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Sample records for neuronal glucose transporter

  1. Difference in transient ischemia-induced neuronal damage and glucose transporter-1 immunoreactivity in the hippocampus between adult and young gerbils

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    Seung Min Park

    2016-05-01

    Full Text Available Objective(s: The alteration of glucose transporters is closely related with the pathogenesis of brain edema. We compared neuronal damage/death in the hippocampus between adult and young gerbils following transient cerebral ischemia/reperfusion and changes of glucose transporter-1(GLUT-1-immunoreactive microvessels in their ischemic hippocampal CA1 region. Materials and Methods: Transient cerebral ischemia was developed by 5-min occlusion of both common carotid arteries. Neuronal damage was examined by cresyl violet staining, NeuN immunohistochemistry and Fluoro-Jade B histofluorescence staining and changes in GLUT-1 expression was carried out by immunohistochemistry. Results: About 90% of pyramidal neurons only in the adult CA1 region were damaged after ischemia/reperfusion; in the young, about 53 % of pyramidal neurons were damaged from 7 days after ischemia/reperfusion. The density of GLUT-1-immunoreactive microvessels was significantly higher in the young sham-group than that in the adult sham-group. In the ischemia-operated-groups, the density of GLUT-1-immunoreactive microvessels was significantly decreased in the adult and young at 1 and 4 days post-ischemia, respectively, thereafter, the density of GLUT-1-immunoreactive microvessels was gradually increased in both groups after ischemia/reperfusion. Conclusion: CA1 pyramidal neurons of the young gerbil were damaged much later than that in the adult and that GLUT-1-immunoreactive microvessels were significantly decreased later in the young. These data indicate that GLUT-1 might differently contribute to neuronal damage according to age after ischemic insults.

  2. Glucose Sensing Neurons in the Ventromedial Hypothalamus

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    Vanessa H. Routh

    2010-10-01

    Full Text Available Neurons whose activity is regulated by glucose are found in a number of brain regions. Glucose-excited (GE neurons increase while glucose-inhibited (GI neurons decrease their action potential frequency as interstitial brain glucose levels increase. We hypothesize that these neurons evolved to sense and respond to severe energy deficit (e.g., fasting that threatens the brains glucose supply. During modern times, they are also important for the restoration of blood glucose levels following insulin-induced hypoglycemia. Our data suggest that impaired glucose sensing by hypothalamic glucose sensing neurons may contribute to the syndrome known as hypoglycemia-associated autonomic failure in which the mechanisms which restore euglycemia following hypoglycemia become impaired. On the other hand, increased responses of glucose sensing neurons to glucose deficit may play a role in the development of Type 2 Diabetes Mellitus and obesity. This review will discuss the mechanisms by which glucose sensing neurons sense changes in interstitial glucose and explore the roles of these specialized glucose sensors in glucose and energy homeostasis.

  3. GDNF pre-treatment aggravates neuronal cell loss in oxygen-glucose deprived hippocampal slice cultures: a possible effect of glutamate transporter up-regulation.

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    Bonde, C; Sarup, A; Schousboe, A; Gegelashvili, G; Noraberg, J; Zimmer, J

    2003-01-01

    Besides its neurotrophic and neuroprotective effects on dopaminergic neurons and spinal motoneurons, glial cell line-derived neurotrophic factor (GDNF) has potent neuroprotective effects in cerebral ischemia. The protective effect has so far been related to reduced activation of N-methyl-D-aspartate receptors (NMDAr). This study tested the effects of GDNF on glutamate transporter expression, with the hypothesis that modulation of glutamate transporter activity would affect the outcome of cerebral ischemia. Organotypic hippocampal slice cultures, derived from 1-week-old rats, were treated with 100 ng/ml GDNF for either 2 or 5 days, followed by Western blot analysis of NMDAr subunit 1 (NR1) and two glutamate transporter subtypes, GLAST and GLT-1. After 5-day exposure to GDNF, expression of GLAST and GLT-1 was up-regulated to 169 and 181% of control values, respectively, whereas NR1 was down-regulated to 64% of control. However, despite these changes that potentially would support neuronal resistance to excitotoxicity, the long-term treatment with GDNF was found to aggravate the neuronal damage induced by oxygen-glucose deprivation (OGD). The increased cell death, assessed by propidium iodide (PI) uptake, occurred not only among the most susceptible CA1 pyramidal cells, but also in CA3 and fascia dentata. Given that glutamate transporters are able to release glutamate by reversed action during energy failure, it is suggested that the observed increase in OGD-induced cell death in the GDNF-pretreated cultures was caused by the build-up of excitotoxic concentrations of extracellular glutamate released through the glutamate transporters, which were up-regulated by GDNF. Although the extent and consequences of glutamate release via reversal of GLAST and GLT-1 transporters seem to vary in different energy failure models, the present findings should be taken into account in clinical trials of GDNF.

  4. Ionotropic glutamate receptors and glutamate transporters are involved in necrotic neuronal cell death induced by oxygen-glucose deprivation of hippocampal slice cultures.

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    Bonde, C; Noraberg, J; Noer, H; Zimmer, J

    2005-01-01

    Organotypic hippocampal slice cultures represent a feasible model for studies of cerebral ischemia and the role of ionotropic glutamate receptors in oxygen-glucose deprivation-induced neurodegeneration. New results and a review of existing data are presented in the first part of this paper. The role of glutamate transporters, with special reference to recent results on inhibition of glutamate transporters under normal and energy-failure (ischemia-like) conditions is reviewed in the last part of the paper. The experimental work is based on hippocampal slice cultures derived from 7 day old rats and grown for about 3 weeks. In such cultures we investigated the subfield neuronal susceptibility to oxygen-glucose deprivation, the type of induced cell death and the involvement of ionotropic glutamate receptors. Hippocampal slice cultures were also used in our studies on glutamate transporters reviewed in the last part of this paper. Neurodegeneration was monitored and/or shown by cellular uptake of propidium iodide, loss of immunocytochemical staining for microtubule-associated protein 2 and staining with Fluoro-Jade B. To distinguish between necrotic vs. apoptotic neuronal cell death we used immunocytochemical staining for active caspase-3 (apoptosis indicator) and Hoechst 33342 staining of nuclear chromatin. Our experimental studies on oxygen-glucose deprivation confirmed that CA1 pyramidal cells were the most susceptible to this ischemia-like condition. Judged by propidium iodide uptake, a selective CA1 lesion, with only minor affection on CA3, occurred in cultures exposed to oxygen-glucose deprivation for 30 min. Nuclear chromatin staining by Hoechst 33342 and staining for active caspase-3 showed that oxygen-glucose deprivation induced necrotic cell death only. Addition of 10 microM of the N-methyl-D-aspartate glutamate receptor antagonist MK-801, and 20 microM of the non-N-methyl-D-aspartate glutamate receptor antagonist 2,3-dihyroxy-6-nitro-7-sulfamoyl

  5. 2-(4-Methoxyphenyl)ethyl-2-acetamido-2-deoxy-β-D-pyranoside confers neuroprotection in cell and animal models of ischemic stroke through calpain1/PKA/CREB-mediated induction of neuronal glucose transporter 3

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    Yu, Shu; Cheng, Qiong; Li, Lu; Liu, Mei; Yang, Yumin; Ding, Fei, E-mail: dingfei@ntu.edu.cn

    2014-06-15

    Salidroside is proven to be a neuroprotective agent of natural origin, and its analog, 2-(4-Methoxyphenyl)ethyl-2-acetamido-2-deoxy-β-D-pyranoside (named SalA-4 g), has been synthesized in our lab. In this study, we showed that SalA-4 g promoted neuronal survival and inhibited neuronal apoptosis in primary hippocampal neurons exposed to oxygen and glucose deprivation (OGD) and in rats subjected to ischemia by transient middle cerebral artery occlusion (MCAO), respectively, and that SalA-4 g was more neuroprotective than salidroside. We further found that SalA-4 g elevated glucose uptake in OGD-injured primary hippocampal neurons and increased the expression and recruitment of glucose transporter 3 (GLUT3) in ischemic brain. Signaling analysis revealed that SalA-4 g triggered the phosphorylation of CREB, and increased the expression of PKA RII in primary hippocampal neurons exposed to OGD injury, while inhibition of PKA/CREB by H-89 alleviated the elevation in glucose uptake and GLUT3 expression, and blocked the protective effects of SalA-4 g. Moreover, SalA-4 g was noted to inhibit intracellular Ca{sup 2+} influx and calpain1 activation in OGD-injured primary hippocampal neurons. Our results suggest that SalA-4 g neuroprotection might be mediated by increased glucose uptake and elevated GLUT3 expression through calpain1/PKA/CREB pathway. - Highlights: • A salidroside (Sal) analog (SalA-4 g) is prepared to be more neuroprotective than Sal. • SalA-4 g protected hippocampal neurons from oxygen and glucose deprivation insult. • SalA-4 g reduced ischemic injury after transient middle cerebral artery occlusion in rats. • Neuroprotection of SalA-4 g was mediated by GLUT3 level via calpain/PKA/CREB pathway.

  6. Glucose transport in adipose tissue

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    Schoonen, AJM; Wientjes, KJC

    2005-01-01

    Based on the well-known extraction equation and the histology of subcutaneous adipose tissue, transport of glucose from capillary to microdialysis probe is described. Results are evaluated of previous studies by our group and others. Arguments are presented for a simple scheme in which the mean

  7. Leptin regulates glutamate and glucose transporters in hypothalamic astrocytes

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    Fuente-Martín, Esther; García-Cáceres, Cristina; Granado, Miriam; de Ceballos, María L.; Sánchez-Garrido, Miguel Ángel; Sarman, Beatrix; Liu, Zhong-Wu; Dietrich, Marcelo O.; Tena-Sempere, Manuel; Argente-Arizón, Pilar; Díaz, Francisca; Argente, Jesús; Horvath, Tamas L.; Chowen, Julie A.

    2012-01-01

    Glial cells perform critical functions that alter the metabolism and activity of neurons, and there is increasing interest in their role in appetite and energy balance. Leptin, a key regulator of appetite and metabolism, has previously been reported to influence glial structural proteins and morphology. Here, we demonstrate that metabolic status and leptin also modify astrocyte-specific glutamate and glucose transporters, indicating that metabolic signals influence synaptic efficacy and glucose uptake and, ultimately, neuronal function. We found that basal and glucose-stimulated electrical activity of hypothalamic proopiomelanocortin (POMC) neurons in mice were altered in the offspring of mothers fed a high-fat diet. In adulthood, increased body weight and fasting also altered the expression of glucose and glutamate transporters. These results demonstrate that whole-organism metabolism alters hypothalamic glial cell activity and suggest that these cells play an important role in the pathology of obesity. PMID:23064363

  8. Methamphetamine inhibits the glucose uptake by human neurons and astrocytes: stabilization by acetyl-L-carnitine.

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    P M Abdul Muneer

    Full Text Available Methamphetamine (METH, an addictive psycho-stimulant drug exerts euphoric effects on users and abusers. It is also known to cause cognitive impairment and neurotoxicity. Here, we hypothesized that METH exposure impairs the glucose uptake and metabolism in human neurons and astrocytes. Deprivation of glucose is expected to cause neurotoxicity and neuronal degeneration due to depletion of energy. We found that METH exposure inhibited the glucose uptake by neurons and astrocytes, in which neurons were more sensitive to METH than astrocytes in primary culture. Adaptability of these cells to fatty acid oxidation as an alternative source of energy during glucose limitation appeared to regulate this differential sensitivity. Decrease in neuronal glucose uptake by METH was associated with reduction of glucose transporter protein-3 (GLUT3. Surprisingly, METH exposure showed biphasic effects on astrocytic glucose uptake, in which 20 µM increased the uptake while 200 µM inhibited glucose uptake. Dual effects of METH on glucose uptake were paralleled to changes in the expression of astrocytic glucose transporter protein-1 (GLUT1. The adaptive nature of astrocyte to mitochondrial β-oxidation of fatty acid appeared to contribute the survival of astrocytes during METH-induced glucose deprivation. This differential adaptive nature of neurons and astrocytes also governed the differential sensitivity to the toxicity of METH in these brain cells. The effect of acetyl-L-carnitine for enhanced production of ATP from fatty oxidation in glucose-free culture condition validated the adaptive nature of neurons and astrocytes. These findings suggest that deprivation of glucose-derived energy may contribute to neurotoxicity of METH abusers.

  9. Oral glucose intake inhibits hypothalamic neuronal activity more effectively than glucose infusion

    NARCIS (Netherlands)

    Smeets, P.A.M.; Vidarsdottir, S.; Graaf, C. de; Stafleu, A.; Osch, M.J.P. van; Viergever, M.A.; Pijl, H.; Grond, J. van der

    2007-01-01

    We previously showed that hypothalamic neuronal activity, as measured by the blood oxygen level-dependent (BOLD) functional MRI signal, declines in response to oral glucose intake. To further explore the mechanism driving changes in hypothalamic neuronal activity in response to an oral glucose load,

  10. Anorexia and impaired glucose metabolism in mice with hypothalamic ablation of Glut4 neurons.

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    Ren, Hongxia; Lu, Taylor Y; McGraw, Timothy E; Accili, Domenico

    2015-02-01

    The central nervous system (CNS) uses glucose independent of insulin. Nonetheless, insulin receptors and insulin-responsive glucose transporters (Glut4) often colocalize in neurons (Glut4 neurons) in anatomically and functionally distinct areas of the CNS. The apparent heterogeneity of Glut4 neurons has thus far thwarted attempts to understand their function. To answer this question, we used Cre-dependent, diphtheria toxin-mediated cell ablation to selectively remove basal hypothalamic Glut4 neurons and investigate the resulting phenotypes. After Glut4 neuron ablation, mice demonstrate altered hormone and nutrient signaling in the CNS. Accordingly, they exhibit negative energy balance phenotype characterized by reduced food intake and increased energy expenditure, without locomotor deficits or gross neuronal abnormalities. Glut4 neuron ablation affects orexigenic melanin-concentrating hormone neurons but has limited effect on neuropeptide Y/agouti-related protein and proopiomelanocortin neurons. The food intake phenotype can be partially normalized by GABA administration, suggesting that it arises from defective GABAergic transmission. Glut4 neuron-ablated mice show peripheral metabolic defects, including fasting hyperglycemia and glucose intolerance, decreased insulin levels, and elevated hepatic gluconeogenic genes. We conclude that Glut4 neurons integrate hormonal and nutritional cues and mediate CNS actions of insulin on energy balance and peripheral metabolism.

  11. Supraoptic oxytocin and vasopressin neurons function as glucose and metabolic sensors

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    Song, Zhilin; Levin, Barry E.; Stevens, Wanida

    2014-01-01

    Neurons in the supraoptic nuclei (SON) produce oxytocin and vasopressin and express insulin receptors (InsR) and glucokinase. Since oxytocin is an anorexigenic agent and glucokinase and InsR are hallmarks of cells that function as glucose and/or metabolic sensors, we evaluated the effect of glucose, insulin, and their downstream effector ATP-sensitive potassium (KATP) channels on calcium signaling in SON neurons and on oxytocin and vasopressin release from explants of the rat hypothalamo-neurohypophyseal system. We also evaluated the effect of blocking glucokinase and phosphatidylinositol 3 kinase (PI3K; mediates insulin-induced mobilization of glucose transporter, GLUT4) on responses to glucose and insulin. Glucose and insulin increased intracellular calcium ([Ca2+]i). The responses were glucokinase and PI3K dependent, respectively. Insulin and glucose alone increased vasopressin release (P sensors to participate in appetite regulation. PMID:24477542

  12. Anorexia and Impaired Glucose Metabolism in Mice With Hypothalamic Ablation of Glut4 Neurons

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    Ren, Hongxia; Lu, Taylor Y.; McGraw, Timothy E.

    2015-01-01

    The central nervous system (CNS) uses glucose independent of insulin. Nonetheless, insulin receptors and insulin-responsive glucose transporters (Glut4) often colocalize in neurons (Glut4 neurons) in anatomically and functionally distinct areas of the CNS. The apparent heterogeneity of Glut4 neurons has thus far thwarted attempts to understand their function. To answer this question, we used Cre-dependent, diphtheria toxin–mediated cell ablation to selectively remove basal hypothalamic Glut4 neurons and investigate the resulting phenotypes. After Glut4 neuron ablation, mice demonstrate altered hormone and nutrient signaling in the CNS. Accordingly, they exhibit negative energy balance phenotype characterized by reduced food intake and increased energy expenditure, without locomotor deficits or gross neuronal abnormalities. Glut4 neuron ablation affects orexigenic melanin-concentrating hormone neurons but has limited effect on neuropeptide Y/agouti-related protein and proopiomelanocortin neurons. The food intake phenotype can be partially normalized by GABA administration, suggesting that it arises from defective GABAergic transmission. Glut4 neuron–ablated mice show peripheral metabolic defects, including fasting hyperglycemia and glucose intolerance, decreased insulin levels, and elevated hepatic gluconeogenic genes. We conclude that Glut4 neurons integrate hormonal and nutritional cues and mediate CNS actions of insulin on energy balance and peripheral metabolism. PMID:25187366

  13. Negative Effects of High Glucose Exposure in Human Gonadotropin-Releasing Hormone Neurons

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    Annamaria Morelli

    2013-01-01

    Full Text Available Metabolic disorders are often associated with male hypogonadotropic hypogonadism, suggesting that hypothalamic defects involving GnRH neurons may impair the reproductive function. Among metabolic factors hyperglycemia has been implicated in the control of the reproductive axis at central level, both in humans and in animal models. To date, little is known about the direct effects of pathological high glucose concentrations on human GnRH neurons. In this study, we investigated the high glucose effects in the human GnRH-secreting FNC-B4 cells. Gene expression profiling by qRT-PCR, confirmed that FNC-B4 cells express GnRH and several genes relevant for GnRH neuron function (KISS1R, KISS1, sex steroid and leptin receptors, FGFR1, neuropilin 2, and semaphorins, along with glucose transporters (GLUT1, GLUT3, and GLUT4. High glucose exposure (22 mM; 40 mM significantly reduced gene and protein expression of GnRH, KISS1R, KISS1, and leptin receptor, as compared to normal glucose (5 mM. Consistent with previous studies, leptin treatment significantly induced GnRH mRNA expression at 5 mM glucose, but not in the presence of high glucose concentrations. In conclusion, our findings demonstrate a deleterious direct contribution of high glucose on human GnRH neurons, thus providing new insights into pathogenic mechanisms linking metabolic disorders to reproductive dysfunctions.

  14. Glucose Transporters in Cardiac Metabolism and Hypertrophy

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    Shao, Dan; Tian, Rong

    2016-01-01

    The heart is adapted to utilize all classes of substrates to meet the high-energy demand, and it tightly regulates its substrate utilization in response to environmental changes. Although fatty acids are known as the predominant fuel for the adult heart at resting stage, the heart switches its substrate preference toward glucose during stress conditions such as ischemia and pathological hypertrophy. Notably, increasing evidence suggests that the loss of metabolic flexibility associated with increased reliance on glucose utilization contribute to the development of cardiac dysfunction. The changes in glucose metabolism in hypertrophied hearts include altered glucose transport and increased glycolysis. Despite the role of glucose as an energy source, changes in other nonenergy producing pathways related to glucose metabolism, such as hexosamine biosynthetic pathway and pentose phosphate pathway, are also observed in the diseased hearts. This article summarizes the current knowledge regarding the regulation of glucose transporter expression and translocation in the heart during physiological and pathological conditions. It also discusses the signaling mechanisms governing glucose uptake in cardiomyocytes, as well as the changes of cardiac glucose metabolism under disease conditions. PMID:26756635

  15. Identification of Glucose Transporters in Aspergillus nidulans

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    dos Reis, Thaila Fernanda; Menino, João Filipe; Bom, Vinícius Leite Pedro; Brown, Neil Andrew; Colabardini, Ana Cristina; Savoldi, Marcela; Goldman, Maria Helena S.; Rodrigues, Fernando; Goldman, Gustavo Henrique

    2013-01-01

    To characterize the mechanisms involved in glucose transport, in the filamentous fungus Aspergillus nidulans, we have identified four glucose transporter encoding genes hxtB-E. We evaluated the ability of hxtB-E to functionally complement the Saccharomyces cerevisiae EBY.VW4000 strain that is unable to grow on glucose, fructose, mannose or galactose as single carbon source. In S. cerevisiae HxtB-E were targeted to the plasma membrane. The expression of HxtB, HxtC and HxtE was able to restore growth on glucose, fructose, mannose or galactose, indicating that these transporters accept multiple sugars as a substrate through an energy dependent process. A tenfold excess of unlabeled maltose, galactose, fructose, and mannose were able to inhibit glucose uptake to different levels (50 to 80 %) in these s. cerevisiae complemented strains. Moreover, experiments with cyanide-m-chlorophenylhydrazone (CCCP), strongly suggest that hxtB, -C, and –E mediate glucose transport via active proton symport. The A. nidulans ΔhxtB, ΔhxtC or ΔhxtE null mutants showed ~2.5-fold reduction in the affinity for glucose, while ΔhxtB and -C also showed a 2-fold reduction in the capacity for glucose uptake. The ΔhxtD mutant had a 7.8-fold reduction in affinity, but a 3-fold increase in the capacity for glucose uptake. However, only the ΔhxtB mutant strain showed a detectable decreased rate of glucose consumption at low concentrations and an increased resistance to 2-deoxyglucose. PMID:24282591

  16. Glucose sensing and signalling; regulation of intestinal glucose transport.

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    Shirazi-Beechey, S P; Moran, A W; Batchelor, D J; Daly, K; Al-Rammahi, M

    2011-05-01

    Epithelial cells lining the inner surface of the intestinal epithelium are in direct contact with a lumenal environment that varies dramatically with diet. It has long been suggested that the intestinal epithelium can sense the nutrient composition of lumenal contents. It is only recently that the nature of intestinal nutrient-sensing molecules and underlying mechanisms have been elucidated. There are a number of nutrient sensors expressed on the luminal membrane of endocrine cells that are activated by various dietary nutrients. We showed that the intestinal glucose sensor, T1R2+T1R3 and the G-protein, gustducin are expressed in endocrine cells. Eliminating sweet transduction in mice in vivo by deletion of either gustducin or T1R3 prevented dietary monosaccharide- and artificial sweetener-induced up-regulation of the Na+/glucose cotransporter, SGLT1 observed in wild-type mice. Transgenic mice, lacking gustducin or T1R3 had deficiencies in secretion of glucagon-like peptide 1 (GLP-1) and, glucose-dependent insulinotrophic peptide (GIP). Furthermore, they had an abnormal insulin profile and prolonged elevation of postprandial blood glucose in response to orally ingested carbohydrates. GIP and GLP-1 increase insulin secretion, while glucagon-like peptide 2 (GLP-2) modulates intestinal growth, blood flow and expression of SGLT1. The receptor for GLP-2 resides in enteric neurons and not in any surface epithelial cells, suggesting the involvement of the enteric nervous system in SGLT1 up-regulation. The accessibility of the glucose sensor and the important role that it plays in regulation of intestinal glucose absorption and glucose homeostasis makes it an attractive nutritional and therapeutic target for manipulation.

  17. DRP1 Suppresses Leptin and Glucose Sensing of POMC Neurons.

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    Santoro, Anna; Campolo, Michela; Liu, Chen; Sesaki, Hiromi; Meli, Rosaria; Liu, Zhong-Wu; Kim, Jung Dae; Diano, Sabrina

    2017-03-07

    Hypothalamic pro-opiomelanocortin (POMC) neurons regulate energy and glucose metabolism. Intracellular mechanisms that enable these neurons to respond to changes in metabolic environment are ill defined. Here we show reduced expression of activated dynamin-related protein (pDRP1), a mitochondrial fission regulator, in POMC neurons of fed mice. These POMC neurons displayed increased mitochondrial size and aspect ratio compared to POMC neurons of fasted animals. Inducible deletion of DRP1 of mature POMC neurons (Drp1(fl/fl)-POMC-cre:ER(T2)) resulted in improved leptin sensitivity and glucose responsiveness. In Drp1(fl/fl)-POMC-cre:ER(T2) mice, POMC neurons showed increased mitochondrial size, ROS production, and neuronal activation with increased expression of Kcnj11 mRNA regulated by peroxisome proliferator-activated receptor (PPAR). Furthermore, deletion of DRP1 enhanced the glucoprivic stimulus in these neurons, causing their stronger inhibition and a greater activation of counter-regulatory responses to hypoglycemia that were PPAR dependent. Together, these data unmasked a role for mitochondrial fission in leptin sensitivity and glucose sensing of POMC neurons. Copyright © 2017 Elsevier Inc. All rights reserved.

  18. Effect of endurance training on glucose transport capacity and glucose transporter expression in rat skeletal muscle

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    Ploug, T; Stallknecht, B M; Pedersen, O

    1990-01-01

    session. Half-times for reversal of contraction-induced glucose transport were similar in trained and untrained muscles. The concentrations of mRNA for GLUT-1 (the erythrocyte-brain-Hep G2 glucose transporter) and GLUT-4 (the adipocyte-muscle glucose transporter) were increased approximately twofold...... by training in fast-twitch red muscle fibers. In parallel to this, Western blot demonstrated a approximately 47% increase in GLUT-1 protein and a approximately 31% increase in GLUT-4 protein. This indicates that the increases in maximum velocity for 3-MG transport in trained muscle is due to an increased...

  19. Effect of endurance training on glucose transport capacity and glucose transporter expression in rat skeletal muscle

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    Ploug, T.; Stallknecht, B.M.; Pedersen, O.; Kahn, B.B.; Ohkuwa, T.; Vinten, J.; Galbo, H. (Panum Institute, Copenhagen (Denmark))

    1990-12-01

    The effect of 10 wk endurance swim training on 3-O-methylglucose (3-MG) uptake (at 40 mM 3-MG) in skeletal muscle was studied in the perfused rat hindquarter. Training resulted in an increase of approximately 33% for maximum insulin-stimulated 3-MG transport in fast-twitch red fibers and an increase of approximately 33% for contraction-stimulated transport in slow-twitch red fibers compared with nonexercised sedentary muscle. A fully additive effect of insulin and contractions was observed both in trained and untrained muscle. Compared with transport in control rats subjected to an almost exhaustive single exercise session the day before experiment both maximum insulin- and contraction-stimulated transport rates were increased in all muscle types in trained rats. Accordingly, the increased glucose transport capacity in trained muscle was not due to a residual effect of the last training session. Half-times for reversal of contraction-induced glucose transport were similar in trained and untrained muscles. The concentrations of mRNA for GLUT-1 (the erythrocyte-brain-Hep G2 glucose transporter) and GLUT-4 (the adipocyte-muscle glucose transporter) were increased approximately twofold by training in fast-twitch red muscle fibers. In parallel to this, Western blot demonstrated a approximately 47% increase in GLUT-1 protein and a approximately 31% increase in GLUT-4 protein. This indicates that the increases in maximum velocity for 3-MG transport in trained muscle is due to an increased number of glucose transporters.

  20. Osteopontin Upregulates the Expression of Glucose Transporters in Osteosarcoma Cells

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    Hsieh, I-Shan; Yang, Rong-Sen; Fu, Wen-Mei

    2014-01-01

    Osteosarcoma is the most common primary malignancy of bone. Even after the traditional standard surgical therapy, metastasis still occurs in a high percentage of patients. Glucose is an important source of metabolic energy for tumor proliferation and survival. Tumors usually overexpress glucose transporters, especially hypoxia-responsive glucose transporter 1 and glucose transporter 3. Osteopontin, hypoxia-responsive glucose transporter 1, and glucose transporter 3 are overexpressed in many types of tumors and have been linked to tumorigenesis and metastasis. In this study, we investigated the regulation of glucose transporters by osteopontin in osteosarcoma. We observed that both glucose transporters and osteopontin were upregulated in hypoxic human osteosarcoma cells. Endogenously released osteopontin regulated the expression of glucose transporter 1 and glucose transporter 3 in osteosarcoma and enhanced glucose uptake into cells via the αvβ3 integrin. Knockdown of osteopontin induced cell death in 20% of osteosarcoma cells. Phloretin, a glucose transporter inhibitor, also caused cell death by treatment alone. The phloretin-induced cell death was significantly enhanced in osteopontin knockdown osteosarcoma cells. Combination of a low dose of phloretin and chemotherapeutic drugs, such as daunomycin, 5-Fu, etoposide, and methotrexate, exhibited synergistic cytotoxic effects in three osteosarcoma cell lines. Inhibition of glucose transporters markedly potentiated the apoptotic sensitivity of chemotherapeutic drugs in osteosarcoma. These results indicate that the combination of a low dose of a glucose transporter inhibitor with cytotoxic drugs may be beneficial for treating osteosarcoma patients. PMID:25310823

  1. Determination of Glucose Utilization Rates in Cultured Astrocytes and Neurons with [(14)C]deoxyglucose: Progress, Pitfalls, and Discovery of Intracellular Glucose Compartmentation.

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    Dienel, Gerald A; Cruz, Nancy F; Sokoloff, Louis; Driscoll, Bernard F

    2017-01-01

    2-Deoxy-D-[(14)C]glucose ([(14)C]DG) is commonly used to determine local glucose utilization rates (CMRglc) in living brain and to estimate CMRglc in cultured brain cells as rates of [(14)C]DG phosphorylation. Phosphorylation rates of [(14)C]DG and its metabolizable fluorescent analog, 2-(N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino)-2-deoxyglucose (2-NBDG), however, do not take into account differences in the kinetics of transport and metabolism of [(14)C]DG or 2-NBDG and glucose in neuronal and astrocytic cells in cultures or in single cells in brain tissue, and conclusions drawn from these data may, therefore, not be correct. As a first step toward the goal of quantitative determination of CMRglc in astrocytes and neurons in cultures, the steady-state intracellular-to-extracellular concentration ratios (distribution spaces) for glucose and [(14)C]DG were determined in cultured striatal neurons and astrocytes as functions of extracellular glucose concentration. Unexpectedly, the glucose distribution spaces rose during extreme hypoglycemia, exceeding 1.0 in astrocytes, whereas the [(14)C]DG distribution space fell at the lowest glucose levels. Calculated CMRglc was greatly overestimated in hypoglycemic and normoglycemic cells because the intracellular glucose concentrations were too high. Determination of the distribution space for [(14)C]glucose revealed compartmentation of intracellular glucose in astrocytes, and probably, also in neurons. A smaller metabolic pool is readily accessible to hexokinase and communicates with extracellular glucose, whereas the larger pool is sequestered from hexokinase activity. A new experimental approach using double-labeled assays with DG and glucose is suggested to avoid the limitations imposed by glucose compartmentation on metabolic assays.

  2. [Glucose transporter type 1 (GLUT-1) deficiency].

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    Cano, A; Ticus, I; Chabrol, B

    2008-11-01

    Impaired glucose transport across the blood brain barrier results in glucose transporter type 1 (GLUT-1) deficiency syndrome, first described in 1991. It is characterized by infantile seizures refractory to anticonvulsive treatments, microcephaly, delays in mental and motor development, spasticity, ataxia, dysarthria and other paroxysmal neurologic phenomena, often occurring prior to meals. Affected infants are normal at birth following an uneventful pregnancy and delivery. Seizures usually begin between the age of one and four months and can be preceded by apneic episodes or abnormal eyes movements. Patients with atypical presentations such as mental retardation and intermittent ataxia without seizures, or movement disorders characterized by choreoathetosis and dystonia, have also been described. Glucose is the principal fuel source for the brain and GLUT-1 is the only vehicle by which glucose enters the brain. In case of GLUT-1 deficiency, the risk of clinical manifestations is increased in infancy and childhood, when the brain glucose demand is maximal. The hallmark of the disease is a low glucose concentration in the cerebrospinal fluid in a presence of normoglycemia (cerebrospinal fluid/blood glucose ratio less than 0.4). The GLUT-1 defect can be confirmed by molecular analysis of the SCL2A1 gene or in erythrocytes by glucose uptake studies and GLUT-1 immunoreactivity. Several heterozygous mutations, with a majority of de novo mutations, resulting in GLUT-1 haploinsufficiency, have been described. Cases with an autosomal dominant transmission have been established and adults can exhibit symptoms of this deficiency. Ketogenic diet is an effective treatment of epileptic manifestations as ketone bodies serve as an alternative fuel for the developing brain. However, this diet is not effective on cognitive impairment and other treatments are being evaluated. The physiopathology of this disorder is partially unclear and its understanding could explain the clinical

  3. The expression and regulation of glucose transporters in tumor cells

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    Pengfei Zhao

    2016-12-01

    Full Text Available Glucose transporter proteins are involved in many physiological and biochemical processes. In particular, the high expressions of sodium-glucose cotransporter and glucose transporter proteins in tumor cells show that these two transporters play a key role in tumor cell metabolism. Studying the crystal structure and conformation of human glucose transporter proteins has enabled the development of drugs based on specific binding sites, opening up a new path towards more effective cancer treatments. This mini review serves to summarize our existing understanding of the metabolic pathways of tumor cells, focusing on the roles of glucose transporter proteins.

  4. Exercise training, glucose transporters, and glucose transport in rat skeletal muscles

    Science.gov (United States)

    Rodnick, K. J.; Henriksen, E. J.; James, D. E.; Holloszy, J. O.

    1992-01-01

    It was previously found that voluntary wheel running induces an increase in the insulin-sensitive glucose transporter, i.e., the GLUT4 isoform, in rat plantaris muscle (K. J. Rodnick, J. O. Holloszy, C. E. Mondon, and D. E. James. Diabetes 39: 1425-1429, 1990). The present study was undertaken to determine whether 1) the increase in muscle GLUT4 protein is associated with an increase in maximally stimulated glucose transport activity, 2) a conversion of type IIb to type IIa or type I muscle fibers plays a role in the increase in GLUT4 protein, and 3) an increase in the GLUT1 isoform is a component of the adaptation of muscle to endurance exercise. Five weeks of voluntary wheel running that resulted in a 33% increase in citrate synthase activity induced a 50% increase in GLUT4 protein in epitrochlearis muscles of female Sprague-Dawley rats. The rate of 2-deoxy-glucose transport maximally stimulated with insulin or insulin plus contractions was increased approximately 40% (P less than 0.05). There was no change in muscle fiber type composition, evaluated by myosin ATPase staining, in the epitrochlearis. There was also no change in GLUT1 protein concentration. We conclude that an increase in GLUT4, but not of GLUT1 protein, is a component of the adaptive response of muscle to endurance exercise and that the increase in GLUT4 protein is associated with an increased capacity for glucose transport.

  5. Direct evidence for activity-dependent glucose phosphorylation in neurons with implications for the astrocyte-to-neuron lactate shuttle.

    Science.gov (United States)

    Patel, Anant B; Lai, James C K; Chowdhury, Golam M I; Hyder, Fahmeed; Rothman, Douglas L; Shulman, Robert G; Behar, Kevin L

    2014-04-01

    Previous (13)C magnetic resonance spectroscopy experiments have shown that over a wide range of neuronal activity, approximately one molecule of glucose is oxidized for every molecule of glutamate released by neurons and recycled through astrocytic glutamine. The measured kinetics were shown to agree with the stoichiometry of a hypothetical astrocyte-to-neuron lactate shuttle model, which predicted negligible functional neuronal uptake of glucose. To test this model, we measured the uptake and phosphorylation of glucose in nerve terminals isolated from rats infused with the glucose analog, 2-fluoro-2-deoxy-D-glucose (FDG) in vivo. The concentrations of phosphorylated FDG (FDG6P), normalized with respect to known neuronal metabolites, were compared in nerve terminals, homogenate, and cortex of anesthetized rats with and without bicuculline-induced seizures. The increase in FDG6P in nerve terminals agreed well with the increase in cortical neuronal glucose oxidation measured previously under the same conditions in vivo, indicating that direct uptake and oxidation of glucose in nerve terminals is substantial under resting and activated conditions. These results suggest that neuronal glucose-derived pyruvate is the major oxidative fuel for activated neurons, not lactate-derived from astrocytes, contradicting predictions of the original astrocyte-to-neuron lactate shuttle model under the range of study conditions.

  6. Palmitate stimulates glucose transport in rat adipocytes by a mechanism involving translocation of the insulin sensitive glucose transporter (GLUT4)

    Science.gov (United States)

    Hardy, R. W.; Ladenson, J. H.; Henriksen, E. J.; Holloszy, J. O.; McDonald, J. M.

    1991-01-01

    In rat adipocytes, palmitate: a) increases basal 2-deoxyglucose transport 129 +/- 27% (p less than 0.02), b) decreases the insulin sensitive glucose transporter (GLUT4) in low density microsomes and increases GLUT4 in plasma membranes and c) increases the activity of the insulin receptor tyrosine kinase. Palmitate-stimulated glucose transport is not additive with the effect of insulin and is not inhibited by the protein kinase C inhibitors staurosporine and sphingosine. In rat muscle, palmitate: a) does not affect basal glucose transport in either the soleus or epitrochlearis and b) inhibits insulin-stimulated glucose transport by 28% (p less than 0.005) in soleus but not in epitrochlearis muscle. These studies demonstrate a potentially important differential role for fatty acids in the regulation of glucose transport in different insulin target tissues.

  7. Glucose transporter 1 localisation throughout pregnancy in the carnivore placenta

    DEFF Research Database (Denmark)

    Wooding, F.B.P.; Dantzer, Vibeke; Klisch, K.

    2007-01-01

    Glucose is one of the major fetal nutrients. Maternofetal transfer requires transport across the several placental membranes. This transfer is mediated by one or more of the fourteen known isoforms of glucose transporter. So far only Glucose Transporters 1 and 3 (GT1, GT3) have been shown...... to be located in placental membranes. GT1 may be the only one on the syncytiotrophoblast (human) or both may be present on the same membrane (rodents) or be required in sequence (ruminants, horses and elephant). This paper shows GT1 to be the only transporter demonstrable by immunocytochemistry in carnivore...

  8. Glucose-induced insulin resistance of skeletal-muscle glucose transport and uptake

    DEFF Research Database (Denmark)

    Richter, Erik; Hansen, B F; Hansen, S A

    1988-01-01

    in the presence of glucose and insulin. The data indicate that exposure to a moderately increased glucose concentration (12 mM) leads to rapidly developing resistance of skeletal-muscle glucose transport and uptake to maximal insulin stimulation. The effect of glucose is enhanced by simultaneous insulin exposure......, whereas exposure for 5 h to insulin itself does not cause measurable resistance to maximal insulin stimulation.......The ability of glucose and insulin to modify insulin-stimulated glucose transport and uptake was investigated in perfused skeletal muscle. Here we report that perfusion of isolated rat hindlimbs for 5 h with 12 mM-glucose and 20,000 microunits of insulin/ml leads to marked, rapidly developing...

  9. Regulation of Intestinal Glucose Absorption by Ion Channels and Transporters

    Directory of Open Access Journals (Sweden)

    Lihong Chen

    2016-01-01

    Full Text Available The absorption of glucose is electrogenic in the small intestinal epithelium. The major route for the transport of dietary glucose from intestinal lumen into enterocytes is the Na+/glucose cotransporter (SGLT1, although glucose transporter type 2 (GLUT2 may also play a role. The membrane potential of small intestinal epithelial cells (IEC is important to regulate the activity of SGLT1. The maintenance of membrane potential mainly depends on the activities of cation channels and transporters. While the importance of SGLT1 in glucose absorption has been systemically studied in detail, little is currently known about the regulation of SGLT1 activity by cation channels and transporters. A growing line of evidence suggests that cytosolic calcium ([Ca2+]cyt can regulate the absorption of glucose by adjusting GLUT2 and SGLT1. Moreover, the absorption of glucose and homeostasis of Ca2+ in IEC are regulated by cation channels and transporters, such as Ca2+ channels, K+ channels, Na+/Ca2+ exchangers, and Na+/H+ exchangers. In this review, we consider the involvement of these cation channels and transporters in the regulation of glucose uptake in the small intestine. Modulation of them may be a potential strategy for the management of obesity and diabetes.

  10. Regulation of Intestinal Glucose Absorption by Ion Channels and Transporters.

    Science.gov (United States)

    Chen, Lihong; Tuo, Biguang; Dong, Hui

    2016-01-14

    The absorption of glucose is electrogenic in the small intestinal epithelium. The major route for the transport of dietary glucose from intestinal lumen into enterocytes is the Na⁺/glucose cotransporter (SGLT1), although glucose transporter type 2 (GLUT2) may also play a role. The membrane potential of small intestinal epithelial cells (IEC) is important to regulate the activity of SGLT1. The maintenance of membrane potential mainly depends on the activities of cation channels and transporters. While the importance of SGLT1 in glucose absorption has been systemically studied in detail, little is currently known about the regulation of SGLT1 activity by cation channels and transporters. A growing line of evidence suggests that cytosolic calcium ([Ca(2+)]cyt) can regulate the absorption of glucose by adjusting GLUT2 and SGLT1. Moreover, the absorption of glucose and homeostasis of Ca(2+) in IEC are regulated by cation channels and transporters, such as Ca(2+) channels, K⁺ channels, Na⁺/Ca(2+) exchangers, and Na⁺/H⁺ exchangers. In this review, we consider the involvement of these cation channels and transporters in the regulation of glucose uptake in the small intestine. Modulation of them may be a potential strategy for the management of obesity and diabetes.

  11. Relationship between cerebral sodium-glucose transporter and hyperglycemia in cerebral ischemia.

    Science.gov (United States)

    Yamazaki, Yui; Harada, Shinichi; Tokuyama, Shogo

    2015-09-14

    Post-ischemic hyperglycemia exacerbates the development of cerebral ischemia. To elucidate this exacerbation mechanism, we focused on sodium-glucose transporter (SGLT) as a mediator that lead hyperglycemia to cerebral ischemia. SGLT transport glucose into the cell, together with sodium ion, using the sodium concentration gradient. We have previously reported that suppression of cerebral SGLT ameliorates cerebral ischemic neuronal damage. However, detail relationship cerebral between SGLT and post-ischemic hyperglycemia remain incompletely defined. Therefore, we examined the involvement of cerebral SGLT on cerebral ischemic neuronal damage with or without hyperglycemic condition. Cell survival rate of primary cultured neurons was assessed by biochemical assay. A mouse model of focal ischemia was generated using a middle cerebral artery occlusion (MCAO). Neuronal damage was assessed with histological and behavioral analyses. Concomitant hydrogen peroxide/glucose treatment exacerbated hydrogen peroxide alone-induced cell death. Although a SGLT family-specific inhibitor, phlorizin had no effect on developed hydrogen peroxide alone-induced cell death, it suppressed cell death induced by concomitant hydrogen peroxide/glucose treatment. α-MG induced a concentration-dependent and significant decrease in neuronal survival. PHZ administered on immediately after reperfusion had no effect, but PHZ given at 6h after reperfusion had an effect. Our in vitro study indicates that SGLT is not involved in neuronal cell death in non-hyperglycemic condition. We have already reported that post-ischemic hyperglycemia begins to develop at 6h after MCAO. Therefore, current our in vivo study show post-ischemic hyperglycemic condition may be necessary for the SGLT-mediated exacerbation of cerebral ischemic neuronal damage.

  12. Mechanical stress and glucose concentration modulate glucose transport in cultured rat podocytes.

    Science.gov (United States)

    Lewko, Barbara; Bryl, Ewa; Witkowski, Jacek M; Latawiec, Elzbieta; Angielski, Stefan; Stepinski, Jan

    2005-02-01

    Recent studies show that mechanical stress modifies both morphology and protein expression in podocytes. Ambient glucose is another factor modulating protein synthesis in these cells. In diabetes, podocytes experience elevated glucose concentrations as well as mechanical strain generated by high intracapillary pressures. Both these factors are responsible for podocyte injury, leading to impairment of kidney glomerular function. In the present study, we examined the effects of glucose concentration and mechanical stress on glucose uptake in podocytes. Following a 24 h pre-incubation in low (2.5 mM, LG), normal (5.6 mM, NG) or high (30 mM, HG) glucose media, cultured rat podocytes were exposed to 4 h mechanical stress. We used the labelled glucose analogue, [3H]2-deoxy-D-glucose, to measure glucose uptake. The distribution of facilitative glucose transporters GLUT2 and GLUT4 was assessed by flow cytometry. In the control (static) cells, glucose uptake was similar in the three glucose groups. In mechanically stressed podocytes, glucose uptake increased 2-fold in the LG and NG groups but increased 3-fold in the HG group. In the NG cells, mechanical load increased the membrane expression of GLUT2 and reduced the membrane-bound GLUT4. In stretched HG cells, the membrane expression of both GLUT2 and GLUT4 was decreased. High glucose decreased the plasma membrane GLUT2 content in the stretched cells, whereas both static and stretched podocytes showed an elevation in GLUT4. Mechanical stress potentiated glucose uptake in podocytes and this effect was enhanced by high ambient glucose. The decreased expression of GLUT2 and GLUT4 on the surface of stretched cells suggests that the activity of other glucose transporters may be regulated by mechanical stress in podocytes.

  13. Biological evaluation of two iodine-123-labeled D-glucose acetals prepared as glucose transporter radioligands

    Energy Technology Data Exchange (ETDEWEB)

    Brunet-Desruet, Marie-Dominique; Ghezzi, Catherine; Morin, Christophe; Comet, Michel; Fagret, Daniel

    1998-07-01

    Two iodinated acetals of D-glucose, 4,6-(R)-O-(2'-iodoethylidene)-{alpha}, {beta}-D-glucose and 4,6-(R)-O-(4'-iodobenzylidene)-{alpha}, {beta}-D-glucose , were prepared and their potential as suitable SPECT radioligands for imaging of glucose transporters was studied. Both are analogs of acetal D-glucose derivatives, which are known to bind to the exofacial sites of the glucose transport protein (GluT). To assess whether iodinated acetals 1 and 2 interacted with the glucose transporter, they were tested in vitro in human erythrocytes (GluT1) and neonatal rat cardiomyocytes (GluT4). The results indicated that 1 and 2 had a very low affinity for the glucose transporter and probably accumulated in cells. Study of their tissue distribution was carried out in the mouse in vivo: Both compounds showed fast tissue clearance with preferential renal elimination. It is concluded that iodinated acetals of D-glucose 1 and 2 are not suitable for GluT targeting in vivo.

  14. Direct neuronal glucose uptake Heralds activity-dependent increases in cerebral metabolism

    DEFF Research Database (Denmark)

    Lundgaard, Iben; Li, Baoman; Xie, Lulu

    2015-01-01

    Metabolically, the brain is a highly active organ that relies almost exclusively on glucose as its energy source. According to the astrocyte-to-neuron lactate shuttle hypothesis, glucose is taken up by astrocytes and converted to lactate, which is then oxidized by neurons. Here we show, using two...

  15. Glucose transport in isolated prosthecae of Asticcacaulis biprosthecum.

    Science.gov (United States)

    Larson, R J; Pate, J L

    1976-04-01

    Active transport of glucose in prosthecae isolated from cells of Asticcacaulis biprosthecum was stimulated by the non-physiological electron donor N, N, N', N'-tetramethyl-p-phenylenediamine dihydrochloride. Glucose uptake was mediated by two transport systems; the apparent Km of the high-affinity system was 1.8 muM and that of the low-affinity system was 34 muM. Free glucose accumulated within prosthecae at a concentration 60 to 200 times above that present externally, depending on the Km of the system being observed. The glucose transport system in prosthecae was stereospecific for D-glucose, and neither methyl alpha-D-glucopyranoside nor 2-deoxyglucose was transported. Uptake of glucose was inhibited by N-ethylmaleimide (NEM) and p-chloromercuribenzoate (PCMB), and the inhibition by PCMB but not by NEM was reversed by dithiothreitol. Glucose uptake was also inhibited by the uncoupling agents 5-chloro-3-t-butyl-2'-nitrosalicylanilide (S-13), 5-chloro-3-(p-chlorophenyl)-4'-chlorosalicylanilide (S-6), and carbonyl-cyanide m-chlorophenylhydrazone (CCCP) and by the respiratory inhibitor KCN. Efflux of glucose from preloaded prosthecae was induced by PCMB and KCN, but not by S-13 or CCCP. Glucose uptake was not affected by arsenate or an inhibitor of membrane-bound adenosine triphosphatases, N, N'-dicyclohexylcarbodiimide. The lack of inhibition by these two compounds, combined with the extremely low levels of adenosine 5'-triphosphate present in prosthecae, indicates that adenosine 5'-triphosphate is not involved in the transport of glucose by prosthecae.

  16. Predominant enhancement of glucose uptake in astrocytes versus neurons during activation of the somatosensory cortex

    OpenAIRE

    Chuquet, Julien; Quilichini, Pascale; Nimchinsky, Esther A.; Buzsáki, György

    2010-01-01

    Glucose is the primary energetic substrate of the brain and measurements of its metabolism are the basis of major functional cerebral imaging methods. Contrary to the general view that neurons are fueled solely by glucose in proportion to their energetic needs, recent in vitro and ex vivo analyses suggest that glucose preferentially feeds astrocytes. However, the cellular fate of glucose in the intact brain has not yet been directly observed. We have used a real-time method for measuring gluc...

  17. Peritoneal transport characteristics with glucose polymer based dialysate.

    Science.gov (United States)

    Ho-dac-Pannekeet, M M; Schouten, N; Langendijk, M J; Hiralall, J K; de Waart, D R; Struijk, D G; Krediet, R T

    1996-09-01

    /PNa+ decreased with 3.86% glucose until 60 minutes, followed by a subsequent increase. The ultrafiltration coefficient (UFC) of the total peritoneal membrane was assessed using 3.86% glucose (0.18 +/- 0.04 ml/min/mm Hg), and the UFC of the small pores was assessed using icodextrin (0.06 +/- 0.008 ml/min/mm Hg). The difference between these represented the UFC through the transcellular pores, which averaged 50.5% of the total UFC, but with a very wide range (0 to 85%). An inverse relation existed between the duration of CAPD treatment and the total ultrafiltration coefficient (r = -0.68, P < 0.04), which could be attributed to a lower UFC of the transcellular pores in long-term patients (r = -0.66, P < 0.05), but not to the UFC of the small pores (r = -0.48, NS). The TCUFRo-60 min through the transcellular pores correlated with the sodium gradient, corrected for diffusion, in the first hour of the dwell (r = 0.69, P < 0.04), indicating that both parameters indeed measure transcellular water transport. It can be concluded that the glucose polymer solution induced sustained ultrafiltration and had no effect on peritoneal membrane characteristics. In addition, the results of the present study support the hypothesis that the glucose polymer solutions exerts its osmotic pressure across intercellular pores with radii of about 40 A. This leads to increased clearances of low molecular weight proteins such as beta 2m that are transported through these pores without sieving of Na+. The latter, as found during 3.86% glucose dialysate, is probably caused by transcellular water transport. The transcellular water transport accounted for 50% of the total ultrafiltration with glucose based dialysis solutions. It was lower in long-term CAPD patients.

  18. Metabolic regulation of lateral hypothalamic glucose-inhibited orexin neurons may influence midbrain reward neurocircuitry.

    Science.gov (United States)

    Sheng, Zhenyu; Santiago, Ammy M; Thomas, Mark P; Routh, Vanessa H

    2014-09-01

    Lateral hypothalamic area (LHA) orexin neurons modulate reward-based feeding by activating ventral tegmental area (VTA) dopamine (DA) neurons. We hypothesize that signals of peripheral energy status influence reward-based feeding by modulating the glucose sensitivity of LHA orexin glucose-inhibited (GI) neurons. This hypothesis was tested using electrophysiological recordings of LHA orexin-GI neurons in brain slices from 4 to 6week old male mice whose orexin neurons express green fluorescent protein (GFP) or putative VTA-DA neurons from C57Bl/6 mice. Low glucose directly activated ~60% of LHA orexin-GFP neurons in both whole cell and cell attached recordings. Leptin indirectly reduced and ghrelin directly enhanced the activation of LHA orexin-GI neurons by glucose decreases from 2.5 to 0.1mM by 53±12% (n=16, PFasting increased activation of LHA orexin-GI neurons by decreased glucose, as would be predicted by these hormonal effects. We also evaluated putative VTA-DA neurons in a novel horizontal slice preparation containing the LHA and VTA. Decreased glucose increased the frequency of spontaneous excitatory post-synaptic currents (sEPSCs; 125 ± 40%, n=9, P<0.05) and action potentials (n=9; P<0.05) in 45% (9/20) of VTA DA neurons. sEPSCs were completely blocked by AMPA and NMDA glutamate receptor antagonists (CNQX 20 μM, n=4; APV 20μM, n=4; respectively), demonstrating that these sEPSCs were mediated by glutamatergic transmission onto VTA DA neurons. Orexin-1 but not 2 receptor antagonism with SB334867 (10μM; n=9) and TCS-OX2-29 (2μM; n=5), respectively, blocks the effects of decreased glucose on VTA DA neurons. Thus, decreased glucose increases orexin-dependent excitatory glutamate neurotransmission onto VTA DA neurons. These data suggest that the glucose sensitivity of LHA orexin-GI neurons links metabolic state and reward-based feeding.

  19. Effects of electroacupuncture on microcirculatory blood flow and glucose transporter function in the hippocampus

    Institute of Scientific and Technical Information of China (English)

    Yan Lu; Bingbing Han; Shijun Wang

    2011-01-01

    Nerve cell metabolism in post brain ischemia depends on increased microcirculation perfusion and transport function of microvascular endothelial cells. In the present study, a rat model of middle cerebral artery occlusion was established to investigate the influence of electroacupuncture(EA)on hippocampal CA1 cerebral blood flow and glucose transporter 1(GLUT1)expression in the microvascular endothelial cp.lls. Following EA at Neiguan(PC 6), the cerebral blood flow in the ischemic hippocampal CA1 region was significantly elevated, the number and microvascular integrated absorbance of the GLUTl-positive cells were significantly increased, nerve cell damage was ameliorated, and GLUT1 protein expression in the ischemic hippocampus was significantly increased. Results demonstrate that EA increased the cerebral blood flow of the hippocampal CA1 region and improved the glucose transport function, thereby attenuating neuronal injuries.

  20. The Effect of Noscapine on Oxygen-Glucose Deprivation on Primary Murine Cortical Neurons in High Glucose Condition.

    Science.gov (United States)

    Vahabzadeh, Gelareh; Ebrahimi, Soltan-Ahmed; Rahbar-Roshandel, Nahid; Mahmoudian, Massoud

    2016-01-01

    In the present work we set out to investigate the neuroprotective effects of noscapine (0.5-2 µM) in presence of D-glucose on primary murine foetal cortical neurons after oxygen-glucose deprivation/24 h. recovery. Cell viability, nitric oxide production and intracellular calcium ((ca(2+))i) levels were evaluated by MTT assay, the modified Griess method and Fura-2 respectively. 25 and 100 mM D-glucose could, in a concentration dependent manner, improve cell viability and decrease NO production and (ca(2+))i level in neuronal cells after ischemic insult. Moreover, pre-incubation of cells with noscapine, noticeably enhanced protective effects of 25 and 100 mM D-glucose compared to similar conditions without noscapine pre-treatment. In fact, noscapine attenuated NO production in a dose-dependent fashion, after 30 minutes (min) OGD, during high-glucose (HG) condition in cortical neurons. Pretreatment with 2 μM noscapine and 25 or 100 mM D-glucose, was shown to decrease the rise in (ca(2+))i induced by Sodium azide/glucose deprivation (chemical OGD) model. These effects were more pronounced than that of 25 or 100 mM D-glucose alone. The present study demonstrated that the neuroprotective effects of HG before an ischemic insult were augmented by pre-treatment with noscapine. Our results also suggested that the neuroprotection offered by both HG and noscapine involve attenuation of NO production and (ca(2+))i levels stimulated by the experimental ischemia in cortical neurons.

  1. The role of cysteine residues in glucose-transporter-GLUT1-mediated transport and transport inhibition.

    Science.gov (United States)

    Wellner, M; Monden, I; Keller, K

    1994-01-01

    The role of cysteine residues in transport function of the glucose transporter GLUT1 was investigated by a mutagenesis-expression strategy. Each of the six cysteine residues was individually replaced by site-directed mutagenesis. Expression of the heterologous wild-type or mutant glucose transporters and transport measurements at two hexose concentrations (50 microM and 5 mM) were undertaken in Xenopus oocytes. The catalytic activity of GLUT1 was retained, despite substitution of each single cysteine residue, which indicated that no individual residue is essential for hexose transport. This finding questions the involvement of oligomerization or intramolecular stabilization by a single disulphide bond as a prerequisite for transporter activation under basal conditions. Application of the impermeant mercurial thiol-group-reactive reagent p-chloromercuribenzenesulphonate (pCMBS) to the external or internal surface of plasma membrane demonstrated that cysteine-429, within the sixth external loop, and cysteine-207, at the beginning of the large intracellular loop which connects transmembrane segments 6 and 7, are the residues which are involved in transport inhibition by impermeant thiol-group-reactive reagents from either side of the cell. These data support the predicted membrane topology of the transport protein by transport measurements. If residues other than the cysteines at positions 429 or 207 are exposed to either side of the plasma membrane by conformational changes, they do not contribute to the transport inhibition by pCMBS. Application of pCMBS to one side of the plasma membrane also inhibited transport from the opposite direction, most likely due to the hindrance of sugar-induced interconversion of transporter conformation. PMID:8192671

  2. Diabetic ketoacidosis, sodium glucose transporter-2 inhibitors and the kidney.

    Science.gov (United States)

    Palmer, Biff F; Clegg, Deborah J; Taylor, Simeon I; Weir, Matthew R

    2016-08-01

    Diabetic ketoacidosis is a serious metabolic condition that may occur in patients with either Type 1 or Type 2 diabetes. The accumulation of ketoacids in the serum is a consequence of insulin deficiency and glucagon excess. Sodium Glucose Transporter 2 (SGLT2) inhibitors are novel therapeutic treatments for improving glucose homeostasis in patients with diabetes. Through reductions in glucose reabsorption by the kidney, they lower serum glucose in patients with Type 2 diabetes and they improve glucose control whether used alone or in combination with other therapies. Mechanistically, these drugs increase serum ketoacids and increase glucagon production, which in some individuals, can lead to formation of diabetic ketoacidosis. This review will first focus in how the kidney normally handles ketoacids, and second will discuss how the SGLT2 inhibitors affect the kidney in such a way so as to enhance the risk for development of ketoacidosis in susceptible individuals.

  3. Regulation of endogenous glucose production in glucose transporter 4 over-expressing mice.

    Directory of Open Access Journals (Sweden)

    Eric D Berglund

    Full Text Available Strategies to amplify whole-body glucose disposal are key therapies to treat type 2 diabetes. Mice that over-express glucose transporter 4 (Glut4 in skeletal muscle, heart, and adipose tissue (G4Tg exhibit increased fasting glucose disposal and thus lowered blood glucose. Intriguingly, G4Tg mice also exhibit improved insulin-stimulated suppression of endogenous glucose production even though Glut4 is not present in the liver. It is unclear, however, if hepatic gluco-regulation is altered in G4Tg mice in the basal, non-insulin-stimulated state. The current studies were performed to examine fasting hepatic glucose metabolism in G4Tg mice and to determine whether gluco-regulatory adaptations exist in the non-insulin-stimulated condition. To test this question, phloridzin-glucose clamps were used to match blood glucose and pancreatic hormone levels while tracer dilution techniques were used to measure glucose flux. These techniques were performed in chronically-catheterized, conscious, and un-stressed 5h-fasted G4Tg and wild-type (WT littermates. Results show reduced blood glucose, hepatic glycogen content, and hepatic glucokinase (GK activity/expression as well as higher endogenous glucose production, glucose disposal, arterial glucagon, and hepatic glucose-6-phosphatase (G6Pase activity/expression in G4Tg mice versus WT controls. Clamping blood glucose for 90 min at ~115 mg/dLin G4Tg and WT mice normalized nearly all variables. Notably, however, net hepatic glycogen synthetic rates were disproportionately elevated compared to changes in blood glucose. In conclusion, these studies demonstrate that basal improvements in glucose tolerance due to increased uptake in extra-hepatic sites provoke important gluco-regulatory adaptations in the liver. Although changes in blood glucose underlie the majority of these adaptations, net hepatic glycogen synthesis is sensitized. These data emphasize that anti-diabetic therapies that target skeletal muscle, heart

  4. Berberine stimulates glucose transport through a mechanism distinct from insulin.

    Science.gov (United States)

    Zhou, Libin; Yang, Ying; Wang, Xiao; Liu, Shangquan; Shang, Wenbin; Yuan, Guoyue; Li, Fengying; Tang, Jinfeng; Chen, Mingdao; Chen, Jialun

    2007-03-01

    Berberine exerts a hypoglycemic effect, but the mechanism remains unknown. In the present study, the effect of berberine on glucose uptake was characterized in 3T3-L1 adipocytes. It was revealed that berberine stimulated glucose uptake in 3T3-L1 adipocytes in a dose- and time-dependent manner with the maximal effect at 12 hours. Glucose uptake was increased by berberine in 3T3-L1 preadipocytes as well. Berberine-stimulated glucose uptake was additive to that of insulin in 3T3-L1 adipocytes, even at the maximal effective concentrations of both components. Unlike insulin, the effect of berberine on glucose uptake was insensitive to wortmannin, an inhibitor of phosphatidylinositol 3-kinase, and SB203580, an inhibitor of p38 mitogen-activated protein kinase. Berberine activated extracellular signal-regulated kinase (ERK) 1/2, but PD98059, an ERK kinase inhibitor, only decreased berberine-stimulated glucose uptake by 32%. Berberine did not induce Ser473 phosphorylation of Akt nor enhance insulin-induced phosphorylation of Akt. Meanwhile, the expression and cellular localization of glucose transporter 4 (GLUT4) were not altered by berberine. Berberine did not increase GLUT1 gene expression. However, genistein, a tyrosine kinase inhibitor, completely blocked berberine-stimulated glucose uptake in 3T3-L1 adipocytes and preadipocytes, suggesting that berberine may induce glucose transport via increasing GLUT1 activity. In addition, berberine increased adenosine monophosphate-activated protein kinase and acetyl-coenzyme A carboxylase phosphorylation. These findings suggest that berberine increases glucose uptake through a mechanism distinct from insulin, and activated adenosine monophosphate-activated protein kinase seems to be involved in the metabolic effect of berberine.

  5. Effects of ketamine on glucose uptake by glucose transporter type 3 expressed in Xenopus oocytes: The role of protein kinase C

    Energy Technology Data Exchange (ETDEWEB)

    Tomioka, Shigemasa, E-mail: tomioka@dent.tokushima-u.ac.jp [Department of Dental Anesthesiology, Institute of Health Biosciences, The University of Tokushima Graduate School, Kuramoto-cho 18-15, Tokushima City, Tokushima 770-8504 (Japan); Kaneko, Miyuki [Department of Dental Anesthesiology, Institute of Health Biosciences, The University of Tokushima Graduate School, Kuramoto-cho 18-15, Tokushima City, Tokushima 770-8504 (Japan); Satomura, Kazuhito [First Department of Oral and Maxillofacial Surgery, Institute of Health Biosciences, The University of Tokushima Graduate School, Kuramoto-cho 18-15, Tokushima City, Tokushima 770-8504 (Japan); Mikyu, Tomiko; Nakajo, Nobuyoshi [Department of Dental Anesthesiology, Institute of Health Biosciences, The University of Tokushima Graduate School, Kuramoto-cho 18-15, Tokushima City, Tokushima 770-8504 (Japan)

    2009-10-09

    We investigated the effects of ketamine on the type 3 facilitative glucose transporter (GLUT3), which plays a major role in glucose transport across the plasma membrane of neurons. Human-cloned GLUT3 was expressed in Xenopus oocytes by injection of GLUT3 mRNA. GLUT3-mediated glucose uptake was examined by measuring oocyte radioactivity following incubation with 2-deoxy-D-[1,2-{sup 3}H]glucose. While ketamine and S(+)-ketamine significantly increased GLUT3-mediated glucose uptake, this effect was biphasic such that higher concentrations of ketamine inhibited glucose uptake. Ketamine (10 {mu}M) significantly increased V{sub max} but not K{sub m} of GLUT3 for 2-deoxy-D-glucose. Although staurosporine (a protein kinase C inhibitor) increased glucose uptake, no additive or synergistic interactions were observed between staurosporine and racemic ketamine or S(+)-ketamine. Treatment with ketamine or S(+)-ketamine partially prevented GLUT3 inhibition by the protein kinase C activator phorbol-12-myrisate-13-acetate. Our results indicate that ketamine increases GLUT3 activity at clinically relevant doses through a mechanism involving PKC inhibition.

  6. Kinetics of glucose transport in rat muscle

    DEFF Research Database (Denmark)

    Ploug, Thorkil; Galbo, H; Vinten, J

    1987-01-01

    -MG concentration exhibited Michaelis-Menten kinetics. Uptake by simple diffusion could not be detected. The maximum 3-O-MG transport velocity (Vmax) was increased more by maximum isometric contractions (10- to 40-fold, depending on fiber type) than by insulin (20,000 microU/ml; 3- to 20-fold) in both red and white...

  7. Brain Glucose Transporter (Glut3) Haploinsufficiency Does Not Impair Mouse Brain Glucose Uptake

    OpenAIRE

    Stuart, Charles A.; Ross, Ian R.; Howell, Mary E. A.; McCurry, Melanie P.; Wood, Thomas G.; Ceci, Jeffrey D.; Kennel, Stephen J.; Wall, Jonathan

    2011-01-01

    Mouse brain expresses three principle glucose transporters. Glut1 is an endothelial marker and is the principal glucose transporter of the blood-brain barrier. Glut3 and Glut6 are expressed in glial cells and neural cells. A mouse line with a null allele for Glut3 has been developed. The Glut3−/− genotype is intrauterine lethal by seven days post-coitis, but the heterozygous (Glut3+/−) littermate survives, exhibiting rapid post-natal weight gain, but no seizures or other behavioral aberration...

  8. Glucose is necessary to maintain neurotransmitter homeostasis during synaptic activity in cultured glutamatergic neurons

    DEFF Research Database (Denmark)

    Bak, Lasse K; Schousboe, Arne; Sonnewald, Ursula

    2006-01-01

    Glucose is the primary energy substrate for the adult mammalian brain. However, lactate produced within the brain might be able to serve this purpose in neurons. In the present study, the relative significance of glucose and lactate as substrates to maintain neurotransmitter homeostasis was inves...

  9. Glucose Levels in Culture Medium Determine Cell Death Mode in MPP(+)-treated Dopaminergic Neuronal Cells.

    Science.gov (United States)

    Yoon, So-Young; Oh, Young J

    2015-09-01

    We previously demonstrated that 1-methyl-4-phenylpyridinium (MPP(+)) causes caspase-independent, non-apoptotic death of dopaminergic (DA) neuronal cells. Here, we specifically examined whether change of glucose concentration in culture medium may play a role for determining cell death modes of DA neurons following MPP(+) treatment. By incubating MN9D cells in medium containing varying concentrations of glucose (5~35 mM), we found that cells underwent a distinct cell death as determined by morphological and biochemical criteria. At 5~10 mM glucose concentration (low glucose levels), MPP(+) induced typical of the apoptotic dell death accompanied with caspase activation and DNA fragmentation as well as cell shrinkage. In contrast, MN9D cells cultivated in medium containing more than 17.5 mM (high glucose levels) did not demonstrate any of these changes. Subsequently, we observed that MPP(+) at low glucose levels but not high glucose levels led to ROS generation and subsequent JNK activation. Therefore, MPP(+)-induced cell death only at low glucose levels was significantly ameliorated following co-treatment with ROS scavenger, caspase inhibitor or JNK inhibitor. We basically confirmed the quite similar pattern of cell death in primary cultures of DA neurons. Taken together, our results suggest that a biochemically distinct cell death mode is recruited by MPP(+) depending on extracellular glucose levels.

  10. Characterization of 6-deoxy-6-iodo-D-glucose: A potential new tool to assess glucose transport

    Energy Technology Data Exchange (ETDEWEB)

    Henry, Christelle; Tanti, Jean-Francois; Gremeaux, Thierry; Morin, Christophe; Van Obberghen, Emmanuel; Comet, Michel; Le Marchand-Brustel, Yannick

    1997-01-01

    6-deoxy-6-iodo-D-glucose (6-DIG) was rapidly taken up by adipocytes. Insulin increased 6-DIG transport in adipocytes isolated from both rats and mice. This stimulation was more important in rat than in mouse adipocytes, in agreement with their respective amount of Glut 4 transporters. In two insulin-resistant states, the biological behavior of 6-DIG and 3-O-methyl-D-glucose was similar. These results indicated that 6-DIG, which was transported into the cells via the glucose transporters, could be potentially useful to measure modifications of glucose transport.

  11. Effect of aspirin on glucose-D transport in intestine of rat

    Institute of Scientific and Technical Information of China (English)

    Mazhar Mushtaq; Farah Deeba Khan; M. Naeem Akhtar; Saghir Ahmad Jafri; Mehboob Bari

    2009-01-01

    Objective: The present study was designed to evaluate the effect a commonly prescribed Non Steroidal Anti In-flammatory Drug (NSAID) i.e. aspirin on brush border membrane in terms of changes in the intestinal transport level of glucose which is monosaccharide with absolute requirement in the body and hence its absorption is directly proportional on the morphology of the intestinal mucosa. Method: Albino rats (Rattus Norvegicus) were divided into two different groups, Group Ⅰ (Control), Group Ⅱ ( aspirin-treated, 50 mg aspirin/kg of body weight). The treatment was continued for 28 days. On the 29th day after o-, vernight fasting, intestine was removed from animals of both groups. Changes in transport of glucose-D in intestine were studied. Result: The results indicated a significant decrease in the transport of glucose-D in aspirin treated group as compared to the con-trol group. Conclusion: Cautious use of NSAID is recommended in commonly observed symptom such as headache and to those patients who are given as a prophylaxis for thrombosis.

