrations lower in MetS, there is no reliable biochemical measure that can capture its various features. We evaluated whether hsCRP, adiponectin, or the ratio of adiponectin or its oligomers, especially the high-molecular-weight (HMW) oligomer, to hsCRP predict MetS in 123 subjects with MetS compared with that in 91 healthy control subjects. MetS subjects had significantly higher hsCRP levels and lower total adiponectin and oligomer levels relative to control subjects (P < .0001). The HMW/total adiponectin and adiponectin/CRP ratios were significantly lower in MetS subjects than control subjects (P < .005). The odds ratio (OR) of MetS using the 75th percentile cutoff for CRP was 3.8 (95% confidence interval [CI], 2.1–6.8) and equivalent to low total adiponectin (OR, 2.5; 95% CI, 1.3–4.5), its oligomers, or the adiponectin/hsCRP ratio (OR, 2.6; 95% CI, 1.5, 4.8). Thus, measurements of CRP, adiponectin, or its oligomers provide robust biomarkers for predicting MetS.
C-reactive protein; Adiponectin; Biomarker; Metabolic syndrome