  12. Endothelial HIF-1α Enables Hypothalamic Glucose Uptake to Drive POMC Neurons.

    Science.gov (United States)

    Varela, Luis; Suyama, Shigetomo; Huang, Yan; Shanabrough, Marya; Tschöp, Matthias H; Gao, Xiao-Bing; Giordano, Frank J; Horvath, Tamas L

    2017-06-01

    Glucose is the primary driver of hypothalamic proopiomelanocortin (POMC) neurons. We show that endothelial hypoxia-inducible factor 1α (HIF-1α) controls glucose uptake in the hypothalamus and that it is upregulated in conditions of undernourishment, during which POMC neuronal activity is decreased. Endothelium-specific knockdown of HIF-1α impairs the ability of POMC neurons to adapt to the changing metabolic environment in vivo, resulting in overeating after food deprivation in mice. The impaired functioning of POMC neurons was reversed ex vivo or by parenchymal glucose administration. These observations indicate an active role for endothelial cells in the central control of metabolism and suggest that central vascular impairments may cause metabolic disorders. © 2017 by the American Diabetes Association.

  13. Berberine acutely activates the glucose transport activity of GLUT1.

    Science.gov (United States)

    Cok, Alexandra; Plaisier, Christina; Salie, Matthew J; Oram, Daniel S; Chenge, Jude; Louters, Larry L

    2011-07-01

    Berberine, which has a long history of use in Chinese medicine, has recently been shown to have efficacy in the treatment of diabetes. While the hypoglycemic effect of berberine has been clearly documented in animal and cell line models, such as 3T3-L1 adipocytes and L6 myotube cells, the mechanism of action appears complex with data implicating activation of the insulin signaling pathway as well as activation of the exercise or AMP kinase-mediated pathway. There have been no reports of the acute affects of berberine on the transport activity of the insulin-insensitive glucose transporter, GLUT1. Therefore, we examined the acute effects of berberine on glucose uptake in L929 fibroblast cells, a cell line that express only GLUT1. Berberine- activated glucose uptake reaching maximum stimulation of five-fold at >40 μM. Significant activation (P berberine effect was not additive to the maximal stimulation by other known stimulants, azide, methylene blue or glucose deprivation, suggesting shared steps between berberine and these stimulants. Berberine significantly reduced the K(m) of glucose uptake from 6.7 ± 1.9 mM to 0.55 ± 0.08 mM, but had no effect on the V(max) of uptake. Compound C, an inhibitor of AMP kinase, did not affect berberine-stimulated glucose uptake, but inhibitors of downstream kinases partially blocked berberine stimulation. SB203580 (inhibitor of p38 MAP kinase) did not affect submaximal berberine activation, but did lower maximal berberine stimulation by 26%, while PD98059 (inhibitor of ERK kinase) completely blocked submaximal berberine activation and decreased the maximal stimulation by 55%. It appears from this study that a portion of the hypoglycemic effects of berberine can be attributed to its acute activation of the transport activity of GLUT1.

  14. Hepatic expression and cellular distribution of the glucose transporter family

    Institute of Scientific and Technical Information of China (English)

    Sumera Karim; David H Adams; Patricia F Lalor

    2012-01-01

    Glucose and other carbohydrates are transported into cells using members of a family of integral membrane glucose transporter (GLUT) molecules.To date 14 members of this family,also called the solute carrier 2A proteins have been identified which are divided on the basis of transport characteristics and sequence similarities into several families (Classes 1 to 3).The expression of these different receptor subtypes varies between different species,tissues and cellular subtypes and each has differential sensitivities to stimuli such as insulin.The liver is a contributor to metabolic carbohydrate homeostasis and is a major site for synthesis,storage and redistribution of carbohydrates.Situations in which the balance of glucose homeostasis is upset such as diabetes or the metabolic syndrome can lead metabolic disturbances that drive chronic organ damage and failure,confirming the importance of understanding the molecular regulation of hepatic glucose homeostasis.There is a considerable literature describing the expression and function of receptors that regulate glucose uptake and release by hepatocytes,the most import cells in glucose regulation and glycogen storage.However there is less appreciation of the roles of GLUTs expressed by non parenchymal cell types within the liver,all of which require carbohydrate to function.A better understanding of the detailed cellular distribution of GLUTs in human liver tissue may shed light on mechanisms underlying disease pathogenesis.This review summarises the available literature on hepatocellular expression of GLUTs in health and disease and highlights areas where further investigation is required.

  15. Near infrared radiation rescues mitochondrial dysfunction in cortical neurons after oxygen-glucose deprivation.

    Science.gov (United States)

    Yu, Zhanyang; Liu, Ning; Zhao, Jianhua; Li, Yadan; McCarthy, Thomas J; Tedford, Clark E; Lo, Eng H; Wang, Xiaoying

    2015-04-01

    Near infrared radiation (NIR) is known to penetrate and affect biological systems in multiple ways. Recently, a series of experimental studies suggested that low intensity NIR may protect neuronal cells against a wide range of insults that mimic diseases such as stroke, brain trauma and neurodegeneration. However, the potential molecular mechanisms of neuroprotection with NIR remain poorly defined. In this study, we tested the hypothesis that low intensity NIR may attenuate hypoxia/ischemia-induced mitochondrial dysfunction in neurons. Primary cortical mouse neuronal cultures were subjected to 4 h oxygen-glucose deprivation followed by reoxygenation for 2 h, neurons were then treated with a 2 min exposure to 810-nm NIR. Mitochondrial function markers including MTT reduction and mitochondria membrane potential were measured at 2 h after treatment. Neurotoxicity was quantified 20 h later. Our results showed that 4 h oxygen-glucose deprivation plus 20 h reoxygenation caused 33.8 ± 3.4 % of neuron death, while NIR exposure significantly reduced neuronal death to 23.6 ± 2.9 %. MTT reduction rate was reduced to 75.9 ± 2.7 % by oxygen-glucose deprivation compared to normoxic controls, but NIR exposure significantly rescued MTT reduction to 87.6 ± 4.5 %. Furthermore, after oxygen-glucose deprivation, mitochondria membrane potential was reduced to 48.9 ± 4.39 % of normoxic control, while NIR exposure significantly ameliorated this reduction to 89.6 ± 13.9 % of normoxic control. Finally, NIR significantly rescued OGD-induced ATP production decline at 20 min after NIR. These findings suggest that low intensity NIR can protect neurons against oxygen-glucose deprivation by rescuing mitochondrial function and restoring neuronal energetics.

  16. Enhancement of glucose transport by selected plant foods in muscle cell line L6.

    Science.gov (United States)

    Noipha, K; Ratanachaiyavong, S; Ninla-Aesong, P

    2010-08-01

    Glucose uptake activity of 11 plant foods was assessed in L6 myotubes. Among them onion and ginger showed potent enhancement of glucose transport. This effect required new protein synthesis of glucose transporters. In addition, onion-induced glucose uptake in L6 myotubes was mediated through activation of phosphoinositide 3-kinase.

  17. Altered DNA methylation of glucose transporter 1 and glucose transporter 4 in patients with major depressive disorder.

    Science.gov (United States)

    Kahl, Kai G; Georgi, Karsten; Bleich, Stefan; Muschler, Marc; Hillemacher, Thomas; Hilfiker-Kleinert, Denise; Schweiger, Ulrich; Ding, Xiaoqi; Kotsiari, Alexandra; Frieling, Helge

    2016-05-01

    Alterations in brain glucose metabolism and in peripheral glucose metabolism have frequently been observed in major depressive disorder (MDD). The insulin independent glucose transporter 1 (GLUT1) plays a key role in brain metabolism while the insulin-dependent GLUT4 is the major glucose transporter for skeletal and cardiac muscle. We therefore examined methylation of GLUT1 and GLUT4 in fifty-two depressed inpatients and compared data to eighteen healthy comparison subjects. DNA methylation of the core promoter regions of GLUT1 and GLUT4 was assessed by bisulfite sequencing. Further factors determined were fasting glucose, cortisol, insulin, interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α). We found significantly increased methylation of the GLUT1 in depressed inpatients compared to healthy comparison subjects (CG). Further findings comprise increased concentrations of fasting cortisol, glucose, insulin, and increased IL-6 and TNF-α. After six weeks of inpatient treatment, significantly lower GLUT1 methylation was observed in remitted patients compared to non-remitters. GLUT4 methylation was not different between depressed patients and CG, and did not differ between remitted and non-remitted patients. Although preliminary we conclude from our results that the acute phase of major depressive disorder is associated with increased GLUT1 methylation and mild insulin resistance. The successful treatment of depression is associated with normalization of GLUT1 methylation in remitters, indicating that this condition may be reversible. Failure of normalization of GLUT1 methylation in non-remitters may point to a possible role of impeded brain glucose metabolism in the maintenance of MDD.

  18. Glucose transport by epithelia prepared from harvested enterocytes

    DEFF Research Database (Denmark)

    Kimura, Yasuhiro; van der Merwe, Marie; Bering, Stine Brandt

    2015-01-01

    , forskolin, and health status exceeding those we have measured using intact tissues. Our findings indicate that epithelia prepared from harvested enterocytes provide an alternative approach for comparative studies of the characteristics of nutrient transport by the upper villus epithelium and the responses...... transporter SGLT-1. Similarly, accumulation of (14)C D-glucose by the epithelia was inhibited by phloridzin, but not phloretin, and was stimulated by pre-exposure to AMP and adenosine, apparently by a microtubule-based mechanism that is disrupted by nocodazole, with the magnitudes of responses to adenosine...

  19. Glucose metabolism and astrocyte-neuron interactions in the neonatal brain.

    Science.gov (United States)

    Brekke, Eva; Morken, Tora Sund; Sonnewald, Ursula

    2015-03-01

    Glucose is essentially the sole fuel for the adult brain and the mapping of its metabolism has been extensive in the adult but not in the neonatal brain, which is believed to rely mainly on ketone bodies for energy supply. However, glucose is absolutely indispensable for normal development and recent studies have shed light on glycolysis, the pentose phosphate pathway and metabolic interactions between astrocytes and neurons in the 7-day-old rat brain. Appropriately (13)C labeled glucose was used to distinguish between glycolysis and the pentose phosphate pathway during development. Experiments using (13)C labeled acetate provided insight into the GABA-glutamate-glutamine cycle between astrocytes and neurons. It could be shown that in the neonatal brain the part of this cycle that transfers glutamine from astrocytes to neurons is operating efficiently while, in contrast, little glutamate is shuttled from neurons to astrocytes. This lack of glutamate for glutamine synthesis is compensated for by anaplerosis via increased pyruvate carboxylation relative to that in the adult brain. Furthermore, compared to adults, relatively more glucose is prioritized to the pentose phosphate pathway than glycolysis and pyruvate dehydrogenase activity. The reported developmental differences in glucose metabolism and neurotransmitter synthesis may determine the ability of the brain at various ages to resist excitotoxic insults such as hypoxia-ischemia.

  20. Crystal structure of the human glucose transporter GLUT1

    Science.gov (United States)

    Deng, Dong; Xu, Chao; Sun, Pengcheng; Wu, Jianping; Yan, Chuangye; Hu, Mingxu; Yan, Nieng

    2014-06-01

    The glucose transporter GLUT1 catalyses facilitative diffusion of glucose into erythrocytes and is responsible for glucose supply to the brain and other organs. Dysfunctional mutations may lead to GLUT1 deficiency syndrome, whereas overexpression of GLUT1 is a prognostic indicator for cancer. Despite decades of investigation, the structure of GLUT1 remains unknown. Here we report the crystal structure of human GLUT1 at 3.2 Å resolution. The full-length protein, which has a canonical major facilitator superfamily fold, is captured in an inward-open conformation. This structure allows accurate mapping and potential mechanistic interpretation of disease-associated mutations in GLUT1. Structure-based analysis of these mutations provides an insight into the alternating access mechanism of GLUT1 and other members of the sugar porter subfamily. Structural comparison of the uniporter GLUT1 with its bacterial homologue XylE, a proton-coupled xylose symporter, allows examination of the transport mechanisms of both passive facilitators and active transporters.

  1. Quantitative measurement of neuronal degeneration in organotypic hippocampal cultures after combined oxygen/glucose deprivation.

    Science.gov (United States)

    Strasser, U; Fischer, G

    1995-04-01

    Organotypic hippocampal cultures were used to study cell degeneration during the recovery period after defined periods (30 and 60 min) of combined oxygen/glucose deprivation mimicking transient ischemic conditions. Staining with the fluorescent dye propidium iodide allowed detection of damaged cells. Fluorescence intensity was measured by an image analysis system and used to quantify cell damage at different time points during the recovery period (up to 22 h). At 30 min of oxygen/glucose deprivation cells in the CA1 area were relatively more sensitive compared to CA3 and dentate gyrus cells, with respect to the time course of degeneration and the percentage of affected cells. Expanding the oxygen/glucose deprivation period from 30 to 60 min drastically increased the percentage of cells dying in all hippocampal areas. Still, however, cells in CA1 degenerated faster compared to those in the CA3 area and dentate gyrus. A histological analysis of toluidine blue as well as MAP2-immunostained sections revealed that almost all neurons degenerated in all hippocampal areas following the 60-min deprivation period, whereas GFAP-stained astrocytes appeared to be unaffected. Therefore, neuronal degeneration could be quantified by taking the fluorescence intensity values 22 h after 60 min of oxygen/glucose deprivation as 100% neuronal damage. The possibility to quantify neuronal damage in organotypic cultures offers a useful tool for detailed studies on mechanisms of neuronal cell death in a cell culture system which is closer to in situ conditions than monolayer cell cultures.

  2. Calpain activation induced by glucose deprivation is mediated by oxidative stress and contributes to neuronal damage.

    Science.gov (United States)

    Páramo, Blanca; Montiel, Teresa; Hernández-Espinosa, Diego R; Rivera-Martínez, Marlene; Morán, Julio; Massieu, Lourdes

    2013-11-01

    The mechanisms leading to neuronal death during glucose deprivation have not been fully elucidated, but a role of oxidative stress has been suggested. In the present study we have investigated whether the production of reactive oxygen species during glucose deprivation, contributes to the activation of calpain, a calcium-dependent protease involved in neuronal injury associated with brain ischemia and cerebral trauma. We have observed a rapid activation of calpain, as monitored by the cleavage of the cytoskeletal protein α-spectrin, after glucose withdrawal, which is reduced by inhibitors of xanthine oxidase, phospholipase A2 and NADPH oxidase. Results suggest that phospholipase A2 and NADPH oxidase contribute to the early activation of calpain after glucose deprivation. In particular NOX2, a member of the NADPH oxidase family is involved, since reduced stimulation of calpain activity is observed after glucose deprivation in hippocampal slices from transgenic mice lacking a functional NOX2. We observed an additive effect of the inhibitors of xanthine oxidase and phospholipase A2 on both ROS production and calpain activity, suggesting a synergistic action of these two enzymes. The present results provide new evidence showing that reactive oxygen species stimulate calpain activation during glucose deprivation and that this mechanism is involved in neuronal death.

  3. Molecular Dynamics Simulations of the Human Glucose Transporter GLUT1.

    Directory of Open Access Journals (Sweden)

    Min-Sun Park

    Full Text Available Glucose transporters (GLUTs provide a pathway for glucose transport across membranes. Human GLUTs are implicated in devastating diseases such as heart disease, hyper- and hypo-glycemia, type 2 diabetes and cancer. The human GLUT1 has been recently crystalized in the inward-facing open conformation. However, there is no other structural information for other conformations. The X-ray structures of E. coli Xylose permease (XylE, a glucose transporter homolog, are available in multiple conformations with and without the substrates D-xylose and D-glucose. XylE has high sequence homology to human GLUT1 and key residues in the sugar-binding pocket are conserved. Here we construct a homology model for human GLUT1 based on the available XylE crystal structure in the partially occluded outward-facing conformation. A long unbiased all atom molecular dynamics simulation starting from the model can capture a new fully opened outward-facing conformation. Our investigation of molecular interactions at the interface between the transmembrane (TM domains and the intracellular helices (ICH domain in the outward- and inward-facing conformation supports that the ICH domain likely stabilizes the outward-facing conformation in GLUT1. Furthermore, inducing a conformational transition, our simulations manifest a global asymmetric rocker switch motion and detailed molecular interactions between the substrate and residues through the water-filled selective pore along a pathway from the extracellular to the intracellular side. The results presented here are consistent with previously published biochemical, mutagenesis and functional studies. Together, this study shed light on the structure and functional relationships of GLUT1 in multiple conformational states.

  4. Glucose deprivation activates diversity of potassium channels in cultured rat hippocampal neurons.

    Science.gov (United States)

    Velasco, Myrian; García, Esperanza; Onetti, Carlos G

    2006-05-01

    1. Glucose is one of the most important substrates for generating metabolic energy required for the maintenance of cellular functions. Glucose-mediated changes in neuronal firing pattern have been observed in the central nervous system of mammals. K(+) channels directly regulated by intracellular ATP have been postulated as a linkage between cellular energetic metabolism and excitability; the functional roles ascribed to these channels include glucose-sensing to regulate energy homeostasis and neuroprotection under energy depletion conditions. The hippocampus is highly sensitive to metabolic insults and is the brain region most sensitive to ischemic damage. Because the identity of metabolically regulated potassium channels present in hippocampal neurons is obscure, we decided to study the biophysical properties of glucose-sensitive potassium channels in hippocampal neurons. 2. The dependence of membrane potential and the sensitivity of potassium channels to glucose and ATP in rat hippocampal neurons were studied in cell-attached and excised inside-out membrane patches. 3. We found that under hypoglycemic conditions, at least three types of potassium channels were activated; their unitary conductance values were 37, 147, and 241 pS in symmetrical K(+), and they were sensitive to ATP. For K(+) channels with unitary conductance of 37 and 241, when the membrane potential was depolarized the longer closed time constant diminished and this produced an increase in the open-state probability; nevertheless, the 147-pS channels were not voltage-dependent. 4. We propose that neuronal glucose-sensitive K(+) channels in rat hippocampus include subtypes of ATP-sensitive channels with a potential role in neuroprotection during short-term or prolonged metabolic stress.

  5. Sweet Taste Receptor Serves to Activate Glucose- and Leptin-Responsive Neurons in the Hypothalamic Arcuate Nucleus and Participates in Glucose Responsiveness

    Science.gov (United States)

    Kohno, Daisuke; Koike, Miho; Ninomiya, Yuzo; Kojima, Itaru; Kitamura, Tadahiro; Yada, Toshihiko

    2016-01-01

    The hypothalamic feeding center plays an important role in energy homeostasis. In the feeding center, whole-body energy signals including hormones and nutrients are sensed, processed, and integrated. As a result, food intake and energy expenditure are regulated. Two types of glucose-sensing neurons exist in the hypothalamic arcuate nucleus (ARC): glucose-excited neurons and glucose-inhibited neurons. While some molecules are known to be related to glucose sensing in the hypothalamus, the mechanisms underlying glucose sensing in the hypothalamus are not fully understood. The sweet taste receptor is a heterodimer of taste type 1 receptor 2 (T1R2) and taste type 1 receptor 3 (T1R3) and senses sweet tastes. T1R2 and T1R3 are distributed in multiple organs including the tongue, pancreas, adipose tissue, and hypothalamus. However, the role of sweet taste receptors in the ARC remains to be clarified. To examine the role of sweet taste receptors in the ARC, cytosolic Ca2+ concentration ([Ca2+]i) in isolated single ARC neurons were measured using Fura-2 fluorescent imaging. An artificial sweetener, sucralose at 10−5–10−2 M dose dependently increased [Ca2+]i in 12–16% of ARC neurons. The sucralose-induced [Ca2+]i increase was suppressed by a sweet taste receptor inhibitor, gurmarin. The sucralose-induced [Ca2+]i increase was inhibited under an extracellular Ca2+-free condition and in the presence of an L-type Ca2+ channel blocker, nitrendipine. Sucralose-responding neurons were activated by high-concentration of glucose. This response to glucose was markedly suppressed by gurmarin. More than half of sucralose-responding neurons were activated by leptin but not ghrelin. Percentages of proopiomelanocortin (POMC) neurons among sucralose-responding neurons and sweet taste receptor expressing neurons were low, suggesting that majority of sucralose-responding neurons are non-POMC neurons. These data suggest that sweet taste receptor-mediated cellular activation mainly

  6. Contribution of Glucose Transport to the Control of the Glycolytic Flux in Trypanosoma brucei

    Science.gov (United States)

    Bakker, Barbara M.; Walsh, Michael C.; Ter Kuile, Benno H.; Mensonides, Femke I. C.; Michels, Paul A. M.; Opperdoes, Fred R.; Westerhoff, Hans V.

    1999-08-01

    The rate of glucose transport across the plasma membrane of the bloodstream form of Trypanosoma brucei was modulated by titration of the hexose transporter with the inhibitor phloretin, and the effect on the glycolytic flux was measured. A rapid glucose uptake assay was developed to measure the transport activity independently of the glycolytic flux. Phloretin proved a competitive inhibitor. When the effect of the intracellular glucose concentration on the inhibition was taken into account, the flux control coefficient of the glucose transporter was between 0.3 and 0.5 at 5 mM glucose. Because the flux control coefficients of all steps in a metabolic pathway sum to 1, this result proves that glucose transport is not the rate-limiting step of trypanosome glycolysis. Under physiological conditions, transport shares the control with other steps. At glucose concentrations much lower than physiological, the glucose carrier assumed all control, in close agreement with model predictions.

  7. Deficient Rab11 activity underlies glucose hypometabolism in primary neurons of Huntington's disease mice

    Energy Technology Data Exchange (ETDEWEB)

    Li, Xueyi, E-mail: xli12@partners.org [Department of Neurology, Massachusetts General Hospital and Harvard Medical School, Charlestown, MA 02129 (United States); Valencia, Antonio; McClory, Hollis; Sapp, Ellen; Kegel, Kimberly B. [Department of Neurology, Massachusetts General Hospital and Harvard Medical School, Charlestown, MA 02129 (United States); DiFiglia, Marian, E-mail: difiglia@helix.mgh.harvard.edu [Department of Neurology, Massachusetts General Hospital and Harvard Medical School, Charlestown, MA 02129 (United States)

    2012-05-18

    Highlights: Black-Right-Pointing-Pointer Primary Huntington's disease neurons are impaired in taking up glucose. Black-Right-Pointing-Pointer Rab11 modulates glucose uptake in neurons. Black-Right-Pointing-Pointer Increasing Rab11 activity attenuates the glucose uptake defect in disease neurons. Black-Right-Pointing-Pointer We provide a novel mechanism for glucose hypometabolism in Huntington's disease. -- Abstract: Huntington's disease (HD) is a progressive neurodegenerative disorder caused by a CAG repeat expansion in the huntingtin gene. Positron emission tomography studies have revealed a decline in glucose metabolism in the brain of patients with HD by a mechanism that has not been established. We examined glucose utilization in embryonic primary cortical neurons of wild-type (WT) and HD knock-in mice, which have 140 CAG repeats inserted in the endogenous mouse huntingtin gene (HD{sup 140Q/140Q}). Primary HD{sup 140Q/140Q} cortical neurons took up significantly less glucose than did WT neurons. Expression of permanently inactive and permanently active forms of Rab11 correspondingly altered glucose uptake in WT neurons, suggesting that normal activity of Rab11 is needed for neuronal uptake of glucose. It is known that Rab11 activity is diminished in HD{sup 140Q/140Q} neurons. Expression of dominant active Rab11 to enhance the activity of Rab11 normalized glucose uptake in HD{sup 140Q/140Q} neurons. These results suggest that deficient activity of Rab11 is a novel mechanism for glucose hypometabolism in HD.

  8. Is contraction-stimulated glucose transport feedforward regulated by Ca2+?

    DEFF Research Database (Denmark)

    Jensen, Thomas Elbenhardt; Angin, Yeliz; Sylow, Lykke

    2014-01-01

    feedforward regulator of the translocation of glucose transporter 4 to the cell surface to facilitate transmembrane glucose transport. This review summarizes the evidence supporting the Ca(2+) feedforward model and its proposed signalling links to regulation of glucose transport in skeletal muscle and other......-stimulated glucose transport. A revised working model is proposed, in which muscle glucose transport during contraction is not directly regulated by SR Ca(2+) release but rather responds exclusively to feedback signals activated secondary to cross-bridge cycling and tension development....

  9. Adipocyte glucose transport regulation by eicosanoid precursors and inhibitors

    Energy Technology Data Exchange (ETDEWEB)

    Lee, H.C.C.

    1987-01-01

    Glucose uptake and free fatty acid release by adipocytes are increased by catecholamines. The mechanism of the stimulatory action of catecholamines on glucose uptake may be via eicosanoid production from release fatty acids. Rats were fed iso-nutrient diets with high or low safflower oil. After one month, 5 rats per diet group were fed diets with aspirin or without aspirin for 2 days. Isolated adipocytes from epididymal fat pads were incubated at 37/sup 0/C, gassed with 95% O/sub 2/-5% CO/sub 2/ in KRB buffer with 3% bovine serum albumin and with or without eicosanoid modifiers; a stimulator (10/sup -5/ M norepinephrine, N), or inhibitors (167 ..mu..l of antiserum to prostaglandin E (AntiE) per 1600 ..mu..l or 23mM Asp), or combinations of these. At 2-, 5-, and 10-min incubation, samples of incubation mixtures were taken to measure 2-deoxy glucose transport using /sup 3/H-2-deoxy glucose, /sup 14/C-inulin, and liquid scintillation counter.

  10. Decreased glucose transporter 1 gene expression and glucose uptake in fetal brain exposed to ethanol

    Energy Technology Data Exchange (ETDEWEB)

    Singh, S.P.; Pullen, G.L.; Srivenugopal, K.S.; Yuan Xiaohua; Snyder, A.K. (Veterans Affairs Medical Center, North Chicago, IL (United States) Chicago Medical School, North Chicago, IL (United States))

    1992-01-01

    Using pregnant rats fed equicaloric liquid diets (AF, ad libitum-fed controls; PF, pair-fed controls; EF, ethanol-fed), the authors have previously shown that maternal alcoholism produces a specific and significant decrease of glucose in the fetal brain, which is accompanied by growth retardation. To further define the mechanisms of ethanol-induced perturbations in fetal fuel supply, they have examined (I) the uptake of 2-deoxyglucose (2-DG) by dissociated brain cells from fetal rats that were exposed to ethanol in utero and (II) the steady-state levels of the glucose transporter-1 (GT-1) mRNA. A 9% decrease in brain weight and a 54.8% reduction in 2-DG uptake into brain cells were found in offspring of EF mothers compared to the AF group. Brain weight correlated with the rate of 2-DG uptake. Northern blot analysis showed a 50% reduction of GT-1 mRNA in EF brain relative to that in the AF and PF groups. They conclude that glucose transport into the brain is an important parameter altered by maternal ethanol ingestion.

  11. Serotonin 2c receptors in pro-opiomelanocortin neurons regulate energy and glucose homeostasis

    Science.gov (United States)

    Energy and glucose homeostasis are regulated by central serotonin 2C receptors. These receptors are attractive pharmacological targets for the treatment of obesity; however, the identity of the serotonin 2C receptor-expressing neurons that mediate the effects of serotonin and serotonin 2C receptor a...

  12. Electroacupuncture regulates glucose-inhibited neurons in treatment of simple obesity

    Institute of Scientific and Technical Information of China (English)

    Zhi Yu; Youbing Xia; Chuanhui Ju; Qinghua Shao; Zhen Mao; Yun Gu; Bin Xu

    2013-01-01

    The glucose-inhibited neurons present in the lateral hypothalamic area are regarded as glucose detectors. This structure is involved in the regulation of food intake through extracellular blood glucose concentrations, and plays a crucial role in obesity onset. In the present study, obesity models established with high fat feeding were treated with electroacupuncture at Zusanli (ST36)/ Inner Court (ST44) on the left side and Tianshu (ST25) bilaterally. We found that electroacupuncture could effectively reduce body weight and the fat-weight ratio, and decrease serum leptin, resistin, tumor necrosis factor alpha, and neuropeptide Y levels, while increase serum adiponectin and cholecystokinin-8 levels. This treatment altered the electrical activity of glucose-inhibited neurons in the lateral hypothalamic area, with electroacupuncture at Zusanli/ Inner Court exerting an inhibitory effect, while electroacupuncture at bilateral Tianshu exerting an excitatory effect. These data suggest that electroacupuncture at the lower limbs and abdominal cavity is an effective means for regulating the activity of glucose-inhibited neurons in the lateral hypothalamic area and for improving the secretory function of adipose tissue.

  13. Glucose transport by epithelia prepared from harvested enterocytes

    DEFF Research Database (Denmark)

    Kimura, Yasuhiro; van der Merwe, Marie; Bering, Stine Brandt;

    2015-01-01

    a simple, novel, and reproducible method for preparing functional epithelia using differentiated enterocytes harvested from the small intestine upper villus of adult mice and preterm pigs with and without necrotizing enterocolitis. Concentrative, rheogenic glucose uptake was used as an indicator...... of epithelial function and was demonstrated by cellular accumulation of tracer (14)C D-glucose and Ussing chamber based short-circuit currents. Assessment of the epithelia by light and immunofluorescent microscopy revealed the harvested enterocytes remain differentiated and establish cell-cell connections......, forskolin, and health status exceeding those we have measured using intact tissues. Our findings indicate that epithelia prepared from harvested enterocytes provide an alternative approach for comparative studies of the characteristics of nutrient transport by the upper villus epithelium and the responses...

  14. Sweet talk: insights into the nature and importance of glucose transport in lung epithelium.

    Science.gov (United States)

    Garnett, James P; Baker, Emma H; Baines, Deborah L

    2012-11-01

    For over 50 years, glucose has been recognised to cross the lung epithelial barrier and be transported by lung epithelial cells. However, until recently, research into these processes focused on their effects on lung liquid volume. Here, we consider a newly identified role for pulmonary glucose transport in maintaining low airway surface liquid (ASL) glucose concentrations and propose that this contributes to lung defence against infection. Glucose diffuses into ASL via paracellular pathways at a rate determined by paracellular permeability and the transepithelial glucose gradient. Glucose is removed from ASL in proximal airways via facilitative glucose transporters, down a concentration gradient generated by intracellular glucose metabolism. In the distal lung, glucose transport via sodium-coupled glucose transporters predominates. These processes vary between species but universally maintain ASL glucose at 3-20-fold lower concentrations than plasma. ASL glucose concentrations are increased in respiratory disease and by hyperglycaemia. Elevated ASL glucose in intensive care patients was associated with increased Staphylococcus aureus infection. Diabetic patients with and without chronic lung disease are at increased risk of respiratory infection. Understanding of mechanisms underlying lung glucose homeostasis could identify new therapeutic targets for control of ASL glucose and prevention and treatment of lung infection.

  15. Bace1 activity impairs neuronal glucose metabolism: rescue by beta-hydroxybutyrate and lipoic acid

    Directory of Open Access Journals (Sweden)

    John A Findlay

    2015-10-01

    Full Text Available Glucose hypometabolism and impaired mitochondrial function in neurons have been suggested to play early and perhaps causative roles in Alzheimer’s disease (AD pathogenesis. Activity of the aspartic acid protease, beta-site amyloid precursor protein (APP cleaving enzyme 1 (BACE1, responsible for beta amyloid peptide generation, has recently been demonstrated to modify glucose metabolism. We therefore examined, using a human neuroblastoma (SH-SY5Y cell line, whether increased BACE1 activity is responsible for a reduction in cellular glucose metabolism. Overexpression of active BACE1, but not a protease-dead mutant BACE1, protein in SH-SY5Y cells reduced glucose oxidation and the basal oxygen consumption rate, which was associated with a compensatory increase in glycolysis. Increased BACE1 activity had no effect on the mitochondrial electron transfer process but was found to diminish substrate delivery to the mitochondria by inhibition of key mitochondrial decarboxylation reaction enzymes. This BACE1 activity-dependent deficit in glucose oxidation was alleviated by the presence of beta hydroxybutyrate or α-lipoic acid. Consequently our data indicate that raised cellular BACE1 activity drives reduced glucose oxidation in a human neuronal cell line through impairments in the activity of specific tricarboxylic acid cycle enzymes. Because this bioenergetic deficit is recoverable by neutraceutical compounds we suggest that such agents, perhaps in conjunction with BACE1 inhibitors, may be an effective therapeutic strategy in the early-stage management or treatment of AD.

  16. BACE1 activity impairs neuronal glucose oxidation: rescue by beta-hydroxybutyrate and lipoic acid.

    Science.gov (United States)

    Findlay, John A; Hamilton, David L; Ashford, Michael L J

    2015-01-01

    Glucose hypometabolism and impaired mitochondrial function in neurons have been suggested to play early and perhaps causative roles in Alzheimer's disease (AD) pathogenesis. Activity of the aspartic acid protease, beta-site amyloid precursor protein (APP) cleaving enzyme 1 (BACE1), responsible for beta amyloid peptide generation, has recently been demonstrated to modify glucose metabolism. We therefore examined, using a human neuroblastoma (SH-SY5Y) cell line, whether increased BACE1 activity is responsible for a reduction in cellular glucose metabolism. Overexpression of active BACE1, but not a protease-dead mutant BACE1, protein in SH-SY5Y cells reduced glucose oxidation and the basal oxygen consumption rate, which was associated with a compensatory increase in glycolysis. Increased BACE1 activity had no effect on the mitochondrial electron transfer process but was found to diminish substrate delivery to the mitochondria by inhibition of key mitochondrial decarboxylation reaction enzymes. This BACE1 activity-dependent deficit in glucose oxidation was alleviated by the presence of beta hydroxybutyrate or α-lipoic acid. Consequently our data indicate that raised cellular BACE1 activity drives reduced glucose oxidation in a human neuronal cell line through impairments in the activity of specific tricarboxylic acid cycle enzymes. Because this bioenergetic deficit is recoverable by neutraceutical compounds we suggest that such agents, perhaps in conjunction with BACE1 inhibitors, may be an effective therapeutic strategy in the early-stage management or treatment of AD.

  17. Ethanolic extract of Allium cepa stimulates glucose transporter typ 4-mediated glucose uptake by the activation of insulin signaling.

    Science.gov (United States)

    Gautam, Sudeep; Pal, Savita; Maurya, Rakesh; Srivastava, Arvind K

    2015-02-01

    The present work was undertaken to investigate the effects and the molecular mechanism of the standardized ethanolic extract of Allium cepa (onion) on the glucose transport for controlling diabetes mellitus. A. cepa stimulates glucose uptake by the rat skeletal muscle cells (L6 myotubes) in both time- and dose-dependent manners. This effect was shown to be mediated by the increased translocation of glucose transporter typ 4 protein from the cytoplasm to the plasma membrane as well as the synthesis of glucose transporter typ 4 protein. The effect of A. cepa extract on glucose transport was stymied by wortmannin, genistein, and AI½. In vitro phosphorylation analysis revealed that, like insulin, A. cepa extract also enhances the tyrosine phosphorylation of the insulin receptor-β, insulin receptor substrate-1, and the serine phosphorylation of Akt under both basal and insulin-stimulated conditions without affecting the total amount of these proteins. Furthermore, it is also shown that the activation of Akt is indispensable for the A. cepa-induced glucose uptake in L6 myotubes. Taken together, these findings provide ample evidence that the ethanolic extract of A. cepa stimulates glucose transporter typ 4 translocation-mediated glucose uptake by the activation of the phosphatidylinositol-4,5-bisphosphate 3-kinase/Akt dependent pathway.

  18. Sodium glucose transporter 2 (SGLT2 inhibition and ketogenesis

    Directory of Open Access Journals (Sweden)

    Sanjay Kalra

    2015-01-01

    Full Text Available Sodium glucose transporter 2 (SGLT2 inhibitors are a recently developed class of drug that have been approved for use in type 2 diabetes. Their unique extra-pancreatic glucuretic mode of action has encouraged their usage in type 1 diabetes as well. At the same time, reports of pseudo ketoacidosis and ketoacidosis related to their use have been published. No clear mechanism for this phenomenon has been demonstrated so far. This communication delves into the biochemical effects of SGLT2 inhibition, discusses the utility of these drugs and proposes steps to maximize safe usage of the molecules.

  19. Screening for Inhibitors of Essential Leishmania Glucose Transporters

    Science.gov (United States)

    2013-07-01

    600 are natural products , and ~400 are other bioactive components. The library was constructed to have a wide range of biological activities and...within each plate. Each well contained 20 µl of ∆lmxgt parasites, complemented with either LmxGT2 or GLUT1, suspended in DME -L medium (7) at an initial...expressing the LmGT2 transporter was grown for 72 hr in DME -L medium containing 5 mM glucose in the presence of compounds (2000) from the MicroSource

  20. Sodium glucose transporter 2 (SGLT2) inhibition and ketogenesis.

    Science.gov (United States)

    Kalra, Sanjay; Sahay, Rakesh; Gupta, Yashdeep

    2015-01-01

    Sodium glucose transporter 2 (SGLT2) inhibitors are a recently developed class of drug that have been approved for use in type 2 diabetes. Their unique extra-pancreatic glucuretic mode of action has encouraged their usage in type 1 diabetes as well. At the same time, reports of pseudo ketoacidosis and ketoacidosis related to their use have been published. No clear mechanism for this phenomenon has been demonstrated so far. This communication delves into the biochemical effects of SGLT2 inhibition, discusses the utility of these drugs and proposes steps to maximize safe usage of the molecules.

  1. Steviol Glycosides Modulate Glucose Transport in Different Cell Types

    Science.gov (United States)

    Rizzo, Benedetta; Zambonin, Laura; Leoncini, Emanuela; Vieceli Dalla Sega, Francesco; Prata, Cecilia; Fiorentini, Diana; Hrelia, Silvana

    2013-01-01

    Extracts from Stevia rebaudiana Bertoni, a plant native to Central and South America, have been used as a sweetener since ancient times. Currently, Stevia extracts are largely used as a noncaloric high-potency biosweetener alternative to sugar, due to the growing incidence of type 2 diabetes mellitus, obesity, and metabolic disorders worldwide. Despite the large number of studies on Stevia and steviol glycosides in vivo, little is reported concerning the cellular and molecular mechanisms underpinning the beneficial effects on human health. The effect of four commercial Stevia extracts on glucose transport activity was evaluated in HL-60 human leukaemia and in SH-SY5Y human neuroblastoma cells. The extracts were able to enhance glucose uptake in both cellular lines, as efficiently as insulin. Our data suggest that steviol glycosides could act by modulating GLUT translocation through the PI3K/Akt pathway since treatments with both insulin and Stevia extracts increased the phosphorylation of PI3K and Akt. Furthermore, Stevia extracts were able to revert the effect of the reduction of glucose uptake caused by methylglyoxal, an inhibitor of the insulin receptor/PI3K/Akt pathway. These results corroborate the hypothesis that Stevia extracts could mimic insulin effects modulating PI3K/Akt pathway. PMID:24327825

  2. Steviol Glycosides Modulate Glucose Transport in Different Cell Types

    Directory of Open Access Journals (Sweden)

    Benedetta Rizzo

    2013-01-01

    Full Text Available Extracts from Stevia rebaudiana Bertoni, a plant native to Central and South America, have been used as a sweetener since ancient times. Currently, Stevia extracts are largely used as a noncaloric high-potency biosweetener alternative to sugar, due to the growing incidence of type 2 diabetes mellitus, obesity, and metabolic disorders worldwide. Despite the large number of studies on Stevia and steviol glycosides in vivo, little is reported concerning the cellular and molecular mechanisms underpinning the beneficial effects on human health. The effect of four commercial Stevia extracts on glucose transport activity was evaluated in HL-60 human leukaemia and in SH-SY5Y human neuroblastoma cells. The extracts were able to enhance glucose uptake in both cellular lines, as efficiently as insulin. Our data suggest that steviol glycosides could act by modulating GLUT translocation through the PI3K/Akt pathway since treatments with both insulin and Stevia extracts increased the phosphorylation of PI3K and Akt. Furthermore, Stevia extracts were able to revert the effect of the reduction of glucose uptake caused by methylglyoxal, an inhibitor of the insulin receptor/PI3K/Akt pathway. These results corroborate the hypothesis that Stevia extracts could mimic insulin effects modulating PI3K/Akt pathway.

  3. Steviol glycosides modulate glucose transport in different cell types.

    Science.gov (United States)

    Rizzo, Benedetta; Zambonin, Laura; Angeloni, Cristina; Leoncini, Emanuela; Dalla Sega, Francesco Vieceli; Prata, Cecilia; Fiorentini, Diana; Hrelia, Silvana

    2013-01-01

    Extracts from Stevia rebaudiana Bertoni, a plant native to Central and South America, have been used as a sweetener since ancient times. Currently, Stevia extracts are largely used as a noncaloric high-potency biosweetener alternative to sugar, due to the growing incidence of type 2 diabetes mellitus, obesity, and metabolic disorders worldwide. Despite the large number of studies on Stevia and steviol glycosides in vivo, little is reported concerning the cellular and molecular mechanisms underpinning the beneficial effects on human health. The effect of four commercial Stevia extracts on glucose transport activity was evaluated in HL-60 human leukaemia and in SH-SY5Y human neuroblastoma cells. The extracts were able to enhance glucose uptake in both cellular lines, as efficiently as insulin. Our data suggest that steviol glycosides could act by modulating GLUT translocation through the PI3K/Akt pathway since treatments with both insulin and Stevia extracts increased the phosphorylation of PI3K and Akt. Furthermore, Stevia extracts were able to revert the effect of the reduction of glucose uptake caused by methylglyoxal, an inhibitor of the insulin receptor/PI3K/Akt pathway. These results corroborate the hypothesis that Stevia extracts could mimic insulin effects modulating PI3K/Akt pathway.

  4. Glial glutamate transporters mediate a functional metabolic crosstalk between neurons and astrocytes in the mouse developing cortex.

    Science.gov (United States)

    Voutsinos-Porche, Brigitte; Bonvento, Gilles; Tanaka, Kohichi; Steiner, Pascal; Welker, Egbert; Chatton, Jean-Yves; Magistretti, Pierre J; Pellerin, Luc

    2003-01-23

    Neuron-glia interactions are essential for synaptic function, and glial glutamate (re)uptake plays a key role at glutamatergic synapses. In knockout mice, for either glial glutamate transporters, GLAST or GLT-1, a classical metabolic response to synaptic activation (i.e., enhancement of glucose utilization) is decreased at an early functional stage in the somatosensory barrel cortex following activation of whiskers. Investigation in vitro demonstrates that glial glutamate transport represents a critical step for triggering enhanced glucose utilization, but also lactate release from astrocytes through a mechanism involving changes in intracellular Na(+) concentration. These data suggest that a metabolic crosstalk takes place between neurons and astrocytes in the developing cortex, which would be regulated by synaptic activity and mediated by glial glutamate transporters.

  5. Domain assembly of the GLUT1 glucose transporter.

    Science.gov (United States)

    Cope, D L; Holman, G D; Baldwin, S A; Wolstenholme, A J

    1994-01-01

    A full-length construct of the glucose transporter isoform GLUT1 has been expressed in Sf9 (Spodoptera frugiperida Clone 9) insect cells, and a photolabelling approach has been used to show that the expressed protein binds the bismannose compound 2-N-4-(1-azi-2,2,2-trifluoroethyl)benzoyl-1,3-bis-(D-mannos- 4-yloxy)-2-propylamine (ATB-BMPA) and cytochalasin B at its exofacial and endofacial binding sites respectively. Constructs of GLUT1 which produce either the N-terminal (amino acids 1-272) or C-terminal (amino acids 254-492) halves are expressed at levels in the plasma membrane which are similar to that of the full-length GLUT1 (approximately 200 pmol/mg of membrane protein), but do not bind either ATB-BMPA or cytochalasin B. When Sf9 cells are doubly infected with virus constructs producing both the C- and N-terminal halves of GLUT1, then the ligand labelling is restored. Only the C-terminal half is labelled, and, therefore, the labelling of this domain is dependent on the presence of the N-terminal half of the protein. These results suggest that the two halves of GLUT1 can assemble to form a stable complex and support the concept of a bilobular structure for the intact glucose transporters in which separate C- and N-domain halves pack together to produce a ligand-binding conformation. Images Figure 1 PMID:8002929

  6. Diabetic Hyperglycemia: Link to Impaired Glucose Transport in Pancreatic β Cells

    Science.gov (United States)

    Unger, Roger H.

    1991-03-01

    Glucose uptake into pancreatic β cells by means of the glucose transporter GLUT-2, which has a high Michaelis constant, is essential for the normal insulin secretory response to hyperglycemia. In both autoimmune and nonautoimmune diabetes, this glucose transport is reduced as a consequence of down-regulation of the normal β-cell transporter. In autoimmune diabetes, circulating immunoglobulins can further impair this glucose transport by inhibiting functionally intact transporters. Insights into mechanisms of the unresponsiveness of β cells to hyperglycemia may improve the management and prevention of diabetes.

  7. Glucose transporters and maximal transport are increased in endurance-trained rat soleus

    Science.gov (United States)

    Slentz, C. A.; Gulve, E. A.; Rodnick, K. J.; Henriksen, E. J.; Youn, J. H.; Holloszy, J. O.

    1992-01-01

    Voluntary wheel running induces an increase in the concentration of the regulatable glucose transporter (GLUT4) in rat plantaris muscle but not in soleus muscle (K. J. Rodnick, J. O. Holloszy, C. E. Mondon, and D. E. James. Diabetes 39: 1425-1429, 1990). Wheel running also causes hypertrophy of the soleus in rats. This study was undertaken to ascertain whether endurance training that induces enzymatic adaptations but no hypertrophy results in an increase in the concentration of GLUT4 protein in rat soleus (slow-twitch red) muscle and, if it does, to determine whether there is a concomitant increase in maximal glucose transport activity. Female rats were trained by treadmill running at 25 m/min up a 15% grade, 90 min/day, 6 days/wk for 3 wk. This training program induced increases of 52% in citrate synthase activity, 66% in hexokinase activity, and 47% in immunoreactive GLUT4 protein concentration in soleus muscles without causing hypertrophy. Glucose transport activity stimulated maximally with insulin plus contractile activity was increased to roughly the same extent (44%) as GLUT4 protein content in soleus muscle by the treadmill exercise training. In a second set of experiments, we examined whether a swim-training program increases glucose transport activity in the soleus in the presence of a maximally effective concentration of insulin. The swimming program induced a 44% increase in immunoreactive GLUT4 protein concentration. Glucose transport activity maximally stimulated with insulin was 62% greater in soleus muscle of the swimmers than in untrained controls. Training did not alter the basal rate of 2-deoxyglucose uptake.(ABSTRACT TRUNCATED AT 250 WORDS).

  8. Automated measurement of fast mitochondrial transport in neurons.

    Science.gov (United States)

    Miller, Kyle E; Liu, Xin-An; Puthanveettil, Sathyanarayanan V

    2015-01-01

    There is growing recognition that fast mitochondrial transport in neurons is disrupted in multiple neurological diseases and psychiatric disorders. However, a major constraint in identifying novel therapeutics based on mitochondrial transport is that the large-scale analysis of fast transport is time consuming. Here we describe methodologies for the automated analysis of fast mitochondrial transport from data acquired using a robotic microscope. We focused on addressing questions of measurement precision, speed, reliably, workflow ease, statistical processing, and presentation. We used optical flow and particle tracking algorithms, implemented in ImageJ, to measure mitochondrial movement in primary cultured cortical and hippocampal neurons. With it, we are able to generate complete descriptions of movement profiles in an automated fashion of hundreds of thousands of mitochondria with a processing time of approximately one hour. We describe the calibration of the parameters of the tracking algorithms and demonstrate that they are capable of measuring the fast transport of a single mitochondrion. We then show that the methods are capable of reliably measuring the inhibition of fast mitochondria transport induced by the disruption of microtubules with the drug nocodazole in both hippocampal and cortical neurons. This work lays the foundation for future large-scale screens designed to identify compounds that modulate mitochondrial motility.

  9. Automated Measurement of Fast Mitochondrial Transport in Neurons

    Directory of Open Access Journals (Sweden)

    Kyle eMiller

    2015-11-01

    Full Text Available There is a growing recognition that fast mitochondrial transport in neurons is disrupted in multiple neurological diseases and psychiatric disorders. However a major constraint in identifying novel therapeutics based on mitochondrial transport is that the large-scale analysis of fast transport is time consuming. Here we describe methodologies for the automated analysis of fast mitochondrial transport from data acquired using a robotic microscope. We focused on addressing questions of measurement precision, speed, reliably, workflow ease, statistical processing and presentation. We used optical flow and particle tracking algorithms, implemented in ImageJ, to measure mitochondrial movement in primary cultured cortical and hippocampal neurons. With it, we are able to generate complete descriptions of movement profiles in an automated fashion of hundred of thousands of mitochondria with a processing time of approximately one hour. We describe the calibration of the parameters of the tracking algorithms and demonstrate that they are capable of measuring the fast transport of a single mitochondrion. We then show that the methods are capable of reliably measuring the inhibition of fast mitochondria transport induced by the disruption of microtubules with the drug nocodazole in both hippocampal and cortical neurons. This work lays the foundation for future large-scale screens designed to identify compounds that modulate mitochondrial motility.

  10. The neuroprotective role and mechanisms of TERT in neurons with oxygen-glucose deprivation.

    Science.gov (United States)

    Li, J; Qu, Y; Chen, D; Zhang, L; Zhao, F; Luo, L; Pan, L; Hua, J; Mu, D

    2013-11-12

    Telomerase reverse transcriptase (TERT) is reported to protect neurons from apoptosis induced by various stresses including hypoxia-ischemia (HI). However, the mechanisms by which TERT exerts its anti-apoptotic role in neurons with HI injury remain unclear. In this study, we examined the protective role and explored the possible mechanisms of TERT in neurons with HI injury in vitro. Primary cultured neurons were exposed to oxygen and glucose deprivation (OGD) for 3h followed by reperfusion to mimic HI injury in vivo. Plasmids containing TERT antisense, sense nucleotides, or mock were transduced into neurons at 48h before OGD. Expression and distribution of TERT were measured by immunofluorescence labeling and western blot. The expression of cleaved caspase 3 (CC3), Bcl-2 and Bax were detected by western blot. Neuronal apoptosis was measured with terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling (TUNEL). The mitochondrial reactive oxygen species (ROS) were measured by MitoSOX Red staining. Fluorescent probe JC-1 was used to measure the mitochondrial membrane potential (ΔΨm). We found that TERT expression increased at 8h and peaked at 24h in neurons after OGD. CC3 expression and neuronal apoptosis were induced and peaked at 24h after OGD. TERT inhibition significantly increased CC3 expression and neuronal apoptosis after OGD treatment. Additionally, TERT inhibition decreased the expression ratio of Bcl-2/Bax, and enhanced ROS production and ΔΨm dissipation after OGD. These data suggest that TERT plays a neuroprotective role via anti-apoptosis in neurons after OGD. The underlying mechanisms may be associated with regulating Bcl-2/Bax expression ratio, attenuating ROS generation, and increasing mitochondrial membrane potential.

  11. The glucose metabolite methylglyoxal inhibits expression of the glucose transporter genes by inactivating the cell surface glucose sensors Rgt2 and Snf3 in yeast.

    Science.gov (United States)

    Roy, Adhiraj; Hashmi, Salman; Li, Zerui; Dement, Angela D; Cho, Kyu Hong; Kim, Jeong-Ho

    2016-03-01

    Methylglyoxal (MG) is a cytotoxic by-product of glycolysis. MG has inhibitory effect on the growth of cells ranging from microorganisms to higher eukaryotes, but its molecular targets are largely unknown. The yeast cell-surface glucose sensors Rgt2 and Snf3 function as glucose receptors that sense extracellular glucose and generate a signal for induction of expression of genes encoding glucose transporters (HXTs). Here we provide evidence that these glucose sensors are primary targets of MG in yeast. MG inhibits the growth of glucose-fermenting yeast cells by inducing endocytosis and degradation of the glucose sensors. However, the glucose sensors with mutations at their putative ubiquitin-acceptor lysine residues are resistant to MG-induced degradation. These results suggest that the glucose sensors are inactivated through ubiquitin-mediated endocytosis and degraded in the presence of MG. In addition, the inhibitory effect of MG on the glucose sensors is greatly enhanced in cells lacking Glo1, a key component of the MG detoxification system. Thus the stability of these glucose sensors seems to be critically regulated by intracellular MG levels. Taken together, these findings suggest that MG attenuates glycolysis by promoting degradation of the cell-surface glucose sensors and thus identify MG as a potential glycolytic inhibitor.

  12. Lipopolysaccharide (LPS) and tumor necrosis factor alpha (TNFα) blunt the response of Neuropeptide Y/Agouti-related peptide (NPY/AgRP) glucose inhibited (GI) neurons to decreased glucose.

    Science.gov (United States)

    Hao, Lihong; Sheng, Zhenyu; Potian, Joseph; Deak, Adam; Rohowsky-Kochan, Christine; Routh, Vanessa H

    2016-10-01

    A population of Neuropeptide Y (NPY) neurons which co-express Agouti-related peptide (AgRP) in the arcuate nucleus of the hypothalamus (ARC) are inhibited at physiological levels of brain glucose and activated when glucose levels decline (e.g. glucose-inhibited or GI neurons). Fasting enhances the activation of NPY/AgRP-GI neurons by low glucose. In the present study we tested the hypothesis that lipopolysaccharide (LPS) inhibits the enhanced activation of NPY/AgRP-GI neurons by low glucose following a fast. Mice which express green fluorescent protein (GFP) on their NPY promoter were used to identify NPY/AgRP neurons. Fasting for 24h and LPS injection decreased blood glucose levels. As we have found previously, fasting increased c-fos expression in NPY/AgRP neurons and increased the activation of NPY/AgRP-GI neurons by decreased glucose. As we predicted, LPS blunted these effects of fasting at the 24h time point. Moreover, the inflammatory cytokine tumor necrosis factor alpha (TNFα) blocked the activation of NPY/AgRP-GI neurons by decreased glucose. These data suggest that LPS and TNFα may alter glucose and energy homeostasis, in part, due to changes in the glucose sensitivity of NPY/AgRP neurons. Interestingly, our findings also suggest that NPY/AgRP-GI neurons use a distinct mechanism to sense changes in extracellular glucose as compared to our previous studies of GI neurons in the adjacent ventromedial hypothalamic nucleus.

  13. Novel Roles for the Insulin-Regulated Glucose Transporter-4 in Hippocampally Dependent Memory.

    Science.gov (United States)

    Pearson-Leary, Jiah; McNay, Ewan C

    2016-11-23

    The insulin-regulated glucose transporter-4 (GluT4) is critical for insulin- and contractile-mediated glucose uptake in skeletal muscle. GluT4 is also expressed in some hippocampal neurons, but its functional role in the brain is unclear. Several established molecular modulators of memory processing regulate hippocampal GluT4 trafficking and hippocampal memory formation is limited by both glucose metabolism and insulin signaling. Therefore, we hypothesized that hippocampal GluT4 might be involved in memory processes. Here, we show that, in male rats, hippocampal GluT4 translocates to the plasma membrane after memory training and that acute, selective intrahippocampal inhibition of GluT4-mediated glucose transport impaired memory acquisition, but not memory retrieval. Other studies have shown that prolonged systemic GluT4 blockade causes insulin resistance. Unexpectedly, we found that prolonged hippocampal blockade of glucose transport through GluT4-upregulated markers of hippocampal insulin signaling prevented task-associated depletion of hippocampal glucose and enhanced both working and short-term memory while also impairing long-term memory. These effects were accompanied by increased expression of hippocampal AMPA GluR1 subunits and the neuronal GluT3, but decreased expression of hippocampal brain-derived neurotrophic factor, consistent with impaired ability to form long-term memories. Our findings are the first to show the cognitive impact of brain GluT4 modulation. They identify GluT4 as a key regulator of hippocampal memory processing and also suggest differential regulation of GluT4 in the hippocampus from that in peripheral tissues. The role of insulin-regulated glucose transporter-4 (GluT4) in the brain is unclear. In the current study, we demonstrate that GluT4 is a critical component of hippocampal memory processes. Memory training increased hippocampal GluT4 translocation and memory acquisition was impaired by GluT4 blockade. Unexpectedly, whereas long

  14. Glucose uptake during contraction in isolated skeletal muscles from neuronal nitric oxide synthase μ knockout mice.

    Science.gov (United States)

    Hong, Yet Hoi; Frugier, Tony; Zhang, Xinmei; Murphy, Robyn M; Lynch, Gordon S; Betik, Andrew C; Rattigan, Stephen; McConell, Glenn K

    2015-05-01

    Inhibition of nitric oxide synthase (NOS) significantly attenuates the increase in skeletal muscle glucose uptake during contraction/exercise, and a greater attenuation is observed in individuals with Type 2 diabetes compared with healthy individuals. Therefore, NO appears to play an important role in mediating muscle glucose uptake during contraction. In this study, we investigated the involvement of neuronal NOSμ (nNOSμ), the main NOS isoform activated during contraction, on skeletal muscle glucose uptake during ex vivo contraction. Extensor digitorum longus muscles were isolated from nNOSμ(-/-) and nNOSμ(+/+) mice. Muscles were contracted ex vivo in a temperature-controlled (30°C) organ bath with or without the presence of the NOS inhibitor N(G)-monomethyl-l-arginine (L-NMMA) and the NOS substrate L-arginine. Glucose uptake was determined by radioactive tracers. Skeletal muscle glucose uptake increased approximately fourfold during contraction in muscles from both nNOSμ(-/-) and nNOSμ(+/+) mice. L-NMMA significantly attenuated the increase in muscle glucose uptake during contraction in both genotypes. This attenuation was reversed by L-arginine, suggesting that L-NMMA attenuated the increase in muscle glucose uptake during contraction by inhibiting NOS and not via a nonspecific effect of the inhibitor. Low levels of NOS activity (~4%) were detected in muscles from nNOSμ(-/-) mice, and there was no evidence of compensation from other NOS isoform or AMP-activated protein kinase which is also involved in mediating muscle glucose uptake during contraction. These results indicate that NO regulates skeletal muscle glucose uptake during ex vivo contraction independently of nNOSμ.

  15. Effect of physical training on glucose transporter protein and mRNA levels in rat adipocytes

    DEFF Research Database (Denmark)

    Stallknecht, B; Andersen, P H; Vinten, J

    1993-01-01

    Physical training increases insulin-stimulated glucose transport and the number of glucose transporters in adipocytes measured by cytochalasin B binding. In the present study we used immunoblotting to measure the abundance of two glucose transporters (GLUT-4, GLUT-1) in white adipocytes from....../or intrinsic activity). GLUT-1 protein and mRNA levels/adipocyte volume did not change with age or training....

  16. Activin A prevents neuron-like PC12 cell apoptosis after oxygen-glucose deprivation

    Institute of Scientific and Technical Information of China (English)

    Guihua Xu; Zhongxin Xu; Jinting He; Hongliang Guo; Chunli Mei; Jiaoqi Wang; Zhongshu Li; Han Chen; Jing Mang; Hong Yang

    2013-01-01

    In this study, PC12 cells were induced to differentiate into neuron-like cells using nerve growth factor, and were subjected to oxygen-glucose deprivation. Cells were treated with 0, 10, 20, 30, 50, 100 ng/mL exogenous Activin A. The 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl tetrazolium bromide assay and Hoechst 33324 staining showed that the survival percentage of PC12 cells significantly decreased and the rate of apoptosis significantly increased after oxygen-glucose deprivation. Exogenous Activin A significantly increased the survival percentage of PC12 cells in a dose-dependent manner. Reverse transcription-PCR results revealed a significant increase in Activin receptor IIA, Smad3 and Smad4 mRNA levels, which are key sites in the Activin A/Smads signaling pathway, in neuron-like cells subjected to oxygen-glucose deprivation, while mRNA expression of the apoptosis-regulation gene caspase-3 decreased. Our experimental findings indicate that exogenous Activin A plays an anti-apoptotic role and protects neurons by means of activating the Activin A/Smads signaling pathway.

  17. Glucose transporter expression in human skeletal muscle fibers

    DEFF Research Database (Denmark)

    Gaster, M; Handberg, A; Beck-Nielsen, H

    2000-01-01

    amplification (TSA) technique to detect the localization of glucose transporter expression in human skeletal muscle. We found expression of GLUT-1, GLUT-3, and GLUT-4 in developing human muscle fibers showing a distinct expression pattern. 1) GLUT-1 is expressed in human skeletal muscle cells during gestation......, but its expression is markedly reduced around birth and is further reduced to undetectable levels within the first year of life; 2) GLUT-3 protein expression appears at 18 wk of gestation and disappears after birth; and 3) GLUT-4 protein is diffusely expressed in muscle cells throughout gestation, whereas...... after birth, the characteristic subcellular localization is as seen in adult muscle fibers. Our results show that GLUT-1, GLUT-3, and GLUT-4 seem to be of importance during muscle fiber growth and development. GLUT-5 protein was undetectable in fetal and adult skeletal muscle fibers. In adult muscle...

  18. Expression, transport, and axonal sorting of neuronal CCL21 in large dense-core vesicles

    NARCIS (Netherlands)

    de Jong, Eiko K.; Vinet, Jonathan; Stanulovic, Vesna S.; Meijer, Michel; Wesseling, Evelyn; Sjollema, Klaas; Boddeke, Hendrikus W. G. M.; Biber, Knut

    2008-01-01

    Neurons are highly polarized cells, and neuron-neuron communication is based on directed transport and release of neurotransmitters, neuropeptides, and neurotrophins. Directed communication may also be attributed to neuron-microglia signaling, since neuronal damage can induce a microglia reaction at

  19. Expression, transport, and axonal sorting of neuronal CCL21 in large dense-core vesicles.

    NARCIS (Netherlands)

    Jong, E.K. de; Vinet, J.; Stanulovic, V.S.; Meijer, Michel; Wesseling, E.; Sjollema, K.; Boddeke, H.W.; Biber, K.

    2008-01-01

    Neurons are highly polarized cells, and neuron-neuron communication is based on directed transport and release of neurotransmitters, neuropeptides, and neurotrophins. Directed communication may also be attributed to neuron-microglia signaling, since neuronal damage can induce a microglia reaction at

  20. A Simple Flow Cytometric Method to Measure Glucose Uptake and Glucose Transporter Expression for Monocyte Subpopulations in Whole Blood.

    Science.gov (United States)

    Palmer, Clovis S; Anzinger, Joshua J; Butterfield, Tiffany R; McCune, Joseph M; Crowe, Suzanne M

    2016-08-12

    Monocytes are innate immune cells that can be activated by pathogens and inflammation associated with certain chronic inflammatory diseases. Activation of monocytes induces effector functions and a concomitant shift from oxidative to glycolytic metabolism that is accompanied by increased glucose transporter expression. This increased glycolytic metabolism is also observed for trained immunity of monocytes, a form of innate immunological memory. Although in vitro protocols examining glucose transporter expression and glucose uptake by monocytes have been described, none have been examined by multi-parametric flow cytometry in whole blood. We describe a multi-parametric flow cytometric protocol for the measurement of fluorescent glucose analog 2-NBDG uptake in whole blood by total monocytes and the classical (CD14(++)CD16(-)), intermediate (CD14(++)CD16(+)) and non-classical (CD14(+)CD16(++)) monocyte subpopulations. This method can be used to examine glucose transporter expression and glucose uptake for total monocytes and monocyte subpopulations during homeostasis and inflammatory disease, and can be easily modified to examine glucose uptake for other leukocytes and leukocyte subpopulations within blood.

  1. Influence of preovulatory estradiol on conceptus survival and uterine glucose transporter expression

    Science.gov (United States)

    Glucose is an essential component of uterine secretions, and is delivered into the uterine lumen by glucose transporters. We have previously reported increased concentrations of glucose in uterine flushes of cows that exhibited estrus. Our objective in the present study was to determine the effects...

  2. Cofilin Inhibition Restores Neuronal Cell Death in Oxygen-Glucose Deprivation Model of Ischemia.

    Science.gov (United States)

    Madineni, Anusha; Alhadidi, Qasim; Shah, Zahoor A

    2016-03-01

    Ischemia is a condition associated with decreased blood supply to the brain, eventually leading to death of neurons. It is associated with a diverse cascade of responses involving both degenerative and regenerative mechanisms. At the cellular level, the changes are initiated prominently in the neuronal cytoskeleton. Cofilin, a cytoskeletal actin severing protein, is known to be involved in the early stages of apoptotic cell death. Evidence supports its intervention in the progression of disease states like Alzheimer's and ischemic kidney disease. In the present study, we have hypothesized the possible involvement of cofilin in ischemia. Using PC12 cells and mouse primary cultures of cortical neurons, we investigated the potential role of cofilin in ischemia in two different in vitro ischemic models: chemical induced oxidative stress and oxygen-glucose deprivation/reperfusion (OGD/R). The expression profile studies demonstrated a decrease in phosphocofilin levels in all models of ischemia, implying stress-induced cofilin activation. Furthermore, calcineurin and slingshot 1L (SSH) phosphatases were found to be the signaling mediators of the cofilin activation. In primary cultures of cortical neurons, cofilin was found to be significantly activated after 1 h of OGD. To delineate the role of activated cofilin in ischemia, we knocked down cofilin by small interfering RNA (siRNA) technique and tested the impact of cofilin silencing on neuronal viability. Cofilin siRNA-treated neurons showed a significant reduction of cofilin levels in all treatment groups (control, OGD, and OGD/R). Additionally, cofilin siRNA-reduced cofilin mitochondrial translocation and caspase 3 cleavage, with a concomitant increase in neuronal viability. These results strongly support the active role of cofilin in ischemia-induced neuronal degeneration and apoptosis. We believe that targeting this protein mediator has a potential for therapeutic intervention in ischemic brain injury and stroke.

  3. The role of SGLT1 and GLUT2 in intestinal glucose transport and sensing.

    Directory of Open Access Journals (Sweden)

    Pia V Röder

    Full Text Available Intestinal glucose absorption is mediated by SGLT1 whereas GLUT2 is considered to provide basolateral exit. Recently, it was proposed that GLUT2 can be recruited into the apical membrane after a high luminal glucose bolus allowing bulk absorption of glucose by facilitated diffusion. Moreover, SGLT1 and GLUT2 are suggested to play an important role in intestinal glucose sensing and incretin secretion. In mice that lack either SGLT1 or GLUT2 we re-assessed the role of these transporters in intestinal glucose uptake after radiotracer glucose gavage and performed Western blot analysis for transporter abundance in apical membrane fractions in a comparative approach. Moreover, we examined the contribution of these transporters to glucose-induced changes in plasma GIP, GLP-1 and insulin levels. In mice lacking SGLT1, tissue retention of tracer glucose was drastically reduced throughout the entire small intestine whereas GLUT2-deficient animals exhibited higher tracer contents in tissue samples than wild type animals. Deletion of SGLT1 resulted also in reduced blood glucose elevations and abolished GIP and GLP-1 secretion in response to glucose. In mice lacking GLUT2, glucose-induced insulin but not incretin secretion was impaired. Western blot analysis revealed unchanged protein levels of SGLT1 after glucose gavage. GLUT2 detected in apical membrane fractions mainly resulted from contamination with basolateral membranes but did not change in density after glucose administration. SGLT1 is unequivocally the prime intestinal glucose transporter even at high luminal glucose concentrations. Moreover, SGLT1 mediates glucose-induced incretin secretion. Our studies do not provide evidence for GLUT2 playing any role in either apical glucose influx or incretin secretion.

  4. Glutamine as an energy substrate in cultured neurons during glucose deprivation.

    Science.gov (United States)

    Peng, Liang; Gu, Li; Zhang, Hongliang; Huang, Xueshi; Hertz, Elna; Hertz, Leif

    2007-11-15

    During glucose deprivation an increase in aspartate formation from glutamine has been observed in different brain preparations, including synaptosomes and cultured astrocytes. To what extent this reaction, which provides a substantial amount of energy, occurs in different types of neurons is unknown. The present study shows that (14)CO(2) formation from [U-(14)C]glutamine in cerebellar granule neurons, a glutamatergic preparation, increased by 60% during glucose deprivation, indicating enhanced aspartate formation or increased complete oxidative degradation of glutamine. In primary cultures of cerebrocortical interneurons, a GABAergic preparation, the rate of (14)CO(2) production from [U-(14) C] glutamine was four times lower and not stimulated by glucose deprivation. During incubation with glutamine (0.8 mM) as the only metabolic substrate, cerebellar granule cells maintained an oxygen consumption rate of 12 nmol/min/mg protein, corresponding to an aspartate formation of 8 nmol/min/mg protein (three oxidations occur between glutamine and aspartate) or to a total oxidative degradation of 3 nmol/min/mg protein. During glucose deprivation, the rate of aspartate formation increased, and during a 20-min incubation in phosphate-buffered saline it amounted to 3.3 nmol/min/mg protein at 0.2 mM glutamine, which might have been more if measured at 0.8 mM glutamine. These values are consistent with the rate of glutamine utilization calculated based on oxygen consumption and leaves open the possibility that some glutamine is completely degraded oxidatively, as has been shown by other authors based on pyruvate recycling and labeling of lactate from aspartate in cerebellar granule neurons.

  5. Trans-anethole protects cortical neuronal cells against oxygen-glucose deprivation/reoxygenation.

    Science.gov (United States)

    Ryu, Sangwoo; Seol, Geun Hee; Park, Hyeon; Choi, In-Young

    2014-10-01

    Trans-anethole has been studied on pharmacological properties such as anti-inflammation, anti-oxidative stress, antifungal and anticancer. However, to date, the anti-ischemic effects of trans-anethole have not been assessed. Therefore, we investigated the neuroprotection of trans-anethole against oxygen-glucose deprivation/reoxygenation (OGD/R)-induced cortical neuronal cell injury, an in vitro model of ischemia. The abilities of trans-anethole to block excitotoxicity, oxidative stress and mitochondrial dysfunction were evaluated in OGD/R-induced neurons. Trans-anethole significantly ameliorated OGD/R-induced neuronal cell injury by attenuating the intracellular calcium overload via the activation of NMDA receptors. Trans-anethole also inhibited OGD/R-induced reactive oxygen species overproduction, which may be derived from the scavenging activity in peroxyl radicals, assessed in an oxygen radical absorbance capacity assay. Furthermore, trans-anethole was shown to attenuate the depolarization of mitochondrial transmembrane. These results indicated that the neuroprotective effect of trans-anethole on OGD/R-induced neuronal injury might be due to its ability to inhibit excitotoxicity, oxidative stress and mitochondrial dysfunction. Considering these multiple pathways causing ischemic neuronal damage, the multi-functional effect of trans-anethole suggested that it may be effective in treating ischemic stroke.

  6. Transport of BMAA into Neurons and Astrocytes by System xc.

    Science.gov (United States)

    Albano, Rebecca; Lobner, Doug

    2017-05-03

    The study of the mechanism of β-N-methylamino-L-alanine (BMAA) neurotoxicity originally focused on its effects at the N-methyl-D-aspartate (NMDA) receptor. In recent years, it has become clear that its mechanism of action is more complicated. First, there are certain cell types, such as motor neurons and cholinergic neurons, where the dominate mechanism of toxicity is through action at AMPA receptors. Second, even in cortical neurons where the primary mechanism of toxicity appears to be activation of NMDA receptors, there are other mechanisms involved. We found that along with NMDA receptors, activation of mGLuR5 receptors and effects on the cystine/glutamate antiporter (system xc-) were involved in the toxicity. The effects on system xc- are of particular interest. System xc- mediates the transport of cystine into the cell in exchange for releasing glutamate into the extracellular fluid. By releasing glutamate, system xc- can potentially cause excitotoxicity. However, through providing cystine to the cell, it regulates the levels of cellular glutathione (GSH), the main endogenous intracellular antioxidant, and in this way may protect cells against oxidative stress. We have previously published that BMAA inhibits cystine uptake leading to GSH depletion and had indirect evidence that BMAA is transported into the cells by system xc-. We now present direct evidence that BMAA is transported into both astrocytes and neurons through system xc-. The fact that BMAA is transported by system xc- also provides a mechanism for BMAA to enter brain cells potentially leading to misincorporation into proteins and protein misfolding.

  7. Effects of noradrenaline on the cell-surface glucose transporters in cultured brown adipocytes: novel mechanism for selective activation of GLUT1 glucose transporters.

    Science.gov (United States)

    Shimizu, Y; Satoh, S; Yano, H; Minokoshi, Y; Cushman, S W; Shimazu, T

    1998-01-01

    Glucose transport into rat brown adipocytes has been shown to be stimulated directly by the sympathetic neurotransmitter, noradrenaline, without a significant increase in the protein content of either GLUT1 or GLUT4 glucose transporter in the plasma membrane [Shimizu, Kielar, Minokoshi and Shimazu (1996) Biochem. J. 314, 485-490]. In the present study, we labelled the exofacial glucose-binding sites of GLUT1 and GLUT4 with a membrane-impermeant photoaffinity reagent, 2-N-[4-(1-azitrifluoroethyl)benzoyl]-[2-3H]1,3-bis- (D-mannos-4-yloxy)-2-propylamine (ATB-[3H]BMPA), to determine which isoform is responsible for the noradrenaline-induced increase in glucose transport into intact brown adipocytes in culture. Insulin stimulated the rate of hexose transport by increasing ATB-[3H]BMPA-labelled cell-surface GLUT4. In contrast, the noradrenaline-induced increase in glucose transport was not accompanied by an increased ATB-[3H]BMPA labelling of GLUT4, nor with an increased amount of GLUT4 in the plasma membrane fraction as assessed by Western blotting, indicating that noradrenaline does not promote the translocation of GLUT4. However, noradrenaline induced an increase in photoaffinity labelling of cell-surface GLUT1 without an apparent increase in the immunoreactive GLUT1 protein in the plasma membrane. This is suggestive of an increased affinity of GLUT1 for the ligand. In fact, the Ki value of non-radioactive ATB-BMPA for 2-deoxy-D-glucose uptake was significantly decreased after treatment of the cells with noradrenaline. The increased photoaffinity labelling of GLUT1 and increased glucose transport caused by noradrenaline were inhibited by a cAMP antagonist, cAMP-S Rp-isomer. These results demonstrate that noradrenaline stimulates glucose transport in brown adipocytes by enhancing the functional activity of GLUT1 through a cAMP-dependent mechanism. PMID:9461536

  8. Central serotonergic neurons activate and recruit thermogenic brown and beige fat and regulate glucose and lipid homeostasis

    DEFF Research Database (Denmark)

    McGlashon, Jacob M; Gorecki, Michelle C; Kozlowski, Amanda E;

    2015-01-01

    diphtheria toxin receptor (DTR) was selectively expressed in central 5-HT neurons. Treatment with diphtheria toxin (DT) eliminated 5-HT neurons and caused loss of thermoregulation, brown adipose tissue (BAT) steatosis, and a >50% decrease in uncoupling protein 1 (Ucp1) expression in BAT and inguinal white...... glucose and lipid homeostasis, in part through recruitment and metabolic activation of brown and beige adipocytes....

  9. Glucose transporter 8 (GLUT8) regulates enterocyte fructose transport and global mammalian fructose utilization.

    Science.gov (United States)

    DeBosch, Brian J; Chi, Maggie; Moley, Kelle H

    2012-09-01

    Enterocyte fructose absorption is a tightly regulated process that precedes the deleterious effects of excess dietary fructose in mammals. Glucose transporter (GLUT)8 is a glucose/fructose transporter previously shown to be expressed in murine intestine. The in vivo function of GLUT8, however, remains unclear. Here, we demonstrate enhanced fructose-induced fructose transport in both in vitro and in vivo models of enterocyte GLUT8 deficiency. Fructose exposure stimulated [(14)C]-fructose uptake and decreased GLUT8 protein abundance in Caco2 colonocytes, whereas direct short hairpin RNA-mediated GLUT8 knockdown also stimulated fructose uptake. To assess GLUT8 function in vivo, we generated GLUT8-deficient (GLUT8KO) mice. GLUT8KO mice exhibited significantly greater jejunal fructose uptake at baseline and after high-fructose diet (HFrD) feeding vs. wild-type mice. Strikingly, long-term HFrD feeding in GLUT8KO mice exacerbated fructose-induced increases in blood pressure, serum insulin, low-density lipoprotein and total cholesterol vs. wild-type controls. Enhanced fructose uptake paralleled with increased abundance of the fructose and glucose transporter, GLUT12, in HFrD-fed GLUT8KO mouse enterocytes and in Caco2 cultures exposed to high-fructose medium. We conclude that GLUT8 regulates enterocyte fructose transport by regulating GLUT12, and that disrupted GLUT8 function has deleterious long-term metabolic sequelae. GLUT8 may thus represent a modifiable target in the prevention and treatment of malnutrition or the metabolic syndrome.

  10. Glucose transporter-1 deficiency syndrome: The expanding clinical and genetic spectrum of a treatable disorder

    NARCIS (Netherlands)

    W.G. Leen (Wilhelmina); J. Klepper (Joerg); M.M. Verbeek (Marcel); M. Leferink (Maike); T. Hofste (Tom); B.G.M. van Engelen (Baziel); R.A. Wevers (Ron); T. Arthur (Todd); N. Bahi-Buisson (Nadia); D. Ballhausen (Diana); J. Bekhof (Jolita); P. van Bogaert (Patrick); I. Carrilho (Inês); B. Chabrol (Brigitte); M.P. Champion (Michael); J. Coldwell (James); P. Clayton (Peter); E. Donner (Elizabeth); A. Evangeliou (Athanasios); F. Ebinger (Friedrich); K. Farrell (Kevin); R.J. Forsyth (Rob); C.G.E.L. de Goede (Christian); S. Gross (Stephanie); S. Grünewald (Sonja); H. Holthausen (Hans); S. Jayawant (Sandeep); K. Lachlan (Katherine); V. Laugel (Vincent); K. Leppig (Kathy); M.J. Lim (Ming); G.M.S. Mancini (Grazia); A.D. Marina; L. Martorell (Loreto); J. McMenamin (Joe); M.E.C. Meuwissen (Marije); H. Mundy (Helen); N.O. Nilsson (Nils); A. Panzer (Axel); B.T. Poll-The; C. Rauscher (Christian); C.M.R. Rouselle (Christophe); I. Sandvig (Inger); T. Scheffner (Thomas); E. Sheridan (Eamonn); N. Simpson (Neil); P. Sykora (Parol); R. Tomlinson (Richard); J. Trounce (John); D.W.M. Webb (David); B. Weschke (Bernhard); H. Scheffer (Hans); M.A. Willemsen (Michél)

    2010-01-01

    textabstractGlucose transporter-1 deficiency syndrome is caused by mutations in the SLC2A1 gene in the majority of patients and results in impaired glucose transport into the brain. From 2004-2008, 132 requests for mutational analysis of the SLC2A1 gene were studied by automated Sanger sequencing an

  11. Glucose transporter-1 deficiency syndrome : the expanding clinical and genetic spectrum of a treatable disorder

    NARCIS (Netherlands)

    Leen, Wilhelmina G.; Klepper, Joerg; Verbeek, Marcel M.; Leferink, Maike; Hofste, Tom; van Engelen, Baziel G.; Wevers, Ron A.; Arthur, Todd; Bahi-Buisson, Nadia; Ballhausen, Diana; Bekhof, Jolita; van Bogaert, Patrick; Carrilho, Ines; Chabrol, Brigitte; Champion, Michael P.; Coldwell, James; Clayton, Peter; Donner, Elizabeth; Evangeliou, Athanasios; Ebinger, Friedrich; Farrell, Kevin; Forsyth, Rob J.; de Goede, Christian G. E. L.; Gross, Stephanie; Grunewald, Stephanie; Holthausen, Hans; Jayawant, Sandeep; Lachlan, Katherine; Laugel, Vincent; Leppig, Kathy; Lim, Ming J.; Mancini, Grazia; Della Marina, Adela; Martorell, Loreto; McMenamin, Joe; Meuwissen, Marije E. C.; Mundy, Helen; Nilsson, Nils O.; Panzer, Axel; Poll-The, Bwee T.; Rauscher, Christian; Rouselle, Christophe M. R.; Sandvig, Inger; Scheffner, Thomas; Sheridan, Eamonn; Simpson, Neil; Sykora, Parol; Tomlinson, Richard; Trounce, John; Webb, David; Weschke, Bernhard; Scheffer, Hans; Willemsen, Michel A.

    2010-01-01

    Glucose transporter-1 deficiency syndrome is caused by mutations in the SLC2A1 gene in the majority of patients and results in impaired glucose transport into the brain. From 2004-2008, 132 requests for mutational analysis of the SLC2A1 gene were studied by automated Sanger sequencing and multiplex

  12. Glucose Transporter Type 1 Deficiency Syndrome with Carbohydrate-Responsive Symptoms but without Epilepsy

    Science.gov (United States)

    Koy, Anne; Assmann, Birgit; Klepper, Joerg; Mayatepek, Ertan

    2011-01-01

    Glucose transporter type 1 deficiency syndrome (GLUT1-DS) is caused by a defect in glucose transport across the blood-brain barrier. The main symptoms are epilepsy, developmental delay, movement disorders, and deceleration of head circumference. A ketogenic diet has been shown to be effective in controlling epilepsy in GLUT1-DS. We report a female…

  13. Role of the AMPKgamma3 isoform in hypoxia-stimulated glucose transport in glycolytic skeletal muscle

    DEFF Research Database (Denmark)

    Deshmukh, Atul S; Glund, Stephan; Tom, Robby Z

    2009-01-01

    Skeletal muscle glucose transport is regulated via the canonical insulin-signaling cascade as well as by energy-sensing signals. 5'-AMP-activated protein kinase (AMPK) has been implicated in the energy status regulation of glucose transport. We determined the role of the AMPKgamma3 isoform...

  14. [High glucose dialysate enhances peritoneal fibrosis through upregulating glucose transporters GLUT1 and SGLT1].

    Science.gov (United States)

    Hong, Mengqi; Nie, Zhenyu; Chen, Zhengyue; Yu, Xiongwei; Bao, Beiyan

    2016-05-25

    Objective: To investigate the role of glucose transporter 1 (GLUT1) and sodium-glucose cotransporter 1 (SGLT1) in high glucose dialysate-induced peritoneal fibrosis. Methods: Thirty six male SD rats were randomly divided into 6 groups (6 in each):normal control group, sham operation group, peritoneal dialysis group (PD group), PD+phloretin group (PD+T group), PD+phlorizin group (PD+Z group), PD+phloretin+phlorizin group (PD+T+Z group). Rat model of uraemia was established using 5/6 nephrotomy, and 2.5% dextrose peritoneal dialysis solution was used in peritoneal dialysis. Peritoneal equilibration test was performed 24 h after dialysis to evaluate transport function of peritoneum in rats; HE staining was used to observe the morphology of peritoneal tissue; and immunohistochemistry was used to detect the expression of GLUT1, SGLT1, TGF-β1 and connective tissue growth factor (CTGF) in peritoneum. Human peritoneal microvascular endothelial cells (HPECs) were divided into 5 groups:normal control group, peritoneal dialysis group (PD group), PD+phloretin group (PD+T group), PD+phlorezin group (PD+Z group), and PD+phloretin+phlorezin group (PD+T+Z group). Real time PCR and Western blotting were used to detect mRNA and protein expressions of GLUT1, SGLT1, TGF-β1, CTGF in peritoneal membrane and HPECs. Results:In vivo, compared with sham operation group, rats in PD group had thickened peritoneum, higher ultrafiltration volume, and the mRNA and protein expressions of GLUT1, SGLT1, CTGF, TGF-β1 were significantly increased (all P<0.05); compared with PD group, thickened peritoneum was attenuated, and the mRNA and protein expressions of GLUT1, SGLT1, CTGF, TGF-β1 were significantly decreased in PD+T, PD+Z and PD+T+Z groups (all P<0.05). Pearson's correlation analysis showed that the expressions of GLUT1, SGLT1 in peritoneum were positively correlated with the expressions of TGF-β1 and CTGF (all P<0.05). In vitro, the mRNA and protein expressions of GLUT1, SGLT1, TGF-β1

  15. Metformin-induced regulation of the intestinal D-glucose transporters.

    OpenAIRE

    Sakar, Yassine; Meddah, Bouchra; El Abbes Faouzi, Moulay; Cherrah, Yahia; Bado, André; Ducroc, Robert

    2010-01-01

    International audience; Metformin is an orally administered drug that lowers blood glucose and improves insulin sensitivity in patients with non insulin-dependent diabetes. Although the antihyperglycemic effect of metformin has been extensively studied, its cellular mechanism(s) of action (including the effect on enterocyte) remains to be defined. This study was designed to examine the effect of metformin on glucose transporters in enterocyte. Na(+)-dependent glucose transporter-1 (SGLT-1) ac...

  16. Adaptive evolution for fast growth on glucose and the effects on the regulation of glucose transport system in Clostridium tyrobutyricum.

    Science.gov (United States)

    Jiang, Ling; Li, Shuang; Hu, Yi; Xu, Qing; Huang, He

    2012-03-01

    Laboratory adaptive evolution of microorganisms offers the possibility of relating acquired mutations to increased fitness of the organism under the conditions used. By combining a fibrous-bed bioreactor, we successfully developed a simple and valuable adaptive evolution strategy in repeated-batch fermentation mode with high initial substrate concentration and evolved Clostridium tyrobutyricum mutant with significantly improved butyric acid volumetric productivity up to 2.25 g/(L h), which is the highest value in batch fermentation reported so far. Further experiments were conducted to pay attention to glucose transport system in consideration of the high glucose consumption rate resulted from evolution. Complete characterization and comparison of the glucose phosphoenolpyruvate (PEP)-dependent phosphotransferase system (PTS) were carried out in the form of toluene-treated cells and cell-free extracts derived from both C. tyrobutyricum wide-type and mutant, while an alternative glucose transport route that requires glucokinase was confirmed by the phenomena of resistance to the glucose analogue 2-deoxyglucose and ATP-dependent glucose phosphorylation. Our results suggest that C. tyrobutyricum mutant is defective in PTS activity and compensates for this defect with enhanced glucokinase activity, resulting in the efficient uptake and consumption of glucose during the whole metabolism.

  17. Stretch-stimulated glucose transport in skeletal muscle is regulated by Rac1

    DEFF Research Database (Denmark)

    Sylow, Lykke; Møller, Lisbeth L V; Kleinert, Maximilian

    2015-01-01

    Alternatives to the canonical insulin signaling pathway for glucose transport are muscle contraction/exercise. Mechanical stress is an integrated part of the muscle contraction/relaxation cycle and passive stretch has been shown to increase muscle glucose transport. However, the signaling mechanism...... regulating stretch-stimulated glucose transport is not well understood. We recently reported that the actin cytoskeleton regulating GTPase, Rac1 was activated in mouse muscle in response to stretching. Rac1 is a regulator of contraction- and insulin-stimulated glucose transport but its role in stretch...... part of the mechanical stress-component of the contraction-stimulus to glucose transport in skeletal muscle. This article is protected by copyright. All rights reserved....

  18. Cerebrospinal lfuid from rats given hypoxic preconditioning protects neurons from oxygen-glucose deprivation-induced injury

    Institute of Scientific and Technical Information of China (English)

    Yan-bo Zhang; Zheng-dong Guo; Mei-yi Li; Si-jie Li; Jing-zhong Niu; Ming-feng Yang; Xun-ming Ji; Guo-wei Lv

    2015-01-01

    Hypoxic preconditioning activates endogenous mechanisms that protect against cerebral isch-emic and hypoxic injury. To better understand these protective mechanisms, adult rats were housed in a hypoxic environment (8% O2/92% N2) for 3 hours, and then in a normal oxygen environment for 12 hours. Their cerebrospinal fluid was obtained to culture cortical neurons from newborn rats for 1 day, and then the neurons were exposed to oxygen-glucose deprivation for 1.5 hours. The cerebrospinal lfuid from rats subjected to hypoxic preconditioning reduced oxygen-glucose deprivation-induced injury, increased survival rate, upregulated Bcl-2 expression and downregulated Bax expression in the cultured cortical neurons, compared with control. These results indicate that cerebrospinal lfuid from rats given hypoxic preconditioning protects against oxygen-glucose deprivation-induced injury by affecting apoptosis-related protein expres-sion in neurons from newborn rats.

  19. Dynamic changes in proprotein convertase 2 activity in cortical neurons after ischemia/reperfusion and oxygen-glucose deprivation

    Institute of Scientific and Technical Information of China (English)

    Shuqin Zhan; An Zhou; Chelsea Piper; Tao Yang

    2013-01-01

    In this study, a rat model of transient focal cerebral ischemia was established by performing 100 minutes of middle cerebral artery occlusion, and an in vitro model of experimental oxygen-glucose deprivation using cultured rat cortical neurons was established. Proprotein convertase 2 activity gradually decreased in the ischemic cortex with increasing duration of reperfusion. In cultured rat cortical neurons, the number of terminal deoxynucleotidyl transferase-mediated 2'-deoxyuridine 5'-triphosphate-biotin nick end labeling-positive neurons significantly increased and proprotein convertase 2 activity also decreased gradually with increasing duration of oxygen-glucose deprivation. These experimental findings indicate that proprotein convertase 2 activity decreases in ischemic rat cortex after reperfusion, as well as in cultured rat cortical neurons after oxygen-glucose deprivation. These changes in enzyme activity may play an important pathological role in brain injury.

  20. Cerebrospinal fluid from rats given hypoxic preconditioning protects neurons from oxygen-glucose deprivation-induced injury.

    Science.gov (United States)

    Zhang, Yan-Bo; Guo, Zheng-Dong; Li, Mei-Yi; Li, Si-Jie; Niu, Jing-Zhong; Yang, Ming-Feng; Ji, Xun-Ming; Lv, Guo-Wei

    2015-09-01

    Hypoxic preconditioning activates endogenous mechanisms that protect against cerebral ischemic and hypoxic injury. To better understand these protective mechanisms, adult rats were housed in a hypoxic environment (8% O2/92% N2) for 3 hours, and then in a normal oxygen environment for 12 hours. Their cerebrospinal fluid was obtained to culture cortical neurons from newborn rats for 1 day, and then the neurons were exposed to oxygen-glucose deprivation for 1.5 hours. The cerebrospinal fluid from rats subjected to hypoxic preconditioning reduced oxygen-glucose deprivation-induced injury, increased survival rate, upregulated Bcl-2 expression and downregulated Bax expression in the cultured cortical neurons, compared with control. These results indicate that cerebrospinal fluid from rats given hypoxic preconditioning protects against oxygen-glucose deprivation-induced injury by affecting apoptosis-related protein expression in neurons from newborn rats.

  1. A novel strategy for the treatment of diabetes mellitus - sodium glucose co-transport inhibitors.

    Science.gov (United States)

    Niazi, Asfandyar Khan; Niazi, Saad Hameed

    2010-12-01

    Diabetes is one of the most common chronic diseases, affecting almost 3 million in Canada alone and is characterized by increased blood glucose levels. Treatment varies from lifestyle changes to oral anti-diabetics and/or insulin. Sodium glucose co-transport inhibitors may offer promising treatment for patients suffering from diabetes. The inhibitors act by increasing the loss of glucose in urine by decreasing the reabsorption of glucose from the proximal tubules of nephrons. The aim of this review was to assess the efficacy of sodium glucose co-transport inhibitors in the treatment of diabetes as well as any adverse effects. Databases such as MEDLINE, COCHRANE and EMBASE were systematically searched for literature on the efficacy of sodium glucose co-transport inhibitors in improving the glycemic control of patients with diabetes. Research showed that sodium glucose co-transport inhibitors significantly decreased blood glucose levels by increasing glucosuria. Due to the diuretic effects of these inhibitors, diabetic patients who were suffering from hypertension showed a decrease in blood pressure. The caloric loss associated with these inhibitors resulted in weight loss as well. The most common adverse effect seen in patients on these medications was mycotic infection of the urinary or genital tract. Sodium glucose co-transport inhibitors may be an effective line of treatment for diabetes. Although short-term research has shown these drugs to be safe and well-tolerated, studies should be conducted to assess the long-term effects of these drugs.

  2. AICAR administration affects glucose metabolism by upregulating the novel glucose transporter, GLUT8, in equine skeletal muscle.

    Science.gov (United States)

    de Laat, M A; Robinson, M A; Gruntmeir, K J; Liu, Y; Soma, L R; Lacombe, V A

    2015-09-01

    Equine metabolic syndrome is characterized by obesity and insulin resistance (IR). Currently, there is no effective pharmacological treatment for this insidious disease. Glucose uptake is mediated by a family of glucose transporters (GLUT), and is regulated by insulin-dependent and -independent pathways, including 5-AMP-activated protein kinase (AMPK). Importantly, the activation of AMPK, by 5-aminoimidazole-4-carboxamide-1-D-ribofuranoside (AICAR) stimulates glucose uptake in both healthy and diabetic humans. However, whether AICAR promotes glucose uptake in horses has not been established. It is hypothesized that AICAR administration would enhance glucose transport in equine skeletal muscle through AMPK activation. In this study, the effect of an intravenous AICAR infusion on blood glucose and insulin concentrations, as well as on GLUT expression and AMPK activation in equine skeletal muscle (quantified by Western blotting) was examined. Upon administration, plasma AICAR rapidly reached peak concentration. Treatment with AICAR resulted in a decrease (P change in lactate concentration. The ratio of phosphorylated to total AMPK was increased (P managing IR requires investigation. Copyright © 2015 Elsevier Ltd. All rights reserved.

  3. Adenovirus-mediated transfection with glucose transporter 3 suppresses PC12 cell apoptosis following ischemic injury

    Institute of Scientific and Technical Information of China (English)

    Junliang Li; Xinke Xu; Shanyi Zhang; Meiguang Zheng; Zhonghua Wu; Yinlun Weng; Leping Ouyang; Jian Yu; Fangcheng Li

    2012-01-01

    In this study, we investigated the effects of adenovirus-mediated transfection of PC12 cells with glucose transporter 3 after ischemic injury. The results of flow cytometry and TUNEL showed that exogenous glucose transporter 3 significantly suppressed PC12 cell apoptosis induced by ischemic injury. The results of isotopic scintiscan and western blot assays showed that, the glucose uptake rate was significantly increased and nuclear factor kappaB expression was significantly decreased after adenovirus-mediated transfection of ischemic PC12 cells with glucose transporter 3. These results suggest that adenovirus-mediated transfection of cells with glucose transporter 3 elevates the energy metabolism of PC12 cells with ischemic injury, and inhibits cell apoptosis.

  4. Arctic ground squirrel neuronal progenitor cells resist oxygen and glucose deprivation-induced death

    Science.gov (United States)

    Drew, Kelly L; Wells, Matthew; McGee, Rebecca; Ross, Austin P; Kelleher-Andersson, Judith

    2016-01-01

    AIM: To investigate the influence of ischemia/reperfusion on arctic ground squirrel (AGS) neuronal progenitor cells (NPCs), we subjected these cultured cells to oxygen and glucose deprivation. METHODS: AGS NPCs were expanded and differentiated into NPCs and as an ischemia vulnerable control, commercially available human NPCs (hNPCs) were seeded from thawed NPCs. NPCs, identified by expression of TUJ1 were seen at 14-21 d in vitro (DIV). Cultures were exposed to control conditions, hypoxia, oxygen and glucose deprivation or glucose deprivation alone or following return to normal conditions to model reperfusion. Cell viability and death were assessed from loss of ATP as well as from measures of alamarBlue® and lactate dehydrogenase in the media and from counts of TUJ1 positive cells using immunocytochemistry. Dividing cells were identified by expression of Ki67 and phenotyped by double labeling with GFAP, MAP2ab or TUJ1. RESULTS: We report that when cultured in NeuraLife™, AGS cells remain viable out to 21 DIV, continue to express TUJ1 and begin to express MAP2ab. Viability of hNPCs assessed by fluorescence alamarBlue (arbitrary units) depends on both glucose and oxygen availability [viability of hNPCs after 24 h oxygen glucose deprivation (OGD) with return of oxygen and glucose decreased from 48151 ± 4551 in control cultures to 43481 ± 2413 after OGD, P < 0.05]. By contrast, when AGS NPCs are exposed to the same OGD with reperfusion at 14 DIV, cell viability assessed by alamarBlue increased from 165305 ± 11719 in control cultures to 196054 ± 13977 after OGD. Likewise AGS NPCs recovered ATP (92766 ± 6089 in control and 92907 ± 4290 after modeled reperfusion; arbitrary luminescence units), and doubled in the ratio of TUJ1 expressing neurons to total dividing cells (0.11 ± 0.04 in control cultures vs 0.22 ± 0.2 after modeled reperfusion, P < 0.05). Maintaining AGS NPCs for a longer time in culture lowered resistance to injury, however, did not impair

  5. Arctic ground squirrel neuronal progenitor cells resist oxygen and glucose deprivation-induced death

    Institute of Scientific and Technical Information of China (English)

    Kelly L Drew; Matthew Wells; Rebecca McGee; Austin P Ross; Judith Kelleher-Andersson

    2016-01-01

    AIM: To investigate the influence of ischemia/reperfusion on arctic ground squirrel(AGS) neuronal progenitor cells(NPCs), we subjected these cultured cells to oxygen and glucose deprivation.METHODS: AGS NPCs were expanded and differentiated into NPCs and as an ischemia vulnerable control, commercially available human NPCs(hNPCs) were seeded from thawed NPCs. NPCs, identified by expression of TUJ1 were seen at 14-21 d in vitro(DIV). Cultures were exposed to control conditions, hypoxia, oxygen and glucose deprivation or glucose deprivation alone or following return to normal conditions to model reperfusion. Cell viability and death were assessed from loss of ATP as well as from measures of alamarB lue and lactate dehydrogenase in the media and from counts of TUJ1 positive cells using immunocytochemistry. Dividing cells were identified by expression of Ki67 and phenotyped by double labeling with GFAP, MAP2 ab or TUJ1. RESULTS: We report that when cultured in NeuraLifeTM, AGS cells remain viable out to 21 DIV, continue to express TUJ1 and begin to express MAP2 ab. Viability of hN PCs assessed by fluorescence alamarB lue(arbitrary units) depends on both glucose and oxygen availability [viability of hNPCs after 24 h oxygen glucose deprivation(OGD) with return of oxygen and glucose decreased from 48151 ± 4551 in control cultures to 43481 ± 2413 after OGD, P < 0.05]. By contrast, when AGS NPCs are exposed to the same OGD with reperfusion at 14 DIV, cell viability assessed by alamar Blue increased from 165305 ± 11719 in control cultures to 196054 ± 13977 after OGD. Likewise AGS NPCs recovered ATP(92766 ± 6089 in control and 92907 ± 4290 after modeled reperfusion; arbitrary luminescence units), and doubled in the ratio of TUJ1 expressing neurons to total dividing cells(0.11 ± 0.04 in control cultures vs 0.22 ± 0.2 after modeled reperfusion, P < 0.05). Maintaining AGS NPCs for a longer time in culture lowered resistance to injury

  6. The Ketone Body, β-Hydroxybutyrate Stimulates the Autophagic Flux and Prevents Neuronal Death Induced by Glucose Deprivation in Cortical Cultured Neurons.

    Science.gov (United States)

    Camberos-Luna, Lucy; Gerónimo-Olvera, Cristian; Montiel, Teresa; Rincon-Heredia, Ruth; Massieu, Lourdes

    2016-03-01

    Glucose is the major energy substrate in brain, however, during ketogenesis induced by starvation or prolonged hypoglycemia, the ketone bodies (KB), acetoacetate and β-hydroxybutyrate (BHB) can substitute for glucose. KB improve neuronal survival in diverse injury models, but the mechanisms by which KB prevent neuronal damage are still not well understood. In the present study we have investigated whether protection by the D isomer of BHB (D-BHB) against neuronal death induced by glucose deprivation (GD), is related to autophagy. Autophagy is a lysosomal-dependent degradation process activated during nutritional stress, which leads to the digestion of damaged proteins and organelles providing energy for cell survival. Results show that autophagy is activated in cortical cultured neurons during GD, as indicated by the increase in the levels of the lipidated form of the microtubule associated protein light chain 3 (LC3-II), and the number of autophagic vesicles. At early phases of glucose reintroduction (GR), the levels of p62 declined suggesting that the degradation of the autophagolysosomal content takes place at this time. In cultures exposed to GD and GR in the presence of D-BHB, the levels of LC3-II and p62 rapidly declined and remained low during GR, suggesting that the KB stimulates the autophagic flux preventing autophagosome accumulation and improving neuronal survival.

  7. Disruption of microtubules in rat skeletal muscle does not inhibit insulin- or contraction-stimulated glucose transport

    DEFF Research Database (Denmark)

    Ai, Hua; Ralston, Evelyn; Lauritzen, Hans P M M

    2003-01-01

    or epitrochlearis muscles. In contrast, nocodazole, another microtubule-disrupting drug, rapidly and dose dependently blocked insulin- and contraction-stimulated glucose transport. A similar discrepancy between colchicine and nocodazole was also found in their ability to block glucose transport in muscle giant...... "ghost" vesicles. This suggests that the ability of insulin and contractions to stimulate glucose transport in muscle does not require an intact microtubule network and that nocodazole inhibits glucose transport independently of its microtubule-disrupting effect....

  8. Involvement of AMPK in regulating the degradation of MAD2B under high glucose in neuronal cells.

    Science.gov (United States)

    Meng, Xianfang; Chu, Guangpin; Ye, Chen; Tang, Hui; Qiu, Ping; Hu, Yue; Li, Man; Zhang, Chun

    2016-12-13

    Although our recent study has demonstrated that mitotic spindle assembly checkpoint protein (MAD2B) mediates high glucose-induced neuronal apoptosis, the mechanisms for MAD2B degradation under hyperglycaemia have not yet been elucidated. In this study, we first found that the activation of adenosine 5'-monophosphate (AMP)-activated protein kinase (AMPK) was decreased in neurons, accompanied with the increased expression of MAD2B. Mechanistically, we demonstrated that activation of AMPK with its activators such as AICAR and metformin decreased the expression of MAD2B, indicating a role of AMPK in regulating the expression of MAD2B. Moreover, activation of AMPK prevented neuronal cells from high glucose-induced injury as demonstrated by the reduced expression of cyclin B1 and percentage of apoptosis as detected by TUNEL. We further found that when total protein synthesis was suppressed by chlorhexidine, the degradation of MAD2B was slower in high glucose-treated neurons and was mainly dependent on the ubiquitin-proteasome system. Finally, it was indicated that high glucose inhibited the ubiquitination of MAD2B, which could be reversed by activation of AMPK. Collectively, this study demonstrates that AMPK acts as a key regulator of MAD2B expression, suggesting that activation of AMPK signalling might be crucial for the treatment of high glucose-induced neuronal injury.

  9. Glucose uptake and growth of glucose-limited chemostat cultures of Aspergillus niger and a disruptant lacking MstA, a high-affinity glucose transporter

    DEFF Research Database (Denmark)

    Jørgensen, Thomas R; vanKuyk, Patricia A; Poulsen, Bjarne R

    2007-01-01

    This is a study of high-affinity glucose uptake in Aspergillus niger and the effect of disruption of a high-affinity monosaccharide-transporter gene, mstA. The substrate saturation constant (K(s)) of a reference strain was about 15 microM in glucose-limited chemostat culture. Disruption of mst......-affinity uptake system of A. niger. The mstA disruptant and a reference strain were cultivated in glucose-limited chemostat cultures at low, intermediate and high dilution rate (D=0.07 h(-1), 0.14 h(-1) and 0.20 h(-1)). Mycelium harvested from steady-state cultures was subjected to glucose uptake assays...

  10. GLP-1 analog raises glucose transport capacity of blood-brain barrier in Alzheimer's disease

    DEFF Research Database (Denmark)

    Gejl, M.; Brock, B.; Egefjord, L.

    2017-01-01

    Objectives: Glucose enters the brain tissue from plasma by facilitated diffusion across the two membranes of the endothelium of the blood-brain barrier (BBB), mediated by the glucose transporter 1 (GLUT1). There is evidence in Alzheimer's disease (AD) of reduction of glucose transport across...... claim that the GLP-1 analog liraglutide may prevent the decline of blood-brain glucose transfer in AD. Methods: In this 26-week test of the hypothesis, we randomized 38 patients with AD to treatment with the GLP-1 analog liraglutide (n = 18) or placebo (n = 20). We determined blood-brain glucose...... transport capacity (Tmax) with [18F]FDG (FDG) (ClinicalTrials.gov NCT01469351). Results: In both groups, the Tmax estimates declined in proportion to the duration of AD. The GLP-1 analog treatment very significantly (P 

  11. The Glucose Sensor-Like Protein Hxs1 Is a High-Affinity Glucose Transporter and Required for Virulence in Cryptococcus neoformans

    Science.gov (United States)

    Baker, Gregory M.; Fahmy, Hany; Jiang, Linghuo; Xue, Chaoyang

    2013-01-01

    Cryptococcus is a major fungal pathogen that frequently causes systemic infection in patients with compromised immunity. Glucose, an important signal molecule and the preferred carbon source for Cryptococcus, plays a critical role in fungal development and virulence. Cryptococcus contains more than 50 genes sharing high sequence homology with hexose transporters in Saccharomyces cerevisiae. However, there is no report on their function in glucose sensing or transport. In this study, we investigated two hexose transporter-like proteins (Hxs1 and Hxs2) in Cryptococcus that share the highest sequence identity with the glucose sensors Snf3 and Rgt2 in S. cerevisiae. The expression of HXS1 is repressed by high glucose, while the HXS2 expression is not regulated by glucose. Functional studies showed that Hxs1 is required for fungal resistance to oxidative stress and fungal virulence. The hxs1Δ mutant exhibited a significant reduction in glucose uptake activity, indicating that Hxs1 is required for glucose uptake. Heterologous expression of Cryptococcus HXS1 rendered the S. cerevisiae mutant lacking all 20 hexose transporters a high glucose uptake activity, demonstrating that Hxs1 functions as a glucose transporter. Heterologous expression of HXS1 in the snf3Δ rgt2Δ double mutant did not complement its growth in YPD medium containing the respiration inhibitor antimycin A, suggesting that Hxs1 may not function as a glucose sensor. Taken together, our results demonstrate that Hxs1 is a high-affinity glucose transporter and required for fungal virulence. PMID:23691177

  12. Neuroprotective effects of (-)-linalool against oxygen-glucose deprivation-induced neuronal injury.

    Science.gov (United States)

    Park, Hyeon; Seol, Geun Hee; Ryu, Sangwoo; Choi, In-Young

    2016-04-01

    (-)-Linalool, a major component of many essential oils, is widely used in cosmetics and flavoring ingredients as well as in traditional medicines. Although various in vitro and in vivo studies have shown that (-)-linalool has anti-convulsant, anti-nociceptive, anti-inflammatory and anti-oxidative properties, its anti-ischemic/hypoxic effects have yet to be determined. This study assessed the neuroprotective effects of (-)-linalool against oxygen-glucose deprivation/reoxygenation (OGD/R)-induced cortical neuronal injury, an in vitro model of ischemic stroke. (-)-Linalool significantly attenuated OGD/R-evoked cortical neuronal injury/death, although it did not inhibit N-methyl-D-aspartate (NMDA)-induced excitotoxicity. (-)-Linalool significantly reduced intracellular oxidative stress during OGD/R-induced injury, as well as scavenging peroxyl radicals (Trolox equivalents or TE = 3.8). This anti-oxidant effect was found to correlate with the restoration of OGD/R-induced decreases in the activities of SOD and catalase. In addition, (-)-linalool inhibited microglial migration induced by monocyte-chemoattractant protein-1 (MCP-1), a chemokine released by OGD/R. These findings show that (-)-linalool has neuroprotective effects against OGD/R-induced neuronal injury, which may be due to its anti-oxidant and anti-inflammatory activities. Detailed examination of the anti-ischemic mechanisms of (-)-linalool may indicate strategies for the development of drugs to treat cerebral ischemic injury.

  13. Expression of hexokinases and glucose transporters in treated and untreated oesophageal adenocarcinoma

    NARCIS (Netherlands)

    Fonteyne, Philippe; Casneuf, Veerle; Pauwels, Patrick; Van Damme, Nancy; Peeters, Marc; Dierckx, Rudi; Van de Wiele, Christophe

    2009-01-01

    The aim of this study was to assess the expression pattern of the high glucose affinity glucose transporters GLUT 1, 2, 3, 4, 8 and 9 and of hexokinases I, II and III in newly diagnosed oesophageal adenocarcinoma by means of immunohistochemistry. Twenty patients eligible to undergo primary surgery a

  14. Glucose Transport into Everted Sacs of the Small Intestine of Mice

    Science.gov (United States)

    Hamilton, Kirk L.; Butt, A. Grant

    2013-01-01

    The Na[superscript +]-glucose cotransporter is a key transport protein that is responsible for absorbing Na[superscript +] and glucose from the luminal contents of the small intestine and reabsorption by the proximal straight tubule of the nephron. Robert K. Crane originally described the cellular model of absorption of Na[superscript +] and…

  15. Expression of hexokinases and glucose transporters in treated and untreated oesophageal adenocarcinoma

    NARCIS (Netherlands)

    Fonteyne, Philippe; Casneuf, Veerle; Pauwels, Patrick; Van Damme, Nancy; Peeters, Marc; Dierckx, Rudi; Van de Wiele, Christophe

    2009-01-01

    The aim of this study was to assess the expression pattern of the high glucose affinity glucose transporters GLUT 1, 2, 3, 4, 8 and 9 and of hexokinases I, II and III in newly diagnosed oesophageal adenocarcinoma by means of immunohistochemistry. Twenty patients eligible to undergo primary surgery a

  16. Peritoneal transport characteristics with glucose polymer-based dialysis fluid in children.

    NARCIS (Netherlands)

    Rusthoven, E.; Krediet, R.T.; Willems, J.L.; Monnens, L.A.H.; Schröder, C.H.

    2004-01-01

    Scarce data are available on the use of glucose polymer-based dialysate in children. The effects of glucose polymer-based dialysate on peritoneal fluid kinetics and solute transport were studied in pediatric patients who were on chronic peritoneal dialysis, and a comparison was made with previously

  17. Mangiferin Upregulates Glyoxalase 1 Through Activation of Nrf2/ARE Signaling in Central Neurons Cultured with High Glucose.

    Science.gov (United States)

    Liu, Yao-Wu; Cheng, Ya-Qin; Liu, Xiao-Li; Hao, Yun-Chao; Li, Yu; Zhu, Xia; Zhang, Fan; Yin, Xiao-Xing

    2016-06-18

    Mangiferin, a natural C-glucoside xanthone, has anti-inflammatory, anti-oxidative, neuroprotective actions. Our previous study showed that mangiferin could attenuate diabetes-associated cognitive impairment of rats by enhancing the function of glyoxalase 1 (Glo-1) in brain. The aim of this study was to investigate whether Glo-1 upregulation by mangiferin in central neurons exposed to chronic high glucose may be related to activation of Nrf2/ARE pathway. Compared with normal glucose (25 mmol/L) culture, Glo-1 protein, mRNA, and activity levels were markedly decreased in primary hippocampal and cerebral cortical neurons cultured with high glucose (50 mmol/L) for 72 h, accompanied by the declined Nrf2 nuclear translocation and protein expression of Nrf2 in cell nucleus, as well as protein expression and mRNA level of γ-glutamylcysteine synthetase (γ-GCS) and superoxide dismutase activity, target genes of Nrf2/ARE signaling. Nonetheless, high glucose cotreating with mangiferin or sulforaphane, a typical inducer of Nrf2 activation, attenuated the above changes in both central neurons. In addition, mangiferin and sulforaphane significantly prevented the formation of advanced glycation end-products (AGEs) reflecting Glo-1 activity, while elevated the level of glutathione, a cofactor of Glo-1 activity and production of γ-GCS, in high glucose cultured central neurons. These findings demonstrated that Glo-1 was greatly downregulated in central neurons exposed to chronic high glucose, which is expected to lead the formation of AGEs and oxidative stress damages. We also proved that mangiferin enhanced the function of Glo-1 under high glucose condition by inducing activation of Nrf2/ARE signaling pathway.

  18. Metformin Protects Neurons against Oxygen-Glucose Deprivation/Reoxygenation -Induced Injury by Down-Regulating MAD2B

    Directory of Open Access Journals (Sweden)

    Xianfang Meng

    2016-11-01

    Full Text Available Background/Aims: Metformin, the common medication for type II diabetes, has protective effects on cerebral ischemia. However, the molecular mechanisms are far from clear. Mitotic arrest deficient 2-like protein 2 (MAD2B, an inhibitor of the anaphase-promoting complex (APC, is widely expressed in hippocampal and cortical neurons and plays an important role in mediating high glucose-induced neurotoxicity. The present study investigated whether metformin modifies the expression of MAD2B and to exert its neuroprotective effects in primary cultured cortical neurons during oxygen-glucose deprivation/reoxygenation (OGD/R, a widely used in vitro model of ischemia/reperfusion. Methods: Primary cortical neurons were cultured, deprived of oxygen-glucose for 1 h, and then recovered with oxygen-glucose for 12 h and 24 h. Cell viability was measured by detecting the levels of lactate dehydrogenase (LDH in culture medium. The levels of MAD2B, cyclin B and p-histone 3 were measured by Western blot. Results: Cell viability of neurons was reduced under oxygen-glucose deprivation/reoxygenation (OGD/R. The expression of MAD2B was increased under OGD/R. The levels of cyclin B1, which is a substrate of APC, were also increased. Moreover, OGD/R up-regulated the phosphorylation levels of histone 3, which is the induction of aberrant re-entry of post-mitotic neurons. However, pretreatment of neurons with metformin alleviated OGD/R-induced injury. Metformin further decreased the expression of MAD2B, cyclin B1 and phosphorylation levels of histone 3. Conclusion: Metformin exerts its neuroprotective effect through regulating the expression of MAD2B in neurons under OGD/R.

  19. Reversible white matter lesions during ketogenic diet therapy in glucose transporter 1 deficiency syndrome.

    Science.gov (United States)

    Shiohama, Tadashi; Fujii, Katsunori; Takahashi, Satoru; Nakamura, Fumito; Kohno, Yoichi

    2013-12-01

    Glucose transporter type 1 deficiency syndrome is caused by brain energy failure resulting from a disturbance in glucose transport. We describe a 4-year-old boy with classical type glucose transporter type 1 deficiency syndrome with a heterozygous splice acceptor site mutation (c.517-2A>G) in the SLCA2A1 gene. We initiated a ketogenic diet at 4 months of age. However, even though his condition was good during ketogenic diet therapy, multiple cerebral white matter and right cerebellum lesions appeared at 9 months of age. The lesions in the cerebral white matter subsequently disappeared, indicating that white matter lesions during diet therapy may be reversible and independent of the ketogenic diet. This is the first report of reversible white matter lesions during ketogenic diet therapy in glucose transporter type 1 deficiency syndrome. Copyright © 2013 Elsevier Inc. All rights reserved.

  20. Oxidant stress and skeletal muscle glucose transport: roles of insulin signaling and p38 MAPK.

    Science.gov (United States)

    Kim, John S; Saengsirisuwan, Vitoon; Sloniger, Julie A; Teachey, Mary K; Henriksen, Erik J

    2006-09-01

    Oxidative stress can impact the regulation of glucose transport activity in a variety of cell lines. In the present study, we assessed the direct effects of an oxidant stress on the glucose transport system in intact mammalian skeletal muscle preparations. Type IIb (epitrochlearis) and type I (soleus) muscles from insulin-sensitive lean Zucker rats were incubated in 8 mM glucose for 2 h in the absence or presence of 100 mU/ml glucose oxidase to produce the oxidant hydrogen peroxide (H(2)O(2)) (60-90 microM). Glucose transport, glycogen synthase activity, and metabolic signaling factors were then assessed. H(2)O(2) significantly (p oxidant stress was prevented by the PI3-kinase inhibitor wortmannin. The oxidant stress also significantly increased phosphorylation of p38 mitogen-activated protein kinase (p38 MAPK) and 5'-AMP-dependent protein kinase. Interestingly, selective inhibition of p38 MAPK using A304000 substantially reduced the activation of glucose transport induced by the oxidant stress. These results support a direct role for oxidative stress in the activation of the glucose transport system in mammalian skeletal muscle and indicate that this process involves engagement of and possible interactions between the PI3-kinase-dependent signaling pathway and activation of p38 MAPK.

  1. Cell surface area regulation in neurons in hippocampal slice cultures is resistant to oxygen-glucose deprivation

    Directory of Open Access Journals (Sweden)

    Natalya Shulyakova

    2010-09-01

    Full Text Available Natalya Shulyakova1,2, Jamie Fong2, Diana Diec2, Adrian Nahirny1,2, Linda R Mills1,21Department of Physiology, University of Toronto, Toronto, ON, Canada, M5T 2S8; 2Toronto Western Hospital Research Institute, University Health Network, 11-430, 399 Bathurst St, Toronto, ON, Canada, M5T 2S8Background: Neurons swell in response to a variety of insults. The capacity to recover, ie, to shrink, is critical for neuronal function and survival. Studies on dissociated neurons have shown that during swelling and shrinking, neurons reorganize their plasma membrane; as neurons swell, in response to hypo-osmotic media, the bilayer area increases. Upon restoration of normo-osmotic media, neurons shrink, forming transient invaginations of the plasma membrane known as vacuole-like dilations (VLDs, to accommodate the decrease in the bilayer.Methods: Here we used confocal microscopy to monitor neuronal swelling and shrinking in the three-dimensional (3D environment of post-natal rat hippocampal slice cultures. To label neurons, we used biolistic transfection, to introduce enhanced green fluorescent protein (eGFP targeted to the cytoplasm; and a membrane targeted GFP (lckGFP, targeted to the plasma membrane.Results: Neurons in slice cultures swelled and shrank in response to hypo-osmotic to normo-osmotic media changes. Oxygen-glucose deprivation (OGD caused sustained neuronal swelling; after reperfusion, some neurons recovered but in others, VLD recovery was stalled. OGD did not impair neuronal capacity to recover from a subsequent osmotic challenge.Conclusion: These results suggest cell surface area regulation (SAR is an intrinsic property of neurons, and that neuronal capacity for SAR may play an important role in the brain’s response to ischemic insults.Keywords: neurons, swelling, ischemia, cell surface area, hippocampal slice culture

  2. A glucose transporter can mediate ribose uptake: definition of residues that confer substrate specificity in a sugar transporter.

    Science.gov (United States)

    Naula, Christina M; Logan, Flora J; Logan, Flora M; Wong, Pui Ee; Barrett, Michael P; Burchmore, Richard J

    2010-09-24

    Sugars, the major energy source for many organisms, must be transported across biological membranes. Glucose is the most abundant sugar in human plasma and in many other biological systems and has been the primary focus of sugar transporter studies in eukaryotes. We have previously cloned and characterized a family of glucose transporter genes from the protozoan parasite Leishmania. These transporters, called LmGT1, LmGT2, and LmGT3, are homologous to the well characterized glucose transporter (GLUT) family of mammalian glucose transporters. We have demonstrated that LmGT proteins are important for parasite viability. Here we show that one of these transporters, LmGT2, is a more effective carrier of the pentose sugar d-ribose than LmGT3, which has a 6-fold lower relative specificity (V(max)/K(m)) for ribose. A pair of threonine residues, located in the putative extracellular loops joining transmembrane helices 3 to 4 and 7 to 8, define a filter that limits ribose approaching the exofacial substrate binding pocket in LmGT3. When these threonines are substituted by alanine residues, as found in LmGT2, the LmGT3 permease acquires ribose permease activity that is similar to that of LmGT2. The location of these residues in hydrophilic loops supports recent suggestions that substrate recognition is separated from substrate binding and translocation in this important group of transporters.

  3. Early alterations in soleus GLUT-4, glucose transport, and glycogen in voluntary running rats

    Science.gov (United States)

    Henriksen, Erik J.; Halseth, Amy E.

    1994-01-01

    Voluntary wheel running (WR) by juvenile female rats was used as a noninterventional model of soleus muscle functional overload to study the regulation of insulin-stimulated glucose transport activity by the glucose transporter (GLUT-4 isoform) protein level and glycogen concentration. Soleus total protein content was significantly greater (+18%;P greater than 0.05) than in age-matched controls after 1 wk of WR, and this hypertrophic response continued in weeks 2-4 (+24-32%). GLUT-4 protein was 39% greater than in controls in 1-wk WR soleus, and this adaptation was accompanied by a similar increase in in vitro insulin-stimulated glucose transport activity(+29%). After 2 and 4 wk of WR, however, insulin-stimulated glucose transport activity had returned to control levels, despite a continued elevation (+25-28%) of GLUT-4 protein. At these two time points, glycogen concentration was significantly enhanced in WR soleus (+21-42%), which coincided with significant reductions in glycogen synthase activity ratios (-23 to-41%). These results indicate that, in this model of soleus muscle functional overload, the GLUT-4 protein level may initially regulate insulin-stimulated glucose transport activity in the absence of changes in other modifying factors. However,this regulation of glucose transport activity by GLUT-4 protein may be subsequently overridden by elevated glycogen concentration.

  4. A karyopherin alpha2 nuclear transport pathway is regulated by glucose in hepatic and pancreatic cells.

    Science.gov (United States)

    Cassany, Aurélia; Guillemain, Ghislaine; Klein, Christophe; Dalet, Véronique; Brot-Laroche, Edith; Leturque, Armelle

    2004-01-01

    We studied the role of the karyopherin alpha2 nuclear import carrier (also known as importin alpha2) in glucose signaling. In mhAT3F hepatoma cells, GFP-karyopherin alpha2 accumulated massively in the cytoplasm within minutes of glucose extracellular addition and returned to the nucleus after glucose removal. In contrast, GFP-karyopherin alpha1 distribution was unaffected regardless of glucose concentration. Glucose increased GFP-karyopherin alpha2 nuclear efflux by a factor 80 and its shuttling by a factor 4. These glucose-induced movements were not due to glycolytic ATP production. The mechanism involved was leptomycin B-insensitive, but phosphatase- and energy-dependent. HepG2 and COS-7 cells displayed no glucose-induced GFP-karyopherin alpha2 movements. In pancreatic MIN-6 cells, the glucose-induced movements of karyopherin alpha2 and the stimulation of glucose-induced gene transcription were simultaneously lost between passages 28 and 33. Thus, extracellular glucose regulates a nuclear transport pathway by increasing the nuclear efflux and shuttling of karyopherin alpha2 in cells in which glucose can stimulate the transcription of sugar-responsive genes.

  5. The insulin-like growth factor I receptor regulates glucose transport by astrocytes.

    Science.gov (United States)

    Hernandez-Garzón, Edwin; Fernandez, Ana M; Perez-Alvarez, Alberto; Genis, Laura; Bascuñana, Pablo; Fernandez de la Rosa, Ruben; Delgado, Mercedes; Angel Pozo, Miguel; Moreno, Estefania; McCormick, Peter J; Santi, Andrea; Trueba-Saiz, Angel; Garcia-Caceres, Cristina; Tschöp, Matthias H; Araque, Alfonso; Martin, Eduardo D; Torres Aleman, Ignacio

    2016-11-01

    Previous findings indicate that reducing brain insulin-like growth factor I receptor (IGF-IR) activity promotes ample neuroprotection. We now examined a possible action of IGF-IR on brain glucose transport to explain its wide protective activity, as energy availability is crucial for healthy tissue function. Using (18) FGlucose PET we found that shRNA interference of IGF-IR in mouse somatosensory cortex significantly increased glucose uptake upon sensory stimulation. In vivo microscopy using astrocyte specific staining showed that after IGF-IR shRNA injection in somatosensory cortex, astrocytes displayed greater increases in glucose uptake as compared to astrocytes in the scramble-injected side. Further, mice with the IGF-IR knock down in astrocytes showed increased glucose uptake in somatosensory cortex upon sensory stimulation. Analysis of underlying mechanisms indicated that IGF-IR interacts with glucose transporter 1 (GLUT1), the main facilitative glucose transporter in astrocytes, through a mechanism involving interactions with the scaffolding protein GIPC and the multicargo transporter LRP1 to retain GLUT1 inside the cell. These findings identify IGF-IR as a key modulator of brain glucose metabolism through its inhibitory action on astrocytic GLUT1 activity. GLIA 2016;64:1962-1971.

  6. Insulin stimulated-glucose transporter Glut 4 is expressed in the retina.

    Directory of Open Access Journals (Sweden)

    Gustavo Sánchez-Chávez

    Full Text Available The vertebrate retina is a very metabolically active tissue whose energy demands are normally met through the uptake of glucose and oxygen. Glucose metabolism in this tissue relies upon adequate glucose delivery from the systemic circulation. Therefore, glucose transport depends on the expression of glucose transporters. Here, we show retinal expression of the Glut 4 glucose transporter in frog and rat retinas. Immunohistochemistry and in situ hybridization studies showed Glut 4 expression in the three nuclear layers of the retina: the photoreceptor, inner nuclear and ganglionar cell layers. In the rat retina immunoprecipitation and Western blot analysis revealed a protein with an apparent molecular mass of 45 kDa. ¹⁴C-glucose accumulation by isolated rat retinas was significantly enhanced by physiological concentrations of insulin, an effect blocked by inhibitors of phosphatidyl-inositol 3-kinase (PI3K, a key enzyme in the insulin-signaling pathway in other tissues. Also, we observed an increase in ³H-cytochalasin binding sites in the presence of insulin, suggesting an increase in transporter recruitment at the cell surface. Besides, insulin induced phosphorylation of Akt, an effect also blocked by PI3K inhibition. Expression of Glut 4 was not modified in retinas of a type 1 diabetic rat model. To our knowledge, our results provide the first evidence of Glut4 expression in the retina, suggesting it as an insulin- responsive tissue.

  7. Effect of pycnogenol on glucose transport in mature 3T3-L1 adipocytes.

    Science.gov (United States)

    Lee, Hee-Hyun; Kim, Kui-Jin; Lee, Ok-Hwan; Lee, Boo-Yong

    2010-08-01

    Pycnogenol, a procyanidins-enriched extract of Pinus maritima bark, possesses antidiabetic properties, which improves the altered parameters of glucose metabolism that are associated with type 2 diabetes mellitus (T2DM). Since the insulin-stimulated antidiabetic activities of natural bioactive compounds are mediated by GLUT4 via the phosphatidylinositol-3-kinase (PI3K) and/or p38 mitogen activated protein kinase (p38-MAPK) pathway, the effects of pycnogenol were examined on the molecular mechanism of glucose uptake by the glucose transport system. 3T3-L1 adipocytes were treated with various concentrations of pycnogenol, and glucose uptake was examined using a non-radioisotope enzymatic assay and by molecular events associated with the glucose transport system using semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR). The results show that pycnogenol increased glucose uptake in fully differentiated 3T3-L1 adipocytes and increased the relative abundance of both GLUT4 and Akt mRNAs through the PI3K pathway in a dose dependent manner. Furthermore, pycnogenol restored the PI3K antagonist-induced inhibition of glucose uptake in the presence of wartmannin, an inhibitor of the PI3K. Overall, these results indicate that pycnogenol may stimulate glucose uptake via the PI3K dependent tyrosine kinase pathways involving Akt. Further the results suggest that pycnogenol might be useful in maintaining blood glucose control.

  8. Protective effects of aloperine on neonatal rat primary cultured hippocampal neurons injured by oxygen-glucose deprivation and reperfusion.

    Science.gov (United States)

    Ma, Ning-Tian; Zhou, Ru; Chang, Ren-Yuan; Hao, Yin-Ju; Ma, Lin; Jin, Shao-Ju; Du, Juan; Zheng, Jie; Zhao, Cheng-Jun; Niu, Yang; Sun, Tao; Li, Wei; Koike, Kazuo; Yu, Jian-Qiang; Li, Yu-Xiang

    2015-10-01

    Aloperine (ALO), one of the alkaloids isolated from Sophora alopecuroides L., is traditionally used for various diseases including neuronal disorders. This study investigated the protective effects of ALO on neonatal rat primary-cultured hippocampal neurons injured by oxygen-glucose deprivation and reperfusion (OGD/RP). Treatment with ALO (25, 50, and 100 mg/l) attenuated neuronal damage (p oxygen species and malondialdehyde production and enhanced the antioxidant enzymatic activities of catalase, superoxide dismutase, glutathione peroxidase and the total antioxidant capacity. The results suggested that ALO has significant neuroprotective effects that can be attributed to anti-oxidative stress.

  9. Interleukin 1 stimulates hexose transport in fibroblasts by increasing the expression of glucose transporters.

    Science.gov (United States)

    Bird, T A; Davies, A; Baldwin, S A; Saklatvala, J

    1990-08-15

    Exposure of quiescent cultures of human gingival fibroblasts (HuGi) and porcine synovicocytes (PSF) to human recombinant interleukin 1 alpha or -beta (IL1 alpha and -beta) enhanced the rate of glycolysis as judged by increased lactate production. The cytokines also increased uptake of [3H]2-deoxyglucose (DG) in a time- and dose-dependent manner. Stimulation of DG uptake was first evident 6-8 h following addition of IL1 and was maximal by 24-30 h. IL1 alpha and -beta were equipotent. Half-maximal stimulation occurred at approximately 1 pM IL1; maximal stimulation (2.5-4.5-fold in HuGi, 3-7-fold in PSF) was obtained with approximately 80 pM IL1. The dose-response curves for lactate production and DG uptake were similar. Increased DG uptake was blocked by specific antisera to IL1 and by inhibitors of protein and RNA synthesis but not by indomethacin, an inhibitor of prostaglandin production. DG uptake was enhanced by IL1 in serum-starved cells in the presence of neutralizing anti-platelet-derived growth factor serum. The effect was therefore not secondary to prostaglandin or platelet-derived growth factor production. No increase in cell cycling was detected in IL1-treated cells under the experimental conditions. Kinetic analysis revealed that the Vmax for DG uptake was increased by IL1 (from 36 to 144 pmol/min/mg of cell protein), whereas the Km was unchanged. HuGi cells were pulse-labeled with [35S]methionine following exposure to IL1. Cell lysates were immunoprecipitated using a specific antiserum raised against human erythrocyte glucose transporter. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis/autoradiography of these immunoprecipitates revealed dose- and time-dependent increases in the net rate of glucose transporter synthesis which mirrored the changes in DG uptake.

  10. Analysis of BH3-only proteins upregulated in response to oxygen/glucose deprivation in cortical neurons identifies Bmf but not Noxa as potential mediator of neuronal injury.

    Science.gov (United States)

    Pfeiffer, S; Anilkumar, U; Chen, G; Ramírez-Peinado, S; Galindo-Moreno, J; Muñoz-Pinedo, C; Prehn, J H M

    2014-10-09

    Stress signaling in response to oxygen/glucose deprivation (OGD) and ischemic injury activates a group of pro-apoptotic genes, the Bcl-2 homology domain 3 (BH3)-only proteins, which are capable of activating the mitochondrial apoptosis pathway. Targeted studies previously identified the BH3-only proteins Puma, Bim and Bid to have a role in ischemic/hypoxic neuronal injury. We here investigated the transcriptional activation of pro-apoptotic BH3-only proteins after OGD-induced injury in murine neocortical neurons. We observed a potent and early upregulation of noxa at mRNA and protein level, and a significant increase in Bmf protein levels during OGD in neocortical neurons and in the ipsilateral cortex of mice subjected to transient middle cerebral artery occlusion (tMCAO). Surprisingly, gene deficiency in noxa reduced neither OGD- nor glutamate-induced neuronal injury in cortical neurons and failed to influence infarct size or neurological deficits after tMCAO. In contrast, bmf deficiency induced significant protection against OGD- or glutamate-induced injury in cultured neurons, and bmf-deficient mice showed reduced neurological deficits after tMCAO in vivo. Collectively, our data not only point to a role of Bmf as a BH3-only protein contributing to excitotoxic and ischemic neuronal injury but also demonstrate that the early and potent induction of noxa does not influence ischemic neuronal injury.

  11. Decreased muscle GLUT-4 and contraction-induced glucose transport after eccentric contractions

    DEFF Research Database (Denmark)

    Kristiansen, S; Asp, Svend; Richter, Erik

    1996-01-01

    Eccentric exercise causes muscle damage and decreased muscle glycogen and glucose transporter isoform (GLUT-4) protein content. We investigated whether the contraction-induced increase in skeletal muscle glucose transport and muscle performance is affected by prior eccentric contractions. The calf...... than in CT rats. In the GW and GR muscle, prior eccentric exercise decreased contraction-induced stimulation of glucose transport compared with CT, ST, and CC rats despite no difference in tension development and oxygen uptake among the groups. There was no change in total GLUT-4 content and glucose...... muscles from rats were stimulated for eccentric (EC) or concentric (CC) contractions or were passively stretched (ST). Muscles from unstimulated control (CT) rats were also studied. Two days later, all rats had their isolated hindlimbs perfused either at rest or during 15 min of isometric muscle...

  12. Neuronal glucose but not lactate utilization is positively correlated with NMDA-induced neurotransmission and fluctuations in cytosolic Ca2+ levels

    DEFF Research Database (Denmark)

    Bak, Lasse K; Walls, Anne B; Schousboe, Arne;

    2009-01-01

    Although the brain utilizes glucose for energy production, individual brain cells may to some extent utilize substrates derived from glucose. Thus, it has been suggested that neurons consume extracellular lactate during synaptic activity. However, the precise role of lactate for fueling neuronal...

  13. Glucose transporter-8 (GLUT8) mediates glucose intolerance and dyslipidemia in high-fructose diet-fed male mice.

    Science.gov (United States)

    DeBosch, Brian J; Chen, Zhouji; Finck, Brian N; Chi, Maggie; Moley, Kelle H

    2013-11-01

    Members of the glucose transporter (GLUT) family of membrane-spanning hexose transporters are subjects of intensive investigation for their potential as modifiable targets to treat or prevent obesity, metabolic syndrome, and type 2 diabetes mellitus. Mounting evidence suggests that the ubiquitously expressed class III dual-specificity glucose and fructose transporter, GLUT8, has important metabolic homeostatic functions. We therefore tested the hypothesis that GLUT8 mediates the deleterious metabolic effects of chronic high-fructose diet exposure. Here we demonstrate resistance to high-fructose diet-induced glucose intolerance and dyslipidemia concomitant with enhanced oxygen consumption and thermogenesis in GLUT8-deficient male mice. Independent of diet, significantly lower systolic blood pressure both at baseline and after high-fructose diet feeding was also observed by tail-cuff plethysmography in GLUT8-deficient mice vs wild-type controls. Resistance to fructose-induced metabolic dysregulation occurred in the context of enhanced hepatic peroxisome proliferator antigen receptor-γ (PPARγ) protein abundance, whereas in vivo hepatic adenoviral GLUT8 overexpression suppressed hepatic PPARγ expression. Taken together, these findings suggest that GLUT8 blockade prevents fructose-induced metabolic dysregulation, potentially by enhancing hepatic fatty acid metabolism through PPARγ and its downstream targets. We thus establish GLUT8 as a promising target in the prevention of diet-induced obesity, metabolic syndrome, and type 2 diabetes mellitus in males.

  14. Short-term caloric restriction normalizes hypothalamic neuronal responsiveness to glucose ingestion in patients with type 2 diabetes

    NARCIS (Netherlands)

    Teeuwisse, W.M.; Widya, R.L.; Paulides, M.; Lamb, H.J.; Smit, J.W.A.; Roos, A. de; Buchem, M.A. van; Pijl, H.; Grond, J. van der

    2012-01-01

    The hypothalamus is critically involved in the regulation of feeding. Previous studies have shown that glucose ingestion inhibits hypothalamic neuronal activity. However, this was not observed in patients with type 2 diabetes. Restoring energy balance by reducing caloric intake and losing weight are

  15. Piracetam and TRH analogues antagonise inhibition by barbiturates, diazepam, melatonin and galanin of human erythrocyte D-glucose transport

    Science.gov (United States)

    Naftalin, Richard J; Cunningham, Philip; Afzal-Ahmed, Iram

    2004-01-01

    Nootropic drugs increase glucose uptake into anaesthetised brain and into Alzheimer's diseased brain. Thyrotropin-releasing hormone, TRH, which has a chemical structure similar to nootropics increases cerebellar uptake of glucose in murine rolling ataxia. This paper shows that nootropic drugs like piracetam (2-oxo 1 pyrrolidine acetamide) and levetiracetam and neuropeptides like TRH antagonise the inhibition of glucose transport by barbiturates, diazepam, melatonin and endogenous neuropeptide galanin in human erythrocytes in vitro. The potencies of nootropic drugs in opposing scopolamine-induced memory loss correlate with their potencies in antagonising pentobarbital inhibition of erythrocyte glucose transport in vitro (PPiracetam and TRH have no direct effects on net glucose transport, but competitively antagonise hypnotic drug inhibition of glucose transport. Other nootropics, like aniracetam and levetiracetam, while antagonising pentobarbital action, also inhibit glucose transport. Analeptics like bemigride and methamphetamine are more potent inhibitors of glucose transport than antagonists of hypnotic action on glucose transport. There are similarities between amino-acid sequences in human glucose transport protein isoform 1 (GLUT1) and the benzodiazepine-binding domains of GABAA (gamma amino butyric acid) receptor subunits. Mapped on a 3D template of GLUT1, these homologies suggest that the site of diazepam and piracetam interaction is a pocket outside the central hydrophilic pore region. Nootropic pyrrolidone antagonism of hypnotic drug inhibition of glucose transport in vitro may be an analogue of TRH antagonism of galanin-induced narcosis. PMID:15148255

  16. Imaging of a glucose analog, calcium and NADH in neurons and astrocytes: dynamic responses to depolarization and sensitivity to pioglitazone.

    Science.gov (United States)

    Pancani, Tristano; Anderson, Katie L; Porter, Nada M; Thibault, Olivier

    2011-12-01

    Neuronal Ca(2+) dyshomeostasis associated with cognitive impairment and mediated by changes in several Ca(2+) sources has been seen in animal models of both aging and diabetes. In the periphery, dysregulation of intracellular Ca(2+) signals may contribute to the development of insulin resistance. In the brain, while it is well-established that type 2 diabetes mellitus is a risk factor for the development of dementia in the elderly, it is not clear whether Ca(2+) dysregulation might also affect insulin sensitivity and glucose utilization. Here we present a combination of imaging techniques testing the disappearance of the fluorescent glucose analog 2-(N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino)-2-deoxyglucose (2-NBDG) as an indication of glycolytic activity in neurons and astrocytes. Our work shows that glucose utilization at rest is greater in neurons compared to astrocytes, and ceases upon activation in neurons with little change in astrocytes. Pretreatment of hippocampal cultures with pioglitazone, a drug used in the treatment of type 2 diabetes, significantly reduced glycolytic activity in neurons and enhanced it in astrocytes. This series of experiments, including Fura-2 and NADH imaging, provides results that are consistent with the idea that Ca(2+) levels may rapidly alter glycolytic activity, and that downstream events beyond Ca(2+) dysregulation with aging, may alter cellular metabolism in the brain.

  17. IL-10 Protects Neurites in Oxygen-Glucose-Deprived Cortical Neurons through the PI3K/Akt Pathway.

    Directory of Open Access Journals (Sweden)

    Longzai Lin

    Full Text Available IL-10, as a cytokine, has an anti-inflammatory cascade following various injuries, but it remains blurred whether IL-10 protects neurites of cortical neurons after oxygen-glucose deprivation injury. Here, we reported that IL-10, in a concentration-dependent manner, reduced neuronal apoptosis and increased neuronal survival in oxygen-glucose-deprived primary cortical neurons, producing an optimal protective effect at 20ng/ml. After staining NF-H and GAP-43, we found that IL-10 significantly protected neurites in terms of axon length and dendrite number by confocal microscopy. Furthermore, it induced the phosphorylation of AKT, suppressed the activation of caspase-3, and up-regulated the protein expression of GAP-43. In contrast, LY294002, a specific inhibitor of PI3K/AKT, reduced the level of AKT phosphorylation and GAP-43 expression, increased active caspase-3 expression and thus significantly weakened IL-10-mediated protective effect in the OGD-induced injury model. IL-10NA, the IL-10 neutralizing antibody, reduced the level of p-PI3K phosphorylation and increased the expression of active caspase-3. These findings suggest that IL-10 provides neuroprotective effects by protecting neurites through PI3K/AKT signaling pathway in oxygen-glucose-deprived primary cortical neurons.

  18. Expression and Localization of Glucose Transporters in Rodent Submandibular Salivary Glands

    Directory of Open Access Journals (Sweden)

    Sibel Cetik

    2014-04-01

    Full Text Available Background/Aim: The submandibular gland is one of the three major salivary glands, producing a mixed secretion; this saliva is hypotonic compared to plasma. It also secretes glucose, but the mechanisms responsible for this process are poorly understood. Our study addressed the question whether glucose transporters are expressed and how are they localized within specific rodent submandibular cells, in order to estimate a possible implication in salivary glucose disposal. Methods: Immunohistochemistry, RT-qPCR and Western blotting were performed to determine the presence/localization of glucose transporters in rodent submandibular glands. Results: GLUT4 was identified in the submandibular salivary gland at both mRNA and protein level. The immunohistochemical analysis revealed its localization preponderantly in the ductal cells of the gland, near to the basolateral. SGLT1 and GLUT1 were highly expressed in submandibular tissues in both acinar and ductal cells, but not GLUT2. These results were confirmed by RT-qPCR. It was also documented that insulin stimulates the net uptake of D-glucose by ductal rings prepared from submandibulary salivary glands, the relative magnitude of such an enhancing action being comparable to that found in hemidiaphragms. Conclusion: At least three major glucose transporters are expressed in the rodent submandibular glands, of which GLUT4 is specifically localized near the basolateral side of ductal structures. This points-out its possible role in regulating glucose uptake from the bloodstream, most likely to sustain ductal cellular metabolism.

  19. IDENTIFICATION OF GLUCOSE TRANSPORTER-1 AND ITS FUNCTIONAL ASSAY IN MOUSE GLOMERULAR MESANGIAL CELLS CULTURED IN VITRO

    Institute of Scientific and Technical Information of China (English)

    章精; 刘志红; 刘栋; 黎磊石

    2001-01-01

    Objective. To evaluate the role of glucose transporter-l (GLUT1) in the glucose uptake of glomerular mesangial cells. Methods. Cultured C57/SJL mouse mesangial cells were used in the study. The expression of GLUT1 mRNA was detected by RT-PCR. The expression of GLUT1 protein was detected by immunofluorescence and flow cytometry. The uptake of glucose and its kinetics were determined by 2-deoxy-[3H] -D-glucose uptake. Results. Both GLUT1 mRNA and protein were found in mouse glomerular mesangial cells. 2-deoxy-D-glucose uptake and kinetics assay showed that this glucose transporter had high affinity for glucose and the glucose uptake specificity was further confirmed by phloretin. Conclusion. Functional GLUT1 did present in mouse mesangial cells cultured in vitro and it might be the predominant transporter mediated the uptake of glucose into mesangial cells.

  20. Characteristics of glucose transport across the microvillous membranes of human term placenta

    Directory of Open Access Journals (Sweden)

    Ravinderjit Kaur Anand

    Full Text Available Transport characteristics of D-glucose were studied in the microvillous vesicles isolated from the human term placenta. Transport occurred by selective and rapid facilitated diffusion system which was inhibitable by phloretin and HgCl2. The transport was dependent on a transmembrane. Na+-gradient indicating a "secondary active transport" system operating. The transport influx was saturable and the kinetic analysis based on Hanes-Woolf plot produced a kt and Jmax value of 1.2 mM and 34 nmoles. mgprotein-1.min-1, respectively. The efflux of D-glucose from the membrane vesicles in a pre-equilibrated assay conditions showed a distinct biphasic pattern differing significantly in the half time efflux. The t1/2 of the fast and slow components was found to be 15 sec and 660 sec, respectively. The transport showed distinct sensitivity to temperature and the Ea values both below and above the transition temperature of 37 ºC, as calculated from the Arrhenius plot were found to be 7600 and 5472 kCa1.mol-1, respectively. Inhibition studies with a number of sugars for hexose transport pathway showed that the glucose epimers, phosphorylated sugars, and even the disaccharides and the pentose sugars competed effectively with D-glucose. The influx was also inhibited by a number of steroids such as progesterone, 17α-hydroxyprogesterone, testosterone and estrogen. Insulin was found to increase glucose transport in a dose- dependent fashion at a concentration of 0.2-1 unit.ml-1. Ouabain, dinitrophenoi and nicotine strongly inhibited D-glucose uptake in the membrane vesicles.

  1. Neuronal Cell Death Induced by Mechanical Percussion Trauma in Cultured Neurons is not Preceded by Alterations in Glucose, Lactate and Glutamine Metabolism

    DEFF Research Database (Denmark)

    Jayakumar, A R; Bak, L K; Rama Rao, K V

    2016-01-01

    contribute to the development of neuronal injury. Cell viability was assayed using the release of the cytoplasmic enzyme lactate dehydrogenase (LDH), together with fluorescence-based cell staining (calcein and ethidium homodimer-1 for live and dead cells, respectively). Trauma had no effect on the LDH...... dysfunction and subsequent energy failure play a role in the pathogenesis of TBI. We therefore examined whether oxidative metabolism of (13)C-labeled glucose, lactate or glutamine is altered early following in vitro mechanical percussion-induced trauma (5 atm) to neurons (4-24 h), and whether such events...

  2. Towards bridging the gap between acid-base transporters and neuronal excitability modulation.

    Science.gov (United States)

    Liu, Ying; Chen, Li-Ming

    2014-01-01

    pH homeostasis is a fundamental regulator of the function of the central nervous system. Dysfunction of acid-base transporters often results in disturbance of neuronal excitability. In a latest issue of Journal of Neuroscience, Jones et al. report that increasing intracellular bicarbonate concentration substantially stimulates the excitability of pyramidal neurons from mouse hippocampus by inhibiting KCNQ potassium channel. The finding shed important new light in understanding the molecular mechanism underlying the regulation of neuronal excitability by acid-base transporters.

  3. Effects of melatonin on streptozotocin-induced retina neuronal apoptosis in high blood glucose rat.

    Science.gov (United States)

    Li, Xiaoyan; Zhang, Maonian; Tang, Weiqiang

    2013-03-01

    One of the main pathological symptoms of early diabetic retinal neuropathy is retina neuronal apoptosis. In the present work we investigated the effects of indoleamine hormone melatonin, a powerful free radical scavenger, on streptozotocin-induced retina neuronal cell apoptosis in high blood glucose rat. After melatonin treatment (10 mg/kg/day), tunel detection was used to monitor the apoptosis rate of neurons in the retinal ganglion cell layer; reversed quantitative PCR was used to measure the mRNA expression of retinal caspase-3, Mn superoxidase dismutase (SOD) and Cu-Zn SOD; and the activities of total SOD (T-SOD) and sub-type SOD was detected using xanthine oxidase enzymatic detection. Our data showed that melatonin treatment leads to a decrease of retinal cell apoptosis and the apoptotic index was (1.67 ± 0.54) % and (7.73 ± 0.95) % at 8 and 12 weeks after treatment. The relative quantitative (RQ) value for caspase-3 mRNA expression was (6.996 ± 1.192) and (7.267 ± 1.178) in melatonin group, which are much lower than the values of diabetic group (12.566 ± 2.272 and (14.297 ± 2.110) at 8 and 12 weeks, respectively) under the same condition. mRNA expression of Mn SOD and Cu-Zn SOD as well as their activities all decreased in the diabetic group compared with the control group. While melatonin treatment induced the expression of Mn SOD mRNA and a continual increase of Mn SOD activity as well as the activity and mRNA expression of Cu-Zn SOD at 12 weeks. Therefore, our results demonstrate that melatonin treatment prevented the decrease in mRNA expression of SOD and the increase in caspase-3 mRNA expression induced by diabetes thus exerts a beneficial effect on retina neuronal apoptosis.

  4. Oat β-glucan depresses SGLT1- and GLUT2-mediated glucose transport in intestinal epithelial cells (IEC-6).

    Science.gov (United States)

    Abbasi, Nazanin N; Purslow, Peter P; Tosh, Susan M; Bakovic, Marica

    2016-06-01

    Oat β-glucan consumption is linked to reduced risk factors associated with diabetes and obesity by lowering glycemic response and serum level of low-density lipoproteins. The purpose of this study was to identify the mechanism of action of oat β-glucan at the interface between the gut wall and the lumen responsible for attenuating glucose levels. We proposed that viscous oat β-glucan acts as a physical barrier to glucose uptake in normally absorptive gut epithelial cells IEC-6 by affecting the expression of intestinal glucose transporters. Concentration and time-dependent changes in glucose uptake were established by using a nonmetabolizable glucose analog 2-[N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino]-2-deoxy-d-glucose. The effectiveness of nutrient transport in IEC-6 cells was shown by significant differences in glucose uptake and corresponding transporter expression. The expressions of glucose transporters sodium-glucose-linked transport protein 1 (SGLT1) and glucose transporter 2 (GLUT2) increased with time (0-60 minutes) and glucose levels (5-25 mmol/L). The suppression of glucose uptake and SGLT1 and GLUT2 expression by increasing concentrations (4-8 mg/mL) of oat β-glucan demonstrated a direct effect of the physical properties of oat β-glucan on glucose transport. These results affirmed oat β-glucan as a dietary agent for minimizing postprandial glucose and showed that modulating the activity of the key intestinal glucose transporters with oat β-glucan could be an effective way of lowering blood glucose levels in patients with diabetes. Copyright © 2016 Elsevier Inc. All rights reserved.

  5. Asymmetric subcellular distribution of glucose transporters in the endothelium of small contractile arteries.

    Science.gov (United States)

    Gaudreault, N; Scriven, D R L; Moore, E D W

    2006-01-01

    The authors have recently reported the presence and asymmetric distribution of the glucose transporters GLUT-1 to -5 and SGLT-1 in the endothelium of rat coronary artery (Gaudreault et al. 2004, Diabetologica, 47, 2081-2092). In the present study the authors investigate and compare the presence and subcellular distribution of the classic glucose transporter isoforms in endothelial cells of cerebral, renal, and mesenteric arteries. The GLUTs and SGLT-1 were examined with immunohistochemistry and wide-field fluorescence microscopy coupled to deconvolution in en face preparation of intact artery. We identified GLUT-1 to -5 and SGLT-1 in the endothelial cells of all three vascular beds. The relative level of expression for each isoform was found comparable amongst arteries. Clusters of the glucose transporter isoforms were found at a high density in proximity to the cell-to-cell junctions. In addition, a consistent asymmetric distribution of GLUT-1 to -5 was found, predominantly located on the abluminal side of the endothelium in all three vascular beds examined (ranging from 68% to 91%, p<.05). The authors conclude that the expression and subcellular distribution of glucose transporters are similar in endothelial cells from vascular beds of comparable diameter and suggest that their subcellular organization may facilitate transendothelial transport of glucose in small contractile arteries.

  6. Role of vitamin D on the expression of glucose transporters in L6 myotubes

    Directory of Open Access Journals (Sweden)

    Bubblu Tamilselvan

    2013-01-01

    Full Text Available Altered expression of glucose transporters is a major characteristic of diabetes. Vitamin D has evolved widespread interest in the pathogenesis and prevention of diabetes. The present study was designed to investigate the effect of vitamin D in the overall regulation of muscle cell glucose transporter expression. L6 cells were exposed to type 1 and type 2 diabetic conditions and the effect of calcitriol (1,25, dihydroxy cholicalciferol on the expression of glucose transporters was studied by real time polymerase chain reaction (RT-PCR. There was a significant decrease in glucose transporter type 1 (GLUT1, GLUT4, vitamin D receptor (VDR, and IR expression in type 1 and 2 diabetic model compared to control group. Treatment of myoblasts with 10-7 M calcitriol for 24 h showed a significant increase in GLUT1, GLUT4, VDR, and insulin receptor (IR expression. The results indicate a potential antidiabetic function of vitamin D on GLUT1, GLUT4, VDR, and IR by improving receptor gene expression suggesting a role for vitamin D in regulation of expression of the glucose transporters in muscle cells.

  7. Sodium Glucose Co-transporter Type 2 (SGLT2) Inhibitors: Targeting the Kidney to Improve Glycemic Control in Diabetes Mellitus

    OpenAIRE

    Bays, Harold

    2013-01-01

    Although hyperglycemia is a key therapeutic focus in the management of patients with type 2 diabetes mellitus (T2DM), many patients experience sub-optimal glycemic control. Current glucose-lowering agents involve the targeting of various body organs. Sodium glucose co-transporter type 2 (SGLT2) inhibitors target the kidney, reduce renal glucose reabsorption, and increase urinary glucose elimination, thus lowering glucose blood levels. This review examines some of the key efficacy and safety d...

  8. Blockade of Ca2+-permeable AMPA/kainate channels decreases oxygen-glucose deprivation-induced Zn2+ accumulation and neuronal loss in hippocampal pyramidal neurons.

    Science.gov (United States)

    Yin, Hong Z; Sensi, Stefano L; Ogoshi, Fumio; Weiss, John H

    2002-02-15

    Synaptic release of Zn2+ and its translocation into postsynaptic neurons probably contribute to neuronal injury after ischemia or epilepsy. Studies in cultured neurons have revealed that of the three major routes of divalent cation entry, NMDA channels, voltage-sensitive Ca2+ channels (VSCCs), and Ca2+-permeable AMPA/kainate (Ca-A/K) channels, Ca-A/K channels exhibit the highest permeability to exogenously applied Zn2+. However, routes through which synaptically released Zn2+ gains entry to postsynaptic neurons have not been characterized in vivo. To model ischemia-induced Zn2+ movement in a system approximating the in vivo situation, we subjected mouse hippocampal slice preparations to controlled periods of oxygen and glucose deprivation (OGD). Timm's staining revealed little reactive Zn2+ in CA1 and CA3 pyramidal neurons of slices exposed in the presence of O2 and glucose. However, 15 min of OGD resulted in marked labeling in both regions. Whereas strong Zn2+ labeling persisted if both the NMDA antagonist MK-801 and the VSCC blocker Gd3+ were present during OGD, the presence of either the Ca-A/K channel blocker 1-naphthyl acetyl spermine (NAS) or the extracellular Zn2+ chelator Ca2+ EDTA substantially decreased Zn2+ accumulation in pyramidal neurons of both subregions. In parallel experiments, slices were subjected to 5 min OGD exposures as described above, followed 4 hr later by staining with the cell-death marker propidium iodide. As in the Timm's staining experiments, substantial CA1 or CA3 pyramidal neuronal damage occurred despite the presence of MK-801 and Gd3+, whereas injury was decreased by NAS or by Ca2+ EDTA (in CA1).

  9. Glucose uptake mediated by glucose transporter 1 is essential for early tooth morphogenesis and size determination of murine molars.

    Science.gov (United States)

    Ida-Yonemochi, Hiroko; Nakatomi, Mitsushiro; Harada, Hidemitsu; Takata, Hiroki; Baba, Otto; Ohshima, Hayato

    2012-03-01

    Glucose is an essential source of energy for body metabolism and is transported into cells by glucose transporters (GLUTs). Well-characterized class I GLUT is subdivided into GLUTs1-4, which are selectively expressed depending on tissue glucose requirements. However, there is no available data on the role of GLUTs during tooth development. This study aims to clarify the functional significance of class I GLUT during murine tooth development using immunohistochemistry and an in vitro organ culture experiment with an inhibitor of GLUTs1/2, phloretin, and Glut1 and Glut2 short interfering RNA (siRNA). An intense GLUT1-immunoreaction was localized in the enamel organ of bud-stage molar tooth germs, where the active cell proliferation occurred. By the bell stage, the expression of GLUT1 in the dental epithelium was dramatically decreased in intensity, and subsequently began to appear in the stratum intermedium at the late bell stage. On the other hand, GLUT2-immunoreactivity was weakly observed in the whole tooth germs throughout all stages. The inhibition of GLUTs1/2 by phloretin in the bud-stage tooth germs induced the disturbance of primary enamel knot formation, resulting in the developmental arrest of the explants and the squamous metaplasia of dental epithelial cells. Furthermore, the inhibition of GLUTs1/2 in cap-to-bell-stage tooth germs reduced tooth size in a dose dependent manner. These findings suggest that the expression of GLUT1 and GLUT2 in the dental epithelial and mesenchymal cells seems to be precisely and spatiotemporally controlled, and the glucose uptake mediated by GLUT1 plays a crucial role in the early tooth morphogenesis and tooth size determination. Copyright © 2011 Elsevier Inc. All rights reserved.

  10. Transport of alpha- and beta-D-glucose by the intact human red cell

    Energy Technology Data Exchange (ETDEWEB)

    Carruthers, A.; Melchior, D.L.

    1985-07-16

    The kinetics of alpha- and beta-D-glucose mutarotation and the transport of these anomers by intact human red cells were determined at 0.6 and 36.6 degrees C. The mutarotation coefficients for alpha- and beta-D-glucose in cell-free tris(hydroxymethyl)aminomethane medium (pH 7.4) at 0.6 degrees C are (2.25 +/- 0.2) and (1.73 +/- 0.42) X 10(-3) min-1, respectively, and at 36.6 degrees C are (69 +/- 12) and (75 +/- 5) X 10(-3) min-1, respectively. These values are in good agreement with previous estimates. At 0.6 degrees C, the red cell contains no detectable mutarotase activity. Initial rates of sugar uptake were measured by using radiolabeled D-glucose and time courses of uptake by turbidimetry. The time courses of alpha- and beta-D-glucose and an equilibrium mixture of alpha- and beta-D-glucose infinite-cis entry are identical at 0.66 degrees C (n = 41) where negligible mutarotation is observed. The apparent Ki values for inhibition of radiolabeled D-glucose initial uptake by unlabeled alpha- or beta-D-glucose at 0.6 degrees C are identical (1.6 mM). The calculated Vmax parameters for uptake of the radiolabeled anomers at this temperature are also indistinguishable. The time courses of infinite-cis alpha- and beta-D-glucose uptake at 36.66 degrees C are identical (n = 40). While D-glucose mutarotation is more rapid at this temperature, the anomers of D-glucose are not transported differently by the red cell hexose transfer system.

  11. Effects of Prolonged Glucose Infusion on Insulin Signal Transduction and Glucose Transport in Rat Skeletal Muscle

    OpenAIRE

    Houdali, Basel

    2000-01-01

    Der Effekt einer in vivo Glucoseinfusion auf die Insulinwirkung im Skelettmuskel der Ratte wurde untersucht. Dazu wurden Dauerkatheter in die rechte Jugularvene implantiert und bis zum rechten Vorhof vorgeschoben. Anschließend wurden die Katheter mit einem Perfusor verbunden. Nach einer zweitägigen Erholungsphase vom Operationsstress, wurde 50%ige Glucose mit einer Infusionsrate von 2 ml/h für zwei oder fünf Tage durchgeführt. Die Kontrolltiere erhielten eine 0,45% Kochsalzinfusion für die...

  12. DAPAGLIFLOZIN: SELECTIVE SODIUM-GLUCOSE CO-TRANSPORTER-2 INHIBITOR IN TYPE 2 DIABETES

    Directory of Open Access Journals (Sweden)

    Sudhakar Pemminati

    2011-11-01

    Full Text Available Dapagliflozin is a promising new drug that targets the so far untapped renal glucose reabsorption. By inhibiting sodium-glucose co-transporter-2 (SGLT2 which is mainly localized in the S1 segment of the proximal tubule, Dapagliflozin promotes renal glucose excretion and reduces hyperglycemia in an insulin-independent manner. Dapagliflozin also produces pronounced weight loss which may be an advantage in patients on sulfonylureas and insulin. Dapagliflozin has the potential to be used as monotherapy, as well as in combination with all approved antidiabetic agents.

  13. Atypical antipsychotic drugs directly impair insulin action in adipocytes: effects on glucose transport, lipogenesis, and antilipolysis.

    Science.gov (United States)

    Vestri, Helliner S; Maianu, Lidia; Moellering, Douglas R; Garvey, W Timothy

    2007-04-01

    Treatment with second-generation antipsychotics (SGAs) has been associated with weight gain and the development of diabetes mellitus, although the mechanisms are unknown. We tested the hypothesis that SGAs exert direct cellular effects on insulin action and substrate metabolism in adipocytes. We utilized two cultured cell models including 3T3-L1 adipocytes and primary cultured rat adipocytes, and tested for effects of SGAs risperidone (RISP), clozapine (CLZ), olanzapine (OLZ), and quetiapine (QUE), together with conventional antipsychotic drugs butyrophenone (BUTY), and trifluoperazine (TFP), over a wide concentration range from 1 to 500 microM. The effects of antipsychotic drugs on basal and insulin-stimulated rates of glucose transport were studied at 3 h, 15 h, and 3 days. Both CLZ and OLZ (but not RISP) at doses as low as 5 microM were able to significantly decrease the maximal insulin-stimulated glucose transport rate by approximately 40% in 3T3-L1 cells, whereas CLZ and RISP reduced insulin-stimulated glucose transport rates in primary cultured rat adipocytes by approximately 50-70%. Conventional drugs (BUTY and TFP) did not affect glucose transport rates. Regarding intracellular glucose metabolism, both SGAs (OLZ, QUE, RISP) and conventional drugs (BUTY and TFP) increased basal and/or insulin-stimulated glucose oxidation rates, whereas rates of lipogenesis were increased by CLZ, OLZ, QUE, and BUTY. Finally, rates of lipolysis in response to isoproterenol were reduced by the SGAs (CLZ, OLZ, QUE, RISP), but not by BUTY or TFP. These experiments demonstrate that antipsychotic drugs can differentially affect insulin action and metabolism through direct cellular effects in adipocytes. However, only SGAs were able to impair the insulin-responsive glucose transport system and to impair lipolysis in adipocytes. Thus, SGAs directly induce insulin resistance and alter lipogenesis and lipolysis in favor of progressive lipid accumulation and adipocyte enlargement. These

  14. Differential regulation of glucose transport activity in yeast by specific cAMP signatures.

    Science.gov (United States)

    Bermejo, Clara; Haerizadeh, Farzad; Sadoine, Mayuri S C; Chermak, Diane; Frommer, Wolf B

    2013-06-15

    Successful colonization and survival in variable environments require a competitive advantage during the initial growth phase after experiencing nutrient changes. Starved yeast cells anticipate exposure to glucose by activating the Hxt5p (hexose transporter 5) glucose transporter, which provides an advantage during early phases after glucose resupply. cAMP and glucose FRET (fluorescence resonance energy transfer) sensors were used to identify three signalling pathways that co-operate in the anticipatory Hxt5p activity in glucose-starved cells: as expected the Snf1 (sucrose nonfermenting 1) AMP kinase pathway, but, surprisingly, the sugar-dependent G-protein-coupled Gpr1 (G-protein-coupled receptor 1)/cAMP/PKA (protein kinase A) pathway and the Pho85 (phosphate metabolism 85)/Plc (phospholipase C) 6/7 pathway. Gpr1/cAMP/PKA are key elements of a G-protein-coupled sugar response pathway that produces a transient cAMP peak to induce growth-related genes. A novel function of the Gpr1/cAMP/PKA pathway was identified in glucose-starved cells: during starvation the Gpr1/cAMP/PKA pathway is required to maintain Hxt5p activity in the absence of glucose-induced cAMP spiking. During starvation, cAMP levels remain low triggering expression of HXT5, whereas cAMP spiking leads to a shift to the high capacity Hxt isoforms.

  15. S100b Counteracts Neurodegeneration of Rat Cholinergic Neurons in Brain Slices after Oxygen-Glucose Deprivation

    Directory of Open Access Journals (Sweden)

    Daniela Serbinek

    2010-01-01

    Full Text Available Alzheimer's disease is a severe chronic neurodegenerative disorder characterized by beta-amyloid plaques, tau pathology, cerebrovascular damage, inflammation, reactive gliosis, and cell death of cholinergic neurons. The aim of the present study is to test whether the glia-derived molecule S100b can counteract neurodegeneration of cholinergic neurons after oxygen-glucose deprivation (OGD in organotypic brain slices of basal nucleus of Meynert. Our data showed that 3 days of OGD induced a marked decrease of cholinergic neurons (60% of control, which could be counteracted by 50 μg/mL recombinant S100b. The effect was dose and time dependent. Application of nerve growth factor or fibroblast growth factor-2 was less protective. C-fos-like immunoreactivity was enhanced 3 hours after OGD indicating metabolic stress. We conclude that S100b is a potent neuroprotective factor for cholinergic neurons during ischemic events.

  16. Hypothalamic neuron projection to autonomic preganglionic levels related with glucose metabolism: a fluorescent labelling study in the rat.

    Science.gov (United States)

    Portillo, F; Carrasco, M; Vallo, J J

    1996-06-01

    The location of hypothalamic paraventricular neurons projecting to sympathetic preganglionic levels and related to the autonomic regulation of various organs involved in glucose metabolism (OGM) was determined by ipsilateral injections of two fluorescent tracers, Diamidino Yellow into the left dorsal motor nucleus of the vagus and Fast Blue into the left intermediolateral cell column of the T8-T9 spinal cord. Hypothalamospinal neurons were mainly located in the dorsal part of the paraventricular hypothalamic nucleus (PVH) and the hypothalamobulbar neurons were most abundant in the ventral, medial and extreme lateral parts of the PVH. No double-labelled neurons were found in the hypothalamus. These results can help the knowledge of the neural hypothalamic network related with the autonomic hypothalamic control.

  17. Flozins, inhibitors of type 2 renal sodium-glucose co-transporter – not only antihyperglycemic drugs

    OpenAIRE

    Mizerski Grzegorz; Kicinski Pawel; Jaroszynski Andrzej

    2015-01-01

    The kidneys play a crucial role in the regulation of the carbohydrate metabolism. In normal physiological conditions, the glucose that filters through the renal glomeruli is subsequently nearly totally reabsorbed in the proximal renal tubules. Two transporters are engaged in this process: sodium-glucose co-transporter type 1 (SGLT1), and sodium-glucose co-transporter type type 2 (SGLT2) - this being located in the luminal membrane of the renal tubular epithelial cells. It was found that the a...

  18. The efficient intracellular sequestration of the insulin-regulatable glucose transporter (GLUT-4) is conferred by the NH2 terminus

    OpenAIRE

    1992-01-01

    GLUT-4 is the major facilitative glucose transporter isoform in tissues that exhibit insulin-stimulated glucose transport. Insulin regulates glucose transport by the rapid translocation of GLUT-4 from an intracellular compartment to the plasma membrane. A critical feature of this process is the efficient exclusion of GLUT-4 from the plasma membrane in the absence of insulin. To identify the amino acid domains of GLUT-4 which confer intracellular sequestration, we analyzed the subcellular dist...

  19. PACSIN3 Overexpression Increases Adipocyte Glucose Transport through GLUT1

    Science.gov (United States)

    Roach, William; Plomann, Markus

    2007-01-01

    PACSIN family members regulate intracellular vesicle trafficking via their ability to regulate cytoskeletal rearrangement. These processes are known to be involved in trafficking of GLUT1 and GLUT4 in adipocytes. In this study PACSIN3 was observed to be the only PACSIN isoform that increases in expression during 3T3-L1 adipocyte differentiation. Overexpression of PACSIN3 in 3T3-L1 adipocytes caused an elevation of glucose uptake. Subcellular fractionation revealed that PACSIN3 overexpression elevated GLUT1 plasma membrane localization without effecting GLUT4 distribution. In agreement with this result, examination of GLUT exofacial presentation at the cell surface by photoaffinity labeling revealed significantly increased GLUT1, but not GLUT4, after overexpression of PACSIN3. These results establish a role for PACSIN3 in regulating glucose uptake in adipocytes via its preferential participation in GLUT1 trafficking. They are consistent with the proposal, which is supported by a recent study, that GLUT1, but not GLUT4, is predominantly endocytosed via the coated pit pathway in unstimulated 3T3-L1 adipocytes. PMID:17320047

  20. New mathematical model for fluid-glucose-albumin transport in peritoneal dialysis

    CERN Document Server

    Cherniha, Roman

    2011-01-01

    A mathematical model for fluid transport in peritoneal dialysis is constructed. The model is based on a three-component nonlinear system of two-dimensional partial differential equations for fluid, glucose and albumin transport with the relevant boundary and initial conditions. Non-constant steady-state solutions of the model are studied. The restrictions on the parameters arising in the model are established with the aim to obtain exact formulae for the non-constant steady-state solutions. As the result, the exact formulae for the fluid fluxes from blood to tissue and across the tissue were constructed together with two linear autonomous ODEs for glucose and albumin concentrations. The analytical results were checked for their applicability for the description of fluid-glucose-albumin transport during peritoneal dialysis.

  1. CREB1 regulates glucose transport of glioma cell line U87 by targeting GLUT1.

    Science.gov (United States)

    Chen, Jiaying; Zhang, Can; Mi, Yang; Chen, Fuxue; Du, Dongshu

    2017-06-23

    Glioma is stemmed from the glial cells in the brain, which is accounted for about 45% of all intracranial tumors. The characteristic of glioma is invasive growth, as well as there is no obvious boundary between normal brain tissue and glioma tissue, so it is difficult to resect completely with worst prognosis. The metabolism of glioma is following the Warburg effect. Previous researches have shown that GLUT1, as a glucose transporter carrier, affected the Warburg effect, but the molecular mechanism is not very clear. CREB1 (cAMP responsive element-binding protein1) is involved in various biological processes, and relevant studies confirmed that CREB1 protein regulated the expression of GLUT1, thus mediating glucose transport in cells. Our experiments mainly reveal that the CREB1 could affect glucose transport in glioma cells by regulating the expression of GLUT1, which controlled the metabolism of glioma and affected the progression of glioma.

  2. Decreased insulin action on muscle glucose transport after eccentric contractions in rats

    DEFF Research Database (Denmark)

    Asp, S; Richter, Erik

    1996-01-01

    We have recently shown that eccentric contractions (Ecc) of rat calf muscles cause muscle damage and decreased glycogen and glucose transporter GLUT-4 protein content in the white (WG) and red gastrocnemius (RG) but not in the soleus (S) (S. Asp, S. Kristiansen, and E. A. Richter. J. Appl. Physiol....... 79: 1338-1345, 1995). To study whether these changes affect insulin action, hindlimbs were perfused at three different insulin concentrations (0, 200, and 20,000 microU/ml) 2 days after one-legged eccentric contractions of the calf muscles. Compared with control, basal glucose transport was slightly...... velocity of glycogen synthase increased similarly with increasing insulin concentrations in Ecc- and control WG and RG. We conclude that insulin action on glucose transport but not glycogen synthase activity is impaired in perfused muscle exposed to prior eccentric contractions....

  3. Influence of blood glucose on the expression of glucose transporter proteins 1 and 3 in the brain of diabetic rats

    Institute of Scientific and Technical Information of China (English)

    HOU Wei-kai; FU Chun-li; ZHANG Wen-wen; CHEN Li; XIAN Yu-xin; ZHANG Li; LAI Hong; HOU Xin-guo; XU Yu-xin; YU Ting; XU Fu-yu; SONG Jun

    2007-01-01

    Background The delivery of glucose from the blood to the brain involves its passage across the endothelial cells of the blood-brain barrier (BBB), which is mediated by the facilitative glucose transporter protein 1 (GLUT1), and then across the neural cell membranes, which is mediated by GLUT3. This study aimed to evaluate the dynamic influence of hyperglycemia on the expression of these GLUTs by measuring their expression in the brain at different blood glucose levels in a rat model of diabetes. This might help to determine the proper blood glucose threshold level in the treatment of diabetic apoplexy.Methods Diabetes mellitus was induced with streptozotocin (STZ) in 30 rats. The rats were randomly divided into 3 groups: diabetic group without blood glucose control (group DM1), diabetic rats treated with low dose insulin (group DM2),and diabetic rats treated with high dose insulin (group DM3). The mRNA and protein levels of GLUT1 and GLUT3 were assayed by reverse transcriptase-polymerase chain reaction (RT-PCR) and immunohistochemistry, respectively.Results Compared with normal control rats, the GLUT1 mRNA was reduced by 46.08%, 29.80%, 19.22% (P<0.01) in DM1, DM2, and DM3 group, respectively; and the GLUT3 mRNA was reduced by 75.00%, 46.75%, and 17.89% (P<0.01)in DM1, DM2, and DM3 group, respectively. The abundance of GLUT1 and GLUT3 proteins had negative correlation with the blood glucose level (P<0.01). The density of microvessels in the brain of diabetic rats did not change significantly compared with normal rats.Conclusions Chronic hyperglycemia downregulates GLUT1 and GLUT3 expression at both mRNA and protein levels in the rat brain, which is not due to the decrease of the density of microvessels. The downregulation of GLUT1 and GLUT3 expression might be the adaptive reaction of the body to prevent excessive glucose entering the cell that may lead to cell damage.

  4. Glycogenolysis in astrocytes supports blood-borne glucose channeling not glycogen-derived lactate shuttling to neurons: evidence from mathematical modeling.

    Science.gov (United States)

    DiNuzzo, Mauro; Mangia, Silvia; Maraviglia, Bruno; Giove, Federico

    2010-12-01

    In this article, we examined theoretically the role of human cerebral glycogen in buffering the metabolic requirement of a 360-second brain stimulation, expanding our previous modeling study of neurometabolic coupling. We found that glycogen synthesis and degradation affects the relative amount of glucose taken up by neurons versus astrocytes. Under conditions of 175:115 mmol/L (∼1.5:1) neuronal versus astrocytic activation-induced Na(+) influx ratio, ∼12% of astrocytic glycogen is mobilized. This results in the rapid increase of intracellular glucose-6-phosphate level on stimulation and nearly 40% mean decrease of glucose flow through hexokinase (HK) in astrocytes via product inhibition. The suppression of astrocytic glucose phosphorylation, in turn, favors the channeling of glucose from interstitium to nearby activated neurons, without a critical effect on the concurrent intercellular lactate trafficking. Under conditions of increased neuronal versus astrocytic activation-induced Na(+) influx ratio to 190:65 mmol/L (∼3:1), glycogen is not significantly degraded and blood glucose is primarily taken up by neurons. These results support a role for astrocytic glycogen in preserving extracellular glucose for neuronal utilization, rather than providing lactate to neurons as is commonly accepted by the current 'thinking paradigm'. This might be critical in subcellular domains during functional conditions associated with fast energetic demands.

  5. Unaltered lactate and glucose transporter levels in the MPTP mouse model of Parkinson's disease

    DEFF Research Database (Denmark)

    Puchades, Maja; Sogn, Carl Johan; Maehlen, Jan

    2013-01-01

    BACKGROUND: Metabolic impairment contributes to development of Parkinson's disease (PD). Mitochondrial dysfunction is involved in degeneration of nigral dopamine neurons. Also, in PD there are alterations in glucose metabolism in nigro-striatal pathways, and increased cerebral lactate levels have...... of MCT1, MCT2 and GLUT1 is not changed following dopaminergic neurodegeneration. This is in contrast to findings in other neurodegenerative disease, such as mesial temporal lobe epilepsy, where there are large alterations in MCT levels....

  6. Glucose transporter type 1 deficiency syndrome effectively treated with modified Atkins diet.

    Science.gov (United States)

    Haberlandt, Edda; Karall, Daniela; Jud, Veronika; Baumgartner, Sara Sigl; Zotter, Sibylle; Rostasy, Kevin; Baumann, Matthias; Scholl-Buergi, Sabine

    2014-04-01

    This is a report on the successful treatment of a 6-year-old girl with genetically proven glucose transporter type 1 deficiency syndrome (GLUT1-DS) with modified Atkins diet (MAD). GLUT1-DS is an inborn disorder of glucose transport across the blood-brain barrier, which leads to energy deficiency of the brain with a broad spectrum of neurological symptoms including therapy-resistant epilepsy. Usually classical ketogenic diet (KD) is the standard treatment for patients with GLUT1-DS. Treatment with MAD, a variant of KD, for an observation period of 17 months resulted in improvement of seizures, alertness, cognitive abilities, and electroencephalography in this patient.

  7. Impairment of brain endothelial glucose transporter by methamphetamine causes blood-brain barrier dysfunction

    Directory of Open Access Journals (Sweden)

    Murrin L Charles

    2011-03-01

    Full Text Available Abstract Background Methamphetamine (METH, an addictive psycho-stimulant drug with euphoric effect is known to cause neurotoxicity due to oxidative stress, dopamine accumulation and glial cell activation. Here we hypothesized that METH-induced interference of glucose uptake and transport at the endothelium can disrupt the energy requirement of the blood-brain barrier (BBB function and integrity. We undertake this study because there is no report of METH effects on glucose uptake and transport across the blood-brain barrier (BBB to date. Results In this study, we demonstrate that METH-induced disruption of glucose uptake by endothelium lead to BBB dysfunction. Our data indicate that a low concentration of METH (20 μM increased the expression of glucose transporter protein-1 (GLUT1 in primary human brain endothelial cell (hBEC, main component of BBB without affecting the glucose uptake. A high concentration of 200 μM of METH decreased both the glucose uptake and GLUT1 protein levels in hBEC culture. Transcription process appeared to regulate the changes in METH-induced GLUT1 expression. METH-induced decrease in GLUT1 protein level was associated with reduction in BBB tight junction protein occludin and zonula occludens-1. Functional assessment of the trans-endothelial electrical resistance of the cell monolayers and permeability of dye tracers in animal model validated the pharmacokinetics and molecular findings that inhibition of glucose uptake by GLUT1 inhibitor cytochalasin B (CB aggravated the METH-induced disruption of the BBB integrity. Application of acetyl-L-carnitine suppressed the effects of METH on glucose uptake and BBB function. Conclusion Our findings suggest that impairment of GLUT1 at the brain endothelium by METH may contribute to energy-associated disruption of tight junction assembly and loss of BBB integrity.

  8. NaHS Protects against the Impairments Induced by Oxygen-Glucose Deprivation in Different Ages of Primary Hippocampal Neurons

    Science.gov (United States)

    Yu, Qian; Wang, Binrong; Zhao, Tianzhi; Zhang, Xiangnan; Tao, Lei; Shi, Jinshan; Sun, Xude; Ding, Qian

    2017-01-01

    Brain ischemia leads to poor oxygen supply, and is one of the leading causes of brain damage and/or death. Neuroprotective agents are thus in great need for treatment purpose. Using both young and aged primary cultured hippocampal neurons as in vitro models, we investigated the effect of sodium hydrosulfide (NaHS), an exogenous donor of hydrogen sulfide, on oxygen-glucose deprivation (OGD) damaged neurons that mimick focal cerebral ischemia/reperfusion (I/R) induced brain injury. NaHS treatment (250 μM) protected both young and aged hippocampal neurons, as indicated by restoring number of primary dendrites by 43.9 and 68.7%, number of dendritic end tips by 59.8 and 101.1%, neurite length by 36.8 and 66.7%, and spine density by 38.0 and 58.5% in the OGD-damaged young and aged neurons, respectively. NaHS treatment inhibited growth-associated protein 43 downregulation, oxidative stress in both young and aged hippocampal neurons following OGD damage. Further studies revealed that NaHS treatment could restore ERK1/2 activation, which was inhibited by OGD-induced protein phosphatase 2 (PP2A) upregulation. Our results demonstrated that NaHS has potent protective effects against neuron injury induced by OGD in both young and aged hippocampal neurons.

  9. Neuronal Rap1 Regulates Energy Balance, Glucose Homeostasis, and Leptin Actions

    Directory of Open Access Journals (Sweden)

    Kentaro Kaneko

    2016-09-01

    Full Text Available The CNS contributes to obesity and metabolic disease; however, the underlying neurobiological pathways remain to be fully established. Here, we show that the small GTPase Rap1 is expressed in multiple hypothalamic nuclei that control whole-body metabolism and is activated in high-fat diet (HFD-induced obesity. Genetic ablation of CNS Rap1 protects mice from dietary obesity, glucose imbalance, and insulin resistance in the periphery and from HFD-induced neuropathological changes in the hypothalamus, including diminished cellular leptin sensitivity and increased endoplasmic reticulum (ER stress and inflammation. Furthermore, pharmacological inhibition of CNS Rap1 signaling normalizes hypothalamic ER stress and inflammation, improves cellular leptin sensitivity, and reduces body weight in mice with dietary obesity. We also demonstrate that Rap1 mediates leptin resistance via interplay with ER stress. Thus, neuronal Rap1 critically regulates leptin sensitivity and mediates HFD-induced obesity and hypothalamic pathology and may represent a potential therapeutic target for obesity treatment.

  10. Central action of FGF19 reduces hypothalamic AGRP/NPY neuron activity and improves glucose metabolism.

    Science.gov (United States)

    Marcelin, Geneviève; Jo, Young-Hwan; Li, Xiaosong; Schwartz, Gary J; Zhang, Ying; Dun, Nae J; Lyu, Rong-Ming; Blouet, Clémence; Chang, Jaw K; Chua, Streamson

    2014-02-01

    Tight control of glucose excursions has been a long-standing goal of treatment for patients with type 2 diabetes mellitus in order to ameliorate the morbidity and mortality associated with hyperglycemia. Fibroblast growth factor (FGF) 19 is a hormone-like enterokine released postprandially that emerged as a potential therapeutic agent for metabolic disorders, including diabetes and obesity. Remarkably, FGF19 treatment has hypoglycemic actions that remain potent in models of genetic and acquired insulin resistance. Here, we provided evidence that the central nervous system responds to FGF19 administered in the periphery. Then, in two mouse models of insulin resistance, leptin-deficiency and high-fat diet feeding, third intra-cerebro-ventricular infusions of FGF19 improved glycemic status, reduced insulin resistance and potentiated insulin signaling in the periphery. In addition, our study highlights a new mechanism of central FGF19 action, involving the suppression of AGRP/NPY neuronal activity. Overall, our work unveils novel regulatory pathways induced by FGF19 that will be useful in the design of novel strategies to control diabetes in obesity.

  11. Mitochondrial swelling impairs the transport of organelles in cerebellar granule neurons.

    Science.gov (United States)

    Kaasik, Allen; Safiulina, Dzhamilja; Choubey, Vinay; Kuum, Malle; Zharkovsky, Alexander; Veksler, Vladimir

    2007-11-09

    Organelle transport in neuronal processes is central to the organization, developmental fate, and functions of neurons. Organelles must be transported through the slender, highly branched neuronal processes, making the axonal transport vulnerable to any perturbation. However, some intracellular structures like mitochondria are able to considerably modify their volume. We therefore hypothesized that swollen mitochondria could impair the traffic of other organelles in neurite shafts. To test this hypothesis, we have investigated the effects of mitochondrial swellers on the organelle traffic. Our data demonstrate that treatment of neurons with potassium ionophore valinomycin led to the fast time-dependent inhibition of organelle movement in cerebellar granule neurons. Similar inhibition was observed in neurons treated with the inhibitors of the mitochondrial respiratory chain, sodium azide and antimycin, which also induced swelling. No decrease in the motility of organelles was observed in cultures treated with inhibitors of ATP production or transport, oligomycin or bongkrekic acid, suggesting that inhibition of the ATP-generating activity itself without swelling does not affect the motility of organelles. The effect of swellers on the traffic was more important in thin processes, thus indicating the role of steric hindrance of swollen mitochondria. We propose that the size and morphology of the transported cargo is also relevant for seamless axonal transport and speculate that mitochondrial swelling could be one of the reasons for impaired organelle transport in neuronal processes.

  12. Oxygen/glucose deprivation induces a reduction in synaptic AMPA receptors on hippocampal CA3 neurons mediated by mGluR1 and adenosine A3 receptors.

    OpenAIRE

    Dennis, Siobhan; Jaafari, Nadia; Cimarosti, Helena; Hanley, Jonathan G.; Henley, Jeremy M.; Mellor, Jack R.

    2011-01-01

    Hippocampal CA1 pyramidal neurons are highly sensitive to ischemic damage, whereas neighboring CA3 pyramidal neurons are less susceptible. It is proposed that switching of AMPA receptor (AMPAR) subunits on CA1 neurons during an in vitro model of ischemia, oxygen/glucose deprivation (OGD), leads to an enhanced permeability of AMPARs to Ca2+, resulting in delayed cell death. However, it is unclear whether the same mechanisms exist in CA3 neurons and whether this underlies the differential sensi...

  13. Oxygen/glucose Deprivation Induces a Reduction in Synaptic AMPA Receptors on Hippocampal CA3 Neurons Mediated by mGluR1 and A3 Receptors

    OpenAIRE

    Dennis, Siobhan H.; Jaafari, Nadia; Cimarosti, Helena; Hanley, Jonathan G.; Henley, Jeremy M.; Mellor, Jack R.

    2011-01-01

    Hippocampal CA1 pyramidal neurons are highly sensitive to ischemic damage, whereas neighbouring CA3 pyramidal neurons are less susceptible. It is proposed that switching of AMPA receptor (AMPAR) subunits on CA1 neurons during an in vitro model of ischemia, oxygen/glucose deprivation (OGD), leads to an enhanced permeability of AMPARs to Ca2+ resulting in delayed cell death. However, it is unclear if the same mechanisms exist in CA3 neurons and whether this underlies the differential sensitivit...

  14. Glucose Transporter 4 (GLUT4) is Not Necessary for Overload-Induced Glucose Uptake or Hypertrophic Growth in Mouse Skeletal Muscle.

    Science.gov (United States)

    McMillin, Shawna L; Schmidt, Denise L; Kahn, Barbara B; Witczak, Carol A

    2017-03-09

    Glucose transporter 4 (GLUT4) is necessary for acute insulin- and contraction-induced skeletal muscle glucose uptake, but its role in chronic muscle loading (overload)-induced glucose uptake is unknown. Our goal was to determine if GLUT4 is required for overload-induced glucose uptake. Overload was induced in mouse plantaris muscle by unilateral synergist ablation. After 5 days, muscle weights and ex vivo [(3)H]-2-deoxy-D-glucose uptake were assessed. Overload-induced muscle glucose uptake and hypertrophic growth were not impaired in muscle-specific GLUT4 knockout mice, demonstrating that GLUT4 is not necessary for these processes. To assess which transporter(s) mediate overload-induced glucose uptake, chemical inhibitors were utilized. The facilitative GLUT inhibitor, cytochalasin B, but not the sodium-dependent glucose-co-transport inhibitor, phloridzin, prevented overload-induced uptake demonstrating that GLUT(s) mediate this effect. To assess which GLUT, hexose competition experiments were performed. Overload-induced [(3)H]-2-deoxy-D-glucose uptake was not inhibited by D-fructose, demonstrating that the fructose-transporting GLUT2, GLUT5, GLUT8, and GLUT12, do not mediate this effect. To assess additional GLUTs, immunoblots were performed. Overload increased GLUT1, GLUT3, GLUT6 and GLUT10 protein levels 2- to 5-fold. Collectively, these results demonstrate that GLUT4 is not necessary for overload-induced muscle glucose uptake or hypertrophic growth, and suggest that GLUT1, GLUT3, GLUT6 and/or GLUT10 mediate overload-induced glucose uptake.

  15. Role of Akt substrate of 160 kDa in insulin-stimulated and contraction-stimulated glucose transport

    DEFF Research Database (Denmark)

    Cartee, Gregory D; Wojtaszewski, Jørgen F P

    2007-01-01

    Insulin and exercise, the most important physiological stimuli to increase glucose transport in skeletal muscle, trigger a redistribution of GLUT4 glucose transporter proteins from the cell interior to the cell surface, thereby increasing glucose transport capacity. The most distal insulin...... signaling protein that has been linked to GLUT4 translocation, Akt substrate of 160 kDa (AS160), becomes phosphorylated in insulin-stimulated 3T3-L1 adipocytes; this is important for insulin-stimulated GLUT4 translocation and glucose transport. Insulin also induces a rapid and dose-dependent increase in AS....../contraction-stimulated glucose uptake is currently inconclusive. The distinct signaling pathways that are stimulated by insulin and exercise/contraction converge at AS160. Although AS160 phosphorylation is apparently important for insulin-stimulated GLUT4 translocation and glucose transport, it is uncertain whether elevated AS...

  16. Tryptic digestion of the human erythrocyte glucose transporter: effects on ligand binding and tryptophan fluorescence.

    Science.gov (United States)

    May, J M; Qu, Z C; Beechem, J M

    1993-09-21

    The conformation of the human erythrocyte glucose transport protein has been shown to determine its susceptibility to enzymatic cleavage on a large cytoplasmic loop. We took the converse approach and investigated the effects of tryptic digestion on the conformational structure of this protein. Exhaustive tryptic digestion of protein-depleted erythrocyte ghosts decreased the affinity of the residual transporter for cytochalasin B by 3-fold but did not affect the total number of binding sites. Tryptic digestion also increased the affinity of the residual transporter for D-glucose and inward-binding sugar phenyl beta-D-glucopyranoside but decreased that for the outward-binding 4,6-O-ethylidene glucose. These results suggest that tryptic cleavage stabilized the remaining transporter in an inward-facing conformation, but one with decreased affinity for cytochalasin B. The steady-state fluorescence emission scan of the purified reconstituted glucose transport protein was unaffected by tryptic digestion. Addition of increasing concentrations of potassium iodide resulted in linear Stern-Volmer plots, which were also unaffected by prior tryptic digestion. The tryptophan oxidant N-bromosuccinimide was investigated to provide a more sensitive measure of tryptophan environment. This agent irreversibly inhibited 3-O-methylglucose transport in intact erythrocytes and cytochalasin B binding in protein-depleted ghosts, with a half-maximal effect observed for each activity at about 0.3-0.4 nM. Treatment of purified glucose transport protein with N-bromosuccinimide resulted in a time-dependent quench of tryptophan fluorescence, which was resolved into two components by nonlinear regression using global analysis. Tryptic digestion retarded the rate of oxidation of the more slowly reacting class of tryptophans. (ABSTRACT TRUNCATED AT 250 WORDS)

  17. Volume regulated anion channel currents of rat hippocampal neurons and their contribution to oxygen-and-glucose deprivation induced neuronal death.

    Directory of Open Access Journals (Sweden)

    Huaqiu Zhang

    Full Text Available Volume-regulated anion channels (VRAC are widely expressed chloride channels that are critical for the cell volume regulation. In the mammalian central nervous system, the physiological expression of neuronal VRAC and its role in cerebral ischemia are issues largely unknown. We show that hypoosmotic medium induce an outwardly rectifying chloride conductance in CA1 pyramidal neurons in rat hippocampal slices. The induced chloride conductance was sensitive to some of the VRAC inhibitors, namely, IAA-94 (300 µM and NPPB (100 µM, but not to tamoxifen (10 µM. Using oxygen-and-glucose deprivation (OGD to simulate ischemic conditions in slices, VRAC activation appeared after OGD induced anoxic depolarization (AD that showed a progressive increase in current amplitude over the period of post-OGD reperfusion. The OGD induced VRAC currents were significantly inhibited by inhibitors for glutamate AMPA (30 µM NBQX and NMDA (40 µM AP-5 receptors in the OGD solution, supporting the view that induction of AD requires an excessive Na(+-loading via these receptors that in turn to activate neuronal VRAC. In the presence of NPPB and DCPIB in the post-OGD reperfusion solution, the OGD induced CA1 pyramidal neuron death, as measured by TO-PRO-3-I staining, was significantly reduced, although DCPIB did not appear to be an effective neuronal VRAC blocker. Altogether, we show that rat hippocampal pyramidal neurons express functional VRAC, and ischemic conditions can initial neuronal VRAC activation that may contribute to ischemic neuronal damage.

  18. dnc-1/dynactin 1 knockdown disrupts transport of autophagosomes and induces motor neuron degeneration.

    Science.gov (United States)

    Ikenaka, Kensuke; Kawai, Kaori; Katsuno, Masahisa; Huang, Zhe; Jiang, Yue-Mei; Iguchi, Yohei; Kobayashi, Kyogo; Kimata, Tsubasa; Waza, Masahiro; Tanaka, Fumiaki; Mori, Ikue; Sobue, Gen

    2013-01-01

    Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease characterized by the progressive loss of motor neurons. We previously showed that the expression of dynactin 1, an axon motor protein regulating retrograde transport, is markedly reduced in spinal motor neurons of sporadic ALS patients, although the mechanisms by which decreased dynactin 1 levels cause neurodegeneration have yet to be elucidated. The accumulation of autophagosomes in degenerated motor neurons is another key pathological feature of sporadic ALS. Since autophagosomes are cargo of dynein/dynactin complexes and play a crucial role in the turnover of several organelles and proteins, we hypothesized that the quantitative loss of dynactin 1 disrupts the transport of autophagosomes and induces the degeneration of motor neuron. In the present study, we generated a Caenorhabditis elegans model in which the expression of DNC-1, the homolog of dynactin 1, is specifically knocked down in motor neurons. This model exhibited severe motor defects together with axonal and neuronal degeneration. We also observed impaired movement and increased number of autophagosomes in the degenerated neurons. Furthermore, the combination of rapamycin, an activator of autophagy, and trichostatin which facilitates axonal transport dramatically ameliorated the motor phenotype and axonal degeneration of this model. Thus, our results suggest that decreased expression of dynactin 1 induces motor neuron degeneration and that the transport of autophagosomes is a novel and substantial therapeutic target for motor neuron degeneration.

  19. dnc-1/dynactin 1 knockdown disrupts transport of autophagosomes and induces motor neuron degeneration.

    Directory of Open Access Journals (Sweden)

    Kensuke Ikenaka

    Full Text Available Amyotrophic lateral sclerosis (ALS is a fatal neurodegenerative disease characterized by the progressive loss of motor neurons. We previously showed that the expression of dynactin 1, an axon motor protein regulating retrograde transport, is markedly reduced in spinal motor neurons of sporadic ALS patients, although the mechanisms by which decreased dynactin 1 levels cause neurodegeneration have yet to be elucidated. The accumulation of autophagosomes in degenerated motor neurons is another key pathological feature of sporadic ALS. Since autophagosomes are cargo of dynein/dynactin complexes and play a crucial role in the turnover of several organelles and proteins, we hypothesized that the quantitative loss of dynactin 1 disrupts the transport of autophagosomes and induces the degeneration of motor neuron. In the present study, we generated a Caenorhabditis elegans model in which the expression of DNC-1, the homolog of dynactin 1, is specifically knocked down in motor neurons. This model exhibited severe motor defects together with axonal and neuronal degeneration. We also observed impaired movement and increased number of autophagosomes in the degenerated neurons. Furthermore, the combination of rapamycin, an activator of autophagy, and trichostatin which facilitates axonal transport dramatically ameliorated the motor phenotype and axonal degeneration of this model. Thus, our results suggest that decreased expression of dynactin 1 induces motor neuron degeneration and that the transport of autophagosomes is a novel and substantial therapeutic target for motor neuron degeneration.

  20. Mammalian glucose permease GLUT1 facilitates transport of arsenic trioxide and methylarsonous acid†

    Science.gov (United States)

    Liu, Zijuan; Sanchez, Marco A.; Jiang, Xuan; Boles, Eckhard; Landfear, Scott M.; Rosen, Barry P.

    2006-01-01

    Arsenic exposure is associated with hypertension, diabetes and cancer. Some mammals methylate arsenic. Saccharomyces cerevisiae hexose permeases catalyze As(OH)3 uptake. Here we report that mammalian glucose transporter GLUT1 catalyzes As(OH)3 and CH3As(OH)2 uptake in yeast or in Xenopus laevis öocytes. Expression of GLUT1 in a yeast lacking other glucose transporters allows for growth on glucose. Yeast expressing yeast HXT1 or rat GLUT1 transport As(OH)3 and CH3As(OH)2. The Km of GLUT1 is to 1.2 mM for CH3As(OH)2, compared to a Km of 3 mM for glucose. Inhibition between glucose and CH3As(OH)2 is noncompetitive, suggesting differences between the translocation pathways of hexoses and arsenicals. Both human and rat GLUT1 catalyze uptake of both As(OH)3 and CH3As(OH)2 in öocytes. Thus GLUT1 may be a major pathway uptake of both inorganic and methylated arsenicals in erythrocytes or the epithelial cells of the blood-brain barrier, contributing to arsenic-related cardiovascular problems and neurotoxicity. PMID:17064664

  1. Pathogenic mutations causing glucose transport defects in GLUT1 transporter: The role of intermolecular forces in protein structure-function.

    Science.gov (United States)

    Raja, Mobeen; Kinne, Rolf K H

    2015-01-01

    Two families of glucose transporter - the Na(+)-dependent glucose cotransporter-1 (SGLT family) and the facilitated diffusion glucose transporter family (GLUT family) - play a crucial role in the translocation of glucose across the epithelial cell membrane. How genetic mutations cause life-threatening diseases like GLUT1-deficiency syndrome (GLUT1-DS) is not well understood. In this review, we have combined previous functional data with our in silico analyses of the bacterial homologue of GLUT members, XylE (an outward-facing, partly occluded conformation) and previously proposed GLUT1 homology model (an inward-facing conformation). A variety of native and mutant side chain interactions were modeled to highlight the potential roles of mutations in destabilizing protein-protein interaction hence triggering structural and functional defects. This study sets the stage for future studies of the structural properties that mediate GLUT1 dysfunction and further suggests that both SGLT and GLUT families share conserved domains that stabilize the transporter structure/function via a similar mechanism.

  2. A tale of two glucose transporters: how GLUT2 re-emerged as a contender for glucose transport into the human beta cell.

    Science.gov (United States)

    van de Bunt, M; Gloyn, A L

    2012-09-01

    Finding novel causes for monogenic forms of diabetes is important as, alongside the clinical implications of such a discovery, it can identify critical proteins and pathways required for normal beta cell function in humans. It is increasingly apparent that there are significant differences between rodent and human islets. One example that has generated interest is the relative importance of the glucose transporter GLUT2 in rodent and human beta cells. The central role of GLUT2 in rodent beta cells is well established, but a number of studies have suggested that other glucose transporters, namely GLUT1 and GLUT3, may play an important role in facilitating glucose transport into human beta cells. In this issue of Diabetologia Sansbury et al (DOI: 10.1007/s00125-012-2595-0 ) report homozygous loss of function mutations in SLC2A2, which encodes GLUT2, as a rare cause of neonatal diabetes. Evidence for a beta cell defect in these subjects comes from very low birthweights, lack of endogenous insulin secretion and a requirement for insulin therapy. Neonatal diabetes is not a consistent feature of SLC2A2 mutations. It is only found in a small percentage of cases (~4%) and the diabetes largely resolves before 18 months of age. This discovery is significant as it suggests that GLUT2 plays an important role in human beta cells, but the interplay and relative roles of other transporters differ from those in rodents. This finding should encourage efforts to delineate the precise role of GLUT2 in the human beta cell at different developmental time points and is a further reminder of critical differences between human and rodent islets.

  3. Glucose uptake and transport in contracting, perfused rat muscle with different pre-contraction glycogen concentrations

    DEFF Research Database (Denmark)

    Hespel, P; Richter, Erik

    1990-01-01

    1. Glucose uptake and transport, muscle glycogen, free glucose and glucose-6-phosphate concentrations were studied in perfused resting and contracting rat skeletal muscle with different pre-contraction glycogen concentrations. Rats were pre-conditioned by a combination of swimming exercise and diet......, resulting in either low (glycogen-depleted rats), normal (control rats) or high (supercompensated rats) muscle glycogen concentrations at the time their hindlimbs were perfused. 2. Compared with control rats, pre-contraction muscle glycogen concentration was approximately 40% lower in glycogen-depleted rats......, whereas it was 40% higher in supercompensated rats. Muscle glycogen break-down correlated positively (r = 0.76; P less than 0.001) with pre-contraction muscle glycogen concentration. 3. Glucose uptake during contractions was approximately 50% higher in glycogen-depleted hindquarters than in control...

  4. Phospho-Rb mediating cell cycle reentry induces early apoptosis following oxygen-glucose deprivation in rat cortical neurons.

    Science.gov (United States)

    Yu, Ying; Ren, Qing-Guo; Zhang, Zhao-Hui; Zhou, Ke; Yu, Zhi-Yuan; Luo, Xiang; Wang, Wei

    2012-03-01

    The aim of this study was to investigate the relationship between cell cycle reentry and apoptosis in cultured cortical neurons following oxygen-glucose deprivation (OGD). We found that the percentage of neurons with BrdU uptake, TUNEL staining, and colocalized BrdU uptake and TUNEL staining was increased relative to control 6, 12 and 24 h after 1 h of OGD. The number of neurons with colocalized BrdU and TUNEL staining was decreased relative to the number of TUNEL-positive neurons at 24 h. The expression of phosphorylated retinoblastoma protein (phospho-Rb) was significantly increased 6, 12 and 24 h after OGD, parallel with the changes in BrdU uptake. Phospho-Rb and TUNEL staining were colocalized in neurons 6 and 12 h after OGD. This colocalization was strikingly decreased 24 h after OGD. Treatment with the cyclin-dependent kinase inhibitor roscovitine (100 μM) decreased the expression of phospho-Rb and reduced neuronal apoptosis in vitro. These results demonstrated that attempted cell cycle reentry with phosphorylation of Rb induce early apoptosis in neurons after OGD and there must be other mechanisms involved in the later stages of neuronal apoptosis besides cell cycle reentry. Phosphoralated Rb may be an important factor which closely associates aberrant cell cycle reentry with the early stages of neuronal apoptosis following ischemia/hypoxia in vitro, and pharmacological interventions for neuroprotection may be useful directed at this keypoint.

  5. Heat stress reduces intestinal barrier integrity and favors intestinal glucose transport in growing pigs.

    Science.gov (United States)

    Pearce, Sarah C; Mani, Venkatesh; Boddicker, Rebecca L; Johnson, Jay S; Weber, Thomas E; Ross, Jason W; Rhoads, Robert P; Baumgard, Lance H; Gabler, Nicholas K

    2013-01-01

    Excessive heat exposure reduces intestinal integrity and post-absorptive energetics that can inhibit wellbeing and be fatal. Therefore, our objectives were to examine how acute heat stress (HS) alters intestinal integrity and metabolism in growing pigs. Animals were exposed to either thermal neutral (TN, 21°C; 35-50% humidity; n=8) or HS conditions (35°C; 24-43% humidity; n=8) for 24 h. Compared to TN, rectal temperatures in HS pigs increased by 1.6°C and respiration rates by 2-fold (Pintestinal integrity was compromised in the HS pigs (ileum and colon TER decreased; PIntestinal permeability was accompanied by an increase in protein expression of myosin light chain kinase (PIntestinal glucose transport and blood glucose were elevated due to HS (Pintestinal integrity and increase intestinal stress and glucose transport.

  6. Regulation of human trophoblast GLUT1 glucose transporter by insulin-like growth factor I (IGF-I.

    Directory of Open Access Journals (Sweden)

    Marc U Baumann

    Full Text Available Glucose transport to the fetus across the placenta takes place via glucose transporters in the opposing faces of the barrier layer, the microvillous and basal membranes of the syncytiotrophoblast. While basal membrane content of the GLUT1 glucose transporter appears to be the rate-limiting step in transplacental transport, the factors regulating transporter expression and activity are largely unknown. In view of the many studies showing an association between IGF-I and fetal growth, we investigated the effects of IGF-I on placental glucose transport and GLUT1 transporter expression. Treatment of BeWo choriocarcinoma cells with IGF-I increased cellular GLUT1 protein. There was increased basolateral (but not microvillous uptake of glucose and increased transepithelial transport of glucose across the BeWo monolayer. Primary syncytial cells treated with IGF-I also demonstrated an increase in GLUT1 protein. Term placental explants treated with IGF-I showed an increase in syncytial basal membrane GLUT1 but microvillous membrane GLUT1 was not affected. The placental dual perfusion model was used to assess the effects of fetally perfused IGF-I on transplacental glucose transport and syncytial GLUT1 content. In control perfusions there was a decrease in transplacental glucose transport over the course of the perfusion, whereas in tissues perfused with IGF-I through the fetal circulation there was no change. Syncytial basal membranes from IGF-I perfused tissues showed an increase in GLUT1 content. These results demonstrate that IGF-I, whether acting via microvillous or basal membrane receptors, increases the basal membrane content of GLUT1 and up-regulates basal membrane transport of glucose, leading to increased transepithelial glucose transport. These observations provide a partial explanation for the mechanism by which IGF-I controls nutrient supply in the regulation of fetal growth.

  7. Glucose transporter-1 (GLUT1) deficiency syndrome: diagnosis and treatment in late childhood.

    NARCIS (Netherlands)

    Gramer, G.; Wolf, N.I.; Vater, D.; Bast, T.; Santer, R.; Kamsteeg, E.J.; Wevers, R.A.; Ebinger, F.

    2012-01-01

    BACKGROUND: Typical cases of glucose transporter-1 deficiency syndrome (GLUT1-DS) present with early-onset epilepsy. We report symptoms, diagnostic results, and effects of therapy in two patients diagnosed with GLUT1-DS at the age of 10 and 15 years, respectively. PATIENTS: Patient 1: After four cer

  8. Wortmannin inhibits both insulin- and contraction-stimulated glucose uptake and transport in rat skeletal muscle

    DEFF Research Database (Denmark)

    Wojtaszewski, Jørgen; Hansen, B F; Ursø, Birgitte

    1996-01-01

    The role of phosphatidylinositol (PI) 3-kinase for insulin- and contraction-stimulated muscle glucose transport was investigated in rat skeletal muscle perfused with a cell-free perfusate. The insulin receptor substrate-1-associated PI 3-kinase activity was increased sixfold upon insulin stimulat...

  9. Glucose transporter-1 (GLUT1) deficiency syndrome: diagnosis and treatment in late childhood.

    NARCIS (Netherlands)

    Gramer, G.; Wolf, N.I.; Vater, D.; Bast, T.; Santer, R.; Kamsteeg, E.J.; Wevers, R.A.; Ebinger, F.

    2012-01-01

    BACKGROUND: Typical cases of glucose transporter-1 deficiency syndrome (GLUT1-DS) present with early-onset epilepsy. We report symptoms, diagnostic results, and effects of therapy in two patients diagnosed with GLUT1-DS at the age of 10 and 15 years, respectively. PATIENTS: Patient 1: After four cer

  10. Glucose transporter-1 (GLUT1) deficiency syndrome: diagnosis and treatment in late childhood.

    NARCIS (Netherlands)

    Gramer, G.; Wolf, N.I.; Vater, D.; Bast, T.; Santer, R.; Kamsteeg, E.J.; Wevers, R.A.; Ebinger, F.

    2012-01-01

    BACKGROUND: Typical cases of glucose transporter-1 deficiency syndrome (GLUT1-DS) present with early-onset epilepsy. We report symptoms, diagnostic results, and effects of therapy in two patients diagnosed with GLUT1-DS at the age of 10 and 15 years, respectively. PATIENTS: Patient 1: After four

  11. Neuronal Cell Death Induced by Mechanical Percussion Trauma in Cultured Neurons is not Preceded by Alterations in Glucose, Lactate and Glutamine Metabolism

    Science.gov (United States)

    Jayakumar, A. R.; Bak, L. K.; Rama Rao, K. V.; Waagepetersen, H.S.; Schousboe, A.; Norenberg, M.D.

    2016-01-01

    Traumatic brain injury (TBI) is a devastating neurological disorder that usually presents in acute and chronic forms. Brain edema and associated increased intracranial pressure in the early phase following TBI are major consequences of acute trauma. On the other hand, neuronal injury, leading to neurobehavioral and cognitive impairments, that usually develop months to years after single or repetitive episodes of head trauma, are major consequences of chronic TBI. The molecular mechanisms responsible for TBI-induced injury, however, are unclear. Recent studies have suggested that early mitochondrial dysfunction and subsequent energy failure play a role in the pathogenesis of TBI. We therefore examined whether oxidative metabolism of 13C-labeled glucose, lactate or glutamine is altered early following in vitro mechanical percussion-induced trauma (5 atm) to neurons (4–24 h), and whether such events contribute to the development of neuronal injury. Cell viability was assayed using the release of the cytoplasmic enzyme lactate dehydrogenase (LDH), together with fluorescence-based cell staining (calcein and ethidium homodimer-1 for live and dead cells, respectively). Trauma had no effect on the LDH release in neurons from 1 h to 18 h. However, a significant increase in LDH release was detected at 24 h after trauma. Similar findings were identified when traumatized neurons were stained with fluorescent markers. Additionally 13C-labeling of glutamate showed a small, but statistically significant decrease at 14 h after trauma. However, trauma had no effect on the cycling ratio of the TCA cycle at any time-period examined. These findings indicate that trauma does not cause a disturbance in oxidative metabolism of any of the substrates used for neurons. Accordingly, such metabolic disturbance does not appear to contribute to the neuronal death in the early stages following trauma. PMID:26729365

  12. Morinda citrifolia fruit juice prevents ischemic neuronal damage through suppression of the development of post-ischemic glucose intolerance.

    Science.gov (United States)

    Harada, Shinichi; Fujita-Hamabe, Wakako; Kamiya, Kohei; Mizushina, Yoshiyuki; Satake, Toshiko; Tokuyama, Shogo

    2010-10-01

    Fruit juice of Morinda citrifolia (Noni juice) is a well-known health drink and has various pharmacological properties including antioxidant and anti-inflammatory effects. We have hitherto found the protective effect of Noni juice on brain damage caused by ischemic stress in mice. In addition, we also recently reported that regulation of post-ischemic glucose intolerance might be important for good prognosis. Here, we focused on the effect of Noni juice on the development of the post-ischemic glucose intolerance as a cerebral protective mechanism. Noni juice was obtained from the mature fruit grown in Okinawa (about 1.5 L/4 kg of fruit; 100% ONJ). Male ddY mice were given 10% ONJ in drinking water for 7 days. Then, mice were subjected to 2 h of middle cerebral artery occlusion (MCAO). Ingestion of 10% ONJ suppressed the development of neuronal damage after MCAO. Interestingly, glucose intolerance observed on the 1st day after MCAO completely disappeared after 10% ONJ administration. Furthermore, ONJ treatment significantly increased serum insulin levels much further than the control group on the 1st day, while serum adiponectin levels were not affected at all. These results suggest that ONJ could facilitate insulin secretion after ischemic stress and may attenuate the development of glucose intolerance. These mechanisms may contribute to the neuronal protective effect of ONJ against ischemic stress.

  13. Protective mechanisms of microRNA-27a against oxygen-glucose deprivation-induced injuries in hippocampal neurons

    Institute of Scientific and Technical Information of China (English)

    Qun Cai; Ting Wang; Wen-jie Yang; Xing Fen

    2016-01-01

    Hypoxic injuries during fetal distress have been shown to cause reduced expression of microRNA-27a (miR-27a), which regulates sensi-tivity of cortical neurons to apoptosis. We hypothesized that miR-27a overexpression attenuates hypoxia-and ischemia-induced neuronal apoptosis by regulating FOXO1, an important transcription factor for regulating the oxidative stress response. miR-27a mimic was transfected into hippocampal neurons to overexpress miR-27a. Results showed increased hippocampal neuronal viability and decreased caspase-3 ex-pression. The luciferase reporter gene system demonstrated that miR-27a directly binded to FOXO1 3′UTR in hippocampal neurons and inhibited FOXO1 expression, suggesting that FOXO1 was the target gene for miR-27a. These ifndings conifrm that miR-27a protects hippo-campal neurons against oxygen-glucose deprivation-induced injuries. The mechanism might be mediated by modulation of FOXO1 and apoptosis-related gene caspase-3 expression.

  14. Taurine Protected Against the Impairments of Neural Stem Cell Differentiated Neurons Induced by Oxygen-Glucose Deprivation.

    Science.gov (United States)

    Xiao, Bo; Liu, Huazhen; Gu, Zeyun; Liu, Sining; Ji, Cheng

    2015-11-01

    Cell transplantation of neural stem cells (NSCs) is a promising approach for neurological recovery both structurally and functionally. However, one big obstacle is to promote differentiation of NSCs into neurons and the followed maturation. In the present study, we aimed to investigate the protective effect of taurine on the differentiation of NSCs and subsequent maturation of their neuronal lineage, when exposed to oxygen-glucose deprivation (OGD). The results suggested that taurine (5-20 mM) promoted the viability and proliferation of NSCs, and it protected against 8 h of OGD induced impairments. Furthermore, 20 mM taurine promoted NSCs to differentiate into neurons after 7 days of culture, and it also protected against the suppressive impairments of 8 h of OGD. Consistently, taurine (20 mM) promoted the neurite sprouting and outgrowth of the NSC differentiated neurons after 14 days of differentiation, which were significantly inhibited by OGD (8 h). At D21, the mushroom spines and spine density were promoted or restored by 20 mM taurine. Taken together, the enhanced viability and proliferation of NSCs, more differentiated neurons and the promoted maturation of neurons by 20 mM taurine support its therapeutic application during stem cell therapy to enhance neurological recovery. Moreover, it protected against the impairments induced by OGD, which may highlight its role for a more direct therapeutic application especially in an ischemic stroke environment.

  15. Silibinin activates AMP-activated protein kinase to protect neuronal cells from oxygen and glucose deprivation-re-oxygenation.

    Science.gov (United States)

    Xie, Zhi; Ding, Sheng-quan; Shen, Ya-fang

    2014-11-14

    In this study, we explored the cytoprotective potential of silibinin against oxygen-glucose deprivation (OGD)-induced neuronal cell damages, and studied underling mechanisms. In vitro model of ischemic stroke was created by keeping neuronal cells (SH-SY5Y cells and primary mouse cortical neurons) in an OGD condition followed by re-oxygenation. Pre-treatment of silibinin significantly inhibited OGD/re-oxygenation-induced necrosis and apoptosis of neuronal cells. OGD/re-oxygenation-induced reactive oxygen species (ROS) production and mitochondrial membrane potential (MMP) reduction were also inhibited by silibinin. At the molecular level, silibinin treatment in SH-SY5Y cells and primary cortical neurons led to significant AMP-activated protein kinase (AMPK) signaling activation, detected by phosphorylations of AMPKα1, its upstream kinase liver kinase B1 (LKB1) and the downstream target acetyl-CoA Carboxylase (ACC). Pharmacological inhibition or genetic depletion of AMPK alleviated the neuroprotective ability of silibinin against OGD/re-oxygenation. Further, ROS scavenging ability by silibinin was abolished with AMPK inhibition or silencing. While A-769662, the AMPK activator, mimicked silibinin actions and suppressed ROS production and neuronal cell death following OGD/re-oxygenation. Together, these results show that silibinin-mediated neuroprotection requires activation of AMPK signaling.

  16. Regulation of glucose transport by ROCK1 differs from that of ROCK2 and is controlled by actin polymerization.

    Science.gov (United States)

    Chun, Kwang-Hoon; Araki, Kazushi; Jee, Yuna; Lee, Dae-Ho; Oh, Byung-Chul; Huang, Hu; Park, Kyong Soo; Lee, Sam W; Zabolotny, Janice M; Kim, Young-Bum

    2012-04-01

    A role of Rho-associated coiled-coil-containing protein kinase (ROCK)1 in regulating whole-body glucose homeostasis has been reported. However, cell-autonomous effects of ROCK1 on insulin-dependent glucose transport in adipocytes and muscle cells have not been elucidated. To determine the specific role of ROCK1 in glucose transport directly, ROCK1 expression in 3T3-L1 adipocytes and L6 myoblasts was biologically modulated. Here, we show that small interfering RNA-mediated ROCK1 depletion decreased insulin-induced glucose transport in adipocytes and myoblasts, whereas adenovirus-mediated ROCK1 expression increased this in a dose-dependent manner, indicating that ROCK1 is permissive for glucose transport. Inhibition of ROCK1 also impaired glucose transporter 4 translocation in 3T3-L1 adipocytes. Importantly, the ED₅₀ of insulin for adipocyte glucose transport was reduced when ROCK1 was expressed, leading to hypersensitivity to insulin. These effects are dependent on actin cytoskeleton remodeling, because inhibitors of actin polymerization significantly decreased ROCK1's effect to promote insulin-stimulated glucose transport. Unlike ROCK2, ROCK1 binding to insulin receptor substrate (IRS)-1 was not detected by immunoprecipitation, although cell fractionation demonstrated both ROCK isoforms localize with IRS-1 in low-density microsomes. Moreover, insulin's ability to increase IRS-1 tyrosine 612 and serine 632/635 phosphorylation was attenuated by ROCK1 suppression. Replacing IRS-1 serine 632/635 with alanine reduced insulin-stimulated phosphatidylinositol 3-kinase activation and glucose transport in 3T3-L1 adipocytes, indicating that phosphorylation of these serine residues of IRS-1, which are substrates of the ROCK2 isoform in vitro, are crucial for maximal stimulation of glucose transport by insulin. Our studies identify ROCK1 as an important positive regulator of insulin action on glucose transport in adipocytes and muscle cells.

  17. The Structure of a Sugar Transporter of the Glucose EIIC Superfamily Provides Insight into the Elevator Mechanism of Membrane Transport.

    Science.gov (United States)

    McCoy, Jason G; Ren, Zhenning; Stanevich, Vitali; Lee, Jumin; Mitra, Sharmistha; Levin, Elena J; Poget, Sebastien; Quick, Matthias; Im, Wonpil; Zhou, Ming

    2016-06-07

    The phosphoenolpyruvate:carbohydrate phosphotransferase systems are found in bacteria, where they play central roles in sugar uptake and regulation of cellular uptake processes. Little is known about how the membrane-embedded components (EIICs) selectively mediate the passage of carbohydrates across the membrane. Here we report the functional characterization and 2.55-Å resolution structure of a maltose transporter, bcMalT, belonging to the glucose superfamily of EIIC transporters. bcMalT crystallized in an outward-facing occluded conformation, in contrast to the structure of another glucose superfamily EIIC, bcChbC, which crystallized in an inward-facing occluded conformation. The structures differ in the position of a structurally conserved substrate-binding domain that is suggested to play a central role in sugar transport. In addition, molecular dynamics simulations suggest a potential pathway for substrate entry from the periplasm into the bcMalT substrate-binding site. These results provide a mechanistic framework for understanding substrate recognition and translocation for the glucose superfamily EIIC transporters.

  18. Neuronal Logistics : Axonal Transport in Development and Disease

    NARCIS (Netherlands)

    R. van den Berg (Robert)

    2016-01-01

    markdownabstractBrain cells are uniquely shaped among the many cell types of the body. While most cells are more or less rounded or square-shaped, neurons grow one or more long axons that can reach lengths of a meter or more. To keep these axons alive and functional, neurons are dependent on an intr

  19. Sucrose nonfermenting AMPK-related kinase (SNARK) mediates contraction-stimulated glucose transport in mouse skeletal muscle

    DEFF Research Database (Denmark)

    Koh, Ho-Jin; Toyoda, Taro; Fujii, Nobuharu;

    2010-01-01

    . Whole-body SNARK heterozygotic knockout mice also had impaired contraction-stimulated glucose transport in skeletal muscle, and knockdown of SNARK in C2C12 muscle cells impaired sorbitol-stimulated glucose transport. SNARK is activated by muscle contraction and is a unique mediator of contraction...

  20. Peritoneal transport dynamics of glucose and icodextrin: the in vitro comparative studies.

    Science.gov (United States)

    Czyzewska, Krystyna; Szary, Beata; Grzelak, Teresa

    2005-01-01

    We performed in vitro experiments with the isolated rabbit parietal peritoneum to evaluate the importance of fluid stirring intensification and of chemical modification of mesothelium and interstitium to the peritoneal transport of glucose and icodextrin. We used a mathematical model of mass transport to calculate the diffusive permeability coefficient, P, in centimeters per second. In control conditions (intact tissue; stirring rate: 11 mL/min), the rate of glucose (2.0 g/dL) transfer remained constant, and no differences were observed for transport from the interstitial to the mesothelial (I-->M) side of the membrane or in the opposite direction (M-->I). The value of P (+/- standard error of the mean) was 2.731 +/- 0.472 x 10(-4) cm/s. In contrast, the icodextrin (7.5 g/dL) I-->M transport rate was higher than that for M-->I (P: 0.319 +/- 0.038 x 10(-4) cm/s and 0.194 +/- 0.035 x 10(-4) cm/s respectively). Dynamics of the icodextrin M-->I transfer were constant, but I-->M increased by 50% over time. The intensification of the stirring rate increased the value of P at varying rates: the increase was greater for icodextrin than for glucose, and greater for the I-->M transport direction than for the M-->I direction for both solutes. Chemical modification (by 2.5 mmol/L sodium deoxycholate) increased glucose and icodextrin I-->M transfer a mean of 41% and 81% respectively, but increased M-->I transfer by 70% and 224% respectively. The dynamics of glucose and icodextrin peritoneal transfer in vitro are different: glucose diffusion is constant, but I-->M icodextrin transfer increases over time and is greater than M-->I transfer Fluid stirring intensification and chemical injury to the peritoneum enhance diffusion of glucose and icodextrin. Glucose and icodextrin M-->I transfer but not I-->M transfer is restricted more by tissue barriers than by stagnant fluid layers.

  1. Subpopulations of rat dorsal root ganglion neurons express active vesicular acetylcholine transporter.

    Science.gov (United States)

    Tata, Ada Maria; De Stefano, M Egle; Tomassy, Giulio Srubek; Vilaró, M Teresa; Levey, Allan I; Biagioni, Stefano

    2004-01-15

    The vesicular acetylcholine transporter (VAChT) is a transmembrane protein required, in cholinergic neurons, for selective storage of acetylcholine into synaptic vesicles. Although dorsal root ganglion (DRG) neurons utilize neuropeptides and amino acids for neurotransmission, we have previously demonstrated the presence of a cholinergic system. To investigate whether, in sensory neurons, the vesicular accumulation of acetylcholine relies on the same mechanisms active in classical cholinergic neurons, we investigated VAChT presence, subcellular distribution, and activity. RT-PCR and Western blot analysis demonstrated the presence of VAChT mRNA and protein product in DRG neurons and in the striatum and cortex, used as positive controls. Moreover, in situ hybridization and immunocytochemistry showed VAChT staining located mainly in the medium/large-sized subpopulation of the sensory neurons. A few small neurons were also faintly labeled by immunocytochemistry. In the electron microscope, immunolabeling was associated with vesicle-like elements distributed in the neuronal cytoplasm and in both myelinated and unmyelinated intraganglionic nerve fibers. Finally, [(3)H]acetylcholine active transport, evaluated either in the presence or in the absence of ATP, also demonstrated that, as previously reported, the uptake of acetylcholine by VAChT is ATP dependent. This study suggests that DRG neurons not only are able to synthesize and degrade ACh and to convey cholinergic stimuli but also are capable of accumulating and, possibly, releasing acetylcholine by the same mechanism used by the better known cholinergic neurons.

  2. 14,15-EET promotes mitochondrial biogenesis and protects cortical neurons against oxygen/glucose deprivation-induced apoptosis

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Lai; Chen, Man; Yuan, Lin; Xiang, Yuting [Department of Physiology and Pathophysiology, School of Basic Medical Sciences, Peking University Health Science Center, Beijing (China); Zheng, Ruimao, E-mail: rmzheng@pku.edu.cn [Department of Anatomy, Histology and Embryology, School of Basic Medical Sciences, Peking University Health Science Center, Beijing (China); Zhu, Shigong, E-mail: sgzhu@bjmu.edu.cn [Department of Physiology and Pathophysiology, School of Basic Medical Sciences, Peking University Health Science Center, Beijing (China)

    2014-07-18

    Highlights: • 14,15-EET inhibits OGD-induced apoptosis in cortical neurons. • Mitochondrial biogenesis of cortical neurons is promoted by 14,15-EET. • 14,15-EET preserves mitochondrial function of cortical neurons under OGD. • CREB mediates effect of 14,15-EET on mitochondrial biogenesis and function. - Abstract: 14,15-Epoxyeicosatrienoic acid (14,15-EET), a metabolite of arachidonic acid, is enriched in the brain cortex and exerts protective effect against neuronal apoptosis induced by ischemia/reperfusion. Although apoptosis has been well recognized to be closely associated with mitochondrial biogenesis and function, it is still unclear whether the neuroprotective effect of 14,15-EET is mediated by promotion of mitochondrial biogenesis and function in cortical neurons under the condition of oxygen–glucose deprivation (OGD). In this study, we found that 14,15-EET improved cell viability and inhibited apoptosis of cortical neurons. 14,15-EET significantly increased the mitochondrial mass and the ratio of mitochondrial DNA to nuclear DNA. Key makers of mitochondrial biogenesis, peroxisome proliferator activator receptor gamma-coactivator 1 alpha (PGC-1α), nuclear respiratory factor 1 (NRF-1) and mitochondrial transcription factor A (TFAM), were elevated at both mRNA and protein levels in the cortical neurons treated with 14,15-EET. Moreover, 14,15-EET markedly attenuated the decline of mitochondrial membrane potential, reduced ROS, while increased ATP synthesis. Knockdown of cAMP-response element binding protein (CREB) by siRNA blunted the up-regulation of PGC-1α and NRF-1 stimulated by 14,15-EET, and consequently abolished the neuroprotective effect of 14,15-EET. Our results indicate that 14,15-EET protects neurons from OGD-induced apoptosis by promoting mitochondrial biogenesis and function through CREB mediated activation of PGC-1α and NRF-1.

  3. A simple method for imaging axonal transport in aging neurons using the adult Drosophila wing.

    Science.gov (United States)

    Vagnoni, Alessio; Bullock, Simon L

    2016-09-01

    There is growing interest in the link between axonal cargo transport and age-associated neuronal dysfunction. The study of axonal transport in neurons of adult animals requires intravital or ex vivo imaging approaches, which are laborious and expensive in vertebrate models. We describe simple, noninvasive procedures for imaging cargo motility within axons using sensory neurons of the translucent Drosophila wing. A key aspect is a method for mounting the intact fly that allows detailed imaging of transport in wing neurons. Coupled with existing genetic tools in Drosophila, this is a tractable system for studying axonal transport over the life span of an animal and thus for characterization of the relationship between cargo dynamics, neuronal aging and disease. Preparation of a sample for imaging takes ∼5 min, with transport typically filmed for 2-3 min per wing. We also document procedures for the quantification of transport parameters from the acquired images and describe how the protocol can be adapted to study other cell biological processes in aging neurons.

  4. Computational Analysis of Axonal Transport: A Novel Assessment of Neurotoxicity, Neuronal Development and Functions

    Directory of Open Access Journals (Sweden)

    Toshiyuki Gotoh

    2012-03-01

    Full Text Available Axonal transport plays a crucial role in neuronal morphogenesis, survival and function. Despite its importance, however, the molecular mechanisms of axonal transport remain mostly unknown because a simple and quantitative assay system for monitoring this cellular process has been lacking. In order to better characterize the mechanisms involved in axonal transport, we formulate a novel computer-assisted monitoring system of axonal transport. Potential uses of this system and implications for future studies will be discussed.

  5. Sodium-glucose co-transporter-2 inhibitors and euglycemic ketoacidosis: Wisdom of hindsight

    OpenAIRE

    Awadhesh Kumar Singh

    2015-01-01

    Sodium-glucose co-transporter-2 inhibitors (SGLT-2i) are newly approved class of oral anti-diabetic drugs, in the treatment of type 2 diabetes, which reduces blood glucose through glucouresis via the kidney, independent, and irrespective of available pancreatic beta-cells. Studies conducted across their clinical development program found, a modest reduction in glycated hemoglobin ranging from −0.5 to −0.8%, without any significant hypoglycemia. Moreover, head-to-head studies versus active com...

  6. Insights from the Fungus Fusarium oxysporum Point to High Affinity Glucose Transporters as Targets for Enhancing Ethanol Production from Lignocellulose

    Science.gov (United States)

    Ali, Shahin S.; Nugent, Brian; Mullins, Ewen; Doohan, Fiona M.

    2013-01-01

    Ethanol is the most-widely used biofuel in the world today. Lignocellulosic plant biomass derived from agricultural residue can be converted to ethanol via microbial bioprocessing. Fungi such as Fusarium oxysporum can simultaneously saccharify straw to sugars and ferment sugars to ethanol. But there are many bottlenecks that need to be overcome to increase the efficacy of microbial production of ethanol from straw, not least enhancement of the rate of fermentation of both hexose and pentose sugars. This research tested the hypothesis that the rate of sugar uptake by F. oxysporum would enhance the ethanol yields from lignocellulosic straw and that high affinity glucose transporters can enhance ethanol yields from this substrate. We characterized a novel hexose transporter (Hxt) from this fungus. The F. oxysporum Hxt represents a novel transporter with homology to yeast glucose signaling/transporter proteins Rgt2 and Snf3, but it lacks their C-terminal domain which is necessary for glucose signalling. Its expression level decreased with increasing glucose concentration in the medium and in a glucose uptake study the Km(glucose) was 0.9 mM, which indicated that the protein is a high affinity glucose transporter. Post-translational gene silencing or over expression of the Hxt in F. oxysporum directly affected the glucose and xylose transport capacity and ethanol yielded by F. oxysporum from straw, glucose and xylose. Thus we conclude that this Hxt has the capacity to transport both C5 and C6 sugars and to enhance ethanol yields from lignocellulosic material. This study has confirmed that high affinity glucose transporters are ideal candidates for improving ethanol yields from lignocellulose because their activity and level of expression is high in low glucose concentrations, which is very common during the process of consolidated processing. PMID:23382943

  7. Insights from the fungus Fusarium oxysporum point to high affinity glucose transporters as targets for enhancing ethanol production from lignocellulose.

    Directory of Open Access Journals (Sweden)

    Shahin S Ali

    Full Text Available Ethanol is the most-widely used biofuel in the world today. Lignocellulosic plant biomass derived from agricultural residue can be converted to ethanol via microbial bioprocessing. Fungi such as Fusarium oxysporum can simultaneously saccharify straw to sugars and ferment sugars to ethanol. But there are many bottlenecks that need to be overcome to increase the efficacy of microbial production of ethanol from straw, not least enhancement of the rate of fermentation of both hexose and pentose sugars. This research tested the hypothesis that the rate of sugar uptake by F. oxysporum would enhance the ethanol yields from lignocellulosic straw and that high affinity glucose transporters can enhance ethanol yields from this substrate. We characterized a novel hexose transporter (Hxt from this fungus. The F. oxysporum Hxt represents a novel transporter with homology to yeast glucose signaling/transporter proteins Rgt2 and Snf3, but it lacks their C-terminal domain which is necessary for glucose signalling. Its expression level decreased with increasing glucose concentration in the medium and in a glucose uptake study the Km((glucose was 0.9 mM, which indicated that the protein is a high affinity glucose transporter. Post-translational gene silencing or over expression of the Hxt in F. oxysporum directly affected the glucose and xylose transport capacity and ethanol yielded by F. oxysporum from straw, glucose and xylose. Thus we conclude that this Hxt has the capacity to transport both C5 and C6 sugars and to enhance ethanol yields from lignocellulosic material. This study has confirmed that high affinity glucose transporters are ideal candidates for improving ethanol yields from lignocellulose because their activity and level of expression is high in low glucose concentrations, which is very common during the process of consolidated processing.

  8. Up-regulation of heme oxygenase-1 by isoflurane preconditioning during tolerance against neuronal injury induced by oxygen glucose deprivation

    Institute of Scientific and Technical Information of China (English)

    Qifang Li; Yesen Zhu; Hong Jiang; Hui Xu; Heping Liu

    2008-01-01

    Heme oxygenase (HO) is the rate-limiting enzyme in the degradation of heme to produce bile pigments and carbon monoxide. The HO-1 isozyme is induced by a variety of factors such as heat, heme, ischemia, and hydrogen peroxide. In recent years, mounting findings have suggested that HO-1 has a neuroprotective activity against ischemic injury. The neuroprotective role of isoflurane, a commonly used anesthetic, has been well documented, but little is known about the underlying mechanisms involved. Recently, isoflurane has been shown to up-regulate HO-1 in the liver. In this study,we show that isoflurane preconditioning promotes the survival of cultured ischemic hippocampal neurons by increasing the number of surviving neurons and their viability. Further study by reverse transcription-polymerase chain reaction and Western blot analysis showed that isoflurane preconditioning significantly increases HO-1 expression in oxygen glucose deprivation (OGD)-induced neuronal injury. Furthermore,inhibition of HO activity by tin protoporphyrin partially abolishes isoflurane preconditioning's protective effect as measured by lactate dehydrogenase release in OGD neurons.These findings indicated that the neuroprotective role of isoflurane preconditioning against OGD-induced injury might be associated with its role in up-regulating HO-1 in ischemic neurons.

  9. Up-regulation of heme oxygenase-1 by isoflurane preconditioning during tolerance against neuronal injury induced by oxygen glucose deprivation.

    Science.gov (United States)

    Li, Qifang; Zhu, Yesen; Jiang, Hong; Xu, Hui; Liu, Heping

    2008-09-01

    Heme oxygenase (HO) is the rate-limiting enzyme in the degradation of heme to produce bile pigments and carbon monoxide. The HO-1 isozyme is induced by a variety of factors such as heat, heme, ischemia, and hydrogen peroxide. In recent years, mounting findings have suggested that HO-1 has a neuroprotective activity against ischemic injury. The neuroprotective role of isoflurane, a commonly used anesthetic, has been well documented, but little is known about the underlying mechanisms involved. Recently, isoflurane has been shown to up-regulate HO-1 in the liver. In this study, we show that isoflurane preconditioning promotes the survival of cultured ischemic hippocampal neurons by increasing the number of surviving neurons and their viability. Further study by reverse transcription-polymerase chain reaction and Western blot analysis showed that isoflurane preconditioning significantly increases HO-1 expression in oxygen glucose deprivation (OGD)-induced neuronal injury. Furthermore, inhibition of HO activity by tin protoporphyrin partially abolishes isoflurane preconditioning's protective effect as measured by lactate dehydrogenase release in OGD neurons. These findings indicated that the neuroprotective role of isoflurane preconditioning against OGD-induced injury might be associated with its role in up-regulating HO-1 in ischemic neurons.

  10. Tracking Quantum-Dot labeled neurotropic factors transport along primary neuronal axons in compartmental microfluidic chambers.

    Science.gov (United States)

    Gluska, Shani; Chein, Michael; Rotem, Nimrod; Ionescu, Ariel; Perlson, Eran

    2016-01-01

    Neurons are highly polarized cells, with very long axons. Neurotrophic factors like the neuronal growth factor (NGF) are secreted from neuronal targets to promote neuron survival and proper function. These neurotrophic factors must undergo retrograde axonal transport towards the cell body, wherein they initiate signaling pathways important for neurons' various functions and overall health. This process of long-distance axonal signaling is conducted by the dynein motor protein, which transmits signaling endosomes of ligand-receptor complexes retrogradely along microtubule tracks. Here we describe step by step the use of polydimethylsiloxane (PDMS) compartmentalized microfluidic chambers for tracking axonal transport of trophic factors, with a focus on labeled NGF. We describe in detail how to fabricate the molds, assemble the PDMS platform, plate neurons and image, as well as analyze NGF transport along the axon. This method is useful for studying molecular communication mechanisms within the neuron's different compartments as well as between the neuron and its diverse microenvironments, both in health and under pathological conditions.

  11. Quantitative analysis of intraneuronal transport in human iPS neurons

    Directory of Open Access Journals (Sweden)

    Haruko Nakamura

    2015-08-01

    Full Text Available Induced pluripotent stem (iPS cells are promising tools to investigate disease mechanism and develop new drugs. Intraneuronal transport, which is fundamental for neuronal survival and function, is vulnerable to various pharmacological and chemical agents and is disrupted in some neurodegenerative disorders. We applied a quantification method for axonal transport by counting CM-DiI–labeled particles traveling along the neurite, which allowed us to monitor and quantitate, for the first time, intraneuronal transport in human neurons differentiated from iPS cells (iCell neurons. We evaluated the acute effects of several anti-neoplastic agents that have been previously shown to affect intraneuronal transport. Vincristine, paclitaxel and oxaliplatin decreased the number of moving particle along neurites. Cisplatin, however, produced no effect on intraneuronal transport, which is in contrast to our previous report indicating that it inhibits transport in chick dorsal root ganglion neurons. Our system may be a useful method for assessing intraneuronal transport and neurotoxicity in human iPS neurons.

  12. Glucose transporters and in vivo glucose uptake in skeletal and cardiac muscle: fasting, insulin stimulation and immunoisolation studies of GLUT1 and GLUT4.

    Science.gov (United States)

    Kraegen, E W; Sowden, J A; Halstead, M B; Clark, P W; Rodnick, K J; Chisholm, D J; James, D E

    1993-01-01

    Our aim was to study glucose transporters GLUT1 and GLUT4 in relation to in vivo glucose uptake in rat cardiac and skeletal muscle. The levels of both transporters were of a similar order of magnitude in whole muscle tissue (GLUT1/GLUT4 ratio varied from 0.1 to 0.6), suggesting that both may have an important physiological role in regulating muscle glucose metabolism. GLUT4 correlated very strongly (r2 = 0.97) with maximal insulin-stimulated glucose uptake (Rg' max., estimated using the glucose clamp plus 2-deoxy[3H]glucose bolus technique) in six skeletal muscles and heart. A distinct difference in regulation of the two transporters was evident in heart: in 5 h-fasted rats, basal glucose uptake and GLUT1 levels in heart were very high and both were reduced, by 90 and 60% respectively, by 48 h fasting. However, in heart (and in red skeletal muscle), neither GLUT4 levels nor Rg' max. were reduced by 48 h fasting. GLUT1 was shown to be specifically expressed in cardiac myocytes, because intracellular vesicles enriched in GLUT4 contained significant levels of GLUT1. In conclusion, the high association of muscle GLUT4 content with insulin responsiveness in different muscles, and the preservation of both with fasting, supports a predominant role of GLUT4 in insulin-mediated glucose uptake. GLUT1 may play an important role in mediating cardiac muscle glucose uptake in the basal metabolic state. Marked changes in GLUT1 expression with alterations in the metabolic state, such as prolonged fasting, may play an important role in cardiac glucose metabolism. Images Figure 1 Figure 2 PMID:8216230

  13. Fibroblast growth factor 10 protects neuron against oxygen–glucose deprivation injury through inducing heme oxygenase-1

    Energy Technology Data Exchange (ETDEWEB)

    Li, Yong-Hua; Yang, Li-Ye; Chen, Wei; Li, Ying-Ke, E-mail: liyingke6f@126.com; Yuan, Hong-Bin, E-mail: yuanhongbin6f@126.com

    2015-01-02

    Highlights: • FGF10 attenuates OGD induced injury in cortical neuron. • FGF10 reduces OGD triggered ROS level in cortical neuron. • FGF10 induces HO-1 expression upon OGD stimuli in cortical neuron. • Knockdown of HO-1 impairs the neuroprotection of FGF10 in OGD model. - Abstract: Fibroblast growth factors (FGFs) are a family of structurally related heparin-binding proteins with diverse biological functions. FGFs participate in mitogenesis, angiogenesis, cell proliferation, development, differentiation and cell migration. Here, we investigated the potential effect of FGF10, a member of FGFs, on neuron survival in oxygen–glucose deprivation (OGD) model. In primary cultured mouse cortical neurons upon OGD, FGF10 treatment (100 and 1000 ng/ml) attenuated the decrease of cell viability and rescued the LDH release. Tuj-1 immunocytochemistry assay showed that FGF10 promoted neuronal survival. Apoptosis assay with Annexin V + PI by flow cytometry demonstrated that FGF10 treatment reduced apoptotic cell proportion. Moreover, immunoblotting showed that FGF10 alleviated the cleaved caspase-3 upregulation caused by OGD. FGF10 treatment also depressed the OGD-induced increase of caspase-3, -8 and -9 activities. At last, we found FGF10 triggered heme oxygenase-1 (HO-1) protein expression rather than hypoxia-inducible factor-1 (HIF-1), AMP-activated protein kinase (AMPK) signaling and extracellular signal-regulated kinases 1/2 (ERK1/2) signaling. Knockdown of HO-1 by siRNA partly abolished the neuroprotection of FGF10 in OGD model. In summary, our observations provide the first evidence for the neuroprotective function of FGF10 against ischemic neuronal injury and suggest that FGF10 may be a promising agent for treatment of ischemic stroke.

  14. Developmental exposure to ethanol increases the neuronal vulnerability to oxygen-glucose deprivation in cerebellar granule cell cultures.

    Science.gov (United States)

    Le Duc, Diana; Spataru, Ana; Ceanga, Mihai; Zagrean, Leon; Schöneberg, Torsten; Toescu, Emil C; Zagrean, Ana-Maria

    2015-07-21

    Prenatal alcohol exposure is associated with microencephaly, cognitive and behavioral deficits, and growth retardation. Some of the mechanisms of ethanol-induced injury, such as high level oxidative stress and overexpression of pro-apoptotic genes, can increase the sensitivity of fetal neurons towards hypoxic/ischemic stress associated with normal labor. Thus, alcohol-induced sequelae may be the cumulative result of direct ethanol toxicity and increased neuronal vulnerability towards metabolic stressors, including hypoxia. We examined the effects of ethanol exposure on the fetal cerebellar granular neurons' susceptibility to hypoxic/hypoglycemic damage. A chronic ethanol exposure covered the entire prenatal period and 5 days postpartum through breastfeeding, a time interval partially extending into the third-trimester equivalent in humans. After a binge-like alcohol exposure at postnatal day 5, glutamatergic cerebellar granule neurons were cultured and grown for 7 days in vitro, then exposed to a 3-h oxygen-glucose deprivation to mimic a hypoxic/ischemic condition. Cellular viability was monitored by dynamic recording of propidium iodide fluorescence over 20 h reoxygenation. We explored differentially expressed genes on microarray data from a mouse embryonic ethanol-exposure model and validated these by real-time PCR on the present model. In the ethanol-treated cerebellar granule neurons we find an increased expression of genes related to apoptosis (Mapk8 and Bax), but also of genes previously described as neuroprotective (Dhcr24 and Bdnf), which might suggest an actively maintained viability. Our data suggest that neurons exposed to ethanol during development are more vulnerable to in vitro hypoxia/hypoglycemia and have higher intrinsic death susceptibility than unexposed neurons.

  15. Expression of glucocorticoid receptor and glucose transporter-1 during placental development in the diabetic rat

    Directory of Open Access Journals (Sweden)

    Ramazan Demir

    2011-07-01

    Full Text Available In various tissues, glucocorticoids (GCs are known to downregulate glucose transport systems; however, their effects on glucose transporters (GLUTs in the placenta of a diabetic rat are unknown. Glucocorticoid hormone action within the cell is regulated by the glucocorticoid receptor (GR. Thus, this study was designed to investigate the relationship between GR and glucose transporter expression in the placenta of the diabetic rat. Our immunohistochemical results indicated that GR and glucose transporter protein 1 (GLUT 1 are expressed ubiquitously in the trophoblast and endothelial cells of the labyrinthine zone, where maternal fetal transport takes place in the rat placenta. Expression of GR in the junctional zone of the rat placenta was detected in giant cells, and in some spongiotrophoblast cells, but not in the glycogen cells. GLUT 1 was present, especially in glycogen cells during early pregnancy, and in the spongiotrophoblast cells of the junctional zone during late pregnancy. Amounts of GR and GLUT 1 protein were increased towards the end of gestation both in the control and the diabetic placenta. However, at days 17 and 19 of gestation, only the placental GR protein was significantly increased in the streptozotocin-induced diabetic rats compared to control rats. Diabetes led to a significant decrease in placental weight at gestation day 15. In contrast, at gestational days 17 and 21, the weights of the diabetic placenta were significantly increased as compared with the controls. Moreover, diabetes induced fetus intrauterine growth retardation at gestational days 13, 17 and 21. In conclusion, the localization pattern of GR and GLUT 1 proteins in the same cell types led us to believe that there might be a relationship between GR and GLUT 1 expressions at the cellular level. GLUT 1 does not play a pivotal role in diabetic pregnancies. However, placental growth abnormalities during diabetic pregnancy may be related to the amount of GR

  16. Glucose and lactate are equally effective in energizing activity-dependent synaptic vesicle turnover in purified cortical neurons.

    Science.gov (United States)

    Morgenthaler, F D; Kraftsik, R; Catsicas, S; Magistretti, P J; Chatton, J-Y

    2006-08-11

    This study examines the role of glucose and lactate as energy substrates to sustain synaptic vesicle cycling. Synaptic vesicle turnover was assessed in a quantitative manner by fluorescence microscopy in primary cultures of mouse cortical neurons. An electrode-equipped perfusion chamber was used to stimulate cells both by electrical field and potassium depolarization during image acquisition. An image analysis procedure was elaborated to select in an unbiased manner synaptic boutons loaded with the fluorescent dye N-(3-triethylammoniumpropyl)-4-(4-(dibutylamino)styryl)pyridinium dibromide (FM1-43). Whereas a minority of the sites fully released their dye content following electrical stimulation, others needed subsequent K(+) depolarization to achieve full release. This functional heterogeneity was not significantly altered by the nature of metabolic substrates. Repetitive stimulation sequences of FM1-43 uptake and release were then performed in the absence of any metabolic substrate and showed that the number of active sites dramatically decreased after the first cycle of loading/unloading. The presence of 1 mM glucose or lactate was sufficient to sustain synaptic vesicle cycling under these conditions. Moreover, both substrates were equivalent for recovery of function after a phase of decreased metabolic substrate availability. Thus, lactate appears to be equivalent to glucose for sustaining synaptic vesicle turnover in cultured cortical neurons during activity.

  17. Tamoxifen mediated estrogen receptor activation protects against early impairment of hippocampal neuron excitability in an oxygen/glucose deprivation brain slice ischemia model

    OpenAIRE

    Zhang, Huaqiu; Xie, Minjie; Gary P. Schools; Feustel, Paul F.; Wang, Wei; Lei, Ting; Kimelberg, Harold K.; Zhou, Min

    2008-01-01

    Pretreatment of ovarectomized rats with estrogen shows long-term protection via activation of the estrogen receptor (ER). However, it remains unknown whether activation of the ER can provide protection against early neuronal damage when given acutely, we simulated ischemic conditions by applying oxygen and glucose deprived (OGD) solution to acute male rat hippocampal slices and examined the neuronal electrophysiological changes. Pyramidal neurons and interneurons showed a time-dependent membr...

  18. Neuronal GLP1R mediates liraglutide’s anorectic but not glucose-lowering effect

    OpenAIRE

    Sisley, Stephanie; Gutierrez-Aguilar, Ruth; Scott, Michael; D’Alessio, David A.; Sandoval, Darleen A.; Seeley, Randy J

    2014-01-01

    Glucose control and weight loss are cornerstones of type 2 diabetes treatment. Currently, only glucagon-like peptide-1 (GLP1) analogs are able to achieve both weight loss and glucose tolerance. Both glucose and body weight are regulated by the brain, which contains GLP1 receptors (GLP1R). Even though the brain is poised to mediate the effects of GLP1 analogs, it remains unclear whether the glucose- and body weight–lowering effects of long-acting GLP1R agonists are via direct action on CNS GLP...

  19. Lack of SLC2A1 (glucose transporter 1) mutations in 30 Italian patients with alternating hemiplegia of childhood.

    Science.gov (United States)

    De Grandis, Elisa; Stagnaro, Michela; Biancheri, Roberta; Giannotta, Melania; Gobbi, Giuseppe; Traverso, Monica; Veneselli, Edvige; Zara, Federico

    2013-07-01

    Alternating hemiplegia of childhood is a rare, predominantly sporadic disorder. Diagnosis is clinical, and little is known about genetics. Glucose transporter 1 deficiency syndrome shares with alternating hemiplegia of childhood paroxysmal and nonparoxysmal symptoms. The aim of the study was to investigate glucose transporter 1 mutations in 30 Italian patients. Genetic material was analyzed by DNA amplification and glucose transporter 1 region sequencing. Mutational analysis findings of the SLC2A1 gene were negative in all patients. The pattern of movement disorders was reviewed. Interictal dystonia and multiple paroxysmal events were typical of alternating hemiplegia of childhood. In conclusion, alternating hemiplegia of childhood is a heterogeneous clinical condition, and although glucose transporter 1 deficiency can represent an undiagnosed cause of this disorder, mutational analysis is not routinely recommended. Alternatively, a careful clinical analysis and the 3-O-methyl-D-glucose uptake test can allow prompt identification of a subgroup of patients with alternating hemiplegia of childhood treatable with a ketogenic diet.

  20. Water transport by the Na+/glucose cotransporter under isotonic conditions

    DEFF Research Database (Denmark)

    Zeuthen, T; Meinild, A K; Klaerke, D A;

    1997-01-01

    Solute cotransport in the Na+/glucose cotransporter is directly coupled to significant water fluxes. The water fluxes are energized by the downhill fluxes of the other substrates by a mechanism within the protein itself. In the present paper we investigate the Na+/glucose cotransporter expressed...... in Xenopus oocytes. We present a method which allows short-term exposures to sugar under voltage clamp conditions. We demonstrate that water is cotransported with the solutes despite no osmotic differences between the external and intracellular solutions. There is a fixed ratio of 195:1 between the number...... of water molecules and the number of Na+ ions transported, equivalent to 390 water molecules per glucose molecule. Unstirred layer effects are ruled out on the basis of experiments on native oocytes incubated with the ionophores gramicidin D or nystatin....

  1. Ca2+ effects on glucose transport and fatty acid oxidation in L6 skeletal muscle cell cultures

    Directory of Open Access Journals (Sweden)

    Darrick Balu

    2016-03-01

    We did find a Ca2+ stimulation (using either caffeine or ionomycin of fatty acid oxidation. This was observed in the absence (but not the presence of added glucose. We conclude that Ca2+ stimulates fatty acid oxidation at a mitochondrial site, secondary to malonyl CoA inhibition (represented by the presence of glucose in our experiments. In summary, the experiments resolve a controversy on Ca2+ stimulation of glucose transport by skeletal muscle, introduce an important experimental consideration for the measurement of glucose transport, and uncover a new site of action for Ca2+ stimulation of fatty acid oxidation.

  2. Description of glucose transport in isolated bovine mammary epithelial cells by a three-compartment model.

    Science.gov (United States)

    Xiao, Changting; Quinton, V Margaret; Cant, John P

    2004-04-01

    Initial rates of glucose entry into isolated bovine mammary epithelial cells display moderate degrees of asymmetry and cooperative interactions between export and import sites. The present study examined the hypothesis that these kinetic features are due to compartmentalization of intracellular glucose. Net uptake of 3-O-methyl-d-[1-(3)H]glucose (3-OMG) by isolated bovine mammary epithelial cells was measured at 37 degrees C. The time course of 3-OMG net uptake was better fitted by a double-exponential equation than by a single- or triple-exponential equation. Compartmental analysis of the time course curve suggested that translocated 3-OMG is distributed into two compartments with fractional volumes of 32.6 +/- 5.7% and 67.4 +/- 5.7%, respectively. The results support the view that glucose transport in bovine mammary epithelial cells is a multistep process consisting of two serial steps: fast, carrier-mediated, symmetric translocation of sugar across the cell plasma membrane into a small compartment and subsequent slow exchange of posttranslocated sugar between two intracellular compartments. A three-compartment model of this system successfully simulated the observed time course of 3-OMG net uptake and the observed dependence of unidirectional entry rates on intra- and extracellular 3-OMG concentrations. Simulations indicated that backflux of radiolabeled sugar from the small compartment to extracellular space during 15 s of incubation gives rise to the apparent asymmetry, trans-stimulation, and cooperativity of mammary glucose transport kinetics. The fixed-site carrier model overestimated the rate of glucose accumulation in cells, and its features can be accounted for by the compartmentalization of intracellular sugar.

  3. A receptor state space model of the insulin signalling system in glucose transport.

    Science.gov (United States)

    Gray, Catheryn W; Coster, Adelle C F

    2015-12-01

    Insulin is a potent peptide hormone that regulates glucose levels in the blood. Insulin-sensitive cells respond to insulin stimulation with the translocation of glucose transporter 4 (GLUT4) to the plasma membrane (PM), enabling the clearance of glucose from the blood. Defects in this process can give rise to insulin resistance and ultimately diabetes. One widely cited model of insulin signalling leading to glucose transport is that of Sedaghat et al. (2002) Am. J. Physiol. Endocrinol. Metab. 283, E1084-E1101. Consisting of 20 deterministic ordinary differential equations (ODEs), it is the most comprehensive model of insulin signalling to date. However, the model possesses some major limitations, including the non-conservation of key components. In the current work, we detail mathematical and sensitivity analyses of the Sedaghat model. Based on the results of these analyses, we propose a reduced state space model of the insulin receptor subsystem. This reduced model maintains the input-output relation of the original model but is computationally more efficient, analytically tractable and resolves some of the limitations of the Sedaghat model.

  4. Oxygen glucose deprivation post-conditioning protects cortical neurons against oxygen-glucose deprivation injury: role of HSP70 and inhibition of apoptosis.

    Science.gov (United States)

    Zhao, Jian-hua; Meng, Xian-li; Zhang, Jian; Li, Yong-li; Li, Yue-juan; Fan, Zhe-ming

    2014-02-01

    In the present study, we examined the effect of oxygen glucose deprivation (OGD) post-conditioning (PostC) on neural cell apoptosis in OGD-PostC model and the protective effect on primary cortical neurons against OGD injury in vitro. Four-h OGD was induced by OGD by using a specialized and humidified chamber. To initiate OGD, culture medium was replaced with de-oxygenated and glucose-free extracellular solution-Locke's medium. After OGD treatment for 4 h, cells were then allowed to recover for 6 h or 20 h. Then lactate dehydrogenase (LDH) release assay, Western blotting and flow cytometry were used to detect cell death, protein levels and apoptotic cells, respectively. For the PostC treatment, three cycles of 15-min OGD, followed by 15 min normal cultivation, were applied immediately after injurious 4-h OGD. Cells were then allowed to recover for 6 h or 20 h, and cell death was assessed by LDH release assay. Apoptotic cells were flow cytometrically evaluated after 4-h OGD, followed by re-oxygenation for 20 h (O4/R20). In addition, Western blotting was used to examine the expression of heat-shock protein 70 (HSP70), Bcl-2 and Bax. The ratio of Bcl-2 expression was (0.44±0.08)% and (0.76±0.10)%, and that of Bax expression was (0.51±0.05)% and (0.39±0.04)%, and that of HSP70 was (0.42±0.031)% and (0.72±0.045)% respectively in OGD group and PostC group. After O4/R6, the rate of neuron death in PostC group and OGD groups was (28.96±3.03)% and (37.02±4.47)%, respectively. Therefore, the PostC treatment could up-regulate the expression of HSP70 and Bcl-2, but down-regulate Bax expression. As compared with OGD group, OGD-induced neuron death and apoptosis were significantly decreased in PostC group (P<0.05). These findings suggest that PostC inhibited OGD-induced neuron death. This neuro-protective effect is likely achieved by anti-apoptotic mechanisms and is associated with over-expression of HSP70.

  5. Quantitative importance of the pentose phosphate pathway determined by incorporation of 13C from [2-13C]- and [3-13C]glucose into TCA cycle intermediates and neurotransmitter amino acids in functionally intact neurons

    DEFF Research Database (Denmark)

    Brekke, Eva Marie; Walls, Anne Byriel; Schousboe, Arne

    2012-01-01

    is known about the PPP in neurons. The activity of the PPP was quantified in cultured cerebral cortical and cerebellar neurons after incubation in the presence of [2-(13)C]glucose or [3-(13)C]glucose. The activity of the PPP was several fold lower than glycolysis in both types of neurons. While metabolism...

  6. Tea catechins modulate the glucose transport system in 3T3-L1 adipocytes.

    Science.gov (United States)

    Ueda, Manabu; Furuyashiki, Takashi; Yamada, Kayo; Aoki, Yukiko; Sakane, Iwao; Fukuda, Itsuko; Yoshida, Ken-Ichi; Ashida, Hitoshi

    2010-11-01

    In this study, we investigated the effects of tea catechins on the translocation of glucose transporter (GLUT) 4 in 3T3-L1 adipocytes. We found that the ethyl acetate fraction of green tea extract, containing abundant catechins, most decreased insulin-induced glucose uptake activity in 3T3-L1 cells. When the cells were treated with 50 μM catechins in the absence or presence of insulin for 30 min, nongallate-type catechins increased glucose uptake activity without insulin, whereas gallate-type catechins decreased insulin-induced glucose uptake activity. (-)-Epicatechin (EC) and (-)-epigallocatechin (EGC), nongallate-type catechins, increased glucose uptake activity in the dose- and time-dependent manner, whereas (-)-catechin 3-gallate (Cg) and (-)-epigallocatechin 3-gallate (EGCg), gallate-type catechins, decreased insulin-induced glucose uptake activity in the dose- and time-dependent manner. When the cells were treated with 50 μM catechins for 30 min, EC and EGC promoted GLUT4 translocation, whereas Cg and EGCg decreased the insulin-induced translocation in the cells. EC and EGC increased phosphorylation of PKCλ/ζ without phosphorylation of insulin receptor (IR) and Akt. Wortmannin and LY294002, inhibitors for phosphatidylinositol 3'-kinase (PI3K), decreased EC- and EGC-induced glucose uptake activity in the cells. Cg and EGCg decreased phosphorylation of PKCλ/ζ in the presence of insulin without affecting insulin-induced phosphorylation of IR, and Akt. Therefore, EC and EGC promote the translocation of GLUT4 through activation of PI3K, and Cg and EGCg inhibit insulin-induced translocation of GLUT4 by the insulin signaling pathway in 3T3-L1 cells.

  7. Noninvasive blood glucose sensing using near infra-red spectroscopy and artificial neural networks based on inverse delayed function model of neuron.

    Science.gov (United States)

    Ramasahayam, Swathi; Koppuravuri, Sri Haindavi; Arora, Lavanya; Chowdhury, Shubhajit Roy

    2015-01-01

    In this paper, a non-invasive blood glucose sensing system is presented using near infra-red(NIR) spectroscopy. The signal from the NIR optodes is processed using artificial neural networks (ANN) to estimate the glucose level in blood. In order to obtain accurate values of the synaptic weights of the ANN, inverse delayed (ID) function model of neuron has been used. The ANN model has been implemented on field programmable gate array (FPGA). Error in estimating glucose levels using ANN based on ID function model of neuron implemented on FPGA, came out to be 1.02 mg/dl using 15 hidden neurons in the hidden layer as against 5.48 mg/dl using ANN based on conventional neuron model.

  8. Sodium glucose co-transporter inhibitors – A new class of old drugs

    Science.gov (United States)

    Malhotra, Aneeta; Kudyar, Surbhi; Gupta, Anil K.; Kudyar, Rattan P.; Malhotra, Pavan

    2015-01-01

    Sodium glucose co-transporter (SGLT) inhibitors are a new class of drugs which are used in the pharmacotherapy of Type-II diabetes, which happens to be a major risk factor for developing both micro as well as macro-vascular complications. These drugs inhibit the glucose reabsorption by inhibiting SGLT, which exhibits a novel and promising mechanism of action by promoting the urinary glucose excretion hence providing a basis of therapeutic intervention. Results of SGLT-II inhibitors are very encouraging as there is a significant elevation of GLP-1 level, which forms the basis of relevance in treatment of diabetes. It targets the HbA1C and keeps a check on its levels. It also exerts other positive benefits such as weight loss, reduction in blood glucose levels, reduction in blood pressure and improvement in insulin resistance and β-cell dysfunction: All contributing to effective glycemic control. SGLT inhibition will develop as effective modality as it has the capability of inhibiting reabsorption of greater percentage of filtered glucose load. PMID:26539362

  9. Glucose transporter 1-mediated glucose uptake is limiting for B-cell acute lymphoblastic leukemia anabolic metabolism and resistance to apoptosis

    Science.gov (United States)

    Liu, T; Kishton, R J; Macintyre, A N; Gerriets, V A; Xiang, H; Liu, X; Abel, E D; Rizzieri, D; Locasale, J W; Rathmell, J C

    2014-01-01

    The metabolic profiles of cancer cells have long been acknowledged to be altered and to provide new therapeutic opportunities. In particular, a wide range of both solid and liquid tumors use aerobic glycolysis to supply energy and support cell growth. This metabolic program leads to high rates of glucose consumption through glycolysis with secretion of lactate even in the presence of oxygen. Identifying the limiting events in aerobic glycolysis and the response of cancer cells to metabolic inhibition is now essential to exploit this potential metabolic dependency. Here, we examine the role of glucose uptake and the glucose transporter Glut1 in the metabolism and metabolic stress response of BCR-Abl+ B-cell acute lymphoblastic leukemia cells (B-ALL). B-ALL cells were highly glycolytic and primary human B-ALL samples were dependent on glycolysis. We show B-ALL cells express multiple glucose transporters and conditional genetic deletion of Glut1 led to a partial loss of glucose uptake. This reduced glucose transport capacity, however, was sufficient to metabolically reprogram B-ALL cells to decrease anabolic and increase catabolic flux. Cell proliferation decreased and a limited degree of apoptosis was also observed. Importantly, Glut1-deficient B-ALL cells failed to accumulate in vivo and leukemic progression was suppressed by Glut1 deletion. Similarly, pharmacologic inhibition of aerobic glycolysis with moderate doses of 2-deoxyglucose (2-DG) slowed B-ALL cell proliferation, but extensive apoptosis only occurred at high doses. Nevertheless, 2-DG induced the pro-apoptotic protein Bim and sensitized B-ALL cells to the tyrosine kinase inhibitor Dasatinib in vivo. Together, these data show that despite expression of multiple glucose transporters, B-ALL cells are reliant on Glut1 to maintain aerobic glycolysis and anabolic metabolism. Further, partial inhibition of glucose metabolism is sufficient to sensitize cancer cells to specifically targeted therapies, suggesting

  10. Comparative study of glucose transporters GLUT-2 and GLUT-5 in ostriches gastrointestinal tract

    Directory of Open Access Journals (Sweden)

    Piret Hussar

    2016-10-01

    Full Text Available The knowledge about transport of sugars in animals and birds gastrointestinal tract is very important for science as carbohydrates are the main energy source of food. Since until now there is few information about the localization of glucose transporters - integral membrane proteins that mediate the transport of glucose and related substances across the cellular membranes - in birds gastrointestinal tract, the aim of the present study was to localize glucose transporters-2 and -5 (GLUT-2 and -5 in three parts of the ostriches gastrointestinal tract – proventriculus, duodenum and ileum - comparatively in ostrich chicken in their early ontogenesis period. Material from the superficial gland zone of the proventriculus, duodenum and terminal zone of the ileum were collected from eight female ostriches (Struthio camelus var. Domesticus: two chickens after hatching, three 7 and three 30-days old ostriches. The material was fixed with 10% formalin, embedded into paraffin, slices 7 μm thick were cut followed by immunohistochemical staining with polyclonal primary antibodies Rabbit anti-GLUT-2 and Rabbit anti-GLUT-5, carried out according to the manufacturers guidelines (IHC kit, Abcam, UK. The results showed that the staining for both antibodies was weaker in all parts of the gastrointestinal tract of ostriches after hatching compared to 7 and 30 days old ostriches showing that the gastrointestinal tract of ostriches immediately after hatching is not entirely capable of transportation of carbohydrates. The results of our study may indicate the possibility of a close relationship between feeding and the ability to transport sugars in the gastrointestinal tract.

  11. Association between coenzyme Q10 and glucose transporter (GLUT1) deficiency

    OpenAIRE

    Yubero, Delia; O’Callaghan, Mar; Montero, Raquel; Ormazabal, Aida; Armstrong, Judith; Espinos, Carmina; Rodríguez, Maria A; Jou, Cristina; Castejon, Esperanza; Aracil, Maria A; Cascajo, Maria V; Gavilan, Angela; Briones, Paz; Jimenez-Mallebrera, Cecilia; Pineda, Mercedes

    2014-01-01

    Background It has been demonstrated that glucose transporter (GLUT1) deficiency in a mouse model causes a diminished cerebral lipid synthesis. This deficient lipid biosynthesis could contribute to secondary CoQ deficiency. We report here, for the first time an association between GLUT1 and coenzyme Q10 deficiency in a pediatric patient. Case presentation We report a 15 year-old girl with truncal ataxia, nystagmus, dysarthria and myoclonic epilepsy as the main clinical features. Blood lactate ...

  12. Molecular cloning of glucose transporter 1 in grouper Epinephelus coioides and effects of an acute hyperglycemia stress on its expression and glucose tolerance.

    Science.gov (United States)

    Liu, Hongyu; Dong, Xiaohui; Chi, Shuyan; Yang, Qihui; Zhang, Shuang; Chen, Liqiao; Tan, Beiping

    2017-02-01

    The glucose transporter family proteins play pivotal roles in glucose metabolism. In this study, we successfully cloned the orange spotted grouper (Epinephelus coioides) glucose transporter 1 (EcGlut1) gene (GenBank accession: JQ623903). The full-length EcGlut1 cDNA was 2126 bp with a 1476 bp ORF, a 437bp5'-UTR and 223bp3'-UTR. EcGlut1 is predicted to encode a 491 amino acid protein with a MW of 53.9 kDa, a pI of 8.66 and a Pfam domain. Bioinformatics analysis revealed that EcGlut1 was evolutionally conserved between fishes with 80-89 % amino acid identities. EcGlut1 was expressed predominantly in heart and liver and at lower levels in muscle, intestine, stomach and brain. We also investigated the effect of acute hyperglycemia stress on EcGlut1 expression. In glucose tolerance test, changes in EcGlut1 mRNA expression in response to glucose injection and glucose metabolism-related indictors were assessed at the same time. Glucose injection significantly suppressed EcGlut1 mRNA expression in liver at 12 h and in brain at 24 h postinjection (P < 0.05). EcGlut1 mRNA levels in heart were increased at 6 h (P < 0.05). Plasma glucose level increased significantly and reached its maximum at 3 h postinjection (P < 0.05). The spatiotemporal expression of EcGlut1 and glucose metabolism suggested that orange spotted grouper might rely on fat anabolism to reduce acute hyperglycemia stress and the delayed transcription of EcGlut1 gene might be one reason for glucose intolerance in E. coioides.

  13. TIMP1 in conditioned media of human adipose stromal cells protects neurons against oxygen-glucose deprivation injury.

    Science.gov (United States)

    Du, Shiwei; Mao, Gengsheng; Zhu, Timothy; Luan, Zuo; Du, Yansheng; Gu, Huiying

    2015-01-01

    Adipose stromal cells (ASC) can protect neurons when administered to brains due to secreted trophic factors. Our previous studies demonstrated that several neurotrophic factors such as brain-derived neurotropic factor (BDNF) and insulin like growth factor-1 (IGF-1) in ASC conditioned media (ASC-CM) can protect brains against hypoxic-ischemic (HI) injury in neonatal rats. In this study, we demonstrated that human ASC-CM potently blockeds caspase-3 mediated cortical neuronal apoptosis under in vitro oxygen-glucose deprivation (OGD). Interestingly, tissue inhibitor of metalloproteinase 1 (TIMP1), a non neurotrophic factor, played a significant role in the ASC-CM-induced neural protection against OGD. Thus, this study establishes the therapeutic potential of TIMP1 together with other neurotrophic factors in ASC-CM for treating cerebral HI disorders.

  14. Ghrelin Facilitates GLUT2-, SGLT1- and SGLT2-mediated Intestinal Glucose Transport in Goldfish (Carassius auratus)

    Science.gov (United States)

    Blanco, Ayelén Melisa; Bertucci, Juan Ignacio; Ramesh, Naresh; Delgado, María Jesús; Valenciano, Ana Isabel; Unniappan, Suraj

    2017-01-01

    Glucose homeostasis is an important biological process that involves a variety of regulatory mechanisms. This study aimed to determine whether ghrelin, a multifunctional gut-brain hormone, modulates intestinal glucose transport in goldfish (Carassius auratus). Three intestinal glucose transporters, the facilitative glucose transporter 2 (GLUT2), and the sodium/glucose co-transporters 1 (SGLT1) and 2 (SGLT2), were studied. Immunostaining of intestinal sections found colocalization of ghrelin and GLUT2 and SGLT2 in mucosal cells. Some cells containing GLUT2, SGLT1 and SGLT2 coexpressed the ghrelin/growth hormone secretagogue receptor 1a (GHS-R1a). Intraperitoneal glucose administration led to a significant increase in serum ghrelin levels, as well as an upregulation of intestinal preproghrelin, ghrelin O-acyltransferase and ghs-r1 expression. In vivo and in vitro ghrelin treatment caused a concentration- and time-dependent modulation (mainly stimulatory) of GLUT2, SGLT1 and SGLT2. These effects were abolished by the GHS-R1a antagonist [D-Lys3]-GHRP-6 and the phospholipase C inhibitor U73122, suggesting that ghrelin actions on glucose transporters are mediated by GHS-R1a via the PLC/PKC signaling pathway. Finally, ghrelin stimulated the translocation of GLUT2 into the plasma membrane of goldfish primary intestinal cells. Overall, data reported here indicate an important role for ghrelin in the modulation of glucoregulatory machinery and glucose homeostasis in fish. PMID:28338019

  15. Neuronal calcium sensor synaptotagmin-9 is not involved in the regulation of glucose homeostasis or insulin secretion.

    Directory of Open Access Journals (Sweden)

    Natalia Gustavsson

    Full Text Available BACKGROUND: Insulin secretion is a complex and highly regulated process. It is well established that cytoplasmic calcium is a key regulator of insulin secretion, but how elevated intracellular calcium triggers insulin granule exocytosis remains unclear, and we have only begun to define the identities of proteins that are responsible for sensing calcium changes and for transmitting the calcium signal to release machineries. Synaptotagmins are primarily expressed in brain and endocrine cells and exhibit diverse calcium binding properties. Synaptotagmin-1, -2 and -9 are calcium sensors for fast neurotransmitter release in respective brain regions, while synaptotagmin-7 is a positive regulator of calcium-dependent insulin release. Unlike the three neuronal calcium sensors, whose deletion abolished fast neurotransmitter release, synaptotagmin-7 deletion resulted in only partial loss of calcium-dependent insulin secretion, thus suggesting that other calcium-sensors must participate in the regulation of insulin secretion. Of the other synaptotagmin isoforms that are present in pancreatic islets, the neuronal calcium sensor synaptotagmin-9 is expressed at the highest level after synaptotagmin-7. METHODOLOGY/PRINCIPAL FINDINGS: In this study we tested whether synaptotagmin-9 participates in the regulation of glucose-stimulated insulin release by using pancreas-specific synaptotagmin-9 knockout (p-S9X mice. Deletion of synaptotagmin-9 in the pancreas resulted in no changes in glucose homeostasis or body weight. Glucose tolerance, and insulin secretion in vivo and from isolated islets were not affected in the p-S9X mice. Single-cell capacitance measurements showed no difference in insulin granule exocytosis between p-S9X and control mice. CONCLUSIONS: Thus, synaptotagmin-9, although a major calcium sensor in the brain, is not involved in the regulation of glucose-stimulated insulin release from pancreatic β-cells.

  16. Xbp1s in Pomc neurons connects ER stress with energy balance and glucose homeostasis

    Science.gov (United States)

    The molecular mechanisms underlying neuronal leptin and insulin resistance in obesity and diabetes remain unclear. Here we show that induction ofthe unfolded protein response transcription factor spliced X-box binding protein 1(Xbp1s) in pro-opio-melanocortin (Pomc) neurons alone is sufficient to pr...

  17. Insulin/PI3K signaling protects dentate neurons from oxygen-glucose deprivation in organotypic slice cultures.

    Science.gov (United States)

    Sun, Xiaolu; Yao, Hang; Douglas, Robert M; Gu, Xiang Q; Wang, Juan; Haddad, Gabriel G

    2010-01-01

    It is known that ischemia/reperfusion induces neurodegeneration in the hippocampus in a subregion-dependent manner. This study investigated the mechanism of selective resistance/vulnerability to oxygen-glucose deprivation (OGD) using mouse organotypic hippocampal cultures. Analysis of propidium iodide uptake showed that OGD-induced duration- and subregion-dependent neuronal injury. When compared with the CA1-3 subregions, dentate neuronal survival was more sensitive to inhibition of phosphatidylinositol 3-kinase (PI3K)/Akt signaling under basal conditions. Dentate neuronal sensitivity to PI3K/Akt signaling activation was inversely related to its vulnerability to OGD-induced injury; insulin/insulin-like growth factor 1 pre-treatment conferred neuroprotection to dentate neurons via activation of PI3K/Akt signaling. In contrast, CA1 and CA3 neurons were less sensitive to disruptions of endogenous PI3K/Akt signaling and protective effects of insulin/insulin-like growth factor 1, but more vulnerable to OGD. OGD-induced injury in CA1 was reduced by inhibition of NMDA receptor or mitogen-activated protein kinase signaling, and was prevented by blocking NMDA receptor in the presence of insulin. The CA2 subregion was distinctive in its response to glutamate, OGD, and insulin, compared with other CA subregions. CA2 neurons were sensitive to the protective effects of insulin against OGD-induced injury, but more resistant to glutamate. Distinctive distribution of insulin receptor beta and basal phospho-Akt was detected in our slice cultures. Our results suggest a role for insulin signaling in subregional resistance/vulnerability to cerebral ischemia.

  18. Icodextrin peritoneal transport in vitro: effect of sodium deoxycholate, glucose, and methylglyoxal.

    Science.gov (United States)

    Szary, Beata; Grzelak, Teresa; Czyzewska, Krystyna

    2007-02-01

    The aim of the in vitro studies was to examine the effect of sodium deoxycholate, glucose, and methylglyoxal on icodextrin peritoneal transfer. The rabbit peritoneum in a modified Ussing chamber was an experimental model. Transport and morphometric analyses were performed. In the first of them, the icodextrin (7.5 g/dL) diffusion from the mesothelial to the interstitial side of the membrane, expressed as a diffusive permeability coefficient (P), was evaluated in the control stage, after chemical modification of the membrane using sodium deoxycholate (104 mg/dL), after the addition of glucose (1.8 g/dL) and methylglyoxal (1 mg/dL), in the separate experimental series. In the second morphometric studies, the thickness and transverse cross-section surface area of native tissue, in 75 min of experiment and after application of sodium deoxycholate, were investigated. In the control conditions, the rate of glucose polymer passage remained constant. A mean value of P +/- SD was 0.194 +/- 0.126 (x10(-4), cm/s) during 120 min of the study. The transfer of icodextrin was enhanced by 224% after 3 min of incubation of the peritoneum with sodium deoxycholate. The introduction of glucose into the circulating medium with icodextrin caused the increase of P values for glucose polymer by 94% during 60 min. In the same conditions, the usage of methylglyoxal did not change transport parameters. Both thickness and transverse cross-section surface area of the native tissue in 75 min of the study did not differ. It was 4.87 microm and 12.50 x 10(2) microm(2) for the mesothelial layer, and 63.83 microm and 208.10 x 10(2) microm(2) for the whole peritoneal membrane. The application of sodium deoxycholate caused the decrease of mesothelium thickness by 20% but the increase of thickness and transverse cross-section surface area of the peritoneum by 37% in comparison with 75 min of experiment. In conclusion, sodium deoxycholate and glucose, but not methylglyoxal, intensify peritoneal

  19. Fibroblast growth factor 10 protects neuron against oxygen-glucose deprivation injury through inducing heme oxygenase-1.

    Science.gov (United States)

    Li, Yong-Hua; Yang, Li-Ye; Chen, Wei; Li, Ying-Ke; Yuan, Hong-Bin

    2015-01-02

    Fibroblast growth factors (FGFs) are a family of structurally related heparin-binding proteins with diverse biological functions. FGFs participate in mitogenesis, angiogenesis, cell proliferation, development, differentiation and cell migration. Here, we investigated the potential effect of FGF10, a member of FGFs, on neuron survival in oxygen-glucose deprivation (OGD) model. In primary cultured mouse cortical neurons upon OGD, FGF10 treatment (100 and 1000 ng/ml) attenuated the decrease of cell viability and rescued the LDH release. Tuj-1 immunocytochemistry assay showed that FGF10 promoted neuronal survival. Apoptosis assay with Annexin V+PI by flow cytometry demonstrated that FGF10 treatment reduced apoptotic cell proportion. Moreover, immunoblotting showed that FGF10 alleviated the cleaved caspase-3 upregulation caused by OGD. FGF10 treatment also depressed the OGD-induced increase of caspase-3, -8 and -9 activities. At last, we found FGF10 triggered heme oxygenase-1 (HO-1) protein expression rather than hypoxia-inducible factor-1 (HIF-1), AMP-activated protein kinase (AMPK) signaling and extracellular signal-regulated kinases 1/2 (ERK1/2) signaling. Knockdown of HO-1 by siRNA partly abolished the neuroprotection of FGF10 in OGD model. In summary, our observations provide the first evidence for the neuroprotective function of FGF10 against ischemic neuronal injury and suggest that FGF10 may be a promising agent for treatment of ischemic stroke.

  20. Quercetin and sesamin protect neuronal PC12 cells from high-glucose-induced oxidation, nitrosative stress, and apoptosis.

    Science.gov (United States)

    Bournival, Julie; Francoeur, Marc-André; Renaud, Justine; Martinoli, Maria-Grazia

    2012-06-01

    Complications of diabetes are now well-known to affect sensory, motor, and autonomic nerves. Diabetes is also thought to be involved in neurodegenerative processes characteristic of several neurodegenerative diseases. Indeed, it has been acknowledged recently that hyperglycemia-induced oxidative stress contributes to numerous cellular reactions typical of central nervous system deterioration. The goal of the present study was to evaluate the effects of the polyphenol quercetin and the lignan sesamin on high-glucose (HG)-induced oxidative damage in an in vitro model of dopaminergic neurons, neuronal PC12 cells. When incubated with HG (13.5 mg/mL), neuronal PC12 cells showed a significant increase of cellular death. Our results revealed that quercetin and sesamin defend neuronal PC12 cells from HG-induced cellular demise. An elevated level of reactive oxygen and nitrogen species is a consequence of improved oxidative stress after HG administration, and we demonstrated that this production diminishes with quercetin and sesamin treatment. We also found that quercetin and sesamin elicited an increment of superoxide dismutase activity. DNA fragmentation, Bax/Bcl-2 ratio, nuclear translocation of apoptosis-inducing factor, as well as poly(adenosine diphosphate [ADP]-ribose) polymerase cleavage were significantly reduced by quercetin and sesamin administration, affirming their antiapoptotic features. Also, HG treatment impacted caspase-3 cleavage, supporting caspase-3-dependent pathways as mechanisms of apoptotic death. Our results indicate a powerful role for these natural dietary compounds and emphasize preventive or complementary nutritional strategies for diabetes control.

  1. Amyloid-Beta Induced Changes in Vesicular Transport of BDNF in Hippocampal Neurons

    OpenAIRE

    Bianca Seifert; Robert Eckenstaler; Raik Rönicke; Julia Leschik; Beat Lutz; Klaus Reymann; Volkmar Lessmann; Tanja Brigadski

    2016-01-01

    The neurotrophin brain derived neurotrophic factor (BDNF) is an important growth factor in the CNS. Deficits in transport of this secretory protein could underlie neurodegenerative diseases. Investigation of disease-related changes in BDNF transport might provide insights into the cellular mechanism underlying, for example, Alzheimer’s disease (AD). To analyze the role of BDNF transport in AD, live cell imaging of fluorescently labeled BDNF was performed in hippocampal neurons of different AD...

  2. Neuronal Activity and Glutamate Uptake Decrease Mitochondrial Mobility in Astrocytes and Position Mitochondria Near Glutamate Transporters

    Science.gov (United States)

    Jackson, Joshua G.; O'Donnell, John C.; Takano, Hajime; Coulter, Douglas A.

    2014-01-01

    Within neurons, mitochondria are nonuniformly distributed and are retained at sites of high activity and metabolic demand. Glutamate transport and the concomitant activation of the Na+/K+-ATPase represent a substantial energetic demand on astrocytes. We hypothesized that mitochondrial mobility within astrocytic processes might be regulated by neuronal activity and glutamate transport. We imaged organotypic hippocampal slice cultures of rat, in which astrocytes maintain their highly branched morphologies and express glutamate transporters. Using time-lapse confocal microscopy, the mobility of mitochondria within individual astrocytic processes and neuronal dendrites was tracked. Within neurons, a greater percentage of mitochondria were mobile than in astrocytes. Furthermore, they moved faster and farther than in astrocytes. Inhibiting neuronal activity with tetrodotoxin (TTX) increased the percentage of mobile mitochondria in astrocytes. Mitochondrial movement in astrocytes was inhibited by vinblastine and cytochalasin D, demonstrating that this mobility depends on both the microtubule and actin cytoskeletons. Inhibition of glutamate transport tripled the percentage of mobile mitochondria in astrocytes. Conversely, application of the transporter substrate d-aspartate reversed the TTX-induced increase in the percentage of mobile mitochondria. Inhibition of reversed Na+/Ca2+ exchange also increased the percentage of mitochondria that were mobile. Last, we demonstrated that neuronal activity increases the probability that mitochondria appose GLT-1 particles within astrocyte processes, without changing the proximity of GLT-1 particles to VGLUT1. These results imply that neuronal activity and the resulting clearance of glutamate by astrocytes regulate the movement of astrocytic mitochondria and suggest a mechanism by which glutamate transporters might retain mitochondria at sites of glutamate uptake. PMID:24478345

  3. Low Red Blood Cell Vitamin C Concentrations Induce Red Blood Cell Fragility: A Link to Diabetes Via Glucose, Glucose Transporters, and Dehydroascorbic Acid

    Directory of Open Access Journals (Sweden)

    Hongbin Tu

    2015-11-01

    Full Text Available Strategies to prevent diabetic microvascular angiopathy focus on the vascular endothelium. Because red blood cells (RBCs are less deformable in diabetes, we explored an original concept linking decreased RBC deformability to RBC ascorbate and hyperglycemia. We characterized ascorbate concentrations from human and mouse RBCs and plasma, and showed an inverse relationship between RBC ascorbate concentrations and deformability, measured by osmotic fragility. RBCs from ascorbate deficient mice were osmotically sensitive, appeared as spherocytes, and had decreased β-spectrin. These aberrancies reversed with ascorbate repletion in vivo. Under physiologic conditions, only ascorbate's oxidation product dehydroascorbic acid (DHA, a substrate for facilitated glucose transporters, was transported into mouse and human RBCs, with immediate intracellular reduction to ascorbate. In vitro, glucose inhibited entry of physiologic concentrations of dehydroascorbic acid into mouse and human RBCs. In vivo, plasma glucose concentrations in normal and diabetic mice and humans were inversely related to respective RBC ascorbate concentrations, as was osmotic fragility. Human RBC β-spectrin declined as diabetes worsened. Taken together, hyperglycemia in diabetes produced lower RBC ascorbate with increased RBC rigidity, a candidate to drive microvascular angiopathy. Because glucose transporter expression, DHA transport, and its inhibition by glucose differed for mouse versus human RBCs, human experimentation is indicated.

  4. Neuronal and non-neuronal GABA transporters as targets for antiepileptic drugs

    DEFF Research Database (Denmark)

    Madsen, Karsten K; White, H Steve; Schousboe, Arne

    2010-01-01

    of such transporters pointing in particular to an interesting role of the transporters located extrasynaptically. It is suggested that the betaine-GABA transporter BGT1 should receive particular interest in this context as the GABA analogue EF 1502 (N-[4,4-bis(3-methyl-2-thienyl)-3-butenyl]-4-(methylamino)-4...

  5. Expression of GLUT1 and GLUT3 glucose transporters in endometrial and breast cancers.

    Science.gov (United States)

    Krzeslak, Anna; Wojcik-Krowiranda, Katarzyna; Forma, Ewa; Jozwiak, Paweł; Romanowicz, Hanna; Bienkiewicz, Andrzej; Brys, Magdalena

    2012-07-01

    Cancer cells have accelerated metabolism and high glucose requirements. The up-regulation of specific glucose transporters may represent a key mechanism by which malignant cells may achieve increased glucose uptake to support the high rate of glycolysis. In present study we analyzed the mRNA and protein expression of GLUT1 and GLUT3 glucose transporters by quantitative real-time polymerase chain reaction (Q-PCR) and Western blotting technique in 76 cases of endometrial carcinoma and 70 cases of breast carcinoma. SLC2A1 and SLCA2A3 mRNAs expression was found, respectively in 100% and 97.4% samples of endometrial cancers and only in 50% and 40% samples of breast cancers. In endometrial cancers GLUT1 and GLUT3 protein expression was identified in 67.1% and 30.3% of cases. Analogously, in breast cancers in 48.7% and 21% of samples, respectively. The results showed that both endometrial and breast poorly differentiated tumors (grade 2 and 3) had significantly higher GLUT1 and GLUT3 expression than well-differentiated tumors (grade 1). Statistically significant association was found between SLCA2A3 mRNA expression and estrogen and progesterone receptors status in breast cancers. GLUT1 has been reported to be involved in the uptake of glucose by endometrial and breast carcinoma cells earlier and the present study determined that GLUT3 expression is also involved. GLUT1 and GLUT3 seem to be important markers in endometrial and breast tumors differentiation.

  6. Placental glucose and amino acid transport in calorie-restricted wild-type and Glut3 null heterozygous mice.

    Science.gov (United States)

    Ganguly, Amit; Collis, Laura; Devaskar, Sherin U

    2012-08-01

    Calorie restriction (CR) decreased placenta and fetal weights in wild-type (wt) and glucose transporter (Glut) 3 heterozygous null (glut3(+/-)) mice. Because placental nutrient transport is a primary energy determinant of placentofetal growth, we examined key transport systems. Maternal CR reduced intra- and transplacental glucose and leucine transport but enhanced system A amino acid transport in wt mice. These transport perturbations were accompanied by reduced placental Glut3 and leucine amino acid transporter (LAT) family member 2, no change in Glut1 and LAT family member 1, but increased sodium coupled neutral amino acid transporter (SNAT) and SNAT2 expression. We also noted decreased total and active phosphorylated forms of mammalian target of rapamycin, which is the intracellular nutrient sensor, the downstream total P70S6 kinase, and pS6 ribosomal protein with no change in total and phosphorylated 4E-binding protein 1. To determine the role of placental Glut3 in mediating CR-induced placental transport changes, we next investigated the effect of gestational CR in glut3(+/-) mice. In glut3(+/-) mice, a key role of placental Glut3 in mediating transplacental and intraplacental glucose transport was established. In addition, reduced Glut3 results in a compensatory increase of leucine and system A transplacental transport. On the other hand, diminished Glut3-mediated intraplacental glucose transport reduced leucine transport and mammalian target of rapamycin and preserved LAT and enhancing SNAT. CR in glut3(+/-) mice further reduced transplacental glucose transport and enhanced system A amino acid transport, although the increased leucine transport was lost. In addition, increased Glut3 was seen and preserved Glut1, LAT, and SNAT. These placental changes collectively protect survival of wt and glut3(+/-) fetuses against maternal CR-imposed reduction of macromolecular nutrients.

  7. Piracetam and TRH analogues antagonise inhibition by barbiturates, diazepam, melatonin and galanin of human erythrocyte D-glucose transport

    OpenAIRE

    2004-01-01

    Nootropic drugs increase glucose uptake into anaesthetised brain and into Alzheimer's diseased brain. Thyrotropin-releasing hormone, TRH, which has a chemical structure similar to nootropics increases cerebellar uptake of glucose in murine rolling ataxia. This paper shows that nootropic drugs like piracetam (2-oxo 1 pyrrolidine acetamide) and levetiracetam and neuropeptides like TRH antagonise the inhibition of glucose transport by barbiturates, diazepam, melatonin and endogenous neuropeptide...

  8. Steady-state brain glucose transport kinetics re-evaluated with a four-state conformational model

    Directory of Open Access Journals (Sweden)

    João M N Duarte

    2009-10-01

    Full Text Available Glucose supply from blood to brain occurs through facilitative transporter proteins. A near linear relation between brain and plasma glucose has been experimentally determined and described by a reversible model of enzyme kinetics. A conformational four-state exchange model accounting for trans-acceleration and asymmetry of the carrier was included in a recently developed multi-compartmental model of glucose transport. Based on this model, we demonstrate that brain glucose (Gbrain as function of plasma glucose (Gplasma can be described by a single analytical equation namely comprising three kinetic compartments: blood, endothelial cells and brain. Transport was described by four parameters: apparent half saturation constant Kt, apparent maximum rate constant Tmax, glucose consumption rate CMRglc, and the iso-inhibition constant Kii that suggests Gbrain as inhibitor of the isomerisation of the unloaded carrier. Previous published data, where Gbrain was quantified as a function of plasma glucose by either biochemical methods or NMR spectroscopy, were used to determine the aforementioned kinetic parameters. Glucose transport was characterized by Kt ranging from 1.5 to 3.5 mM, Tmax/CMRglc from 4.6 to 5.6, and Kii from 51 to 149 mM. It was noteworthy that Kt was on the order of a few mM, as previously determined from the reversible model. The conformational four-state exchange model of glucose transport into the brain includes both efflux and transport inhibition by Gbrain, predicting that Gbrain eventually approaches a maximum concentration. However, since Kii largely exceeds Gplasma, iso-inhibition is unlikely to be of substantial importance for plasma glucose below 25 mM. As a consequence, the reversible model can account for most experimental observations under euglycaemia and moderate cases of hypo- and hyperglycaemia.

  9. Neuroprotective effects of salvianolic acid B against oxygen-glucose deprivation/reperfusion damage in primary rat cortical neurons

    Institute of Scientific and Technical Information of China (English)

    WANG Yun; JIANG Yu-feng; HUANG Qi-fu; GE Gui-ling; CUI Wei

    2010-01-01

    Background Cerebral ischemia-reperfusion injury is the main reason for the loss of neurons in the ischemic cerebrovascular disease. Therefore, to deeply understand its pathogenesis and find a new target is the key issue to be solved. This research aimed to investigate the neuroprotective effects of salvianolic acid B (SalB) against oxygen-glucose deprivation/reperfusion (OGD/RP) damage in primary rat cortical neurons.Methods The primary cultures of neonatal Wister rats were randomly divided into the control group, the OGD/RP group and the SalB-treatment group (10 mg/L). The cell model was established by depriving of oxygen and glucose for 3 hours and reperfusion for 3 hours and 24 hours, respectively. The neuron viability was determined by MTT assay. The level of cellular reactive oxygen species (ROS) was detected by fluorescent labeling method and spin trapping technique respectively. The activities of neuronal Mn-superoxide dismutase (Mn-SOD), catalase (CAT) and glutathione peroxidase (GSH-PX) were assayed by chromatometry. The mitochondria membrane potential (△ψm) was quantitatively analyzed by flow cytometry. The release rate of cytochrome c was detected by Western blotting. The neuronal ultrastructure was observed by transmission electron microscopy. Statistical significance was evaluated by analysis of variance (ANOVA)followed by Student-Newman-Keuls test.Results OGD/RP increased the level of cellular ROS, but decreased the cell viability and the activities of Mn-SOD, CAT and GSH-PX; SalB treatment significantly reduced the level of ROS (P <0.05); and enhanced the cell viability (P <0.05)and the activities of these antioxidases (P <0.05). Additionally, OGD/RP induced the fluorescence value of △ψm to diminish and the release rate of cytochrome c to rise notably; SalB markedly elevated the level of △ψm (P <0.01) and depressed the release rate of cytochrome c (P <0.05); it also ameliorated the neuronal morphological injury.Conclusion The

  10. Myocardial glucose transporters and glycolytic metabolism during ischemia in hyperglycemic diabetic swine.

    Science.gov (United States)

    Stanley, W C; Hall, J L; Smith, K R; Cartee, G D; Hacker, T A; Wisneski, J A

    1994-01-01

    We assessed the effects of 4 weeks of streptozocin-induced diabetes on regional myocardial glycolytic metabolism during ischemia in anesthetized open-chest domestic swine. Diabetic animals were hyperglycemic (12.0 +/- 2.1 v 6.6 +/- .5 mmol/L), and had lower fasting insulin levels (27 +/- 8 v 79 +/- 19 pmol/L). Myocardial glycolytic metabolism was studied with coronary flow controlled by an extracorporeal perfusion circuit. Left anterior descending coronary artery (LAD) flow was decreased by 50% for 45 minutes and left circumflex (CFX) flow was constant. Myocardial glucose uptake and extraction were measured with D-[6-3H]-2-deoxyglucose (DG) and myocardial blood flow was measured with microspheres. The rate of glucose conversion to lactate and lactate uptake and output were assessed with a continuous infusion of [6-14C]glucose and [U-13C]lactate into the coronary perfusion circuit. Both diabetic and nondiabetic animals had sharp decreases in subendocardial blood flow during ischemia (from 1.21 +/- .10 to 0.43 +/- .08 mL.g-1.min-1 in the nondiabetic group, and from 1.30 +/- .15 to 0.55 +/- .11 in the diabetic group). Diabetes had no significant effect on myocardial glucose uptake or glucose conversion to lactate under either well-perfused or ischemic conditions. Forty-five minutes of ischemia resulted in significant glycogen depletion in the subendocardium in both nondiabetic and diabetic animals, with no differences between the two groups. Glycolytic metabolism is not impaired in hyperglycemic diabetic swine after 1 month of the disease when compared with that in normoglycemic nondiabetic animals. The myocardial content of the insulin-regulatable glucose transporter (GLUT 4) was measured in left ventricular biopsies.(ABSTRACT TRUNCATED AT 250 WORDS)

  11. Towards bridging the gap between acid-base transporters and neuronal excitability modulation

    OpenAIRE

    2014-01-01

    pH homeostasis is a fundamental regulator of the function of the central nervous system. Dysfunction of acid-base transporters often results in disturbance of neuronal excitability. In a latest issue of Journal of Neuroscience, Jones et al. report that increasing intracellular bicarbonate concentration substantially stimulates the excitability of pyramidal neurons from mouse hippocampus by inhibiting KCNQ potassium channel. The finding shed important new light in understanding the molecular m...

  12. SAT1, a glutamine transporter, is preferentially expressed in GABAergic neurons

    Directory of Open Access Journals (Sweden)

    Tom Tallak Solbu

    2010-02-01

    Full Text Available Subsets of GABAergic neurons are able to maintain high frequency discharge patterns, which requires efficient replenishment of the releasable pool of GABA. Although glutamine is considered a preferred precursor of GABA, the identity of transporters involved in glutamine uptake by GABAergic neurons remains elusive. Molecular analyses revealed that SAT1 (Slc38a1 features system A characteristics with a preferential affinity for glutamine, and that SAT1 mRNA expression is associated with GABAergic neurons. By generating specific antibodies against SAT1 we show that this glutamine carrier is particularly enriched in GABAergic neurons. Cellular SAT1 distribution resembles that of GAD67, an essential GABA synthesis enzyme, suggesting that SAT1 can be involved in translocating glutamine into GABAergic neurons to facilitate inhibitory neurotransmitter generation.

  13. Live-cell imaging of post-golgi transport vesicles in cultured hippocampal neurons

    DEFF Research Database (Denmark)

    Jensen, Camilla Stampe; Misonou, Hiroaki

    2015-01-01

    The subcellular localization of neuronal membrane signaling molecules such as receptors and ion channels depends on intracellular trafficking mechanisms. Essentially, vesicular trafficking mechanisms ensure that a large number of membrane proteins are correctly targeted to different subcellular...... compartments of neurons. In the past two decades, the establishment and advancement of fluorescent protein technology have provided us with opportunities to study how proteins are trafficked in living cells. However, live imaging of trafficking processes in neurons necessitate imaging tools to distinguish...... the several different routes that neurons use for protein trafficking. Here we provide a novel protocol to selectively visualize post-Golgi transport vesicles carrying fluorescent-labeled ion channel proteins in living neurons. Further, we provide a number of analytical tools we developed to quantify...

  14. Diet effects on glucose absorption in the small intestine of neonatal calves: importance of intestinal mucosal growth, lactase activity, and glucose transporters.

    Science.gov (United States)

    Steinhoff-Wagner, Julia; Zitnan, Rudolf; Schönhusen, Ulrike; Pfannkuche, Helga; Hudakova, Monika; Metges, Cornelia C; Hammon, Harald M

    2014-10-01

    Colostrum (C) feeding in neonatal calves improves glucose status and stimulates intestinal absorptive capacity, leading to greater glucose absorption when compared with milk-based formula feeding. In this study, diet effects on gut growth, lactase activity, and glucose transporters were investigated in several gut segments of the small intestine. Fourteen male German Holstein calves received either C of milkings 1, 3, and 5 (d 1, 2, and 3 in milk) or respective formulas (F) twice daily from d 1 to d 3 after birth. Nutrient content, and especially lactose content, of C and respective F were the same. On d 4, calves were fed C of milking 5 or respective F and calves were slaughtered 2h after feeding. Tissue samples from duodenum and proximal, mid-, and distal jejunum were taken to measure villus size and crypt depth, mucosa and brush border membrane vesicles (BBMV) were taken to determine protein content, and mRNA expression and activity of lactase and mRNA expression of sodium-dependent glucose co-transporter-1 (SGLT1) and facilitative glucose transporter (GLUT2) were determined from mucosal tissue. Additionally, protein expression of SGLT1 in BBMV and GLUT2 in crude mucosal membranes and BBMV were determined, as well as immunochemically localized GLUT2 in the intestinal mucosa. Villus circumference, area, and height were greater, whereas crypt depth was smaller in C than in F. Lactase activity tended to be greater in C than in F. Protein expression of SGLT1 was greater in F than in C. Parameters of villus size, lactase activity, SGLT1 protein expression, as well as apical and basolateral GLUT2 localization in the enterocytes differed among gut segments. In conclusion, C feeding, when compared with F feeding, enhances glucose absorption in neonatal calves primarily by stimulating mucosal growth and increasing absorptive capacity in the small intestine, but not by stimulating abundance of intestinal glucose transporters.

  15. Effect of fiber type and nutritional state on AICAR- and contraction-stimulated glucose transport in rat muscle

    DEFF Research Database (Denmark)

    Ai, Hua; Ihlemann, Jacob; Hellsten, Ylva

    2002-01-01

    )- and alpha(2)-isoforms of AMPK. Expression of both isoforms varied with fiber types, and alpha(2) was highly expressed in nuclei. In conclusion, AICAR-stimulated glucose transport varies with muscle fiber type and nutritional state. AMPK is unlikely to be the sole mediator of contraction-stimulated glucose...

  16. Effect of vanadate on glucose transporter (GLUT4) intrinsic activity in skeletal muscle plasma membrane giant vesicles

    DEFF Research Database (Denmark)

    Kristiansen, S; Youn, J; Richter, Erik

    1996-01-01

    for 1.5 h with concentrations of vanadate ranging from 3 to 40 mmol l-1 at 34 degrees C before being used for determination of glucose transport. The dose-response curve showed that vanadate decreased the specific D-glucose uptake by a maximum of 70% compared with a control preparation. The vanadate...

  17. Glucose Transport in the Extremely Thermoacidophilic Sulfolobus solfataricus Involves a High-Affinity Membrane-Integrated Binding Protein

    NARCIS (Netherlands)

    Albers, Sonja-V.; Elferink, Marieke G.L.; Charlebois, Robert L.; Sensen, Christoph W.; Driessen, Arnold J.M.; Konings, Wil N.

    1999-01-01

    The archaeon Sulfolobus solfataricus grows optimally at 80°C and pH 2.5 to 3.5 on carbon sources such as yeast extracts, tryptone, and various sugars. Cells rapidly accumulate glucose. This transport activity involves a membrane-bound glucose-binding protein that interacts with its substrate with

  18. Kinesin KIF4A transports integrin β1 in developing axons of cortical neurons.

    Science.gov (United States)

    Heintz, Tristan G; Heller, Janosch P; Zhao, Rongrong; Caceres, Alfredo; Eva, Richard; Fawcett, James W

    2014-11-01

    CNS axons have poor regenerative ability compared to PNS axons, and mature axons regenerate less well than immature embryonic axons. The loss of regenerative ability with maturity is accompanied by the setting up of a selective transport filter in axons, restricting the types of molecule that are present. We confirm that integrins (represented by subunits β1 and α5) are present in early cortical axons in vitro but are excluded from mature axons. Ribosomal protein and L1 show selective axonal transport through association with kinesin kif4A; we have therefore examined the hypothesis that integrin transport might also be in association with kif4A. Kif4A is present in all processes of immature cortical neurons cultured at E18, then downregulated by 14days in vitro, coinciding with the exclusion of integrin from axons. Kif4a co-localises with β1 integrin in vesicles in neurons and non-neuronal cells, and the two molecules co-immunoprecipitate. Knockdown of KIF4A expression with shRNA reduced the level of integrin β1 in axons of developing neurons and reduced neurite elongation on laminin, an integrin-dependent substrate. Overexpression of kif4A triggered apoptosis in neuronal and non-neuronal cells. In mature neurons expression of kif4A-GFP at a modest level did not kill the cells, and the kif4A was detectable in their axons. However this was not accompanied by an increase in integrin β1 axonal transport, suggesting that kif4A is not the only integrin transporter, and that integrin exclusion from axons is controlled by factors other than the kif4A level. Copyright © 2014 Elsevier Inc. All rights reserved.

  19. Effects of deprivation of oxygen or glucose on the neural activity in the guinea pig hippocampal slice--intracellular recording study of pyramidal neurons.

    Science.gov (United States)

    Takata, T; Okada, Y

    1995-06-12

    The block of synaptic transmission and neural activity during deprivation of oxygen or glucose has been simply attributed to the lack of energy due to the disorder of energy production. To clarify the interrelation between neural activity and energy metabolism during hypoxia or glucose deprivation, we studied the changes in ATP levels and electrical events of pyramidal neurons in the CA3 region and [Ca2+]i mobilization of the dendritic and cellular region of CA3 area, using guinea pig hippocampal slices. The studies of field potentials and intracellular recording from the pyramidal cell of CA3 area during hypoxia or glucose deprivation revealed that the cessation of synaptic activity and the depolarization of resting potential occurred earlier than during glucose deprivation while the increase of [Ca2+]i was slow during hypoxia but rapid during glucose deprivation although the ATP level of CA3 area was maintained at its original level for 20 min during both conditions. When glucose was replaced by lactate, ATP concentration was not reduced but the electrical activity decayed and [Ca2+]i increased with the similar time course as observed during lack of glucose, only. These results suggest that different mechanisms underlie the block of synaptic transmission in the CA3 pyramidal neurons during hypoxia and glucose deprivation and that lactate cannot substitute for glucose in the maintenance of neural activity.

  20. Ribosomal S6K1 in POMC and AgRP Neurons Regulates Glucose Homeostasis but Not Feeding Behavior in Mice

    Directory of Open Access Journals (Sweden)

    Mark A. Smith

    2015-04-01

    Full Text Available Hypothalamic ribosomal S6K1 has been suggested as a point of convergence for hormonal and nutrient signals in the regulation of feeding behavior, bodyweight, and glucose metabolism. However, the long-term effects of manipulating hypothalamic S6K1 signaling on energy homeostasis and the cellular mechanisms underlying these roles are unclear. We therefore inactivated S6K1 in pro-opiomelanocortin (POMC and agouti-related protein (AgRP neurons, key regulators of energy homeostasis, but in contrast to the current view, we found no evidence that S6K1 regulates food intake and bodyweight. In contrast, S6K1 signaling in POMC neurons regulated hepatic glucose production and peripheral lipid metabolism and modulated neuronal excitability. S6K1 signaling in AgRP neurons regulated skeletal muscle insulin sensitivity and was required for glucose sensing by these neurons. Our findings suggest that S6K1 signaling is not a general integrator of energy homeostasis in the mediobasal hypothalamus but has distinct roles in the regulation of glucose homeostasis by POMC and AgRP neurons.

  1. Ribosomal S6K1 in POMC and AgRP Neurons Regulates Glucose Homeostasis but Not Feeding Behavior in Mice.

    Science.gov (United States)

    Smith, Mark A; Katsouri, Loukia; Irvine, Elaine E; Hankir, Mohammed K; Pedroni, Silvia M A; Voshol, Peter J; Gordon, Matthew W; Choudhury, Agharul I; Woods, Angela; Vidal-Puig, Antonio; Carling, David; Withers, Dominic J

    2015-04-21

    Hypothalamic ribosomal S6K1 has been suggested as a point of convergence for hormonal and nutrient signals in the regulation of feeding behavior, bodyweight, and glucose metabolism. However, the long-term effects of manipulating hypothalamic S6K1 signaling on energy homeostasis and the cellular mechanisms underlying these roles are unclear. We therefore inactivated S6K1 in pro-opiomelanocortin (POMC) and agouti-related protein (AgRP) neurons, key regulators of energy homeostasis, but in contrast to the current view, we found no evidence that S6K1 regulates food intake and bodyweight. In contrast, S6K1 signaling in POMC neurons regulated hepatic glucose production and peripheral lipid metabolism and modulated neuronal excitability. S6K1 signaling in AgRP neurons regulated skeletal muscle insulin sensitivity and was required for glucose sensing by these neurons. Our findings suggest that S6K1 signaling is not a general integrator of energy homeostasis in the mediobasal hypothalamus but has distinct roles in the regulation of glucose homeostasis by POMC and AgRP neurons. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.

  2. [Neuroprotective effects of the effective components group of xiaoshuantongluo against oxygen-glucose deprivation in primary cultured rat cortical neurons].

    Science.gov (United States)

    Xie, Xin-Mei; Pang, Xiao-Bin; Zhao, Yan; Wang, Bao-Quan; Chen, Ruo-Yun; Du, Guan-Hua

    2014-08-01

    This study is to investigate the effect of the effective components group of Xiaoshuantongluo (XECG) on neuronal injury induced by oxygen-glucose deprivation (OGD) in primary cortical cultures isolated from SD rat cortex at day 3 and the possible mechanism. Cells were divided into control group, OGD model group and XECG group (1, 3 and 10 mg x L(-1)). The cell viability was assessed with MTT assay and the LDH release rate was measured by enzyme label kit. The cell apoptosis was analyzed using Hoechst staining. RT-PCR was applied to detect the mRNA levels of JAK2 and STAT3. Western blotting was used to detect the expressions of Bcl-2, Bax, p-JAK2 and p-STAT3 proteins. Results showed that XECG resulted in an obvious resistance to oxygen-glucose deprivation-induced cell apoptosis and decrement of cell viability, decrease the cell LDH release rate. XECG could adjust the expression of Bcl-2 and Bax proteins and increase Bcl-2/Bax ratio, up-regulate the expression of p-JAK2 and p-STAT3. In conclusion, XECG could protect against the neuronal injury cells exposed to OGD, which may be relevant to the promotion of JAK2/STAT3 signaling pathway, and impact the expression of Bax and Bcl-2.

  3. Curcumin pretreatment and post-treatment both improve the antioxidative ability of neurons with oxygen-glucose deprivation

    Institute of Scientific and Technical Information of China (English)

    Jing-xian Wu; Lu-yu Zhang; Yan-lin Chen; Shan-shan Yu; Yong Zhao; Jing Zhao

    2015-01-01

    Recent studies have shown that induced expression of endogenous antioxidative enzymes thr-ough activation of the antioxidant response element/nuclear factor erythroid 2-related factor 2 (Nrf2) pathway may be a neuroprotective strategy. In this study, rat cerebral cortical neurons culturedin vitrowere pretreated with 10 µM curcumin or post-treated with 5 µM curcumin, respectively before or after being subjected to oxygen-glucose deprivation and reoxygenation for 24 hours. Both pretreatment and post-treatment resulted in a signiifcant decrease of cell injury as indicated by propidium iodide/Hoechst 33258 staining, a prominent increase of Nrf2 protein expression as indicated by western blot analysis, and a remarkable increase of protein expression and enzyme activity in whole cell lysates of thioredoxin before ischemia, after ischemia, and after reoxygenation. In addition, post-treatment with curcumin inhibited early DNA/RNA oxidation as indicated by immunocytochemistry and increased nuclear Nrf2 protein by inducing nuclear accumulation of Nrf2. These findings suggest that curcumin activates the expression of thi-oredoxin, an antioxidant protein in the Nrf2 pathway, and protects neurons from death caused by oxygen-glucose deprivation in anin vitro model of ischemia/reperfusion. We speculate that pharmacologic stimulation of antioxidant gene expression may be a promising approach to neu-roprotection after cerebral ischemia.

  4. Curcumin pretreatment and post-treatment both improve the antioxidative ability of neurons with oxygen-glucose deprivation

    Directory of Open Access Journals (Sweden)

    Jing-xian Wu

    2015-01-01

    Full Text Available Recent studies have shown that induced expression of endogenous antioxidative enzymes thr-ough activation of the antioxidant response element/nuclear factor erythroid 2-related factor 2 (Nrf2 pathway may be a neuroprotective strategy. In this study, rat cerebral cortical neurons cultured in vitro were pretreated with 10 μM curcumin or post-treated with 5 μM curcumin, respectively before or after being subjected to oxygen-glucose deprivation and reoxygenation for 24 hours. Both pretreatment and post-treatment resulted in a significant decrease of cell injury as indicated by propidium iodide/Hoechst 33258 staining, a prominent increase of Nrf2 protein expression as indicated by western blot analysis, and a remarkable increase of protein expression and enzyme activity in whole cell lysates of thioredoxin before ischemia, after ischemia, and after reoxygenation. In addition, post-treatment with curcumin inhibited early DNA/RNA oxidation as indicated by immunocytochemistry and increased nuclear Nrf2 protein by inducing nuclear accumulation of Nrf2. These findings suggest that curcumin activates the expression of thioredoxin, an antioxidant protein in the Nrf2 pathway, and protects neurons from death caused by oxygen-glucose deprivation in an in vitro model of ischemia/reperfusion. We speculate that pharmacologic stimulation of antioxidant gene expression may be a promising approach to neuroprotection after cerebral ischemia.

  5. 14,15-EET promotes mitochondrial biogenesis and protects cortical neurons against oxygen/glucose deprivation-induced apoptosis.

    Science.gov (United States)

    Wang, Lai; Chen, Man; Yuan, Lin; Xiang, Yuting; Zheng, Ruimao; Zhu, Shigong

    2014-07-18

    14,15-Epoxyeicosatrienoic acid (14,15-EET), a metabolite of arachidonic acid, is enriched in the brain cortex and exerts protective effect against neuronal apoptosis induced by ischemia/reperfusion. Although apoptosis has been well recognized to be closely associated with mitochondrial biogenesis and function, it is still unclear whether the neuroprotective effect of 14,15-EET is mediated by promotion of mitochondrial biogenesis and function in cortical neurons under the condition of oxygen-glucose deprivation (OGD). In this study, we found that 14,15-EET improved cell viability and inhibited apoptosis of cortical neurons. 14,15-EET significantly increased the mitochondrial mass and the ratio of mitochondrial DNA to nuclear DNA. Key makers of mitochondrial biogenesis, peroxisome proliferator activator receptor gamma-coactivator 1 alpha (PGC-1α), nuclear respiratory factor 1 (NRF-1) and mitochondrial transcription factor A (TFAM), were elevated at both mRNA and protein levels in the cortical neurons treated with 14,15-EET. Moreover, 14,15-EET markedly attenuated the decline of mitochondrial membrane potential, reduced ROS, while increased ATP synthesis. Knockdown of cAMP-response element binding protein (CREB) by siRNA blunted the up-regulation of PGC-1α and NRF-1 stimulated by 14,15-EET, and consequently abolished the neuroprotective effect of 14,15-EET. Our results indicate that 14,15-EET protects neurons from OGD-induced apoptosis by promoting mitochondrial biogenesis and function through CREB mediated activation of PGC-1α and NRF-1.

  6. Epigenetic Regulation of Glucose Transporters in Non-Small Cell Lung Cancer

    Energy Technology Data Exchange (ETDEWEB)

    O' Byrne, Kenneth J.; Baird, Anne-Marie; Kilmartin, Lisa; Leonard, Jennifer; Sacevich, Calen; Gray, Steven G., E-mail: sgray@stjames.ie [Department of Clinical Medicine, Thoracic Oncology Research Group, Institute of Molecular Medicine, Trinity Centre for Health Sciences, St James' s Hospital, Dublin 8 (Ireland)

    2011-03-25

    Due to their inherently hypoxic environment, cancer cells often resort to glycolysis, or the anaerobic breakdown of glucose to form ATP to provide for their energy needs, known as the Warburg effect. At the same time, overexpression of the insulin receptor in non-small cell lung cancer (NSCLC) is associated with an increased risk of metastasis and decreased survival. The uptake of glucose into cells is carried out via glucose transporters or GLUTs. Of these, GLUT-4 is essential for insulin-stimulated glucose uptake. Following treatment with the epigenetic targeting agents histone deacetylase inhibitors (HDACi), GLUT-3 and GLUT-4 expression were found to be induced in NSCLC cell lines, with minimal responses in transformed normal human bronchial epithelial cells (HBECs). Similar results for GLUT-4 were observed in cells derived from liver, muscle, kidney and pre-adipocytes. Bioinformatic analysis of the promoter for GLUT-4 indicates that it may also be regulated by several chromatin binding factors or complexes including CTCF, SP1 and SMYD3. Chromatin immunoprecipitation studies demonstrate that the promoter for GLUT-4 is dynamically remodeled in response to HDACi. Overall, these results may have value within the clinical setting as (a) it may be possible to use this to enhance fluorodeoxyglucose (18F) positron emission tomography (FDG-PET) imaging sensitivity; (b) it may be possible to target NSCLC through the use of HDACi and insulin mediated uptake of the metabolic targeting drugs such as 2-deoxyglucose (2-DG); or (c) enhance or sensitize NSCLC to chemotherapy.

  7. Septin 7 forms a complex with CD2AP and nephrin and regulates glucose transporter trafficking.

    Science.gov (United States)

    Wasik, Anita A; Polianskyte-Prause, Zydrune; Dong, Meng-Qiu; Shaw, Andrey S; Yates, John R; Farquhar, Marilyn G; Lehtonen, Sanna

    2012-09-01

    Podocytes are insulin-sensitive and take up glucose in response to insulin. This requires nephrin, which interacts with vesicle-associated membrane protein 2 (VAMP2) on GLUT4 storage vesicles (GSVs) and facilitates their fusion with the plasma membrane. In this paper, we show that the filament-forming GTPase septin 7 is expressed in podocytes and associates with CD2-associated protein (CD2AP) and nephrin, both essential for glomerular ultrafiltration. In addition, septin 7 coimmunoprecipitates with VAMP2. Subcellular fractionation of cultured podocytes revealed that septin 7 is found in both cytoplasmic and membrane fractions, and immunofluorescence microscopy showed that septin 7 is expressed in a filamentous pattern and is also found on vesicles and the plasma membrane. The filamentous localization of septin 7 depends on CD2AP and intact actin organization. A 2-deoxy-d-glucose uptake assay indicates that depletion of septin 7 by small interfering RNA or alteration of septin assembly by forchlorfenuron facilitates glucose uptake into cells and further, knockdown of septin 7 increased the interaction of VAMP2 with nephrin and syntaxin 4. The data indicate that septin 7 hinders GSV trafficking and further, the interaction of septin 7 with nephrin in glomeruli suggests that septin 7 may participate in the regulation of glucose transport in podocytes.

  8. DHHC7 Palmitoylates Glucose Transporter 4 (Glut4) and Regulates Glut4 Membrane Translocation.

    Science.gov (United States)

    Du, Keyong; Murakami, Shoko; Sun, Yingmin; Kilpatrick, Casey L; Luscher, Bernhard

    2017-02-17

    Insulin-dependent translocation of glucose transporter 4 (Glut4) to the plasma membrane plays a key role in the dynamic regulation of glucose homeostasis. We recently showed that this process is critically dependent on palmitoylation of Glut4 at Cys-223. To gain further insights into the regulation of Glut4 palmitoylation, we set out to identify the palmitoyl acyltransferase (PAT) involved. Here we report that among 23 mammalian DHHC proteins, DHHC7 is the major Glut4 PAT, based on evidence that ectopic expression of DHHC7 increased Glut4 palmitoylation, whereas DHHC7 knockdown in 3T3-L1 adipocytes and DHHC7 KO in adipose tissue and muscle decreased Glut4 palmitoylation. Moreover, inactivation of DHHC7 suppressed insulin-dependent Glut4 membrane translocation in both 3T3-L1 adipocytes and primary adipocytes. Finally, DHHC7 KO mice developed hyperglycemia and glucose intolerance, thereby confirming that DHHC7 represents the principal PAT for Glut4 and that this mechanism is essential for insulin-regulated glucose homeostasis.

  9. Metabolic Control of Type 2 Diabetes by Targeting the GLUT4 Glucose Transporter: Intervention Approaches.

    Science.gov (United States)

    Alam, Fahmida; Islam, Md Asiful; Khalil, Md Ibrahim; Gan, Siew Hua

    2016-01-01

    Type 2 diabetes mellitus (T2DM), the most common form of diabetes, is characterized by insulin resistance in the hepatic and peripheral tissues. Glucose transporter 4 (GLUT4) plays a major role in the pathophysiology of T2DM. Its defective expression or translocation to the peripheral cell plasma membrane in T2DM patients hinders the entrance of glucose into the cell for energy production. In addition to suitable drugs, an appropriate diet and/or exercise can be implemented to target the increase in GLUT4 expression, GLUT4 concentrations and GLUT4 translocation to the cell surface when managing the glucose metabolism of T2DM patients. In this review, we discussed successful intervention strategies that were individually administered or coupled with diet and/or exercise and affected the expression and translocation of GLUT4 in T2DM while reducing the excess glucose load from the blood. Additionally, some potentially good synthetic and natural compounds, which can activate the insulin-independent GLUT4 signaling pathways for the efficient management of T2DM, are highlighted as possible targets or emerging alternative sources for future anti-diabetic drug development.

  10. MAPT Genetic Variation and Neuronal Maturity Alter Isoform Expression Affecting Axonal Transport in iPSC-Derived Dopamine Neurons

    Directory of Open Access Journals (Sweden)

    Joel E. Beevers

    2017-08-01

    Full Text Available The H1 haplotype of the microtubule-associated protein tau (MAPT locus is genetically associated with neurodegenerative diseases, including Parkinson's disease (PD, and affects gene expression and splicing. However, the functional impact on neurons of such expression differences has yet to be fully elucidated. Here, we employ extended maturation phases during differentiation of induced pluripotent stem cells (iPSCs into mature dopaminergic neuronal cultures to obtain cultures expressing all six adult tau protein isoforms. After 6 months of maturation, levels of exon 3+ and exon 10+ transcripts approach those of adult brain. Mature dopaminergic neuronal cultures display haplotype differences in expression, with H1 expressing 22% higher levels of MAPT transcripts than H2 and H2 expressing 2-fold greater exon 3+ transcripts than H1. Furthermore, knocking down adult tau protein variants alters axonal transport velocities in mature iPSC-derived dopaminergic neuronal cultures. This work links haplotype-specific MAPT expression with a biologically functional outcome relevant for PD.

  11. Glucose transporter-1 (GLUT-1) immunoreactivity in benign, premalignant and malignant lesions of the gallbladder.

    Science.gov (United States)

    Legan, Mateja; Tevžič, Spela; Tolar, Ana; Luzar, Boštjan; Marolt, Vera Ferlan

    2011-03-01

    GLUT-1 is a transmembrane glucose transport protein that allows the facilitated transport of glucose into cells, normally expressed in tissues which depend mainly on glucose metabolism. Enhanced expression of GLUT-1 can also be found in a large spectrum of carcinomas. This study aimed to investigate GLUT-1 expression in gallbladder tissue: from normal tissue samples, hyperplasias, low-grade and high-grade dysplasias to gallbladder carcinomas. In all, 115 archived samples of gallbladder tissue from 68 patients, presented after cholecystectomy, were immunohistochemically stained for GLUT-1. According to the intensity of GLUT-1 immunoreactivity, samples were divided into negative (stained 0-10% of cells stained), positive with weak to moderate (10-50%) and positive with strong (>50%) GLUT-1 expression. The GLUT-1 immunoreactivity of the samples showed a characteristic increase from premalignant lesions to carcinomas. Normal gallbladder tissue samples did not express GLUT-1 (100%). Weak expression was shown only focally in hyperplasias, but to a greater extent with low-grade dysplasias (20%), high-grade dysplasias (40%) and carcinomas (51.8%). Normal gallbladder tissue is GLUT-1 negative. GLUT-1 expression in carcinoma tissue is significantly higher than in dysplastic lesions. Strong GLUT-1 expression indicates 100% specificity for detecting gallbladder carcinomas. Therefore, GLUT-1 is a candidate as a diagnostic as well as a tissue prognostic marker in gallbladder carcinoma patients.

  12. 27-Hydroxycholesterol impairs neuronal glucose uptake through an IRAP/GLUT4 system dysregulation.

    Science.gov (United States)

    Ismail, Muhammad-Al-Mustafa; Mateos, Laura; Maioli, Silvia; Merino-Serrais, Paula; Ali, Zeina; Lodeiro, Maria; Westman, Eric; Leitersdorf, Eran; Gulyás, Balázs; Olof-Wahlund, Lars; Winblad, Bengt; Savitcheva, Irina; Björkhem, Ingemar; Cedazo-Mínguez, Angel

    2017-02-17

    Hypercholesterolemia is associated with cognitively deteriorated states. Here, we show that excess 27-hydroxycholesterol (27-OH), a cholesterol metabolite passing from the circulation into the brain, reduced in vivo brain glucose uptake, GLUT4 expression, and spatial memory. Furthermore, patients exhibiting higher 27-OH levels had reduced (18)F-fluorodeoxyglucose uptake. This interplay between 27-OH and glucose uptake revealed the engagement of the insulin-regulated aminopeptidase (IRAP). 27-OH increased the levels and activity of IRAP, countered the IRAP antagonist angiotensin IV (AngIV)-mediated glucose uptake, and enhanced the levels of the AngIV-degrading enzyme aminopeptidase N (AP-N). These effects were mediated by liver X receptors. Our results reveal a molecular link between cholesterol, brain glucose, and the brain renin-angiotensin system, all of which are affected in some neurodegenerative diseases. Thus, reducing 27-OH levels or inhibiting AP-N maybe a useful strategy in the prevention of the altered glucose metabolism and memory decline in these disorders.

  13. Differential effects of glutamate transporter inhibitors on the global electrophysiological response of astrocytes to neuronal stimulation.

    Science.gov (United States)

    Bernardinelli, Yann; Chatton, Jean-Yves

    2008-11-13

    Astrocytes are responsible for regulating extracellular levels of glutamate and potassium during neuronal activity. Glutamate clearance is handled by glutamate transporter subtypes glutamate transporter 1 and glutamate-aspartate transporter in astrocytes. DL-threo-beta-benzyloxyaspartate (TBOA) and dihydrokainate (DHK) are extensively used as inhibitors of glial glutamate transport activity. Using whole-cell recordings, we characterized the effects of both transporter inhibitors on afferent-evoked astrocyte currents in acute cortical slices of 3-week-old rats. When neuronal afferents were stimulated, passive astrocytes responded by a rapid inward current followed by a persistent tail current. The first current corresponded to a glutamate transporter current. This current was inhibited by both inhibitors and by tetrodotoxin. The tail current is an inward potassium current as it was blocked by barium. Besides inhibiting transporter currents, TBOA strongly enhanced the tail current. This effect was barium-sensitive and might be due to a rise in extracellular potassium level and increased glial potassium uptake. Unlike TBOA, DHK did not enhance the tail current but rather inhibited it. This result suggests that, in addition to inhibiting glutamate transport, DHK prevents astrocyte potassium uptake, possibly by blockade of inward-rectifier channels. This study revealed that, in brain slices, glutamate transporter inhibitors exert complex effects that cannot be attributed solely to glutamate transport inhibition.

  14. Oxygen-glucose deprivation of neurons transfected with toll-like receptor 3-siRNA Determination of an optimal transfection sequence

    Institute of Scientific and Technical Information of China (English)

    Guiyun Cui; Xiaopeng Wang; Xinchun Ye; Jie Zu; Kun Zan; Fang Hua

    2013-01-01

    Tol-like receptor 3 protein expression has been shown to be upregulated during cerebral ische-mia/reperfusion injury in rats. In this study, rat primary cortical neurons were subjected to oxy-gen-glucose deprivation to simulate cerebral ischemia/reperfusion injury. Chemical y synthesized smal interfering RNA (siRNA)-1280,-1724 and-418 specific to tol-like receptor 3 were transfected into oxygen-glucose deprived cortical neurons to suppress the upregulation of tol-like receptor 3 protein expression. Western blotting demonstrated that after transfection with siRNA, tol-like ceptor 3 protein expression reduced, especial y in the tol-like receptor 3-1724 group. These results suggested that siRNA-1724 is an optimal sequence for inhibiting tol-like receptor 3 expression in cortical neurons fol owing oxygen-glucose deprivation.

  15. Daucosterol protects neurons against oxygen-glucose deprivation/reperfusion-mediated injury by activating IGF1 signaling pathway.

    Science.gov (United States)

    Jiang, Li-hua; Yuan, Xiao-lin; Yang, Nian-yun; Ren, Li; Zhao, Feng-ming; Luo, Ban-xin; Bian, Yao-yao; Xu, Jian-ya; Lu, Da-xiang; Zheng, Yuan-yuan; Zhang, Chuan-juan; Diao, Yuan-ming; Xia, Bao-mei; Chen, Gang

    2015-08-01

    We previously reported that daucosterol (a sterolin) up-regulates the expression of insulin-like growth factor I (IGF1)(1) protein in neural stem cells. In this study, we investigated the effects of daucosterol on the survival of cultured cortical neurons after neurons were subjected to oxygen and glucose deprivation and simulated reperfusion (OGD/R)(2), and determined the corresponding molecular mechanism. The results showed that post-treatment of daucosterol significantly reduced neuronal loss, as well as apoptotic rate and caspase-3 activity, displaying the neuroprotective activity. We also found that daucosterol increased the expression level of IGF1 protein, diminished the down-regulation of p-AKT(3) and p-GSK-3β(4), thus activating the AKT(5) signal pathway. Additionally, it diminished the down-regulation of the anti-apoptotic proteins Mcl-1(6) and Bcl-2(7), and decreased the expression level of the pro-apoptotic protein Bax(8), thus raising the ratio of Bcl-2/Bax. The neuroprotective effect of daucosterol was inhibited in the presence of picropodophyllin (PPP)(9), the inhibitor of insulin-like growth factor I receptors (IGF1R)(10). Our study provided information about daucosterol as an efficient and inexpensive neuroprotectants, to which the IGF1-like activity of daucosterol contributes. Daucosterol could be potentially developed as a medicine for ischemic stroke treatment.

  16. CAR-associated vesicular transport of an adenovirus in motor neuron axons.

    Science.gov (United States)

    Salinas, Sara; Bilsland, Lynsey G; Henaff, Daniel; Weston, Anne E; Keriel, Anne; Schiavo, Giampietro; Kremer, Eric J

    2009-05-01

    Axonal transport is responsible for the movement of signals and cargo between nerve termini and cell bodies. Pathogens also exploit this pathway to enter and exit the central nervous system. In this study, we characterised the binding, endocytosis and axonal transport of an adenovirus (CAV-2) that preferentially infects neurons. Using biochemical, cell biology, genetic, ultrastructural and live-cell imaging approaches, we show that interaction with the neuronal membrane correlates with coxsackievirus and adenovirus receptor (CAR) surface expression, followed by endocytosis involving clathrin. In axons, long-range CAV-2 motility was bidirectional with a bias for retrograde transport in nonacidic Rab7-positive organelles. Unexpectedly, we found that CAR was associated with CAV-2 vesicles that also transported cargo as functionally distinct as tetanus toxin, neurotrophins, and their receptors. These results suggest that a single axonal transport carrier is capable of transporting functionally distinct cargoes that target different membrane compartments in the soma. We propose that CAV-2 transport is dictated by an innate trafficking of CAR, suggesting an unsuspected function for this adhesion protein during neuronal homeostasis.

  17. CAR-associated vesicular transport of an adenovirus in motor neuron axons.

    Directory of Open Access Journals (Sweden)

    Sara Salinas

    2009-05-01

    Full Text Available Axonal transport is responsible for the movement of signals and cargo between nerve termini and cell bodies. Pathogens also exploit this pathway to enter and exit the central nervous system. In this study, we characterised the binding, endocytosis and axonal transport of an adenovirus (CAV-2 that preferentially infects neurons. Using biochemical, cell biology, genetic, ultrastructural and live-cell imaging approaches, we show that interaction with the neuronal membrane correlates with coxsackievirus and adenovirus receptor (CAR surface expression, followed by endocytosis involving clathrin. In axons, long-range CAV-2 motility was bidirectional with a bias for retrograde transport in nonacidic Rab7-positive organelles. Unexpectedly, we found that CAR was associated with CAV-2 vesicles that also transported cargo as functionally distinct as tetanus toxin, neurotrophins, and their receptors. These results suggest that a single axonal transport carrier is capable of transporting functionally distinct cargoes that target different membrane compartments in the soma. We propose that CAV-2 transport is dictated by an innate trafficking of CAR, suggesting an unsuspected function for this adhesion protein during neuronal homeostasis.

  18. Inhibition of protein kinase CbetaII increases glucose uptake in 3T3-L1 adipocytes through elevated expression of glucose transporter 1 at the plasma membrane.

    NARCIS (Netherlands)

    Bosch, R.R.; Bazuine, M.; Wake, M.M.; Span, P.N.; Olthaar, A.J.; Schurmann, A.; Maassen, J.A.; Hermus, A.R.M.M.; Willems, P.H.G.M.; Sweep, C.G.J.

    2003-01-01

    The mechanism via which diacylglycerol-sensitive protein kinase Cs (PKCs) stimulate glucose transport in insulin-sensitive tissues is poorly defined. Phorbol esters, such as phorbol-12-myristate-13-acetate (PMA), are potent activators of conventional and novel PKCs. Addition of PMA increases the rat

  19. Progesterone treatment before experimental hypoxia-ischemia enhances the expression of glucose transporter proteins GLUT1 and GLUT3 in neonatal rats

    Institute of Scientific and Technical Information of China (English)

    Xinjuan Li; Hua Han; Ruanling Hou; Linyu Wei; Guohong Wang; Chaokun Li; Dongliang Li

    2013-01-01

    Progesterone is an efficient candidate for treating stroke and traumatic brain damage.The current study was designed to investigate the effects of progesterone on glucose transporter proteins (GLUT1 and GLUT3) during hypoxic-ischemic injury in a neonatal rat model.We demonstrated strong staining for GLUT1 in the walls of blood vessels and GLUT3 immunoreactivity in hippocampal neurons after hypoxiaischemia.Hypoxia-ischemia elevated GLUT1 and GLUT3 at both the mRNA and protein levels in the hippocampus,and pre-treatment with progesterone (8 mg/kg) further enhanced their accumulation until 24 h after hypoxic-ischemic injury.These results showed that progesterone treatment induced the accumulation of both GLUT1 and GLUT3 transporters,and an energy-compensation mechanism may be involved in the neuroprotective effect of progesterone during hypoxic-ischemic injury after cerebral ischemic attacks.

  20. Analysis of metabolism of 6FDG: a PET glucose transport tracer

    Energy Technology Data Exchange (ETDEWEB)

    Muzic, Raymond F., E-mail: raymond.muzic@case.edu [Department of Radiology, Case Western Reserve University, Cleveland, OH 44106 (United States); Department of Biomedical Engineering, Case Western Reserve University, Cleveland, OH 44106 (United States); Chandramouli, Visvanathan [Department of Radiology, Case Western Reserve University, Cleveland, OH 44106 (United States); Huang, Hsuan-Ming [Department of Biomedical Engineering, Case Western Reserve University, Cleveland, OH 44106 (United States); Wu Chunying; Wang Yanming [Department of Radiology, Case Western Reserve University, Cleveland, OH 44106 (United States); Ismail-Beigi, Faramarz [Department of Medicine, Case Western Reserve University, Cleveland, OH 44106 (United States)

    2011-07-15

    Introduction: We are developing {sup 18}F-labeled 6-fluoro-6-deoxy-D-glucose ([{sup 18}F]6FDG) as a tracer of glucose transport. As part of this process it is important to characterize and quantify putative metabolites. In contrast to the ubiquitous positron emission tomography (PET) tracer {sup 18}F-labeled 2-fluoro-2-deoxy-D-glucose ([{sup 18}F]2FDG) which is phosphorylated and trapped intracellularly, the substitution of fluorine for a hydroxyl group at carbon-6 in [{sup 18}F]6FDG should prevent its phosphorylation. Consequently, [{sup 18}F]6FDG has the potential to trace the transport step of glucose metabolism without the confounding effects of phosphorylation and subsequent steps of metabolism. Herein the focus is to determine whether, and the degree to which, [{sup 18}F]6FDG remains unchanged following intravenous injection. Methods: Biodistribution studies were performed using 6FDG labeled with {sup 18}F or with the longer-lived radionuclides {sup 3}H and {sup 14}C. Tissues were harvested at 1, 6, and 24 h following intravenous administration and radioactivity was extracted from the tissues and analyzed using a combination of ion exchange columns, high-performance liquid chromatography, and chemical reactivity. Results: At the 1 h time-point, the vast majority of radioactivity in the liver, brain, heart, skeletal muscle, and blood was identified as 6FDG. At the 6-h and 24-h time points, there was evidence of a minor amount of radioactive material that appeared to be 6-fluoro-6-deoxy-D-sorbitol and possibly 6-fluoro-6-deoxy-D-gluconic acid. Conclusion: On the time scale typical of PET imaging studies radioactive metabolites of [{sup 18}F]6FDG are negligible.

  1. Expression, purification, and functional characterization of the insulin-responsive facilitative glucose transporter GLUT4.

    Science.gov (United States)

    Kraft, Thomas E; Hresko, Richard C; Hruz, Paul W

    2015-12-01

    The insulin-responsive facilitative glucose transporter GLUT4 is of fundamental importance for maintenance of glucose homeostasis. Despite intensive effort, the ability to express and purify sufficient quantities of structurally and functionally intact protein for biophysical analysis has previously been exceedingly difficult. We report here the development of novel methods to express, purify, and functionally reconstitute GLUT4 into detergent micelles and proteoliposomes. Rat GLUT4 containing FLAG and His tags at the amino and carboxy termini, respectively, was engineered and stably transfected into HEK-293 cells. Overexpression in suspension culture yielded over 1.5 mg of protein per liter of culture. Systematic screening of detergent solubilized GLUT4-GFP fusion protein via fluorescent-detection size exclusion chromatography identified lauryl maltose neopentyl glycol (LMNG) as highly effective for isolating monomeric GLUT4 micelles. Preservation of structural integrity and ligand binding was demonstrated via quenching of tryptophan fluorescence and competition of ATB-BMPA photolabeling by cytochalasin B. GLUT4 was reconstituted into lipid nanodiscs and proper folding was confirmed. Reconstitution of purified GLUT4 with amphipol A8-35 stabilized the transporter at elevated temperatures for extended periods of time. Functional activity of purified GLUT4 was confirmed by reconstitution of LMNG-purified GLUT4 into proteoliposomes and measurement of saturable uptake of D-glucose over L-glucose. Taken together, these data validate the development of an efficient means to generate milligram quantities of stable and functionally intact GLUT4 that is suitable for a wide array of biochemical and biophysical analyses.

  2. Postnatal changes in glucose transporter 3 expression in the dentate gyrus of the C57BL/6 mouse model.

    Science.gov (United States)

    Jung, Hyo Young; Yim, Hee Sun; Yoo, Dae Young; Kim, Jong Whi; Chung, Jin Young; Seong, Je Kyung; Yoon, Yeo Sung; Kim, Dae Won; Hwang, In Koo

    2016-03-01

    In this study, we observed the ontogenetic changes in glucose transporter 3 (GLUT3) immunoreactivity, a major neuronal GLUT, in the dentate gyrus of mouse brains at various ages: postnatal day (P) 1, 7, 14, 28, and 56. At P1, cresyl violet staining showed abundant neurons in the dentate gyrus, whereas the granule cell layer was ill-defined. At P7, the granule cell layer was observed, and cresyl violet-positive cells were dispersed throughout the polymorphic layer. At P14, the granule cell layer was well-defined, and cresyl violet positive cells were detected abundantly in the polymorphic layer. At P28 and P56, cresyl violet-positive cells were observed in the granule cell layer, as well as in the polymorphic layer. At P1, GLUT3 immunoreactivity was detected in the dentate gyrus. At P7, GLUT3 immunoreactive cells were scattered in the polymorphic and molecular layer. However, at P14, GLUT3 immunoreactivity was observed in the polymorphic layer as well as subgranular zone of the dentate gyrus. At P28, GLUT3 immunoreactivity was detected in the polymorphic layer of the dentate gyrus. At P56, GLUT3 immunoreactivity was observed predominantly in the subgranular zone of the dentate gyrus. GLUT3 immunoreactive cells were mainly colocalized with doublecortin, which is a marker for differentiated neuroblasts, in the polymorphic layer and subgranular zone of dentate gyrus at P14 and P56. These results suggest that the expression of GLUT3 is closely associated with postnatal development of the dentate gyrus and adult neurogenesis.

  3. Modulation of glucose transporter 1 (GLUT1 expression levels alters mouse mammary tumor cell growth in vitro and in vivo.

    Directory of Open Access Journals (Sweden)

    Christian D Young

    Full Text Available Tumor cells exhibit an altered metabolism characterized by elevated aerobic glycolysis and lactate secretion which is supported by an increase in glucose transport and consumption. We hypothesized that reducing or eliminating the expression of the most prominently expressed glucose transporter(s would decrease the amount of glucose available to breast cancer cells thereby decreasing their metabolic capacity and proliferative potential.Of the 12 GLUT family glucose transporters expressed in mice, GLUT1 was the most abundantly expressed at the RNA level in the mouse mammary tumors from MMTV-c-ErbB2 mice and cell lines examined. Reducing GLUT1 expression in mouse mammary tumor cell lines using shRNA or Cre/Lox technology reduced glucose transport, glucose consumption, lactate secretion and lipid synthesis in vitro without altering the concentration of ATP, as well as reduced growth on plastic and in soft agar. The growth of tumor cells with reduced GLUT1 expression was impaired when transplanted into the mammary fat pad of athymic nude mice in vivo. Overexpression of GLUT1 in a cell line with low levels of endogenous GLUT1 increased glucose transport in vitro and enhanced growth in nude mice in vivo as compared to the control cells with very low levels of GLUT1.These studies demonstrate that GLUT1 is the major glucose transporter in mouse mammary carcinoma models overexpressing ErbB2 or PyVMT and that modulation of the level of GLUT1 has an effect upon the growth of mouse mammary tumor cell lines in vivo.

  4. Pharmacokinetic and pharmacodynamic profile of empagliflozin, a sodium glucose co-transporter 2 inhibitor.

    Science.gov (United States)

    Scheen, André J

    2014-03-01

    Empagliflozin is an orally active, potent and selective inhibitor of sodium glucose co-transporter 2 (SGLT2), currently in clinical development to improve glycaemic control in adults with type 2 diabetes mellitus (T2DM). SGLT2 inhibitors, including empagliflozin, are the first pharmacological class of antidiabetes agents to target the kidney in order to remove excess glucose from the body and, thus, offer new options for T2DM management. SGLT2 inhibitors exert their effects independently of insulin. Following single and multiple oral doses (0.5-800 mg), empagliflozin was rapidly absorbed and reached peak plasma concentrations after approximately 1.33-3.0 h, before showing a biphasic decline. The mean terminal half-life ranged from 5.6 to 13.1 h in single rising-dose studies, and from 10.3 to 18.8 h in multiple-dose studies. Following multiple oral doses, increases in exposure were dose-proportional and trough concentrations remained constant after day 6, indicating a steady state had been reached. Oral clearance at steady state was similar to corresponding single-dose values, suggesting linear pharmacokinetics with respect to time. No clinically relevant alterations in pharmacokinetics were observed in mild to severe hepatic impairment, or in mild to severe renal impairment and end-stage renal disease. Clinical studies did not reveal any relevant drug-drug interactions with several other drugs commonly prescribed to patients with T2DM, including warfarin. Urinary glucose excretion (UGE) rates were higher with empagliflozin versus placebo and increased with dose, but no relevant impact on 24-h urine volume was observed. Increased UGE resulted in proportional reductions in fasting plasma glucose and mean daily glucose concentrations.

  5. Continuous-culture study of the regulation of glucose and fructose transport in Kluyveromyces marxianus CBS 6556.

    Science.gov (United States)

    Postma, E; Van den Broek, P J

    1990-01-01

    Regulation of transport of D-glucose and D-fructose was studied in Kluyveromyces marxianus grown in continuous culture. Both substrates could be transported by at least two different transport systems, low-affinity transport and high-affinity proton-sugar symport. The low-affinity transporter, specific for both glucose and fructose, was constitutively present and was apparently not regulated by carbon catabolite repression. Regulation of the activity of the glucose- and fructose-specific proton symport systems appeared to proceed mainly through catabolite repression. Activation of symport did not need the presence of specific inductor molecules in the medium. Nevertheless, the capacities of the proton-sugar symporters varied in cells grown on a wide variety of carbon sources. The possibility that the control of proton symport activity is related to the presence of specific intracellular metabolites is discussed. PMID:2160928

  6. Forskolin photoaffinity labels with specificity for adenylyl cyclase and the glucose transporter

    Energy Technology Data Exchange (ETDEWEB)

    Morris, D.I.; Robbins, J.D.; Ruoho, A.E.; Sutkowski, E.M.; Seamon, K.B. (Division of Biochemistry and Biophysics, Food and Drug Administration, Bethesda, MD (USA))

    1991-07-15

    Two photolabels, N-(3-(4-azido-3-125I-phenyl)-propionamide)-6- aminoethylcarbamylforskolin(125I-6-AIPP-Fsk) and N-(3-(4-azido-3-125I-phenyl)propionamide)-7-aminoethylcarbamyl-7- desacetylforskolin (125I-7-AIPP-Fsk) were synthesized with specific activities of 2200 Ci/mmol and used to label adenylyl cyclase and the glucose transporter. The affinities of the photolabels for adenylyl cyclase were determined by their inhibition of (3H)forskolin binding to bovine brain membranes. 6-AIPP-Fsk and 7-AIPP-Fsk inhibited (3H)forskolin binding with IC50 values of 15 nM and 200 nM, respectively. 125I-6-AIPP-Fsk labeled a 115-kDa protein in control and GTP {gamma} S-preactivated bovine brain membranes. This labeling was inhibited by forskolin but not by 1,9-dideoxyforskolin or cytochalasin B. 125I-6-AIPP-Fsk labeling of partially purified adenylyl cyclase was inhibited by forskolin but not by 1,9-dideoxyforskolin. 125I-7-AIPP-Fsk specifically labeled a 45-kDa protein and not a 115-kDa protein in control and GTP {gamma} S-preactivated brain membranes. This labeling was inhibited by forskolin, 1,9-dideoxyforskolin, cytochalasin B, and D-glucose but not cytochalasin E or L-glucose. Human erythrocyte membranes were photolyzed with 125I-6-AIPP-Fsk and 125I-7-AIPP-Fsk. 125I-7-AIPP-Fsk, but not 125I-6-AIPP-Fsk, strongly labeled a broad 45-70-kDa band. Forskolin, 7-bromoacetyl-7-desacetylforskolin, 1,9-dideoxyforskolin, cytochalasin B, and D-glucose, but not cytochalasin E or L-glucose, inhibited 125I-7-AIPP-Fsk labeling of the 45-70-kDa band. 125I-6-AIPP-Fsk and 125I-7-AIPP-Fsk are high affinity photolabels with specificity for adenylyl cyclase and the glucose transporter, respectively.

  7. Reduction of glucose uptake through inhibition of hexose transporters and enhancement of their endocytosis by methylglyoxal in Saccharomyces cerevisiae.

    Science.gov (United States)

    Yoshida, Aya; Wei, Dandan; Nomura, Wataru; Izawa, Shingo; Inoue, Yoshiharu

    2012-01-02

    Diabetes mellitus is characterized by an impairment of glucose uptake even though blood glucose levels are increased. Methylglyoxal is derived from glycolysis and has been implicated in the development of diabetes mellitus, because methylglyoxal levels in blood and tissues are higher in diabetic patients than in healthy individuals. However, it remains to be elucidated whether such factors are a cause, or consequence, of diabetes. Here, we show that methylglyoxal inhibits the activity of mammalian glucose transporters using recombinant Saccharomyces cerevisiae cells genetically lacking all hexose transporters but carrying cDNA for human GLUT1 or rat GLUT4. We found that methylglyoxal inhibits yeast hexose transporters also. Glucose uptake was reduced in a stepwise manner following treatment with methylglyoxal, i.e. a rapid reduction within 5 min, followed by a slow and gradual reduction. The rapid reduction was due to the inhibitory effect of methylglyoxal on hexose transporters, whereas the slow and gradual reduction seemed due to endocytosis, which leads to a decrease in the amount of hexose transporters on the plasma membrane. We found that Rsp5, a HECT-type ubiquitin ligase, is responsible for the ubiquitination of hexose transporters. Intriguingly, Plc1 (phospholipase C) negatively regulated the endocytosis of hexose transporters in an Rsp5-dependent manner, although the methylglyoxal-induced endocytosis of hexose transporters occurred irrespective of Plc1. Meanwhile, the internalization of hexose transporters following treatment with methylglyoxal was delayed in a mutant defective in protein kinase C.

  8. Topology mapping of insulin-regulated glucose transporter GLUT4 using computational biology.

    Science.gov (United States)

    Chakraborty, Chiranjib; Bandyopadhyay, Sanghamitra; Maulik, Ujjwal; Agoramoorthy, Govindasamy

    2013-01-01

    The type 2 diabetes is increasing rapidly around the globe. The primary cause for this is insulin resistance due to the disruption of the insulin signal transduction mechanism. Insulin signal transduction stimulates glucose transport through the glucose transporter GLUT4, by promoting the exocytosis process. Understanding the structural topology of GLUT4 mechanism will increase our understanding of the dynamic activities about glucose transport and its regulation in the membrane environment. However, little is known about the topology of GLUT4. In this article, we have determined the amino acid composition, disulfide topology, structure conformation pattern of GLUT4. The amino acid composition portrays that leucine composition is the highest contributing to 15.5% among all other amino acids. Three cysteine residues such as Cys223, Cys361, and Cys363 were observed and the last two were associated with one disulfide bond formation. We have generated surface cavities to know the clefts/pockets on the surface of this protein that showed few irregular cavities placed mostly in the transmembrane-helical part. Besides, topology mapping of 12 transmembrane-helixes was done to predict N- and O-glycosylation sites and to show the highly glycosylated GLUT4 that includes both N- and O-glycosylation sites. Furthermore, hydrophobic segment and molecular charge distribution were analyzed. This article shows that bioinformatics tools can provide a rapid methodology to predict the topology of GLUT4. It also provides insights into the structural details and structural functioning relationships in the human GLUT4. The results can be of great help to advance future drug development research using GLUT4 as a target protein.

  9. Availability of neurotransmitter glutamate is diminished when beta-hydroxybutyrate replaces glucose in cultured neurons

    DEFF Research Database (Denmark)

    Lund, Trine Meldgaard; Risa, Øystein; Sonnewald, Ursula

    2009-01-01

    Ketone bodies serve as alternative energy substrates for the brain in cases of low glucose availability such as during starvation or in patients treated with a ketogenic diet. The ketone bodies are metabolized via a distinct pathway confined to the mitochondria. We have compared metabolism of [2...

  10. Availability of neurotransmitter glutamate is diminished when beta-hydroxybutyrate replaces glucose in cultured neurons

    DEFF Research Database (Denmark)

    Lund, Trine Meldgaard; Risa, Øystein; Sonnewald, Ursula;

    2009-01-01

    Ketone bodies serve as alternative energy substrates for the brain in cases of low glucose availability such as during starvation or in patients treated with a ketogenic diet. The ketone bodies are metabolized via a distinct pathway confined to the mitochondria. We have compared metabolism of [2...

  11. Interaction of Peptide Transporter 1 With D-Glucose and L-Glutamic Acid; Possible Involvement of Taste Receptors.

    Science.gov (United States)

    Arakawa, Hiroshi; Ohmachi, Taichi; Ichiba, Kiko; Kamioka, Hiroki; Tomono, Takumi; Kanagawa, Masahiko; Idota, Yoko; Hatano, Yasuko; Yano, Kentaro; Morimoto, Kaori; Ogihara, Takuo

    2016-01-01

    We investigated the influence of sweet and umami (savory) tastants on the intestinal absorption of cephalexin (CEX), a substrate of peptide transporter 1 (PEPT1, SLC15A1) in rats. After oral administration of glucose or mannitol to rats, CEX was administered together with a second dose of glucose or mannitol. Western blot analysis indicated that expression of PEPT1 in rat jejunum membrane was decreased by glucose, compared to mannitol. Furthermore, the maximum plasma concentration (Cmax) of orally administered CEX was reduced by glucose compared to mannitol. The effect of glucose was diminished by nifedipine, a L-type Ca(2+) channel blocker. We also found that Cmax of orally administered CEX was reduced by treatment with L-glutamic acid, compared to D-glutamic acid. Thus, excessive intake of glucose and L-glutamic acid may impair oral absorption of PEPT1 substrates.

  12. Cryo electron tomography of herpes simplex virus during axonal transport and secondary envelopment in primary neurons.

    Directory of Open Access Journals (Sweden)

    Iosune Ibiricu

    2011-12-01

    Full Text Available During herpes simplex virus 1 (HSV1 egress in neurons, viral particles travel from the neuronal cell body along the axon towards the synapse. Whether HSV1 particles are transported as enveloped virions as proposed by the 'married' model or as non-enveloped capsids suggested by the 'separate' model is controversial. Specific viral proteins may form a recruitment platform for microtubule motors that catalyze such transport. However, their subviral location has remained elusive. Here we established a system to analyze herpesvirus egress by cryo electron tomography. At 16 h post infection, we observed intra-axonal transport of progeny HSV1 viral particles in dissociated hippocampal neurons by live-cell fluorescence microscopy. Cryo electron tomography of frozen-hydrated neurons revealed that most egressing capsids were transported independently of the viral envelope. Unexpectedly, we found not only DNA-containing capsids (cytosolic C-capsids, but also capsids lacking DNA (cytosolic A-/B-capsids in mid-axon regions. Subvolume averaging revealed lower amounts of tegument on cytosolic A-/B-capsids than on C-capsids. Nevertheless, all capsid types underwent active axonal transport. Therefore, even few tegument proteins on the capsid vertices seemed to suffice for transport. Secondary envelopment of capsids was observed at axon terminals. On their luminal face, the enveloping vesicles were studded with typical glycoprotein-like spikes. Furthermore, we noted an accretion of tegument density at the concave cytosolic face of the vesicle membrane in close proximity to the capsids. Three-dimensional analysis revealed that these assembly sites lacked cytoskeletal elements, but that filamentous actin surrounded them and formed an assembly compartment. Our data support the 'separate model' for HSV1 egress, i.e. progeny herpes viruses being transported along axons as subassemblies and not as complete virions within transport vesicles.

  13. Characterization of the avian GLUT1 glucose transporter: differential regulation of GLUT1 and GLUT3 in chicken embryo fibroblasts.

    Science.gov (United States)

    Wagstaff, P; Kang, H Y; Mylott, D; Robbins, P J; White, M K

    1995-01-01

    Vertebrate cells that are transformed by oncogenes such as v-src or are stimulated by mitogens have increased rates of glucose uptake. In rodent cells, the mechanisms whereby glucose transport is up-regulated are well understood. Stimulation of glucose transport involves an elevation in mRNA encoding the GLUT1 glucose transporter that is controlled at the levels of both transcription and mRNA stability. Cloning and sequencing of chicken GLUT1 cDNA showed that it shares 95% amino acid sequence similarity to mammalian GLUT1s. Nevertheless, unlike mammalian GLUT1 mRNA, it was not induced by v-src, serum addition, or treatment with the tumor promoter 12-O-tetradecanoylphorbol 13-acetate in chicken embryo fibroblasts. Rather, the induction of glucose transport in chicken embryo fibroblasts by v-src, serum, and 12-O-tetradecanoylphorbol 13-acetate was associated with induction of GLUT3 mRNA level and GLUT3 transcription. Rat fibroblasts were also found to express both GLUT1 and GLUT3 isoforms, but v-src induced GLUT1 and not GLUT3. This suggests that animal cells require both a basal and an upregulatable glucose transporter and that these functions have been subsumed by different GLUT isoforms in avian and mammalian cells. Images PMID:8589457

  14. Impaired axonal transport in motor neurons correlates with clinical prion disease.

    Directory of Open Access Journals (Sweden)

    Vladimir Ermolayev

    2009-08-01

    Full Text Available Prion diseases are fatal neurodegenerative disorders causing motor dysfunctions, dementia and neuropathological changes such as spongiosis, astroglyosis and neuronal loss. The chain of events leading to the clinical disease and the role of distinct brain areas are still poorly understood. The role of nervous system integrity and axonal properties in prion pathology are still elusive. There is no evidence of both the functional axonal impairments in vivo and their connection with prion disease. We studied the functional axonal impairments in motor neurons at the onset of clinical prion disease using the combination of tracing as a functional assay for axonal transport with immunohistochemistry experiments. Well-established and novel confocal and ultramicroscopy techniques were used to image and quantify labeled neurons. Despite profound differences in the incubation times, 30% to 45% of neurons in the red nucleus of different mouse lines showed axonal transport impairments at the disease onset bilaterally after intracerebral prion inoculation and unilaterally -- after inoculation into the right sciatic nerve. Up to 94% of motor cortex neurons also demonstrated transport defects upon analysis by alternative imaging methods. Our data connect axonal transport impairments with disease symptoms for different prion strains and inoculation routes and establish further insight on the development of prion pathology in vivo. The alterations in localization of the proteins involved in the retrograde axonal transport allow us to propose a mechanism of transport disruption, which involves Rab7-mediated cargo attachment to the dynein-dynactin pathway. These findings suggest novel targets for therapeutic and diagnostic approaches in the early stages of prion disease.

  15. Overexpression of glucose transporter-1 (GLUT-1) predicts poor prognosis in epithelial ovarian cancer.

    Science.gov (United States)

    Cho, Hanbyoul; Lee, You Sun; Kim, Julie; Chung, Joon-Yong; Kim, Jae-Hoon

    2013-11-01

    Illumina microarray was used to identify differentially expressed genes in three epithelial ovarian cancer (EOC) cells. To validate the microarray data, mRNA and protein level of glucose transporter-1 (GLUT-1) was examined. GLUT-1 had an EOC/normal cells ratio of 5.51 based on microarray. Real-time PCR and immunohistochemistry demonstrated that GLUT-1 expression was significantly increased in EOC (p = .029 and p GLUT-1 overexpression (HR = 4.80, p = .027) and lymph node metastases (HR = 8.35, p = .016) conferred a significantly worse overall survival. In conclusion, GLUT-1 expression is remarkably upregulated in EOC and predicts a poor overall survival.

  16. Water transport by the Na+/glucose cotransporter under isotonic conditions

    DEFF Research Database (Denmark)

    Zeuthen, T; Meinild, A K; Klaerke, D A

    1997-01-01

    in Xenopus oocytes. We present a method which allows short-term exposures to sugar under voltage clamp conditions. We demonstrate that water is cotransported with the solutes despite no osmotic differences between the external and intracellular solutions. There is a fixed ratio of 195:1 between the number...... of water molecules and the number of Na+ ions transported, equivalent to 390 water molecules per glucose molecule. Unstirred layer effects are ruled out on the basis of experiments on native oocytes incubated with the ionophores gramicidin D or nystatin....

  17. β-Hydroxybutyrate Boosts Mitochondrial and Neuronal Metabolism but is not Preferred Over Glucose Under Activated Conditions.

    Science.gov (United States)

    Achanta, Lavanya B; Rowlands, Benjamin D; Thomas, Donald S; Housley, Gary D; Rae, Caroline D

    2017-06-01

    The ketone body, β-hydroxybutyrate (βOHB), is metabolised by the brain alongside the mandatory brain fuel glucose. To examine the extent and circumstances by which βOHB can supplement glucose metabolism, we studied guinea pig cortical brain slices using increasing concentrations of [U-(13)C]D-βOHB in conjunction with [1-(13)C]D-glucose under conditions of normo- and hypoglycaemia, as well as under high potassium (40 mmol/L K(+)) depolarization in normo- and hypoglycaemic conditions. The contribution of βOHB to synthesis of GABA was also probed by inhibiting the synthesis of glutamine, a GABA precursor, with methionine sulfoximine (MSO). [U-(13)C]D-βOHB at lower concentrations (0.25 and 1.25 mmol/L) stimulated mitochondrial metabolism, producing greater total incorporation of label into glutamate and GABA but did not have a similar effect in the cytosolic compartment where labelling of glutamine was reduced at 1.25 mmol/L [U-(13)C]D-βOHB. At higher concentrations (2.5 mmol/L) [U-(13)C]D-βOHB inhibited metabolism of [1-(13)C]D-glucose, and reduced total label incorporation and total metabolite pools. When glucose levels were reduced, βOHB was able to partially restore the loss of glutamate and GABA caused by hypoglycaemia, but was not able to supplement levels of lactate, glutamine or alanine or to prevent the increase in aspartate. Under depolarizing conditions glucose was the preferred substrate over βOHB, even in hypoglycaemic conditions where comparatively less βOHB was incorporated except into aspartate isotopomers. Inhibition of glutamine synthesis with MSO had no significant effect on incorporation of label from [U-(13)C]D-βOHB into GABA C2,1 indicating that the majority of this GABA was synthesized in GABAergic neurons from [U-(13)C]D-βOHB rather than from Gln C4,5 imported from astrocytes.

  18. Preferential Transport and Metabolism of Glucose in Bergmann Glia over Purkinje Cells: A Multiphoton Study of Cerebellar Slices

    Institute of Scientific and Technical Information of China (English)

    L.F.BARROS; R.COURJARET; P.JAKOBY; A.LOAIZA; C.LOHR; J.W.DEITMER

    2009-01-01

    horizontally across the molecular layer, presumably through gap junctions between Bergmann glial cells. Our main conclusion is that in acute cerebellar slices, the glucose transport capacity and glycolytic rate of Bergmann glia are several-fold higher than those of Purkinje cells. Given that the cerebellum is largely fueled by glucose and Purkinje neurons are estimated to spend more energy than Bergmann glial cells, these results suggest substantial shuttling of an energy-rich metabolite like

  19. Effects of glucose and insulin on the H9c2 (2-1) cell proliferation may be mediated through regulating glucose transporter 4 expression

    Institute of Scientific and Technical Information of China (English)

    LIU Qian; HUANG Qing-xian; LOU Fu-chen; ZHANG Li; WANG Kun; YU Shan; XU Hua

    2013-01-01

    Background The change of glucose transporter 4 (GLUT4) expression could influence glucose uptake in the myocardial cells and then effect myocardial metabolism,which maybe one of the factor for the diabetes cardiovascular disease.This study aimed to explore the influence of glucose and insulin at different concentrations on H9c2 (2-1) cell proliferation and its GLUT4 expression in vitro,and evaluate the correlation between myocardial cells proliferation and GLUT4 expression.This might be helpful for understanding the relationship between glucose metabolism and cardiovascular disease.Methods According to glucose concentrations in culture medium,cultured H9c2 rat myocardial cells were divided into five groups:control group (NC,glucose concentration 5.0 mmol/L),low glucose group (LG,glucose concentration 0.1 mmol/L),high glucose group 1 (HG1,glucose concentration 10 mmol/L),high glucose group 2 (HG2,glucose concentration 15 mmol/L),high glucose group 3 (HG3,glucose concentration 20 mmol/L).Then according to different insulin concentrations in culture medium,each group was further divided into two subgroups:normal insulin subgroup (INSc,insulin concentration 3.8 mU/L),high insulin subgroup (INSh,insulin concentration 7.6 mU/L).H9c2 (2-1) cells were cultured for 1,2,3 days,the proliferation of cells were assayed by cell counting Kit-8 assay,the expressions of GLUT4 mRNA and protein were detected with RT-PCR and Western Blotting technique,and the relation between myocardial cells proliferation and GLUT4 expression was evaluated.Results Compared with NC group,cell proliferation (OD value) was lower in LG,HG2,HG3 group but higher in HG1 group on the second and the third day (P <0.05).There was a negative correlation between OD value and the glucose level in HG1,HG2,HG3 groups (P <0.05).OD value in INSc subgroups was lower than that in INSh subgroups (P <0.05).GLUT4 mRNA was lower in LG,HG2,HG3 groups than that in NC group (P <0.05).Compared with NC group,GLUT4 m

  20. Gadd45b prevents autophagy and apoptosis against rat cerebral neuron oxygen-glucose deprivation/reperfusion injury.

    Science.gov (United States)

    He, Guoqian; Xu, Wenming; Tong, Linyan; Li, Shuaishuai; Su, Shiceng; Tan, Xiaodan; Li, Changqing

    2016-04-01

    Autophagic (type II) cell death has been suggested to play pathogenetic roles in cerebral ischemia. Growth arrest and DNA damage response 45b (Gadd45b) has been shown to protect against rat brain ischemia injury through inhibiting apoptosis. However, the relationship between Gadd45b and autophagy in cerebral ischemia/reperfusion (I/R) injury remains uncertain. The aim of this study is to investigate the effect of Gadd45b on autophagy. We adopt the oxygen-glucose deprivation and reperfusion (OGD/R) model of rat primary cortex neurons, and lentivirus interference used to silence Gadd45b expression. Cell viability and injury assay were performed using CCK-8 and LDH kit. Autophagy activation was monitored by expression of ATG5, LC3, Beclin-1, ATG7 and ATG3. Neuron apoptosis was monitored by expression of Bcl-2, Bax, cleaved caspase3, p53 and TUNEL assay. Neuron neurites were assayed by double immunofluorescent labeling with Tuj1 and LC3B. Here, we demonstrated that the expression of Gadd45b was strongly up-regulated at 24 h after 3 h OGD treatment. ShRNA-Gadd45b increased the expression of autophagy related proteins, aggravated OGD/R-induced neuron cell apoptosis and neurites injury. ShRNA-Gadd45b co-treatment with autophagy inhibitor 3-methyladenine (3-MA) or Wortmannin partly inhibited the ratio of LC3II/LC3I, and slightly ameliorated neuron cell apoptosis under OGD/R. Furthermore, shRNA-Gadd45b inhibited the p-p38 level involved in autophagy, but increased the p-JNK level involved in apoptosis. ShRNA-Gadd45b co-treatment with p38 inhibitor obviously induced autophagy. ShRNA-Gadd45b co-treatment with JNK inhibitor alleviated neuron cell apoptosis. In conclusion, our data suggested that Gadd45b inhibited autophagy and apoptosis under OGD/R. Gadd45b may be a common regulatory protein to control autophagy and apoptosis.

  1. Elevated skeletal muscle glucose transporter levels in exercise-trained middle-aged men.

    Science.gov (United States)

    Houmard, J A; Egan, P C; Neufer, P D; Friedman, J E; Wheeler, W S; Israel, R G; Dohm, G L

    1991-10-01

    Exercise training has been proposed to improve whole body insulin sensitivity through a postreceptor adaptation in skeletal muscle. This study examined if levels of the insulin-responsive muscle glucose transporter protein (GLUT-4) were associated with improved insulin sensitivity in trained vs. sedentary middle-aged individuals. Muscle GLUT-4 levels and oral glucose tolerance test (OGTT) responses were obtained in age-matched trained and sedentary men (n = 11). Plasma insulin levels during the OGTT were significantly lower (P less than 0.01) in the trained men, whereas no differences were seen in plasma glucose responses. GLUT-4 protein content was approximately twofold higher in the trained men (2.41 +/- 0.17 vs. 1.36 +/- 0.11 micrograms standard, P less than 0.001). OGTT responses and GLUT-4 levels were not altered 15-18 h after a standard exercise bout in six representative sedentary subjects. These data suggest that GLUT-4 levels are increased in conjunction with insulin sensitivity in chronically exercise-trained middle-aged men. This finding suggests a possible mechanism for the improved insulin sensitivity observed with exercise training in humans.

  2. Interference of the noradrenergic neurotoxin DSP4 with neuronal and nonneuronal monoamine transporters.

    Science.gov (United States)

    Wenge, Birger; Bönisch, Heinz

    2009-12-01

    The haloalkylamine DSP4 (N[-2-chloroethyl]-N-ethyl-2-bromobenzylamine) is a noradrenergic neurotoxin, which is used for the chemical denervation of noradrenergic neurons, and it has been proposed to be a selective substrate for the neuronal, Na(+)- and Cl(-)-dependent noradrenaline transporter (NAT). In the present study, we investigated whether DSP4 not only interacts with the human NAT (hNAT) but also with other neuronal monoamine transporters such as the transporters for dopamine (hDAT) and serotonin (hSERT) or with nonneuronal (Na(+)-independent) monoamine transporters also known as organic cation transporters (OCTs), such as hOCT(1), hOCT(2), and hOCT(3). Using human embryonic kidney HEK293 cells heterologously expressing the corresponding transporter, we show that DSP4 irreversibly inhibits the hNAT, hDAT, hSERT, and hOCT(3). However, this inhibition includes a reversible component at the hDAT, hSERT, and hOCT(3) but not at the hNAT. The inhibitory potency of DSP4 at the neuronal transporters was highest at the hNAT (IC(50) about 5 microM), and it was about five and 40 times lower at the hSERT and hDAT, respectively. DSP4 inhibited all three hOCTs with high potency (IC(50) about 1 microM) but in a completely reversible manner at hOCT(1) and hOCT(2). Cytotoxicity by 24-h exposure of hNAT- or hOCT-expressing cells to low DSP4 concentrations (DSP4's high-affinity uptake through the NAT together with its completely irreversible mode of interaction with the NAT may contribute to its selectivity as noradrenergic neurotoxin.

  3. Effect of puerarin on the P13K pathway for glucose transportation and insulin signal transduction in adipocytes

    Institute of Scientific and Technical Information of China (English)

    ZHAO Ying; ZHOU You; YIN Hui-jun; ZHANG Ying

    2009-01-01

    To explore the effect of puerarin on insulin receptor (IR), insulin receptor substrate-1 (IRS-1) and protein expression of protein kinase B(PKB) in the P13K pathway of the glucose consumption, transportation and insulin signal transduction in 3T3-L1 adipocytes with insulin resistance. The insulin resistance 3T3-L1 adiocytes model was established by free fatty acid induction. The model cells were managed with puerarin in different concentrations. Glucose consumption was detected with glucose oxidase method, glucose transportation rate was determined by 2-deoxy-3 H glucose ingesting method, and the IR, IRS-1 and PKB expression were determined by Western blot. Glucose consumption and transportation were significantly decreased in the model adipocytes, but increased after treated with puerarin (P < 0. 01 ). Moreover, the level of tyrosine phosphorylation of IR subunit βwas higher in the puerarin treated groups, and that of IRS-1 was higher in the group treated with low dose puerarin than that in the model group. The 3T3-L1 adipocytes of insulin resistance model could be induced by free fatty acid successfully, puerarin could promote the glucose utilization in them to alleviate the insulin resistance, which may be related with the action in advancing the tyrosine phosphorylation of IR and IRS-1.

  4. Ischemic preconditioning decreases intracellular zinc accumulation induced by oxygen-glucose deprivation in gerbil hippocampal CA1 neurons.

    Science.gov (United States)

    Miyawaki, Takahiro; Yokota, Hidenori; Oguro, Keiji; Kato, Kengo; Shimazaki, Kuniko

    2004-05-27

    In normal gerbils, intracellular zinc ions ([Zn2+]i) and calcium ions ([Ca2+]i) accumulate in hippocampal CA1 neurons after global ischemia. We examined whether ischemic preconditioning modifies these changes in gerbil hippocampal slices. In normal slices, large increases in [Zn2+]i and [Ca2+]i were observed in the stratum radiatum of the CA1 area after oxygen-glucose deprivation. In preconditioned slices, there were significantly decreased peak levels of [Zn2+]i and [Ca2+]i in CA1. However, there were no differences in the peak levels of these ions in CA3 and dentate gyrus. These results suggest that modified [Zn2+]i and [Ca2+]i accumulation after an ischemic insult might be important for the mechanisms of ischemic tolerance induced by preconditioning.

  5. Zinc transporter 5 and zinc transporter 7 induced by high glucose protects peritoneal mesothelial cells from undergoing apoptosis.

    Science.gov (United States)

    Zhang, Xiuli; Liang, Dan; Guo, Baolei; Deng, Wenyan; Chi, Zhi-Hong; Cai, Yuan; Wang, Lining; Ma, Jianfei

    2013-04-01

    Zinc is an essential micronutrient and cytoprotectant involved in many types of apoptosis. The zinc transporter family SLC30A (ZnTs) is an important factor in the regulation of zinc homeostasis; however, its function in apoptosis in peritoneal mesothelial cells (PMCs) remains unknown. This study explores the regulation of zinc transporters and how they play a role in cell survival, particularly in rat peritoneal mesothelial cells (RPMCs), surrounding glucose concentrations, and the molecular mechanism involved. The messenger RNA (mRNA) transcripts were quantitatively measured by real-time polymerase chain reaction for all known nine zinc transport exporters (SLC30A1-8,10), as well as in primary RPMCs and the cells cultured under nonstimulated and HG-stimulated conditions. While many zinc transporters were constitutively expressed, ZnT5 mRNA and ZnT7 mRNA were strongly induced by HG. Overexpression of ZnT5 and ZnT7 respectively resulted in a decrease in the expression of caspace 3, caspace 8, BAX, and AIF and coincided with cell survival in the presence of HG. Inhibition of ZnT5 and ZnT7 expression using considerable siRNA-mediated knockdown of RPMCs was examined and, afterwards, the impact on cell apoptosis was investigated. Increased levels of apoptosis were observed after knockdown of ZnT5 and ZnT7. Furthermore, overexpression of ZnT5 and ZnT7 is accompanied by activation of PI3K/Akt pathway and inhibiting HG-induced apoptosis. This study suggests that the zinc transporting system in RPMCs is influenced by exposure to HG, particularly ZnT5 and ZnT7. This may account for the inhibition of HG-induced RPMC apoptosis and peritoneum injury, likely through targeting PI3K/Akt pathway-mediated cell survival.

  6. Endoproteolytic cleavage of TUG protein regulates GLUT4 glucose transporter translocation.

    Science.gov (United States)

    Bogan, Jonathan S; Rubin, Bradley R; Yu, Chenfei; Löffler, Michael G; Orme, Charisse M; Belman, Jonathan P; McNally, Leah J; Hao, Mingming; Cresswell, James A

    2012-07-06

    To promote glucose uptake into fat and muscle cells, insulin causes the translocation of GLUT4 glucose transporters from intracellular vesicles to the cell surface. Previous data support a model in which TUG traps GLUT4-containing vesicles and tethers them intracellularly in unstimulated cells and in which insulin mobilizes this pool of vesicles by releasing this tether. Here we show that TUG undergoes site-specific endoproteolytic cleavage, which separates a GLUT4-binding, N-terminal region of TUG from a C-terminal region previously suggested to bind an intracellular anchor. Cleavage is accelerated by insulin stimulation in 3T3-L1 adipocytes and is highly dependent upon adipocyte differentiation. The N-terminal TUG cleavage product has properties of a novel 18-kDa ubiquitin-like modifier, which we call TUGUL. The C-terminal product is observed at the expected size of 42 kDa and also as a 54-kDa form that is released from membranes into the cytosol. In transfected cells, intact TUG links GLUT4 to PIST and also binds Golgin-160 through its C-terminal region. PIST is an effector of TC10α, a GTPase previously shown to transmit an insulin signal required for GLUT4 translocation, and we show using RNAi that TC10α is required for TUG proteolytic processing. Finally, we demonstrate that a cleavage-resistant form of TUG does not support highly insulin-responsive GLUT4 translocation or glucose uptake in 3T3-L1 adipocytes. Together with previous results, these data support a model whereby insulin stimulates TUG cleavage to liberate GLUT4 storage vesicles from the Golgi matrix, which promotes GLUT4 translocation to the cell surface and enhances glucose uptake.

  7. Immunohistochemical Evaluation of Glucose Transporter Type 1 in Epithelial Dysplasia and Oral Squamous Cell Carcinoma.

    Science.gov (United States)

    Pereira, Karuza Maria Alves; Feitosa, Sthefane Gomes; Lima, Ana Thayssa Tomaz; Luna, Ealber Carvalho Macedo; Cavalcante, Roberta Barroso; de Lima, Kenio Costa; Chaves, Filipe Nobre; Costa, Fábio Wildson Gurgel

    2016-01-01

    Oral squamous cell carcinoma (OSCC) is the most common malignancy of the oral cavity and some of these have been documented in association or preceded by oral epithelial dysplasia (OED). Aggressive cancers with fast growth have demonstrated overexpression of some glucose transporters (GLUTs). Thus, the aim of this study was to analyze the immunohistochemical expression of the glucose transporter, GLUT-1, in OEDs and OSCCs, seeking to better elucidate the biological behavior of neoplasias. Fifteen cases were selected this research of both lesions. Five areas were analyzed from each case by counting the percentage of positive cells at 400x magnification. Immunoreactivity of GLUT-1 was observed in 100% of the samples ranging from 54.2% to 86.2% for the OSCC and 73.9% to 97.4% for the OED. Statistical test revealed that there was greater overexpression of GLUT-1 in OED than the OSCC (p=0.01). It is believed the high expression of GLUT-1 may reflect the involvement of GLUT-1 in early stages of oral carcinogenesis.

  8. Paraoxonase 2 Facilitates Pancreatic Cancer Growth and Metastasis by Stimulating GLUT1-Mediated Glucose Transport.

    Science.gov (United States)

    Nagarajan, Arvindhan; Dogra, Shaillay Kumar; Sun, Lisha; Gandotra, Neeru; Ho, Thuy; Cai, Guoping; Cline, Gary; Kumar, Priti; Cowles, Robert A; Wajapeyee, Narendra

    2017-08-17

    Metabolic deregulation is a hallmark of human cancers, and the glycolytic and glutamine metabolism pathways were shown to be deregulated in pancreatic ductal adenocarcinoma (PDAC). To identify new metabolic regulators of PDAC tumor growth and metastasis, we systematically knocked down metabolic genes that were overexpressed in human PDAC tumor samples using short hairpin RNAs. We found that p53 transcriptionally represses paraoxonase 2 (PON2), which regulates GLUT1-mediated glucose transport via stomatin. The loss of PON2 initiates the cellular starvation response and activates AMP-activated protein kinase (AMPK). In turn, AMPK activates FOXO3A and its transcriptional target, PUMA, which induces anoikis to suppress PDAC tumor growth and metastasis. Pharmacological or genetic activation of AMPK, similar to PON2 inhibition, blocks PDAC tumor growth. Collectively, our results identify PON2 as a new modulator of glucose transport that regulates a pharmacologically tractable pathway necessary for PDAC tumor growth and metastasis. Copyright © 2017 Elsevier Inc. All rights reserved.

  9. Modified Atkins diet therapy for a case with glucose transporter type 1 deficiency syndrome.

    Science.gov (United States)

    Ito, Susumu; Oguni, Hirokazu; Ito, Yasushi; Ishigaki, Keiko; Ohinata, Junko; Osawa, Makiko

    2008-03-01

    Glucose transporter type 1 deficiency syndrome (GLUT-1 DS), giving rise to impaired glucose transport across the blood-brain barrier, is characterized by infantile seizures, complex motor disorders, global developmental delay, acquired microcephaly, and hypoglycorrhachia. GLUT-1 DS can be treated effectively with a ketogenic diet because it can provide an alternative fuel for brain metabolism; however, the excessive restriction of food intake involved frequently makes it difficult for patients to initiate or continue the diet. Recently, the modified Atkins diet, which is much less restrictive in terms of the total calorie and protein intake than the classical ketogenic diet, has been shown to be effective and well tolerated in children with intractable epilepsy. We successfully introduced the modified Atkins diet to a 7-year-old boy with GLUT-1 DS, whose caregivers refused ketogenic diet treatment because of strong concerns over restricting the diet. The modified Atkins diet should be considered for patients with GLUT-1 DS as an alternative to the traditional ketogenic diet.

  10. Sites of glucose transporter-4 vesicle fusion with the plasma membrane correlate spatially with microtubules.

    Directory of Open Access Journals (Sweden)

    Jennine M Dawicki-McKenna

    Full Text Available In adipocytes, vesicles containing glucose transporter-4 (GLUT4 redistribute from intracellular stores to the cell periphery in response to insulin stimulation. Vesicles then fuse with the plasma membrane, facilitating glucose transport into the cell. To gain insight into the details of microtubule involvement, we examined the spatial organization and dynamics of microtubules in relation to GLUT4 vesicle trafficking in living 3T3-L1 adipocytes using total internal reflection fluorescence (TIRF microscopy. Insulin stimulated an increase in microtubule density and curvature within the TIRF-illuminated region of the cell. The high degree of curvature and abrupt displacements of microtubules indicate that substantial forces act on microtubules. The time course of the microtubule density increase precedes that of the increase in intensity of fluorescently-tagged GLUT4 in this same region of the cell. In addition, portions of the microtubules are highly curved and are pulled closer to the cell cortex, as confirmed by Parallax microscopy. Microtubule disruption delayed and modestly reduced GLUT4 accumulation at the plasma membrane. Quantitative analysis revealed that fusions of GLUT4-containing vesicles with the plasma membrane, detected using insulin-regulated aminopeptidase with a pH-sensitive GFP tag (pHluorin, preferentially occur near microtubules. Interestingly, long-distance vesicle movement along microtubules visible at the cell surface prior to fusion does not appear to account for this proximity. We conclude that microtubules may be important in providing spatial information for GLUT4 vesicle fusion.

  11. A Protein Kinase C Phosphorylation Motif in GLUT1 Affects Glucose Transport and is Mutated in GLUT1 Deficiency Syndrome.

    Science.gov (United States)

    Lee, Eunice E; Ma, Jing; Sacharidou, Anastasia; Mi, Wentao; Salato, Valerie K; Nguyen, Nam; Jiang, Youxing; Pascual, Juan M; North, Paula E; Shaul, Philip W; Mettlen, Marcel; Wang, Richard C

    2015-06-04

    Protein kinase C has been implicated in the phosphorylation of the erythrocyte/brain glucose transporter, GLUT1, without a clear understanding of the site(s) of phosphorylation and the possible effects on glucose transport. Through in vitro kinase assays, mass spectrometry, and phosphospecific antibodies, we identify serine 226 in GLUT1 as a PKC phosphorylation site. Phosphorylation of S226 is required for the rapid increase in glucose uptake and enhanced cell surface localization of GLUT1 induced by the phorbol ester 12-O-tetradecanoyl-phorbol-13-acetate (TPA). Endogenous GLUT1 is phosphorylated on S226 in primary endothelial cells in response to TPA or VEGF. Several naturally occurring, pathogenic mutations that cause GLUT1 deficiency syndrome disrupt this PKC phosphomotif, impair the phosphorylation of S226 in vitro, and block TPA-mediated increases in glucose uptake. We demonstrate that the phosphorylation of GLUT1 on S226 regulates glucose transport and propose that this modification is important in the physiological regulation of glucose transport.

  12. Molecular mechanisms beyond glucose transport in diabetes-related male infertility.

    Science.gov (United States)

    Alves, M G; Martins, A D; Rato, L; Moreira, P I; Socorro, S; Oliveira, P F

    2013-05-01

    Diabetes mellitus (DM) is one of the greatest public health threats in modern societies. Although during a few years it was suggested that DM had no significant effect in male reproductive function, this view has been challenged in recent years. The increasing incidence of DM worldwide will inevitably result in a higher prevalence of this pathology in men of reproductive age and subfertility or infertility associated with DM is expected to dramatically rise in upcoming years. From a clinical perspective, the evaluation of semen parameters, as well as spermatozoa deoxyribonucleic acid (DNA) integrity, are often studied due to their direct implications in natural and assisted conception. Nevertheless, recent studies based on the molecular mechanisms beyond glucose transport in testicular cells provide new insights in DM-induced alterations in male reproductive health. Testicular cells have their own glucose sensing machinery that react to hormonal fluctuations and have several mechanisms to counteract hyper- and hypoglycemic events. Moreover, the metabolic cooperation between testicular cells is crucial for normal spermatogenesis. Sertoli cells (SCs), which are the main components of blood-testis barrier, are not only responsible for the physical support of germ cells but also for lactate production that is then metabolized by the developing germ cells. Any alteration in this tied metabolic cooperation may have a dramatic consequence in male fertility potential. Therefore, we present an overview of the clinical significance of DM in the male reproductive health with emphasis on the molecular mechanisms beyond glucose fluctuation and transport in testicular cells. Copyright © 2013 Elsevier B.V. All rights reserved.

  13. Glucose and Intermediary Metabolism and Astrocyte-Neuron Interactions Following Neonatal Hypoxia-Ischemia in Rat.

    Science.gov (United States)

    Brekke, Eva; Berger, Hester Rijkje; Widerøe, Marius; Sonnewald, Ursula; Morken, Tora Sund

    2017-01-01

    Neonatal hypoxia-ischemia (HI) and the delayed injury cascade that follows involve excitotoxicity, oxidative stress and mitochondrial failure. The susceptibility to excitotoxicity of the neonatal brain may be related to the capacity of astrocytes for glutamate uptake. Furthermore, the neonatal brain is vulnerable to oxidative stress, and the pentose phosphate pathway (PPP) may be of particular importance for limiting this kind of injury. Also, in the neonatal brain, neurons depend upon de novo synthesis of neurotransmitters via pyruvate carboxylase in astrocytes to increase neurotransmitter pools during normal brain development. Several recent publications describing intermediary brain metabolism following neonatal HI have yielded interesting results: (1) Following HI there is a prolonged depression of mitochondrial metabolism in agreement with emerging evidence of mitochondria as vulnerable targets in the delayed injury cascade. (2) Astrocytes, like neurons, are metabolically impaired following HI, and the degree of astrocytic malfunction may be an indicator of the outcome following hypoxic and hypoxic-ischemic brain injury. (3) Glutamate transfer from neurons to astrocytes is not increased following neonatal HI, which may imply that astrocytes fail to upregulate glutamate uptake in response to the massive glutamate release during HI, thus contributing to excitotoxicity. (4) In the neonatal brain, the activity of the PPP is reduced following HI, which may add to the susceptibility of the neonatal brain to oxidative stress. The present review aims to discuss the metabolic temporal alterations observed in the neonatal brain following HI.

  14. The carcinine transporter CarT is required in Drosophila photoreceptor neurons to sustain histamine recycling.

    Science.gov (United States)

    Stenesen, Drew; Moehlman, Andrew T; Krämer, Helmut

    2015-12-14

    Synaptic transmission from Drosophila photoreceptors to lamina neurons requires recycling of histamine neurotransmitter. Synaptic histamine is cleared by uptake into glia and conversion into carcinine, which functions as transport metabolite. How carcinine is transported from glia to photoreceptor neurons remains unclear. In a targeted RNAi screen for genes involved in this pathway, we identified carT, which encodes a member of the SLC22A transporter family. CarT expression in photoreceptors is necessary and sufficient for fly vision and behavior. Carcinine accumulates in the lamina of carT flies. Wild-type levels are restored by photoreceptor-specific expression of CarT, and endogenous tagging suggests CarT localizes to synaptic endings. Heterologous expression of CarT in S2 cells is sufficient for carcinine uptake, demonstrating the ability of CarT to utilize carcinine as a transport substrate. Together, our results demonstrate that CarT transports the histamine metabolite carcinine into photoreceptor neurons, thus contributing an essential step to the histamine-carcinine cycle.

  15. The carcinine transporter CarT is required in Drosophila photoreceptor neurons to sustain histamine recycling

    Science.gov (United States)

    Stenesen, Drew; Moehlman, Andrew T; Krämer, Helmut

    2015-01-01

    Synaptic transmission from Drosophila photoreceptors to lamina neurons requires recycling of histamine neurotransmitter. Synaptic histamine is cleared by uptake into glia and conversion into carcinine, which functions as transport metabolite. How carcinine is transported from glia to photoreceptor neurons remains unclear. In a targeted RNAi screen for genes involved in this pathway, we identified carT, which encodes a member of the SLC22A transporter family. CarT expression in photoreceptors is necessary and sufficient for fly vision and behavior. Carcinine accumulates in the lamina of carT flies. Wild-type levels are restored by photoreceptor-specific expression of CarT, and endogenous tagging suggests CarT localizes to synaptic endings. Heterologous expression of CarT in S2 cells is sufficient for carcinine uptake, demonstrating the ability of CarT to utilize carcinine as a transport substrate. Together, our results demonstrate that CarT transports the histamine metabolite carcinine into photoreceptor neurons, thus contributing an essential step to the histamine–carcinine cycle. DOI: http://dx.doi.org/10.7554/eLife.10972.001 PMID:26653853

  16. Concentration of membrane antigens by forward transport and trapping in neuronal growth cones.

    Science.gov (United States)

    Sheetz, M P; Baumrind, N L; Wayne, D B; Pearlman, A L

    1990-04-20

    Formation of the nervous system requires that neuronal growth cones follow specific paths and then stop at recognition signals, sensed at the growth cone's leading edge. We used antibody-coated gold particles viewed by video-enhanced differential interference contrast microscopy to observe the distribution and movement of two cell surface molecules, N-CAM and the 2A1 antigen, on growth cones of cultured cortical neurons. Gold particles are occasionally transported forward at 1-2 microns/s to the leading edge where they are trapped but continue to move. Concentration at the edge persists after cytochalasin D treatment or ATP depletion, but active movements to and along edges cease. We also observed a novel outward movement of small cytoplasmic aggregates at 1.8 microns/s in filopodia. We suggest that active forward transport and trapping involve reversible attachment of antigens to and transport along cytoskeletal elements localized to edges of growth cones.

  17. FDA-approved drugs that protect mammalian neurons from glucose toxicity slow aging dependent on cbp and protect against proteotoxicity.

    Directory of Open Access Journals (Sweden)

    Alex Lublin

    Full Text Available Screening a library of drugs with known safety profiles in humans yielded 30 drugs that reliably protected mammalian neurons against glucose toxicity. Subsequent screening demonstrated that 6 of these 30 drugs increase lifespan in C. elegans: caffeine, ciclopirox olamine, tannic acid, acetaminophen, bacitracin, and baicalein. Every drug significantly reduced the age-dependent acceleration of mortality rate. These protective effects were blocked by RNAi inhibition of cbp-1 in adults only, which also blocks protective effects of dietary restriction. Only 2 drugs, caffeine and tannic acid, exhibited a similar dependency on DAF-16. Caffeine, tannic acid, and bacitracin also reduced pathology in a transgenic model of proteotoxicity associated with Alzheimer's disease. These results further support a key role for glucose toxicity in driving age-related pathologies and for CBP-1 in protection against age-related pathologies. These results also provide novel lead compounds with known safety profiles in human for treatment of age-related diseases, including Alzheimer's disease and diabetic complications.

  18. Protection from neuronal damage induced by combined oxygen and glucose deprivation in organotypic hippocampal cultures by glutamate receptor antagonists.

    Science.gov (United States)

    Strasser, U; Fischer, G

    1995-07-31

    Organotypic hippocampal cultures were exposed to defined periods (30 and 60 min) of combined oxygen and glucose deprivation, mimicking transient ischemic conditions. The involvement of different glutamate receptors in individual hippocampal subfields (CA1, CA3 and dentate gyrus) was studied using antagonists of NMDA (dizocilpine) and AMPA/kainate receptors (CNQX and GYKI 52466). Staining with the fluorescent dye propidium iodide (PI) allowed detection of damaged cells. For quantitative determination of neuronal damage, fluorescence intensity was measured after a 22 h recovery period and was related to maximal fluorescence intensity measured after fixation and PI restaining of the cultures at the end of the experiment. Dizocilpine (10 microM), CNQX (100 microM) and GYKI 52466 (100 microM) provided complete protection in CA1, CA3 and dentate gyrus following the moderate ischemic insult, when the antagonists were present permanently. This indicates that none of the ionotropic glutamate receptor subtypes dominated toxicity in the most sensitive subpopulation of neurons. When applied only during the recovery period protection with dizocilpine (10 microM) or CNQX (100 microM) was drastically reduced by about 60% in the most sensitive area (CA1), but only slightly by 15% in CA3. Therefore the onset of irreversible damage seems to occur earlier in CA1 than in CA3. Blockade of AMPA/kainate receptors by GYKI 52466 (100 microM) offered no neuroprotection if the compound was applied only during the recovery period.(ABSTRACT TRUNCATED AT 250 WORDS)

  19. Amyloid-Beta Induced Changes in Vesicular Transport of BDNF in Hippocampal Neurons

    Directory of Open Access Journals (Sweden)

    Bianca Seifert

    2016-01-01

    Full Text Available The neurotrophin brain derived neurotrophic factor (BDNF is an important growth factor in the CNS. Deficits in transport of this secretory protein could underlie neurodegenerative diseases. Investigation of disease-related changes in BDNF transport might provide insights into the cellular mechanism underlying, for example, Alzheimer’s disease (AD. To analyze the role of BDNF transport in AD, live cell imaging of fluorescently labeled BDNF was performed in hippocampal neurons of different AD model systems. BDNF and APP colocalized with low incidence in vesicular structures. Anterograde as well as retrograde transport of BDNF vesicles was reduced and these effects were mediated by factors released from hippocampal neurons into the extracellular medium. Transport of BDNF was altered at a very early time point after onset of human APP expression or after acute amyloid-beta(1-42 treatment, while the activity-dependent release of BDNF remained unaffected. Taken together, extracellular cleavage products of APP induced rapid changes in anterograde and retrograde transport of BDNF-containing vesicles while release of BDNF was unaffected by transgenic expression of mutated APP. These early transport deficits might lead to permanently impaired brain functions in the adult brain.

  20. Intracellular pH regulation by acid/base transporters in mammalian neurons

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    Vernon A. Ruffin

    2014-02-01

    Full Text Available Intracellular pH (pHi regulation in the brain is important in both physiological and physiopathological conditions because changes in pHi generally result in altered neuronal excitability. In this review, we will cover 4 major areas: [1] The effect of pHi on cellular processes in the brain, including channel activity and neuronal excitability. [2] pHi homeostasis and how it is determined by the balance between rates of acid loading (JL and extrusion (JE. The balance between JE and JL determine steady-state pHi, as well as the ability of the cell to defend pHi in the face of extracellular acid-base disturbances (e.g. metabolic acidosis. [3] The properties and importance of members of the SLC4 and SLC9 families of acid-base transporters expressed in the brain that contribute to JL (namely the Cl-HCO3 exchanger AE3 and JE (the Na-H exchangers NHE1, NHE3 and NHE5 as well as the Na+- coupled HCO3- transporters NBCe1, NBCn1, NDCBE, and NBCn2. [4] The effect of acid-base disturbances on neuronal function and the roles of acid-base transporters in defending neuronal pHi under physiopathologic conditions.

  1. Characterization of cerebral glucose dynamics in vivo with a four-state conformational model of transport at the blood-brain barrier.

    Science.gov (United States)

    Duarte, João M N; Gruetter, Rolf

    2012-05-01

    Determination of brain glucose transport kinetics in vivo at steady-state typically does not allow distinguishing apparent maximum transport rate (T(max)) from cerebral consumption rate. Using a four-state conformational model of glucose transport, we show that simultaneous dynamic measurement of brain and plasma glucose concentrations provide enough information for independent and reliable determination of the two rates. In addition, although dynamic glucose homeostasis can be described with a reversible Michaelis-Menten model, which is implicit to the large iso-inhibition constant (K(ii)) relative to physiological brain glucose content, we found that the apparent affinity constant (K(t)) was better determined with the four-state conformational model of glucose transport than with any of the other models tested. Furthermore, we confirmed the utility of the present method to determine glucose transport and consumption by analysing the modulation of both glucose transport and consumption by anaesthesia conditions that modify cerebral activity. In particular, deep thiopental anaesthesia caused a significant reduction of both T(max) and cerebral metabolic rate for glucose consumption. In conclusion, dynamic measurement of brain glucose in vivo in function of plasma glucose allows robust determination of both glucose uptake and consumption kinetics.

  2. Experimental lead poisoning and intestinal transport of glucose, amino acids, and sodium.

    Science.gov (United States)

    Wapnir, R A; Exeni, R A; McVicar, M; Lipshitz, F

    1977-03-01

    Juvenile rats fed a diet containing 1% lead acetate for 7 weeks, in addition to an impaired growth rate and renal function derangements, suffered malabsorption of glucose and certain amino acids, as assessed by an in vivo perfusion technique. The reduction in glucose absorption ranged between 10% and 31% when the carbohydrate was pumped in concentrations of 2-80 mM. This alteration was compatible with a noncompetitive type of transport inhibition. The intestinal absorption of glycine, lysine, and phenylalanine were, respectively, decreased 22, 18, and 15% when these amino acids were present at 1 mM levels. Sodium transport was severely reduced (57.6 +/- 17.9 (SEM) vs. 124.2 +/- 17.4 muEq/min-cm) and intestinal mucosa (Na+-K+)-ATPase was concomitantly lower in the lead-intoxicated rats (186.4 +/- 19.0 vs 268.4 +/- 29.8 nmol P/min-mg protein). However, this enzyme was not altered in liver and kidney. Furthermore, intestinal mucosa fructose-1,6-diphosphatase, succinic dehydrogenase, pyruvate kinase, and tryptophan hydroxylase were not different in experimental and control animals. These studies substantiate the presence of functional and biochemical abnormalities in the intestinal mucosa of young rats when fed substantial amounts of a soluble lead salt. It is, therefore, reasonable to accept the possibility that physiologic damage occurs in tissues directly subjected to high and persistent levels of a toxic agents, as it occurs in other organs, underscoring the parallelism between transport mechanisms at the renal and intestinal levels.

  3. Prolactin-induced Subcellular Targeting of GLUT1 Glucose Transporter in Living Mammary Epithelial Cells

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    Arieh Riskin

    2015-10-01

    Full Text Available Background: Studying the biological pathways involved in mammalian milk production during lactation could have many clinical implications. The mammary gland is unique in its requirement for transport of free glucose into the cell for the synthesis of lactose, the primary carbohydrate in milk. Objective: To study GLUT1 trafficking and subcellular targeting in living mammary epithelial cells (MEC in culture. Methods: Immunocytochemistry was used to study GLUT1 hormonally regulated subcellular targeting in human MEC (HMEC. To study GLUT1 targeting and recycling in living mouse MEC (MMEC in culture, we constructed fusion proteins of GLUT1 and green fluorescent protein (GFP and expressed them in CIT3 MMEC. Cells were maintained in growth medium (GM, or exposed to secretion medium (SM, containing prolactin. Results: GLUT1 in HMEC localized primarily to the plasma membrane in GM. After exposure to prolactin for 4 days, GLUT1 was targeted intracellularly and demonstrated a perinuclear distribution, co-localizing with lactose synthetase. The dynamic trafficking of GFP-GLUT1 fusion proteins in CIT3 MMEC suggested a basal constitutive GLUT1 recycling pathway between an intracellular pool and the cell surface that targets most GLUT1 to the plasma membrane in GM. Upon exposure to prolactin in SM, GLUT1 was specifically targeted intracellularly within 90–110 minutes. Conclusions: Our studies suggest intracellular targeting of GLUT1 to the central vesicular transport system upon exposure to prolactin. The existence of a dynamic prolactin-induced sorting machinery for GLUT1 could be important for transport of free glucose into the Golgi for lactose synthesis during lactation.

  4. The effects of sodium-glucose co-transporter 2 inhibitors in patients with type 2 diabetes

    DEFF Research Database (Denmark)

    Storgaard, Heidi; Gluud, Lise Lotte; Christensen, Mikkel

    2014-01-01

    to the knowledge regarding the beneficial and harmful effects of SGLT-2i in patients with type 2 diabetes. We plan to publish the study irrespective of the results. RESULTS: The study will be disseminated by peer-review publication and conference presentation. TRIAL REGISTRATION NUMBER: PROSPERO CRD42014008960......INTRODUCTION: Sodium-glucose co-transporter 2 inhibitors (SGLT-2i) increase urinary glucose excretion through a reduced renal glucose reabsorption. We plan to perform a systematic review of SGLT-2i for treatment of type 2 diabetes. METHODS AND ANALYSIS: A systematic review with meta......-analyses of randomised clinical trials on SGLT-2i versus placebo, other oral glucose lowering drugs or insulin for patients with type 2 diabetes will be performed. The primary end point will be the glycated haemoglobin. Secondary end points will include changes in body weight, body mass index, fasting plasma glucose...

  5. In Vitro Infection of Trypanosoma cruzi Causes Decrease in Glucose Transporter Protein-1 (GLUT1 Expression in Explants of Human Placental Villi Cultured under Normal and High Glucose Concentrations

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    Luciana Mezzano

    2012-01-01

    Full Text Available Trypanosoma cruzi, the etiologic Chagas' disease agent, induces changes in protein pattern of the human placenta syncytiotrophoblast. The glucose transporter protein-1 (GLUT1 is the primary isoform involved in transplacental glucose transport. We carried out in vitro assays to determine if T. cruzi infection would induce changes in placental GLUT1 protein expression under normal and high concentration of glucose. Using Western blot and immunohistological techniques, GLUT1 expression was determined in normal placental villi cultured under normal or high concentrations of glucose, with or without in vitro T. cruzi infection, for 24 and 48 hours. High glucose media or T. cruzi infection alone reduced GLUT1 expression. A yet more accentuated reduction was observed when infection and high glucose condition took place together. We inform, for the first time, that T. cruzi infection may induce reduction of GLUT1 expression under normal and high glucose concentrations, and this effect is synergic to high glucose concentrations.

  6. Insulin-sensitive regulation of glucose transport and GLUT4 translocation in skeletal muscle of GLUT1 transgenic mice.

    Science.gov (United States)

    Etgen, G J; Zavadoski, W J; Holman, G D; Gibbs, E M

    1999-01-01

    Skeletal muscle glucose transport was examined in transgenic mice overexpressing the glucose transporter GLUT1 using both the isolated incubated-muscle preparation and the hind-limb perfusion technique. In the absence of insulin, 2-deoxy-d-glucose uptake was increased approximately 3-8-fold in isolated fast-twitch muscles of GLUT1 transgenic mice compared with non-transgenic siblings. Similarly, basal glucose transport activity was increased approximately 4-14-fold in perfused fast-twitch muscles of transgenic mice. In non-transgenic mice insulin accelerated glucose transport activity approximately 2-3-fold in isolated muscles and to a much greater extent ( approximately 7-20-fold) in perfused hind-limb preparations. The observed effect of insulin on glucose transport in transgenic muscle was similarly dependent upon the technique used for measurement, as insulin had no effect on isolated fast-twitch muscle from transgenic mice, but significantly enhanced glucose transport in perfused fast-twitch muscle from transgenic mice to approximately 50-75% of the magnitude of the increase observed in non-transgenic mice. Cell-surface glucose transporter content was assessed via 2-N-4-(l-azi-2,2,2-trifluoroethyl)benzoyl-1,3-bis-(d -mannos-4-yloxy)-2-propylamine photolabelling methodology in both isolated and perfused extensor digitorum longus (EDL). Cell-surface GLUT1 was enhanced by as much as 70-fold in both isolated and perfused EDL of transgenic mice. Insulin did not alter cell-surface GLUT1 in either transgenic or non-transgenic mice. Basal levels of cell-surface GLUT4, measured in either isolated or perfused EDL, were similar in transgenic and non-transgenic mice. Interestingly, insulin enhanced cell-surface GLUT4 approximately 2-fold in isolated EDL and approximately 6-fold in perfused EDL of both transgenic and non-transgenic mice. In summary, these results reveal differences between isolated muscle and perfused hind-limb techniques, with the latter method showing a

  7. Expression of conventional and novel glucose transporters, GLUT1, -9, -10, and -12, in vascular smooth muscle cells

    OpenAIRE

    Pyla, Rajkumar; Poulose, Ninu; Jun, John Y.; Segar, Lakshman

    2013-01-01

    Intimal hyperplasia is characterized by exaggerated proliferation of vascular smooth muscle cells (VSMCs). Enhanced VSMC growth is dependent on increased glucose uptake and metabolism. Facilitative glucose transporters (GLUTs) are comprised of conventional GLUT isoforms (GLUT1–5) and novel GLUT isoforms (GLUT6–14). Previous studies demonstrate that GLUT1 overexpression or GLUT10 downregulation contribute to phenotypic changes in VSMCs. To date, the expression profile of all 14 GLUT isoforms h...

  8. Xylose and xylose/glucose co-fermentation by recombinant Saccharomyces cerevisiae strains expressing individual hexose transporters.

    Science.gov (United States)

    Gonçalves, Davi L; Matsushika, Akinori; de Sales, Belisa B; Goshima, Tetsuya; Bon, Elba P S; Stambuk, Boris U

    2014-09-01

    Since the uptake of xylose is believed to be one of the rate-limiting steps for xylose ethanol fermentation by recombinant Saccharomyces cerevisiae strains, we transformed a hxt-null strain lacking the major hexose transporters (hxt1Δ-hxt7Δ and gal2Δ) with an integrative plasmid to overexpress the genes for xylose reductase (XYL1), xylitol dehydrogenase (XYL2) and xylulokinase (XKS1), and analyzed the impact that overexpression of the HXT1, HXT2, HXT5 or HXT7 permeases have in anaerobic batch fermentations using xylose, glucose, or xylose plus glucose as carbon sources. Our results revealed that the low-affinity HXT1 permease allowed the maximal consumption of sugars and ethanol production rates during xylose/glucose co-fermentations, but was incapable to allow xylose uptake when this sugar was the only carbon source. The moderately high-affinity HXT5 permease was a poor glucose transporter, and it also did not allow significant xylose uptake by the cells. The moderately high-affinity HXT2 permease allowed xylose uptake with the same rates as those observed during glucose consumption, even under co-fermentation conditions, but had the drawback of producing incomplete fermentations. Finally, the high-affinity HXT7 permease allowed efficient xylose fermentation, but during xylose/glucose co-fermentations this permease showed a clear preference for glucose. Thus, our results indicate that approaches to engineer S. cerevisiae HXT transporters to improve second generation bioethanol production need to consider the composition of the biomass sugar syrup, whereby the HXT1 transporter seems more suitable for hydrolysates containing xylose/glucose blends, whereas the HXT7 permease would be a better choice for xylose-enriched sugar streams.

  9. Contraction-stimulated glucose transport in muscle is controlled by AMPK and mechanical stress but not sarcoplasmatic reticulum Ca2+ release

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    Thomas E. Jensen

    2014-10-01

    Full Text Available Understanding how muscle contraction orchestrates insulin-independent muscle glucose transport may enable development of hyperglycemia-treating drugs. The prevailing concept implicates Ca2+ as a key feed forward regulator of glucose transport with secondary fine-tuning by metabolic feedback signals through proteins such as AMPK. Here, we demonstrate in incubated mouse muscle that Ca2+ release is neither sufficient nor strictly necessary to increase glucose transport. Rather, the glucose transport response is associated with metabolic feedback signals through AMPK, and mechanical stress-activated signals. Furthermore, artificial stimulation of AMPK combined with passive stretch of muscle is additive and sufficient to elicit the full contraction glucose transport response. These results suggest that ATP-turnover and mechanical stress feedback are sufficient to fully increase glucose transport during muscle contraction, and call for a major reconsideration of the established Ca2+ centric paradigm.

  10. The biological activity of Coccinia indica on glucose transporter 1 (GLUT1 promoter

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    Juntipa Purintrapiban

    2009-08-01

    Full Text Available Plant derivatives with purported hypoglycemic properties have been used in traditional medicine around the world. Coccinia indica (ivy gourd is used in traditional medicine to treat diabetics in many countries. C. indica is able to cause a reduction in blood glucose level and has shown hypoglycemic activity in vitro and in vivo. However, the mechanism of this effect remains unknown. In this study, we generated the pGL3-glucose transporter 1 (GLUT1 promoter to elucidate the molecular mechanism of the regulation of GLUT1 gene expression in response to a water extract of C. indica stem (CIextract. A fragment of 2.1 kb of rat GLUT1 promoter, located at -2,106 to +134, was linked to firefly luciferase. The regulating transcription was analyzed in transient expression assay after transfection and exposure of L6 myocytes with the GLUT1 promoter system and CI extract, respectively. Under normal condition (5 mM glucose, promoter activity induced by 0.15 mg CI extract was markedly increased by 5.71 fold from the basal value. CI extract was more effective than 2 mM metformin. Surprisingly, promoter activity in hyperglycemic condition (15 mM glucose induced by 0.50 mg CI was increased by 1.63 fold from the basal value. In addition, CI extract increased the 2-deoxyglucose (2-DG uptake in L6myocytes in a dose-dependent manner in both conditions, 5 mM and 15 mM glucose. GLUT1 protein was determined by Western blot analysis and the level also increased in a dose-dependent fashion. Interestingly, the activity of the -273 to +134 of GLUT1 promoter was increased by 2.12 fold from the basal value. This site is the transcription initiation site containing GC box and TATA box. These observations suggest that the hypoglycemic action of C. indica may regulate through the activation of GLUT1 promoter resulting in an increase of the GLUT1 protein expression.

  11.  The role of glucose transporter 1 (GLUT1 in the diagnosis and therapy of tumors

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    Paweł Jóźwiak

    2012-03-01

    Full Text Available  Malignant cells are known to enhance glucose metabolism, to increase glucose uptake and to inhibit the process of oxidative phosphorylation. Accelerated glycolysis is one of the biochemical characteristics of cancer cells that allow them to compensate the inefficient extraction of energy from glucose in order to continue their uncontrolled growth and proliferation. Upregulation of glucose transport across the plasma membrane is mediated by a family of facilitated glucose transporter proteins named GLUT. Overexpression of GLUTs, especially the hypoxia-responsive GLUT1, has been frequently observed in various human carcinomas. Many studies have reported a correlation between GLUT1 expression level and the grade of tumor aggressiveness, which suggests that GLUT1 expression may be of prognostic significance. Therefore, GLUT1 is a key rate-limiting factor in the transport and glucose metabolism in cancer cells. This paper presents the current state of knowledge on GLUT1 regulation as well as its utility in the diagnosis and therapy of cancers.

  12. Immunohistochemical expression of the glucose transporters Glut-1 and Glut-3 in human malignant melanomas and benign melanocytic lesions

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    Parente Paola

    2008-09-01

    Full Text Available Abstract Background Reported data indicate that cancer cells have increased rates of glucose metabolism, as determined by 18FDG-PET imaging in patients with malignancies. The results of many studies have demonstrated that the expression of glucose transporters, especially Glut-1, is increased in a variety of malignancies. This study was undertaken to assess the differential expression of Glut-1 and Glut-3 by benign and malignant melanocytic lesions. Methods Immunohistochemical staining for Glut-1 and Glut-3 was performed on paraffin-embedded tissue sections prepared from melanocytic nevi (12 cases, Spitz nevi (12 cases and primary cutaneous malignant melanomas (20 cases. Results We observed immunoreactivity for Glut-1 in all melanocytic nevi, 9 of the 12 Spitz nevi and in 9 of the 20 malignant melanomas, whereas Glut-3 was expressed in all the melanocytic lesions, both benign and malignant. Conclusion These findings indicate that the glucose transporters Glut-1 and Glut-3 play a role in the glucose metabolism of melanocytic cells. Glut-1 was present in the majority of benign nevi, whereas its expression was downregulated in 55% of malignant melanomas. Our results suggest that glucose transporter Glut-1 expression can significantly discriminate between human malignant melanoma and benign melanocytic nevi, and support the idea that additional mechanisms other than Glut-1 may contribute to glucose uptake in melanomas.

  13. The Influence of Initial Peritoneal Transport Characteristics, Inflammation, and High Glucose Exposure on Prognosis for Peritoneal Membrane Function

    Science.gov (United States)

    Fernández-Reyes, M. José; Bajo, M. Auxiliadora; Del Peso, Gloria; Ossorio, Marta; Díaz, Raquel; Carretero, Beatriz; Selgas, Rafael

    2012-01-01

    ♦ Background: Fast transport status, acquired with time on peritoneal dialysis (PD), is a pathology induced by peritoneal exposure to bioincompatible solutions. Fast transport has important clinical consequences and should be prevented. ♦ Objective: We analyzed the repercussions of initial peritoneal transport characteristics on the prognosis for peritoneal membrane function, and also whether the influence of peritonitis and high exposure to glucose are different according to the initial peritoneal transport characteristics or the moment when such events occur. ♦ Methods: The study included 275 peritoneal dialysis patients with at least 2 peritoneal function studies (at baseline and 1 year). Peritoneal kinetic studies were performed at baseline and annually. Those studies consist of a 4-hour dwell with glucose (1.5% during 1981 - 1990, and 2.27% during 1991 - 2002) to calculate the peritoneal mass transfer coefficients of urea and creatinine (milliliters per minute) using a previously described mathematical model. ♦ Results: Membrane prognosis and technique survival were independent of baseline transport characteristics. Fast transport and ultrafiltration (UF) failure are reversible conditions, provided that peritonitis and high glucose exposure are avoided during the early dialysis period. The first year on PD is a main determining factor for the membrane’s future, and the mass transfer coefficient of creatinine at year 1 is the best functional predictor of future PD history. After 5 years on dialysis, permeability frequently increases, and UF decreases. Icodextrin is associated with peritoneal protection. ♦ Conclusions: Peritoneal membrane prognosis is independent of baseline transport characteristics. Intrinsic fast transport and low UF are reversible conditions when peritonitis and high glucose exposure are avoided during the early dialysis period. Icodextrin helps in glucose avoidance and is associated with peritoneal protection. PMID:22473036

  14. The sentrin-conjugating enzyme mUbc9 interacts with GLUT4 and GLUT1 glucose transporters and regulates transporter levels in skeletal muscle cells

    Science.gov (United States)

    Giorgino, Francesco; de Robertis, Ottilia; Laviola, Luigi; Montrone, Carmela; Perrini, Sebastio; McCowen, Karen C.; Smith, Robert J.

    2000-01-01

    Glucose transport in insulin-regulated tissues is mediated by the GLUT4 and GLUT1 transporters. Using the yeast two-hybrid system, we have cloned the sentrin-conjugating enzyme mUbc9 as a protein that interacts with the GLUT4 COOH-terminal intracellular domain. The mUbc9 enzyme was found to bind directly to GLUT4 and GLUT1 through an 11-aa sequence common to the two transporters and to modify both transporters covalently by conjugation with the mUbc9 substrate, sentrin. Overexpression of mUbc9 in L6 skeletal muscle cells decreased GLUT1 transporter abundance 65%, resulting in decreased basal glucose transport. By contrast, mUbc9 overexpression increased GLUT4 abundance 8-fold, leading to enhanced transport stimulation by insulin. A dominant-negative mUbc9 mutant lacking catalytic activity had effects opposite to those of wild-type mUbc9. The regulation of GLUT4 and GLUT1 was specific, as evidenced by an absence of mUbc9 interaction with or regulation of the GLUT3 transporter isoform in L6 skeletal muscle cells. The mUbc9 sentrin-conjugating enzyme represents a novel regulator of GLUT1 and GLUT4 protein levels with potential importance as a determinant of basal and insulin-stimulated glucose uptake in normal and pathophysiological states. PMID:10655495

  15. Intestinal D-glucose transport and membrane fluidity along crypt-villus axis of streptozocin-induced diabetic rats.

    Science.gov (United States)

    Dudeja, P K; Wali, R K; Klitzke, A; Brasitus, T A

    1990-10-01

    Diabetes was induced in male Lewis rats by a single injection of streptozocin (50 mg/kg body wt ip). After 10-14 days, diabetic and age- and sex-matched control animals were killed, and their proximal small intestines were removed. Villus-tip, mid-villus, and lower-villus enterocytes were harvested from each group with a method that combined divalent cation chelation with mild mechanical dissociation. These fractions were used as starting material to prepare brush-border membrane vesicles. Preparations from each of these fractions were then analyzed and compared with respect to their Na(+)-gradient-dependent and Na(+)-independent D-glucose transport, lipid fluidity, and lipid composition. The results of these experiments demonstrated that 1) maximum rates of Na(+)-gradient-dependent D-glucose transport (Vmax) were greatest in membrane vesicles prepared from mature cells (villus tip and mid villus) of control rats; 2) the glucose concentration producing half-maximal rates of transport (Km), however, was significantly lower in lower-villus membrane vesicles of control rats, suggesting that a distinct glucose transporter existed in the membranes of these relatively immature enterocytes; 3) Na(+)-gradient-dependent, but not Na(+)-independent, D-glucose uptake was greater in diabetic membrane vesicles prepared from mid-villus and lower-villus fractions but not in vesicles prepared from villus-tip cells; and 4) no obvious relationship between alterations in membrane lipid fluidity and enhanced uptake of Na(+)-gradient-dependent D-glucose by these transporter(s) could be established in this experimental model of acute diabetes mellitus.

  16. Phylogenetic analysis and tissue distribution of elasmobranch glucose transporters and their response to feeding

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    Courtney A. Deck

    2016-03-01

    Full Text Available Elasmobranch diets consist of high quantities of protein and lipids, but very low levels of carbohydrates including glucose. Reflecting this diet, most tissues use lipids and ketone bodies as their main metabolic fuel. However, the rectal gland has been shown to be dependent on glucose as a fuel, so we hypothesized that glucose transporters (GLUTs would be present and upregulated in the gland during times of activation (e.g. following a meal. In this study, we searched for and identified putative class I GLUTs in three elasmobranchs and a holocephalan using transcriptomes, and used these to reconstruct a Bayesian phylogeny. We determined that each of the four species possessed three of the four class I GLUT sequences, but the identities of the isoforms present in each species differed between the elasmobranchs (GLUT1, 3 and 4 and the holocephalan (GLUT1, 2 and 3. We then used qPCR to measure mRNA levels of these GLUTs in the rectal gland, liver, intestine, and muscle of fed and starved spiny dogfish (Squalus suckleyi. The rectal gland data showed higher mRNA levels of GLUT4 in the starved relative to the fed fish. In the muscle, both GLUT1 and 4 were significantly elevated at 24 h post-feeding, as was the case for GLUT4 in the liver. In the intestine on the other hand, GLUT4 was significantly elevated by 6 h post-feeding, remaining elevated through 48 h. We suggest that GLUT4 has taken on the role of GLUT2 in elasmobranchs as the expression patterns observed in the liver and intestine are representative of GLUT2 in other vertebrates.

  17. Conditioning causes an increase in glucose transporter-4 levels in mononuclear cells in sled dogs.

    Science.gov (United States)

    Schnurr, Theresia M; Reynolds, Arleigh J; Gustafson, Sally J; Duffy, Lawrence K; Dunlap, Kriya L

    2014-10-01

    This study was designed to investigate the effects of physical conditioning on the expression of the insulin sensitive glucose transporter-4 protein (GLUT4) on mononuclear cells and HOMA-IR levels in dogs and compared to results reported in human skeletal muscle and the skeletal muscle of rodent models. Blood was sampled from conditioned dogs (n = 8) and sedentary dogs (n = 8). The conditioned dogs were exercised four months prior the experiment and were following a uniform training protocol, whereas the sedentary dogs were not. GLUT4 expression in mononuclear cells and plasma insulin levels were measured using commercially available enzyme-linked immunosorbent assay (ELISA). Blood glucose levels were determined using blood plasma. HOMA-IR was calculated using plasma insulin and blood glucose levels using the linear approximation formula. Our results indicate that the state of conditioning had a significant effect on the GLUT4 expression at the surface of mononuclear cells. HOMA-IR was also affected by conditioning in dogs. GLUT4 levels in mononuclear cells of sled dogs were inversely correlated with the homeostasis model assessment of insulin sensitivity. This study demonstrates that conditioning increases GLUT4 levels in mononuclear cells of sled dogs as it has been previously reported in skeletal muscle. Our results support the potential of white blood cells as a proxy tissue for studying insulin signaling and may lead to development of a minimally invasive and direct marker of insulin resistance. This may be the first report of GLUT4 in mononuclear cells in response to exercise and measured with ELISA.

  18. Familial Dysautonomia (FD Human Embryonic Stem Cell Derived PNS Neurons Reveal that Synaptic Vesicular and Neuronal Transport Genes Are Directly or Indirectly Affected by IKBKAP Downregulation.

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    Sharon Lefler

    Full Text Available A splicing mutation in the IKBKAP gene causes Familial Dysautonomia (FD, affecting the IKAP protein expression levels and proper development and function of the peripheral nervous system (PNS. Here we found new molecular insights for the IKAP role and the impact of the FD mutation in the human PNS lineage by using a novel and unique human embryonic stem cell (hESC line homozygous to the FD mutation originated by pre implantation genetic diagnosis (PGD analysis. We found that IKBKAP downregulation during PNS differentiation affects normal migration in FD-hESC derived neural crest cells (NCC while at later stages the PNS neurons show reduced intracellular colocalization between vesicular proteins and IKAP. Comparative wide transcriptome analysis of FD and WT hESC-derived neurons together with the analysis of human brains from FD and WT 12 weeks old embryos and experimental validation of the results confirmed that synaptic vesicular and neuronal transport genes are directly or indirectly affected by IKBKAP downregulation in FD neurons. Moreover we show that kinetin (a drug that corrects IKBKAP alternative splicing promotes the recovery of IKAP expression and these IKAP functional associated genes identified in the study. Altogether, these results support the view that IKAP might be a vesicular like protein that might be involved in neuronal transport in hESC derived PNS neurons. This function seems to be mostly affected in FD-hESC derived PNS neurons probably reflecting some PNS neuronal dysfunction observed in FD.

  19. Familial Dysautonomia (FD) Human Embryonic Stem Cell Derived PNS Neurons Reveal that Synaptic Vesicular and Neuronal Transport Genes Are Directly or Indirectly Affected by IKBKAP Downregulation.

    Science.gov (United States)

    Lefler, Sharon; Cohen, Malkiel A; Kantor, Gal; Cheishvili, David; Even, Aviel; Birger, Anastasya; Turetsky, Tikva; Gil, Yaniv; Even-Ram, Sharona; Aizenman, Einat; Bashir, Nibal; Maayan, Channa; Razin, Aharon; Reubinoff, Benjamim E; Weil, Miguel

    2015-01-01

    A splicing mutation in the IKBKAP gene causes Familial Dysautonomia (FD), affecting the IKAP protein expression levels and proper development and function of the peripheral nervous system (PNS). Here we found new molecular insights for the IKAP role and the impact of the FD mutation in the human PNS lineage by using a novel and unique human embryonic stem cell (hESC) line homozygous to the FD mutation originated by pre implantation genetic diagnosis (PGD) analysis. We found that IKBKAP downregulation during PNS differentiation affects normal migration in FD-hESC derived neural crest cells (NCC) while at later stages the PNS neurons show reduced intracellular colocalization between vesicular proteins and IKAP. Comparative wide transcriptome analysis of FD and WT hESC-derived neurons together with the analysis of human brains from FD and WT 12 weeks old embryos and experimental validation of the results confirmed that synaptic vesicular and neuronal transport genes are directly or indirectly affected by IKBKAP downregulation in FD neurons. Moreover we show that kinetin (a drug that corrects IKBKAP alternative splicing) promotes the recovery of IKAP expression and these IKAP functional associated genes identified in the study. Altogether, these results support the view that IKAP might be a vesicular like protein that might be involved in neuronal transport in hESC derived PNS neurons. This function seems to be mostly affected in FD-hESC derived PNS neurons probably reflecting some PNS neuronal dysfunction observed in FD.

  20. Exercise-induced increase in glucose transport, GLUT-4, and VAMP-2 in plasma membrane from human muscle

    DEFF Research Database (Denmark)

    Kristiansen, S; Hargreaves, Mark; Richter, Erik

    1996-01-01

    contractions may induce trafficking of GLUT-4-containing vesicles via a mechanism similar to neurotransmitter release. Our results demonstrate for the first time exercise-induced translocation of GLUT-4 and VAMP-2 to the plasma membrane of human muscle and increased sarcolemmal glucose transport.......A major effect of muscle contractions is an increase in sarcolemmal glucose transport. We have used a recently developed technique to produce sarcolemmal giant vesicles from human muscle biopsy samples obtained before and after exercise. Six men exercised for 10 min at 50% maximal O2 uptake (Vo2max...

  1. Sodium-glucose co-transporter-2 inhibitors and euglycemic ketoacidosis: Wisdom of hindsight

    Directory of Open Access Journals (Sweden)

    Awadhesh Kumar Singh

    2015-01-01

    Full Text Available Sodium-glucose co-transporter-2 inhibitors (SGLT-2i are newly approved class of oral anti-diabetic drugs, in the treatment of type 2 diabetes, which reduces blood glucose through glucouresis via the kidney, independent, and irrespective of available pancreatic beta-cells. Studies conducted across their clinical development program found, a modest reduction in glycated hemoglobin ranging from −0.5 to −0.8%, without any significant hypoglycemia. Moreover, head-to-head studies versus active comparators yielded comparable efficacy. Interestingly, weight and blood pressure reduction were additionally observed, which was not only consistent but significantly superior to active comparators, including metformin, sulfonylureas, and dipeptydylpeptide-4 inhibitors. Indeed, these additional properties makes this class a promising oral anti-diabetic drug. Surprisingly, a potentially fatal unwanted side effect of diabetic ketoacidosis has been noted with its widespread use, albeit rarely. Nevertheless, this has created a passé among the clinicians. This review is an attempt to pool those ketosis data emerging with SGLT-2i, and put a perspective on its implicated mechanism.

  2. A potential role for glucose transporters in the evolution of human brain size.

    Science.gov (United States)

    Fedrigo, Olivier; Pfefferle, Adam D; Babbitt, Courtney C; Haygood, Ralph; Wall, Christine E; Wray, Gregory A

    2011-01-01

    Differences in cognitive abilities and the relatively large brain are among the most striking differences between humans and their closest primate relatives. The energy trade-off hypothesis predicts that a major shift in energy allocation among tissues occurred during human origins in order to support the remarkable expansion of a metabolically expensive brain. However, the molecular basis of this adaptive scenario is unknown. Two glucose transporters (SLC2A1 and SLC2A4) are promising candidates and present intriguing mutations in humans, resulting, respectively, in microcephaly and disruptions in whole-body glucose homeostasis. We compared SLC2A1 and SLC2A4 expression between humans, chimpanzees and macaques, and found compensatory and biologically significant expression changes on the human lineage within cerebral cortex and skeletal muscle, consistent with mediating an energy trade-off. We also show that these two genes are likely to have undergone adaptation and participated in the development and maintenance of a larger brain in the human lineage by modulating brain and skeletal muscle energy allocation. We found that these two genes show human-specific signatures of positive selection on known regulatory elements within their 5'-untranslated region, suggesting an adaptation of their regulation during human origins. This study represents the first case where adaptive, functional and genetic lines of evidence implicate specific genes in the evolution of human brain size. Copyright © 2011 S. Karger AG, Basel.

  3. Sodium-glucose co-transporter-2 inhibitors and euglycemic ketoacidosis: Wisdom of hindsight

    Science.gov (United States)

    Singh, Awadhesh Kumar

    2015-01-01

    Sodium-glucose co-transporter-2 inhibitors (SGLT-2i) are newly approved class of oral anti-diabetic drugs, in the treatment of type 2 diabetes, which reduces blood glucose through glucouresis via the kidney, independent, and irrespective of available pancreatic beta-cells. Studies conducted across their clinical development program found, a modest reduction in glycated hemoglobin ranging from −0.5 to −0.8%, without any significant hypoglycemia. Moreover, head-to-head studies versus active comparators yielded comparable efficacy. Interestingly, weight and blood pressure reduction were additionally observed, which was not only consistent but significantly superior to active comparators, including metformin, sulfonylureas, and dipeptydylpeptide-4 inhibitors. Indeed, these additional properties makes this class a promising oral anti-diabetic drug. Surprisingly, a potentially fatal unwanted side effect of diabetic ketoacidosis has been noted with its widespread use, albeit rarely. Nevertheless, this has created a passé among the clinicians. This review is an attempt to pool those ketosis data emerging with SGLT-2i, and put a perspective on its implicated mechanism. PMID:26693421

  4. Glucose transporter 1 (GLUT1) expression is associated with intestinal type of gastric carcinoma.

    Science.gov (United States)

    Kim, W. S.; Kim, Y. Y.; Jang, S. J.; Kimm, K.; Jung, M. H.

    2000-01-01

    Increased expression of glucose transporter1 (GLUT1) has been reported in many human cancers. We hypothesized that the degree of GLUT1 might provide a useful biological information in gastric adenocarcinoma. RT-PCR and immunostaining were used to analyze GLUT1 expression in gastric cancer. RT-PCR showed GLUT1 expression was not largely detected in normal gastric tissue but was detected in cancerous gastric tissue of counterpart. By immunohistochemistry, GLUT1 protein was absent in normal gastric epithelium and intestinal metaplasia. 11 of 65 patients with gastric adenocarcinoma had specific GLUT1 immunostaining in a plasma membrane pattern with varied intensities. GLUT1 protein did not show any significant correlation with tumor stage and nodal metastasis (p>0.05 by Mann-Whitney test). However, the positive immunostaining for GLUT1 is associated with intestinal differentiation (p=0.003). Our results suggest that GLUT1 protein is associated with intestinal type of gastric cancer. PMID:10983690

  5. Unusual phenotype of glucose transport protein type 1 deficiency syndrome: A case report and literature review

    Directory of Open Access Journals (Sweden)

    Annio Posar

    2014-01-01

    Full Text Available The glucose transport protein type 1 (GLUT1 deficit causes a chronic brain energy failure. The classic phenotype of GLUT1 deficiency syndrome is characterized by: Mild to severe motor delay and mental retardation; infantile-onset epilepsy; head growth deceleration; movement disorders (ataxia, dystonia, spasticity; and non-epileptic paroxysmal events (intermittent ataxia, periodic confusion, recurrent headaches. During last years the classic phenotype of this syndrome, as originally reported, has expanded. We report the atypical phenotype of a boy with GLUT1 deficiency syndrome, characterized by mild mental retardation and drug-resistant absence seizures with onset at the age of 6 years, without movement disorders nor decrease of head circumference. A prompt diagnosis of this disorder is mandatory since the ketogenic diet might represent an effective treatment.

  6. In vivo measurements of brain glucose transport using the reversible Michaelis-Menten model and simultaneous measurements of cerebral blood flow changes during hypoglycemia.

    Science.gov (United States)

    Choi, I Y; Lee, S P; Kim, S G; Gruetter, R

    2001-06-01

    Glucose is the major substrate that sustains normal brain function. When the brain glucose concentration approaches zero, glucose transport across the blood-brain barrier becomes rate limiting for metabolism during, for example, increased metabolic activity and hypoglycemia. Steady-state brain glucose concentrations in alpha-chloralose anesthetized rats were measured noninvasively as a function of plasma glucose. The relation between brain and plasma glucose was linear at 4.5 to 30 mmol/L plasma glucose, which is consistent with the reversible Michaelis-Menten model. When the model was fitted to the brain glucose measurements, the apparent Michaelis-Menten constant, Kt, was 3.3 +/- 1.0 mmol/L, and the ratio of the maximal transport rate relative to CMRglc, Tmax/CMRglc, was 2.7 +/- 0.1. This Kt is comparable to the authors' previous human data, suggesting that glucose transport kinetics in humans and rats are similar. Cerebral blood flow (CBF) was simultaneously assessed and constant above 2 mmol/L plasma glucose at 73 +/- 6 mL 100 g(-1) min(-1). Extrapolation of the reversible Michaelis-Menten model to hypoglycemia correctly predicted the plasma glucose concentration (2.1 +/- 0.6 mmol/L) at which brain glucose concentrations approached zero. At this point, CBF increased sharply by 57% +/- 22%, suggesting that brain glucose concentration is the signal that triggers defense mechanisms aimed at improving glucose delivery to the brain during hypoglycemia.

  7. Electrophysiological evidence for glial-subtype glutamate transporter functional expression in rat cerebellar granule neurons

    Directory of Open Access Journals (Sweden)

    Mafra R.A.

    2003-01-01

    Full Text Available A glutamate-sensitive inward current (Iglu is described in rat cerebellar granule neurons and related to a glutamate transport mechanism. We examined the features of Iglu using the patch-clamp technique. In steady-state conditions the Iglu measured 8.14 ± 1.9 pA. Iglu was identified as a voltage-dependent inward current showing a strong rectification at positive potentials. L-Glutamate activated the inward current in a dose-dependent manner, with a half-maximal effect at about 18 µM and a maximum increase of 51.2 ± 4.4%. The inward current was blocked by the presence of dihydrokainate (0.5 mM, shown by others to readily block the GLT1 isoform. We thus speculate that Iglu could be attributed to the presence of a native glutamate transporter in cerebellar granule neurons.

  8. Interactions of androgens, green tea catechins and the antiandrogen flutamide with the external glucose-binding site of the human erythrocyte glucose transporter GLUT1

    Science.gov (United States)

    Naftalin, Richard J; Afzal, Iram; Cunningham, Philip; Halai, Mansur; Ross, Clare; Salleh, Naguib; Milligan, Stuart R

    2003-01-01

    This study investigates the effects of androgens, the antiandrogen flutamide and green tea catechins on glucose transport inhibition in human erythrocytes. These effects may relate to the antidiabetogenic effects of green tea. Testosterone, 4-androstene-3,17-dione, dehydroepiandrosterone (DHEA) and DHEA-3-acetate inhibit glucose exit from human erythrocytes with half-maximal inhibitions (Ki) of 39.2±8.9, 29.6±3.7, 48.1±10.2 and 4.8±0.98 μM, respectively. The antiandrogen flutamide competitively relieves these inhibitions and of phloretin. Dehydrotestosterone has no effect on glucose transport, indicating the differences between androgen interaction with GLUT1 and human androgen receptor (hAR). Green tea catechins also inhibit glucose exit from erythrocytes. Epicatechin 3-gallate (ECG) has a Ki ECG of 0.14±0.01 μM, and epigallocatechin 3-gallate (EGCG) has a Ki EGCG of 0.97±0.13 μM. Flutamide reverses these effects. Androgen-screening tests show that the green tea catechins do not act genomically. The high affinities of ECG and EGCG for GLUT1 indicate that this might be their physiological site of action. There are sequence homologies between GLUT1 and the ligand-binding domain (LBD) of hAR containing the amino-acid triads Arg 126, Thr 30 and Asn 288, and Arg 126, Thr 30 and Asn 29, with similar 3D topology to the polar groups binding 3-keto and 17-β OH steroid groups in hAR LBD. These triads are appropriately sited for competitive inhibition of glucose import at the external opening of the hydrophilic pore traversing GLUT1. PMID:12970085

  9. Calpain-dependent disruption of nucleo-cytoplasmic transport in ALS motor neurons

    Science.gov (United States)

    Yamashita, Takenari; Aizawa, Hitoshi; Teramoto, Sayaka; Akamatsu, Megumi; Kwak, Shin

    2017-01-01

    Nuclear dysfunction in motor neurons has been hypothesized to be a principal cause of amyotrophic lateral sclerosis (ALS) pathogenesis. Here, we investigated the mechanism by which the nuclear pore complex (NPC) is disrupted in dying motor neurons in a mechanistic ALS mouse model (adenosine deaminase acting on RNA 2 (ADAR2) conditional knockout (AR2) mice) and in ALS patients. We showed that nucleoporins (Nups) that constituted the NPC were cleaved by activated calpain via a Ca2+-permeable AMPA receptor-mediated mechanism in dying motor neurons lacking ADAR2 expression in AR2 mice. In these neurons, nucleo-cytoplasmic transport was disrupted, and the level of the transcript elongation enzyme RNA polymerase II phosphorylated at Ser2 was significantly decreased. Analogous changes were observed in motor neurons lacking ADAR2 immunoreactivity in sporadic ALS patients. Therefore, calpain-dependent NPC disruption may participate in ALS pathogenesis, and inhibiting Ca2+-mediated cell death signals may be a therapeutic strategy for ALS. PMID:28045133

  10. Distribution of neutral amino acid transporter ASCT1 in the non-neuronal tissued of mice

    OpenAIRE

    Hashimoto, Yoshiharu; Sadamoto, Yoshihiro; Konno, Akihiro; Kon, Yasuhiro

    2004-01-01

    Distribution of ASCT1, a neurtral amino acid transporter, in non-neuronal peripheral tissues of adult and developong mice was examines by immunohistochemistry and immunoelectron microscopy. Immunoreactivity for ASCT1 in the digestive system was localized in basal cells of stratified squamous epithelia from oral parietes to nonglandular region of the stomach, chief cells of the glandular stomach, acinar cells of the salivary gland and exocrine pancreas, and Paneth's cells of the small intestin...

  11. Glucose elevates NITRATE TRANSPORTER2.1 protein levels and nitrate transport activity independently of its HEXOKINASE1-mediated stimulation of NITRATE TRANSPORTER2.1 expression.

    Science.gov (United States)

    de Jong, Femke; Thodey, Kate; Lejay, Laurence V; Bevan, Michael W

    2014-01-01

    Mineral nutrient uptake and assimilation is closely coordinated with the production of photosynthate to supply nutrients for growth. In Arabidopsis (Arabidopsis thaliana), nitrate uptake from the soil is mediated by genes encoding high- and low-affinity transporters that are transcriptionally regulated by both nitrate and photosynthate availability. In this study, we have studied the interactions of nitrate and glucose (Glc) on gene expression, nitrate transport, and growth using glucose-insensitive2-1 (gin2-1), which is defective in sugar responses. We confirm and extend previous work by showing that HEXOKINASE1-mediated oxidative pentose phosphate pathway (OPPP) metabolism is required for Glc-mediated NITRATE TRANSPORTER2.1 (NRT2.1) expression. Treatment with pyruvate and shikimate, two products derived from intermediates of the OPPP that are destined for amino acid production, restores wild-type levels of NRT2.1 expression, suggesting that metabolites derived from OPPP metabolism can, together with Glc, directly stimulate high levels of NRT2.1 expression. Nitrate-mediated NRT2.1 expression is not influenced by gin2-1, showing that Glc does not influence NRT2.1 expression through nitrate-mediated mechanisms. We also show that Glc stimulates NRT2.1 protein levels and transport activity independently of its HEXOKINASE1-mediated stimulation of NRT2.1 expression, demonstrating another possible posttranscriptional mechanism influencing nitrate uptake. In gin2-1 plants, nitrate-responsive biomass growth was strongly reduced, showing that the supply of OPPP metabolites is essential for assimilating nitrate for growth.

  12. Muscle Mitochondrial ATP Synthesis and Glucose Transport/Phosphorylation in Type 2 Diabetes

    OpenAIRE

    Julia Szendroedi; Schmid, Albrecht I; Marek Chmelik; Christian Toth; Attila Brehm; Martin Krssak; Peter Nowotny; Michael Wolzt; Werner Waldhausl; Michael Roden

    2007-01-01

    Editors' Summary Background. Diabetes mellitus is an increasingly common chronic disease characterized by high blood sugar (glucose) levels. In normal individuals, blood sugar levels are maintained by the hormone insulin. Insulin is released by the pancreas when blood glucose levels rise after eating (glucose is produced by the digestion of food) and “instructs” insulin-responsive muscle and fat cells to take up glucose from the bloodstream. The cells then use glucose as a fuel or convert it ...

  13. Electron transport phosphorylation in rumen butyrivibrios: unprecedented ATP yield for glucose fermentation to butyrate

    Directory of Open Access Journals (Sweden)

    Timothy eHackmann

    2015-06-01

    Full Text Available From a genomic analysis of rumen butyrivibrios (Butyrivibrio and Pseudobutyrivibrio spp., we have re-evaluated the contribution of electron transport phosphorylation to ATP formation in this group. This group is unique in that most (76% genomes were predicted to possess genes for both Ech and Rnf transmembrane ion pumps. These pumps act in concert with the NifJ and Bcd-Etf to form a electrochemical potential (ΔμH+ and ΔμNa+, which drives ATP synthesis by electron transport phosphorylation. Of the 62 total butyrivibrio genomes currently available from the Hungate 1000 project, all 62 were predicted to possess NifJ, which reduces oxidized ferredoxin (Fdox during pyruvate conversion to acetyl-CoA. All 62 possessed all subunits of Bcd-Etf, which reduces Fdox and oxidizes reduced NAD (NADred during crotonyl-CoA reduction. Additionally, 61 genomes possessed all subunits of the Rnf, which generates ΔμH+ or ΔμNa+ from oxidation of reduced Fd and reduction of oxidized NAD (NADox. Further, 47 genomes possessed all 6 subunits of the Ech, which generates ΔμH+ from oxidation of reduced Fd (Fdred. For glucose fermentation to butyrate and H2, the electrochemical potential established should drive synthesis of ~1.5 ATP by the F0F1-ATP synthase (possessed by all 62 genomes. The total yield is ~4.5 ATP/glucose after accounting for 3 ATP formed by classic substrate-level phosphorylation, and it is one the highest yields for any glucose fermentation. The yield was the same when unsaturated fatty acid bonds, not H+, served as the electron acceptor (as during biohydrogenation. Possession of both Ech and Rnf had been previously documented in only a few sulfate-reducers, was rare in other rumen prokaryotic genomes in our analysis, and may confer an energetic advantage to rumen butyrivibrios. This unique energy conservation system might enhance the butyrivibrios’ ability to overcome growth inhibition by unsaturated fatty acids, as postulated herein.

  14. Cannabidiol attenuates OGD/R-induced damage by enhancing mitochondrial bioenergetics and modulating glucose metabolism via pentose-phosphate pathway in hippocampal neurons

    Directory of Open Access Journals (Sweden)

    Shanshan Sun

    2017-04-01

    Full Text Available Deficient bioenergetics and diminished redox conservation have been implicated in the development of cerebral ischemia/reperfusion injury. In this study, the mechanisms underlying the neuroprotective effects of cannabidiol (CBD, a nonpsychotropic compound derived from Cannabis sativa with FDA-approved antiepilepsy properties, were studied in vitro using an oxygen–glucose-deprivation/reperfusion (OGD/R model in a mouse hippocampal neuronal cell line. CBD supplementation during reperfusion rescued OGD/R-induced cell death, attenuated intracellular ROS generation and lipid peroxidation, and simultaneously reversed the abnormal changes in antioxidant biomarkers. Using the Seahorse XFe24 Extracellular Flux Analyzer, we found that CBD significantly improved basal respiration, ATP-linked oxygen consumption rate, and the spare respiratory capacity, and augmented glucose consumption in OGD/R-injured neurons. The activation of glucose 6-phosphate dehydrogenase and the preservation of the NADPH/NADP+ ratio implies that the pentose-phosphate pathway is stimulated by CBD, thus protecting hippocampal neurons from OGD/R injury. This study is the first to document the neuroprotective effects of CBD against OGD/R insult, which depend in part on attenuating oxidative stress, enhancing mitochondrial bioenergetics, and modulating glucose metabolism via the pentose-phosphate pathway, thus preserving both energy and the redox balance.

  15. Uptake and phloem transport of glucose-fipronil conjugate in Ricinus communis involve a carrier-mediated mechanism.

    Science.gov (United States)

    Wu, Han-Xiang; Yang, Wen; Zhang, Zhi-Xiang; Huang, Ting; Yao, Guang-Kai; Xu, Han-Hong

    2012-06-20

    Some compounds containing glucose are absorbed via the monosaccharide transporters of the plasma membrane. A glucose-fipronil conjugate, N-[3-cyano-1-[2,6-dichloro-4-(trifluoromethyl)phenyl]-4-[(trifluoromethyl)sulfinyl]-1H-pyrazol-5-yl]-1-(β-d-glucopyranosyl)-1H-1,2,3-triazole-4-methanamine (GTF), has been synthesized in our previous work. GTF exhibits moderate phloem mobility in Ricinus communis. In the current paper, we demonstrate that the uptake of GTF by Ricinus seedling cotyledon discs is partly mediated by an active carrier system (K(m)1 = 0.17 mM; V(max)1 = 2.2 nmol cm(-2) h(-1)). Four compounds [d-glucose, sucrose, phloridzin, and carbonyl cyanide m-chlorophenylhydrazone (CCCP)] were examined for their effect on GTF uptake. Phloridzin as well as CCCP markedly inhibit GTF uptake, and d-glucose weakly competes with it. The phloem transport of GTF in Ricinus seedlings is found to involve an active carrier-mediated mechanism that effectively contributes to the GTF phloem loading. The results prove that adding a glucose core is a reasonable and feasible approach to confer phloem mobility to fipronil by utilizing plant monosaccharide transporters.

  16. Zinc finger protein 407 (ZFP407) regulates insulin-stimulated glucose uptake and glucose transporter 4 (Glut4) mRNA.

    Science.gov (United States)

    Buchner, David A; Charrier, Alyssa; Srinivasan, Ethan; Wang, Li; Paulsen, Michelle T; Ljungman, Mats; Bridges, Dave; Saltiel, Alan R

    2015-03-06

    The glucose transporter GLUT4 facilitates insulin-stimulated glucose uptake in peripheral tissues including adipose, muscle, and heart. GLUT4 function is impaired in obesity and type 2 diabetes leading to hyperglycemia and an increased risk of cardiovascular disease and neuropathy. To better understand the regulation of GLUT4 function, a targeted siRNA screen was performed and led to the discovery that ZFP407 regulates insulin-stimulated glucose uptake in adipocytes. The decrease in insulin-stimulated glucose uptake due to ZFP407 deficiency was attributed to a reduction in GLUT4 mRNA and protein levels. The decrease in GLUT4 was due to both decreased transcription of Glut4 mRNA and decreased efficiency of Glut4 pre-mRNA splicing. Interestingly, ZFP407 coordinately regulated this decrease in transcription with an increase in the stability of Glut4 mRNA, resulting in opposing effects on steady-state Glut4 mRNA levels. More broadly, transcriptome analysis revealed that ZFP407 regulates many peroxisome proliferator-activated receptor (PPAR) γ target genes beyond Glut4. ZFP407 was required for the PPARγ agonist rosiglitazone to increase Glut4 expression, but was not sufficient to increase expression of a PPARγ target gene reporter construct. However, ZFP407 and PPARγ co-overexpression synergistically activated a PPARγ reporter construct beyond the level of PPARγ alone. Thus, ZFP407 may represent a new modulator of the PPARγ signaling pathway.

  17. Protective effect of telmisartan against oxidative damage induced by high glucose in neuronal PC12 cell.

    Science.gov (United States)

    Eslami, Habib; Sharifi, Ali M; Rahimi, Hamzeh; Rahati, Maryam

    2014-01-13

    Telmisartan is an angiotensin II type 1 receptor blocker and partial agonist of peroxisome proliferator-activated receptor gamma (PPAR-γ). Here, we investigated the protective capacity of telmisartan against high glucose (HG)-elicited oxidative damage in PC12 cells. The activity of lactate dehydrogenase (LDH), NADPH oxidase (NOX), superoxide dismutase (SOD), catalase (CAT) as well as the levels of malondialdehyde (MDA), glutathione (GSH), intracellular reactive oxygen species (ROS), cell viability and DNA fragmentation were measured in HG-treated PC12 cells with and without telmisartan co-treatment. Moreover, the direct antioxidant effect of telmisartan was determined by 2,2-azinobis-(3-ethylbenzothiazoline-6-sulphonic acid) (ABTS) assay and protein expression of Bax, Bcl-2, cleaved caspase-3 and NOX subunit p47phox by western blotting. Telmisartan exhibited antioxidant activity in the ABTS assay with the IC50 value of 37.5 μM. Pretreatment of PC12 cells with telmisartan, prior to HG exposure, was associated with a marked diminution in cleaved caspase-3 expression, DNA fragmentation, Bax/Bcl-2 ratio, intracellular ROS and MDA levels. Additionally, the cell viability, GSH level, SOD and CAT activity were notably elevated by telmisartan, whereas the activity and the protein expression of NADPH oxidase subunit p47phox were attenuated. Interestingly, co-treatment with GW9662, a PPAR-γ antagonist, partially inhibited the beneficial effects of telmisartan. These findings suggest that telmisartan has protective effects on HG-induced neurotoxicity in PC12 cells, which may be related to its antioxidant action and inhibition of NADPH oxidase. Furthermore, the results show that PPAR-γ activation is involved in the neuroprotective effects of telmisartan. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

  18. Sodium-glucose co-transporter 2 (SGLT2 inhibitors: a growing class of anti-diabetic agents

    Directory of Open Access Journals (Sweden)

    Eva M Vivian

    2014-12-01

    Full Text Available Although several treatment options are available to reduce hyperglycemia, only about half of individuals with diagnosed diabetes mellitus (DM achieve recommended glycemic targets. New agents that reduce blood glucose concentrations by novel mechanisms and have acceptable safety profiles are needed to improve glycemic control and reduce the complications associated with type 2 diabetes mellitus (T2DM. The renal sodium-glucose co-transporter 2 (SGLT2 is responsible for reabsorption of most of the glucose filtered by the kidney. Inhibitors of SGLT2 lower blood glucose independent of the secretion and action of insulin by inhibiting renal reabsorption of glucose, thereby promoting the increased urinary excretion of excess glucose. Canagliflozin, dapagliflozin, and empagliflozin are SGLT2 inhibitors approved as treatments for T2DM in the United States, Europe, and other countries. Canagliflozin, dapagliflozin, and empagliflozin increase renal excretion of glucose and improve glycemic parameters in patients with T2DM when used as monotherapy or in combination with other antihyperglycemic agents. Treatment with SGLT2 inhibitors is associated with weight reduction, lowered blood pressure, and a low intrinsic propensity to cause hypoglycemia. Overall, canagliflozin, dapagliflozin, and empagliflozin are well tolerated. Cases of genital infections and, in some studies, urinary tract infections have been more frequent in canagliflozin-, dapagliflozin-, and empagliflozin-treated patients compared with those receiving placebo. Evidence from clinical trials suggests that SGLT2 inhibitors are a promising new treatment option for T2DM.

  19. Fat gain with physical detraining is correlated with increased glucose transport and oxidation in periepididymal white adipose tissue in rats

    Energy Technology Data Exchange (ETDEWEB)

    Sertié, R.A.L.; Andreotti, S. [Departamento de Fisiologia e Biofísica, Instituto de Ciências Biomédicas, Universidade de São Paulo, São Paulo, SP (Brazil); Proença, A.R.G. [Laboratório de Biotecnologia, Faculdade de Ciências Aplicadas, Universidade Estadual de Campinas, Limeira, SP (Brazil); Campaña, A.B.; Lima, F.B. [Departamento de Fisiologia e Biofísica, Instituto de Ciências Biomédicas, Universidade de São Paulo, São Paulo, SP (Brazil)

    2015-05-26

    As it is a common observation that obesity tends to occur after discontinuation of exercise, we investigated how white adipocytes isolated from the periepididymal fat of animals with interrupted physical training transport and oxidize glucose, and whether these adaptations support the weight regain seen after 4 weeks of physical detraining. Male Wistar rats (45 days old, weighing 200 g) were divided into two groups (n=10): group D (detrained), trained for 8 weeks and detrained for 4 weeks; and group S (sedentary). The physical exercise was carried out on a treadmill for 60 min/day, 5 days/week for 8 weeks, at 50-60% of the maximum running capacity. After the training protocol, adipocytes isolated from the periepididymal adipose tissue were submitted to glucose uptake and oxidation tests. Adipocytes from detrained animals increased their glucose uptake capacity by 18.5% compared with those from sedentary animals (P<0.05). The same cells also showed a greater glucose oxidation capacity in response to insulin stimulation (34.55%) compared with those from the S group (P<0.05). We hypothesize that, owing to the more intense glucose entrance into adipose cells from detrained rats, more substrate became available for triacylglycerol synthesis. Furthermore, this increased glucose oxidation rate allowed an increase in energy supply for triacylglycerol synthesis. Thus, physical detraining might play a role as a possible obesogenic factor for increasing glucose uptake and oxidation by adipocytes.

  20. Regulation of the GLUT1 glucose transporter in cultured myocytes: total number and subcellular distribution as determined by photoaffinity labelling.

    Science.gov (United States)

    el-Kebbi, I M; Roser, S; Pollet, R J; Cushman, S W; Wilson, C M

    1994-01-01

    We have used the impermeant photoaffinity label 2-N-4-(1-azi-2,2,2-trifluoroethyl)benzoyl-[2-3H] 1,3-bis-(D-mannos-4-yloxy)-2-propylamine (ATB-[2-3H]BMPA) to identify and quantify the glucose transporters on the surface of BC3H-1 cells, a continuously cultured skeletal-muscle cell line lacking the MyoD transcription factor required for cell fusion. ATB-[2-3H]BMPA was used in combination with immunoprecipitation of the GLUT1 glucose transporter, the only isoform expressed in these cells. The total cellular GLUT1 content was also determined by photolabelling and immunoprecipitation after cell permeabilization with digitonin (0.025%). In glucose-starved cells, 85% of the glucose transporters were present at the cell surface in the basal state, with little change in response to insulin (200 nM), correlating with lack of additional 2-deoxyglucose uptake in response to insulin. Feeding the cells with glucose (25 mM) for 24 h resulted in an 80% decrease in the total GLUT1 content relative to starved cells, of which only 25% were present on the cell surface. This was associated with an 85% decrease in 2-deoxyglucose uptake. In addition, acute stimulation of the fed cells with insulin or phorbol 12-myristate 13-acetate (PMA) led to an increase in GLUT1 at the cell surface, and, in correspondence, an increase in 2-deoxyglucose uptake by approx. 2- and 4-fold respectively. We conclude that exofacial photoaffinity labelling of glucose transporters with ATB-[2-3H]BMPA in the presence and absence of digitonin, followed by specific immunoprecipitation, provides an accurate measure of total and cell-surface glucose transporters in differentiated BC3H-1 muscle cells. This technique demonstrates that glucose pre-feeding (1) decreases the total number of GLUT1 and (2) redistributes the majority of the remaining transporters to an intracellular site, where they can now be translocated to the cell surface in response to insulin and PMA. PMID:8037688

  1. The metal transporter SMF-3/DMT-1 mediates aluminum-induced dopamine neuron degeneration.

    Science.gov (United States)

    VanDuyn, Natalia; Settivari, Raja; LeVora, Jennifer; Zhou, Shaoyu; Unrine, Jason; Nass, Richard

    2013-01-01

    Aluminum (Al(3+)) is the most prevalent metal in the earth's crust and is a known human neurotoxicant. Al(3+) has been shown to accumulate in the substantia nigra of patients with Parkinson's disease (PD), and epidemiological studies suggest correlations between Al(3+) exposure and the propensity to develop both PD and the amyloid plaque-associated disorder Alzheimer's disease (AD). Although Al(3+) exposures have been associated with the development of the most common neurodegenerative disorders, the molecular mechanism involved in Al(3+) transport in neurons and subsequent cellular death has remained elusive. In this study, we show that a brief exposure to Al(3+) decreases mitochondrial membrane potential and cellular ATP levels, and confers dopamine (DA) neuron degeneration in the genetically tractable nematode Caenorhabditis elegans (C. elegans). Al(3+) exposure also exacerbates DA neuronal death conferred by the human PD-associated protein α-synuclein. DA neurodegeneration is dependent on SMF-3, a homologue to the human divalent metal transporter (DMT-1), as a functional null mutation partially inhibits the cell death. We also show that SMF-3 is expressed in DA neurons, Al(3+) exposure results in a significant decrease in protein levels, and the neurodegeneration is partially dependent on the PD-associated transcription factor Nrf2/SKN-1 and caspase Apaf1/CED-4. Furthermore, we provide evidence that the deletion of SMF-3 confers Al(3+) resistance due to sequestration of Al(3+) into an intracellular compartment. This study describes a novel model for Al(3+)-induced DA neurodegeneration and provides the first molecular evidence of an animal Al(3+) transporter.

  2. Exercise-induced increase in glucose transport, GLUT-4, and VAMP-2 in plasma membrane from human muscle

    DEFF Research Database (Denmark)

    Kristiansen, S; Hargreaves, Mark; Richter, Erik

    1996-01-01

    ) and then to fatigue at 100% Vo2max (5.7 +/- 0.2 min). Vesicle glucose transport at 5 mM increased from 3.3 +/- 0.6 pmol.microgram-1.min-1 at rest to 6.6 +/- 1.0 pmol.microgram-1.min-1 at fatigue (mean +/- SE, n = 6, P increase in glucose transport was associated with a 1.6-fold increase in vesicle GLUT......A major effect of muscle contractions is an increase in sarcolemmal glucose transport. We have used a recently developed technique to produce sarcolemmal giant vesicles from human muscle biopsy samples obtained before and after exercise. Six men exercised for 10 min at 50% maximal O2 uptake (Vo2max......-4 protein content. Glucose transport normalized to GLUT-4 protein content also increased with exercise, suggesting increased intrinsic activity of GLUT-4. Furthermore, exercise resulted in a 1.4-fold increase in sarcolemmal vesicle-associated membrane protein (VAMP-2) content, suggesting that muscle...

  3. In Silico Modeling-based Identification of Glucose Transporter 4 (GLUT4)-selective Inhibitors for Cancer Therapy.

    Science.gov (United States)

    Mishra, Rama K; Wei, Changyong; Hresko, Richard C; Bajpai, Richa; Heitmeier, Monique; Matulis, Shannon M; Nooka, Ajay K; Rosen, Steven T; Hruz, Paul W; Schiltz, Gary E; Shanmugam, Mala

    2015-06-05

    Tumor cells rely on elevated glucose consumption and metabolism for survival and proliferation. Glucose transporters mediating glucose entry are key proximal rate-limiting checkpoints. Unlike GLUT1 that is highly expressed in cancer and more ubiquitously expressed in normal tissues, GLUT4 exhibits more limited normal expression profiles. We have previously determined that insulin-responsive GLUT4 is constitutively localized on the plasma membrane of myeloma cells. Consequently, suppression of GLUT4 or inhibition of glucose transport with the HIV protease inhibitor ritonavir elicited growth arrest and/or apoptosis in multiple myeloma. GLUT4 inhibition also caused sensitization to metformin in multiple myeloma and chronic lymphocytic leukemia and a number of solid tumors suggesting the broader therapeutic utility of targeting GLUT4. This study sought to identify selective inhibitors of GLUT4 to develop a more potent cancer chemotherapeutic with fewer potential off-target effects. Recently, the crystal structure of GLUT1 in an inward open conformation was reported. Although this is an important achievement, a full understanding of the structural biology of facilitative glucose transport remains elusive. To date, there is no three-dimensional structure for GLUT4. We have generated a homology model for GLUT4 that we utilized to screen for drug-like compounds from a library of 18 million compounds. Despite 68% homology between GLUT1 and GLUT4, our virtual screen identified two potent compounds that were shown to target GLUT4 preferentially over GLUT1 and block glucose transport. Our results strongly bolster the utility of developing GLUT4-selective inhibitors as anti-cancer therapeutics.

  4. Brazilein inhibits neuronal inflammation induced by cerebral ischemia and oxygen-glucose deprivation through targeting NOD2 expression.

    Science.gov (United States)

    Yan, Xiao-Jin; Chai, Yu-Shuang; Yuan, Zhi-Yi; Wang, Xin-Pei; Jiang, Jing-Fei; Lei, Fan; Xing, Dong-Ming; DU, Li-Jun

    2016-05-01

    Brazilein is reported to have immunosuppressive effect on cardiovascular and cerebral-vascular diseases. The essential roles of innate immunity in cerebral ischemia are increasingly identified, but no studies concerning the influence of brazilein on the innate immunity receptors have been reported. The present study was designed to investigate the regulation of NOD2 (Nucleotide-binding oligomerization domain-containing protein 2) by brazilein for its protection of neuron in cerebral ischemia in vivo and oxygen-glucose deprivation in vitro. The results showed that brazilein could reverse the elevated expression of NOD2 and TNFα (tumor necrosis factor alpha) elicited by cerebral ischemia and reperfusion. This reduction could also be detected in normal mice and C17.2 cells, indicating that this suppressive effect of brazilein was correlated with NOD2. The results from GFP reporter plasmid assay suggested brazilein inhibited NOD2 gene transcription. In conclusion, brazilein could attenuate NOD2 and TNFα expression in cerebral ischemia and NOD2 may be one possible target of brazilein for its immune suppressive effect in neuro-inflammation.

  5. Neuroprotective and Anti-Apoptotic Effects of CSP-1103 in Primary Cortical Neurons Exposed to Oxygen and Glucose Deprivation

    Science.gov (United States)

    Porrini, Vanessa; Sarnico, Ilenia; Benarese, Marina; Branca, Caterina; Mota, Mariana; Lanzillotta, Annamaria; Bellucci, Arianna; Parrella, Edoardo; Faggi, Lara; Spano, Pierfranco; Imbimbo, Bruno Pietro; Pizzi, Marina

    2017-01-01

    CSP-1103 (formerly CHF5074) has been shown to reverse memory impairment and reduce amyloid plaque as well as inflammatory microglia activation in preclinical models of Alzheimer’s disease. Moreover, it was found to improve cognition and reduce brain inflammation in patients with mild cognitive impairment. Recent evidence suggests that CSP-1103 acts through a single molecular target, the amyloid precursor protein intracellular domain (AICD), a transcriptional regulator implicated in inflammation and apoptosis. We here tested the possible anti-apoptotic and neuroprotective activity of CSP-1103 in a cell-based model of post-ischemic injury, wherein the primary mouse cortical neurons were exposed to oxygen-glucose deprivation (OGD). When added after OGD, CSP-1103 prevented the apoptosis cascade by reducing cytochrome c release and caspase-3 activation and the secondary necrosis. Additionally, CSP-1103 limited earlier activation of p38 and nuclear factor κB (NF-κB) pathways. These results demonstrate that CSP-1103 is neuroprotective in a model of post-ischemic brain injury and provide further mechanistic insights as regards its ability to reduce apoptosis and potential production of pro-inflammatory cytokines. In conclusion, these findings suggest a potential use of CSP-1103 for the treatment of brain ischemia. PMID:28106772

  6. Comparative Study of the Effect of Baicalin and Its Natural Analogs on Neurons with Oxygen and Glucose Deprivation Involving Innate Immune Reaction of TLR2/TNFα

    OpenAIRE

    Li, Hui-Ying; Hu, Jun; Zhao, Shuang; Yuan, Zhi-Yi; Wan, Hong-Jiao; Lei, Fan; Ding, Yi; Xing, Dong-Ming; Du, Li-Jun

    2012-01-01

    This work is to study the baicalin and its three analogs, baicalin, wogonoside, and wogonin, on the protective effect of neuron from oxygen-glucose deprivation (OGD) and toll-like receptor 2 (TLR2) expression in OGD damage. The results showed that baicalin and its three analogs did protect neurons from OGD damage and downregulated protein level of TLR2. D-Glucopyranosiduronic acid on site 7 in the structure played a core of cytotoxicity of these flavonoid analogs. The methoxyl group on carbon...

  7. Aqueous extract of tamarind seeds selectively increases glucose transporter-2, glucose transporter-4, and islets' intracellular calcium levels and stimulates β-cell proliferation resulting in improved glucose homeostasis in rats with streptozotocin-induced diabetes mellitus.

    Science.gov (United States)

    Sole, Sushant Shivdas; Srinivasan, B P

    2012-08-01

    Tamarindus indica Linn. has been in use for a long time in Asian food and traditional medicine for different diseases including diabetes and obesity. However, the molecular mechanisms of these effects have not been fully understood. In view of the multidimensional activity of tamarind seeds due to their having high levels of polyphenols and flavonoids, we hypothesized that the insulin mimetic effect of aqueous tamarind seed extract (TSE) might increase glucose uptake through improvement in the expression of genes of the glucose transporter (GLUT) family and sterol regulatory element-binding proteins (SREBP) 1c messenger RNA (mRNA) in the liver. Daily oral administration of TSE to streptozotocin (STZ)-induced (90 mg/kg intraperitoneally) type 2 diabetic male Wistar rats at different doses (120 and 240 mg/kg body weight) for 4 weeks showed positive correlation with intracellular calcium and insulin release in isolated islets of Langerhans. Tamarind seed extract supplementation significantly improved the GLUT-2 protein and SREBP-1c mRNA expression in the liver and GLUT-4 protein and mRNA expression in the skeletal muscles of diabetic rats. The elevated levels of serum nitric oxide (NO), glycosylated hemoglobin level (hemoglobin (A1c)) and tumor necrosis factor α (TNF-α) decreased after TSE administration. Immunohistochemical findings revealed that TSE abrogated STZ-induced apoptosis and increased β-cell neogenesis, indicating its effect on islets and β-cell mass. In conclusion, it was found that the antidiabetic effect of TSE on STZ-induced diabetes resulted from complex mechanisms of β-cell neogenesis, calcium handling, GLUT-2, GLUT-4, and SREBP-1c. These findings show the scope for formulating a new herbal drug for diabetes therapy.

  8. Caffeamide 36-13 Regulates the Antidiabetic and Hypolipidemic Signs of High-Fat-Fed Mice on Glucose Transporter 4, AMPK Phosphorylation, and Regulated Hepatic Glucose Production

    Directory of Open Access Journals (Sweden)

    Yueh-Hsiung Kuo

    2014-01-01

    Full Text Available This study was to investigate the antidiabetic and antihyperlipidemic effects of (E-3-[3, 4-dihydroxyphenyl-1-(piperidin-1-ylprop-2-en-1-one] (36-13 (TS, one of caffeic acid amide derivatives, on high-fat (HF- fed mice. The C57BL/6J mice were randomly divided into the control (CON group and the experimental group, which was firstly fed a HF diet for 8 weeks. Then, the HF group was subdivided into four groups and was given TS orally (including two doses or rosiglitazone (Rosi or vehicle for 4 weeks. Blood, skeletal muscle, and tissues were examined by measuring glycaemia and dyslipidemia-associated events. TS effectively prevented HF diet-induced increases in the levels of blood glucose, triglyceride, insulin, leptin, and free fatty acid (FFA and weights of visceral fa; moreover, adipocytes in the visceral depots showed a reduction in size. TS treatment significantly increased the protein contents of glucose transporter 4 (GLUT4 in skeletal muscle; TS also significantly enhanced Akt phosphorylation in liver, whereas it reduced the expressions of phosphoenolpyruvate carboxykinase (PEPCK and glucose-6-phosphatase (G6Pase. Moreover, TS enhanced phosphorylation of AMP-activated protein kinase (phospho-AMPK both in skeletal muscle and liver tissue. Therefore, it is possible that the activation of AMPK by TS resulted in enhanced glucose uptake in skeletal muscle, contrasting with diminished gluconeogenesis in liver. TS exhibits hypolipidemic effect by decreasing the expressions of fatty acid synthase (FAS. Thus, antidiabetic properties of TS occurred as a result of decreased hepatic glucose production by PEPCK and G6Pase downregulation and improved insulin sensitization. Thus, amelioration of diabetic and dyslipidemic state by TS in HF-fed mice occurred by regulation of GLUT4, G6Pase, and FAS and phosphorylation of AMPK.

  9. Effects of acute hyperinsulinemia on insulin signal transduction and glucose transporters in ovine fetal skeletal muscle.

    Science.gov (United States)

    Anderson, Marianne S; Thamotharan, M; Kao, Doris; Devaskar, Sherin U; Qiao, Liping; Friedman, Jacob E; Hay, William W

    2005-02-01

    To test the effects of acute fetal hyperinsulinemia on the pattern and time course of insulin signaling in ovine fetal skeletal muscle, we measured selected signal transduction proteins in the mitogenic, protein synthetic, and metabolic pathways in the skeletal muscle of normally growing fetal sheep in utero. In experiment 1, 4-h hyperinsulinemic-euglycemic clamps were conducted in anesthetized twin fetuses to produce selective fetal hyperinsulinemia-euglycemia in one twin and euinsulinemia-euglycemia in the other. Serial skeletal muscle biopsies were taken from each fetus during the clamp and assayed by Western blot for selected insulin signal transduction proteins. Tyrosine phosphorylation of the insulin receptor, insulin receptor substrate-1, and the p85 subunit of phosphatidylinositol 3-kinase doubled at 30 min and gradually returned to control values by 240 min. Phosphorylation of extracellular signal-regulated kinase 1,2 was increased fivefold through 120 min of insulin infusion and decreased to control concentration by 240 min. Protein kinase B phosphorylation doubled at 30 min and remained elevated throughout the study. Phosphorylation of p70 S6K increased fourfold at 30, 60, and 120 min. In the second experiment, a separate group of nonanesthetized singleton fetuses was clamped to intermediate and high hyperinsulinemic-euglycemic conditions for 1 h. GLUT4 increased fourfold in the plasma membrane at 1 h, and hindlimb glucose uptake increased significantly at the higher insulin concentration. These data demonstrate that an acute increase in fetal plasma insulin concentration stimulates a unique pattern of insulin signal transduction proteins in intact skeletal muscle, thereby increasing pathways for mRNA translation, glucose transport, and cell growth.

  10. Aspartate-444 is essential for productive substrate interactions in a neuronal glutamate transporter.

    Science.gov (United States)

    Teichman, Shlomit; Kanner, Baruch I

    2007-06-01

    In the central nervous system, electrogenic sodium- and potassium-coupled glutamate transporters terminate the synaptic actions of this neurotransmitter. In contrast to acidic amino acids, dicarboxylic acids are not recognized by glutamate transporters, but the related bacterial DctA transporters are capable of transporting succinate and other dicarboxylic acids. Transmembrane domain 8 contains several residues that differ between these two types of transporters. One of these, aspartate-444 of the neuronal glutamate transporter EAAC1, is conserved in glutamate transporters, but a serine residue occupies this position in DctA transporters. When aspartate-444 is mutated to serine, cysteine, alanine, or even to glutamate, uptake of D-[(3)H]-aspartate as well as the inwardly rectifying steady-state currents induced by acidic amino acids is impaired. Even though succinate was not capable of inducing any steady-state transport currents, the dicarboxylic acid inhibited the sodium-dependent transient currents by the mutants with a neutral substitution at position 444. In the neutral substitution mutants inhibition of the transients was also observed with acidic amino acids. In the D444E mutant, acidic amino acids were potent inhibitors of the transient currents, whereas the apparent affinity for succinate was lower by at least three orders of magnitude. Even though L-aspartate could bind to D444E with a high apparent affinity, this binding resulted in inhibition rather than stimulation of the uncoupled anion conductance. Thus, a carboxylic acid-containing side chain at position 444 prevents the interaction of glutamate transporters with succinate, and the presence of aspartate itself at this position is crucial for productive substrate binding compatible with substrate translocation.

  11. Ginsenosides, ingredients of the root of Panax ginseng, are not substrates but inhibitors of sodium-glucose transporter 1.

    Science.gov (United States)

    Gao, Shengli; Kushida, Hirotaka; Makino, Toshiaki

    2017-01-01

    Recent pharmacokinetic studies have revealed that ginsenosides, the major ingredients of ginseng (the roots of Panax ginseng), are present in the plasma collected from subjects receiving ginseng, and speculated that ginsenosides might be actively transported via glucose transporters. We evaluated whether ginsenosides Rb1 and Rg1, and their metabolites from enteric bacteria act as substrates of sodium-glucose cotransporter (SGLT) 1, the major glucose transporter expressed on the apical side of intestinal epithelial cells. First, we evaluated the competing effects of ginseng extract and ginsenosides on the uptake of [(14)C]methyl-glucose, a substrate of SGLT1, by SGLT1-overexpressing HEK293 cells. A boiling water extract of ginseng inhibited SGLT1 in a concentration-dependent manner with an IC50 value of 0.85 mg/ml. By activity-guided fractionation, we determined that the fraction containing ginsenosides displayed an inhibitory effect on SGLT1. Of the ginsenosides evaluated, protopanaxatriol-type ginsenosides were not found to inhibit SGLT1, whereas protopanaxadiol-type ginsenosides, including ginsenosides Rd, Rg3, Rh2, F2 and compound K, exhibited significant inhibitory effects on SGLT1, with ginsenoside F2 having the highest activity with an IC50 value of 23.0 µM. Next, we measured the uptake of ginsenoside F2 and compound K into Caco-2 cells, a cell line frequently used to evaluate the intestinal absorption of drugs. The uptake of ginsenoside F2 and compound K into Caco-2 cells was not competitively inhibited by glucose. Furthermore, the uptake of ginsenoside F2 and compound K into SGLT1-overexpressing HEK293 cells was not significantly higher than into mock cells. Ginsenoside F2 and compound K did not appear to be substrates of SGLT1, although these compounds could inhibit SGLT1. Ginsenosides might be absorbed by passive diffusion through the intestinal membrane or actively transported via unknown transporters other than SGLT1.

  12. Vesicular stomatitis virus with the rabies virus glycoprotein directs retrograde transsynaptic transport among neurons in vivo

    Directory of Open Access Journals (Sweden)

    Kevin T Beier

    2013-02-01

    Full Text Available Defining the connections among neurons is critical to our understanding of the structure and function of the nervous system. Recombinant viruses engineered to transmit across synapses provide a powerful approach for the dissection of neuronal circuitry in vivo. We recently demonstrated that recombinant vesicular stomatitis virus (VSV can be endowed with anterograde or retrograde synaptic tracing ability by providing the virus with different glycoproteins. Here we extend the characterization of the transmission and gene expression of VSV with the rabies virus glycoprotein (RABV-G, and provide examples of its activity relative to the anterograde tracer form of rVSV. rVSV with RABV-G was found to drive strong expression of transgenes and to spread rapidly from neuron to neuron in only a retrograde manner. Depending upon how the RABV-G was delivered, VSV served as a polysynaptic or monosynaptic tracer, or was able to define projections through axonal uptake and retrograde transport. In animals co-infected with rVSV in its anterograde form, rVSV with RABV-G could be used to begin to characterize the similarities and differences in connections to a given area. rVSV with RABV-G provides a flexible, rapid, and versatile tracing tool that complements the previously described VSV-based anterograde transsynaptic tracer.

  13. Quantitative importance of the pentose phosphate pathway determined by incorporation of 13C from [2-13C]- and [3-13C]glucose into TCA cycle intermediates and neurotransmitter amino acids in functionally intact neurons

    DEFF Research Database (Denmark)

    Brekke, Eva Marie; Walls, Anne Byriel; Schousboe, Arne

    2012-01-01

    of (13)C-labeled glucose via the PPP does not appear to contribute to the production of releasable lactate, it contributes to labeling of tricarboxylic acid (TCA) cycle intermediates and related amino acids. Based on glutamate isotopomers, it was calculated that PPP activity accounts for ~6% of glucose...... metabolism in cortical neurons and ~4% in cerebellar neurons. This is the first demonstration that pyruvate generated from glucose via the PPP contributes to the synthesis of acetyl CoA for oxidation in the TCA cycle. Moreover, the fact that (13)C labeling from glucose is incorporated into glutamate proves...

  14. Alternating Carrier Models of Asymmetric Glucose Transport Violate the Energy Conservation Laws

    Science.gov (United States)

    Naftalin, Richard J

    2008-01-01

    Alternating access transporters with high-affinity externally facing sites and low-affinity internal sites relate substrate transit directly to the unliganded asymmetric “carrier” (Ci) distribution. When both bathing solutions contain equimolar concentrations of ligand, zero net flow of the substrate-carrier complex requires a higher proportion of unliganded low-affinity inside sites (\\documentclass[10pt]{article} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{pmc} \\usepackage[Euler]{upgreek} \\pagestyle{empty} \\oddsidemargin -1.0in \\begin{document} \\begin{equation*}{\\propto}1/K_{{\\mathrm{D}}}^{{\\mathrm{in}}}\\end{equation*}\\end{document}) and slower unliganded “free” carrier transit from inside to outside than in the reverse direction. However, asymmetric rates of unliganded carrier movement, kij, imply that an energy source, ΔGcarrier = RT ln (koi/kio) = RT ln (Cin/Cout) = RT ln (\\documentclass[10pt]{article} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{pmc} \\usepackage[Euler]{upgreek} \\pagestyle{empty} \\oddsidemargin -1.0in \\begin{document} \\begin{equation*}K_{{\\mathrm{D}}}^{{\\mathrm{in}}}/K_{{\\mathrm{D}}}^{{\\mathrm{out}}}\\end{equation*}\\end{document}), where R is the universal gas constant (8.314 Joules/M/K°), and T is the temperature, assumed here to be 300 K°, sustains the asymmetry. Without this invalid assumption, the constraints of carrier path cyclicity, combined with asymmetric ligand affinities and equimolarity at equilibrium, are irreconcilable, and any passive asymmetric uniporter or cotransporter model system, e.g., Na-glucose cotransporters, espousing this fundamental error is untenable. With glucose transport via GLUT1, the higher maximal rate and Km of net ligand exit compared to net ligand entry is only properly

  15. Replacement of both tryptophan residues at 388 and 412 completely abolished cytochalasin B photolabelling of the GLUT1 glucose transporter.

    Science.gov (United States)

    Inukai, K; Asano, T; Katagiri, H; Anai, M; Funaki, M; Ishihara, H; Tsukuda, K; Kikuchi, M; Yazaki, Y; Oka, Y

    1994-01-01

    A mutated GLUT1 glucose transporter, a Trp-388, 412 mutant whose tryptophans 388 and 412 were both replaced by leucines, was constructed by site-directed mutagenesis and expressed in Chinese hamster ovary cells. Glucose transport activity was decreased to approx. 30% in the Trp-388, 412 mutant compared with that in the wild type, a similar decrease in transport activity had been observed previously in the Trp-388 mutant and the Trp-412 mutant which had leucine at 388 and 412 respectively. Cytochalasin B labelling of the Trp-388 mutant was only decreased rather than abolished, a result similar to that obtained previously for the Trp-412 mutant. Cytochalasin B labelling was finally abolished completely in the Trp-388, 412 mutant, while cytochalasin B binding to this mutant was decreased to approx. 30% of that of the wild-type GLUT1 at the concentration used for photolabelling. This level of binding is thought to be adequate to detect labelling, assuming that the labelling efficiency of these transporters is similar. These findings suggest that cytochalasin B binds to the transmembrane domain of the glucose transporter in the vicinity of helix 10-11, and is inserted covalently by photoactivation at either the 388 or the 412 site. Images Figure 1 Figure 2 PMID:8092986

  16. Non-invasive, in-vivo monitoring of neuronal transport impairment in a mouse model of tauopathy using MEMRI

    Science.gov (United States)

    Bertrand, Anne; Khan, Umer; Hoang, Dung M.; Novikov, Dmitry S.; Krishnamurthy, Pavan; Rajamohamed Sait, Hameetha B.; Little, Benjamin W.; Sigurdsson, Einar M.; Wadghiri, Youssef Z.

    2012-01-01

    The impairment of axonal transport by overexpression or hyperphosphorylation of tau is well documented for in vitro conditions; however, only a few studies on this phenomenon have been conducted in vivo, using invasive procedures, and with contradictory results. Here we used the non-invasive, Manganese-Enhanced Magnetic Resonance Imaging technique (MEMRI), to study for the first time a pure model of tauopathy, the JNPL3 transgenic mouse line, which overexpresses a mutated (P301L) form of the human tau protein. We show progressive impairment in neuronal transport as tauopathy advances. These findings are further supported by a significant correlation between the severity of the impairment in neuronal transport assessed by MEMRI, and the degree of abnormal tau assessed by histology. Unlike conventional techniques that focus on axonal transport measurement, MEMRI can provide a global analysis of neuronal transport, i.e. from dendrites to axons and at the macroscopic scale of fiber tracts. Neuronal transport impairment has been shown to be a key pathogenic process in Alzheimer’s disease and numerous other neurodegenerative disorders. Hence, MEMRI provides a promising set of functional biomarkers to be used during preclinical trials to facilitate the selection of new drugs aimed at restoring neuronal transport in neurodegenerative diseases. PMID:22960250

  17. Protection against Oxygen-Glucose Deprivation/Reperfusion Injury in Cortical Neurons by Combining Omega-3 Polyunsaturated Acid with Lyciumbarbarum Polysaccharide

    Directory of Open Access Journals (Sweden)

    Zhe Shi

    2016-01-01

    Full Text Available Ischemic stroke, characterized by the disturbance of the blood supply to the brain, is a severe worldwide health threat with high mortality and morbidity. However, there is no effective pharmacotherapy for ischemic injury. Currently, combined treatment is highly recommended for this devastating injury. In the present study, we investigated neuroprotective effects of the combination of omega-3 polyunsaturated fatty acids (ω-3 PUFAs and Lyciumbarbarum polysaccharide (LBP on cortical neurons using an in vitro ischemic model. Our study demonstrated that treatment with docosahexaenoic acid (DHA, a major component of the ω-3 PUFAs family, significantly inhibited the increase of intracellular Ca2+ in cultured wild type (WT cortical neurons subjected to oxygen-glucose deprivation/reperfusion (OGD/R injury and promoted their survival compared with the vehicle-treated control. The protective effects were further confirmed in cultured neurons with high endogenous ω-3 PUFAs that were isolated from fat-1 mice, in that a higher survival rate was found in fat-1 neurons compared with wild-type neurons after OGD/R injury. Our study also found that treatment with LBP (50 mg/L activated Trk-B signaling in cortical neurons and significantly attenuated OGD/R-induced cell apoptosis compared with the control. Notably, both combining LBP treatment with ω-3 PUFAs administration to WT neurons and adding LBP to fat-1 neurons showed enhanced effects on protecting cortical neurons against OGD/R injury via concurrently regulating the intracellular calcium overload and neurotrophic pathway. The results of the study suggest that ω-3 PUFAs and LBP are promising candidates for combined pharmacotherapy for ischemic stroke.

  18. Protection against Oxygen-Glucose Deprivation/Reperfusion Injury in Cortical Neurons by Combining Omega-3 Polyunsaturated Acid with Lyciumbarbarum Polysaccharide.

    Science.gov (United States)

    Shi, Zhe; Wu, Di; Yao, Jian-Ping; Yao, Xiaoli; Huang, Zhijian; Li, Peng; Wan, Jian-Bo; He, Chengwei; Su, Huanxing

    2016-01-13

    Ischemic stroke, characterized by the disturbance of the blood supply to the brain, is a severe worldwide health threat with high mortality and morbidity. However, there is no effective pharmacotherapy for ischemic injury. Currently, combined treatment is highly recommended for this devastating injury. In the present study, we investigated neuroprotective effects of the combination of omega-3 polyunsaturated fatty acids (ω-3 PUFAs) and Lyciumbarbarum polysaccharide (LBP) on cortical neurons using an in vitro ischemic model. Our study demonstrated that treatment with docosahexaenoic acid (DHA), a major component of the ω-3 PUFAs family, significantly inhibited the increase of intracellular Ca(2+) in cultured wild type (WT) cortical neurons subjected to oxygen-glucose deprivation/reperfusion (OGD/R) injury and promoted their survival compared with the vehicle-treated control. The protective effects were further confirmed in cultured neurons with high endogenous ω-3 PUFAs that were isolated from fat-1 mice, in that a higher survival rate was found in fat-1 neurons compared with wild-type neurons after OGD/R injury. Our study also found that treatment with LBP (50 mg/L) activated Trk-B signaling in cortical neurons and significantly attenuated OGD/R-induced cell apoptosis compared with the control. Notably, both combining LBP treatment with ω-3 PUFAs administration to WT neurons and adding LBP to fat-1 neurons showed enhanced effects on protecting cortical neurons against OGD/R injury via concurrently regulating the intracellular calcium overload and neurotrophic pathway. The results of the study suggest that ω-3 PUFAs and LBP are promising candidates for combined pharmacotherapy for ischemic stroke.

  19. Effects of oxygen and glucose deprivation on the expression and distribution of neuronal and inducible nitric oxide synthases and on protein nitration in rat cerebral cortex.

    Science.gov (United States)

    Alonso, David; Serrano, Julia; Rodríguez, Ignacio; Ruíz-Cabello, Jesús; Fernández, Ana Patricia; Encinas, Juan Manuel; Castro-Blanco, Susana; Bentura, María Luisa; Santacana, María; Richart, Ana; Fernández-Vizarra, Paula; Uttenthal, Lars Otto; Rodrigo, José

    2002-02-04

    Changes in the nitric oxide (NO) system of the rat cerebral cortex were investigated by immunohistochemistry, immunoblotting, NO synthase (NOS) activity assay, and magnetic resonance imaging (MRI) in an experimental model of global cerebral ischemia and reperfusion. Brains were perfused transcardially with an oxygenated plasma substitute and subjected to 30 minutes of oxygen and glucose deprivation, followed by reperfusion for up to 12 hours with oxygenated medium containing glucose. A sham group was perfused without oxygen or glucose deprivation, and a further group was treated with the NOS inhibitor N(omega)-nitro-L-arginine methyl ester (L-NAME) before and during perfusion. Global ischemia led to cerebrocortical injury as shown by diffusion MRI. This was accompanied by increasing morphologic changes in the large type I interneurons expressing neuronal NOS (nNOS) and the appearance of nNOS immunoreactivity in small type II neurons. The nNOS-immunoreactive band and calcium-dependent NOS activity showed an initial increase, followed by a fall after 6 hours of reperfusion. Inducible NOS immunoreactivity appeared in neurons, especially pyramidal cells of layers IV-V, after 4 hours of reperfusion, with corresponding changes on immunoblotting and in calcium-independent NOS activity. Immunoreactive protein nitrotyrosine, present in the nuclear area of neurons in nonperfused controls and sham-perfused animals, showed changes in intensity and distribution, appearing in the neuronal processes during the reperfusion period. Prior and concurrent L-NAME administration blocked the changes on diffusion MRI and attenuated the morphologic changes, suggesting that NO and consequent peroxynitrite formation during ischemia-reperfusion contributes to cerebral injury.

  20. High-Affinity Glucose Transport in Aspergillus nidulans Is Mediated by the Products of Two Related but Differentially Expressed Genes

    Science.gov (United States)

    Ventura, Luisa; González, Ramón; Ramón, Daniel; MacCabe, Andrew P.

    2014-01-01

    Independent systems of high and low affinity effect glucose uptake in the filamentous fungus Aspergillus nidulans. Low-affinity uptake is known to be mediated by the product of the mstE gene. In the current work two genes, mstA and mstC, have been identified that encode high-affinity glucose transporter proteins. These proteins' primary structures share over 90% similarity, indicating that the corresponding genes share a common origin. Whilst the function of the paralogous proteins is little changed, they differ notably in their patterns of expression. The mstC gene is expressed during the early phases of germination and is subject to CreA-mediated carbon catabolite repression whereas mstA is expressed as a culture tends toward carbon starvation. In addition, various pieces of genetic evidence strongly support allelism of mstC and the previously described locus sorA. Overall, our data define MstC/SorA as a high-affinity glucose transporter expressed in germinating conidia, and MstA as a high-affinity glucose transporter that operates in vegetative hyphae under conditions of carbon limitation. PMID:24751997

  1. High-affinity glucose transport in Aspergillus nidulans is mediated by the products of two related but differentially expressed genes.

    Directory of Open Access Journals (Sweden)

    Josep V Forment

    Full Text Available Independent systems of high and low affinity effect glucose uptake in the filamentous fungus Aspergillus nidulans. Low-affinity uptake is known to be mediated by the product of the mstE gene. In the current work two genes, mstA and mstC, have been identified that encode high-affinity glucose transporter proteins. These proteins' primary structures share over 90% similarity, indicating that the corresponding genes share a common origin. Whilst the function of the paralogous proteins is little changed, they differ notably in their patterns of expression. The mstC gene is expressed during the early phases of germination and is subject to CreA-mediated carbon catabolite repression whereas mstA is expressed as a culture tends toward carbon starvation. In addition, various pieces of genetic evidence strongly support allelism of mstC and the previously described locus sorA. Overall, our data define MstC/SorA as a high-affinity glucose transporter expressed in germinating conidia, and MstA as a high-affinity glucose transporter that operates in vegetative hyphae under conditions of carbon limitation.

  2. Insulin Resistance in Nondiabetic Peritoneal Dialysis Patients: Associations with Body Composition, Peritoneal Transport, and Peritoneal Glucose Absorption.

    Science.gov (United States)

    Bernardo, Ana Paula; Oliveira, Jose C; Santos, Olivia; Carvalho, Maria J; Cabrita, Antonio; Rodrigues, Anabela

    2015-12-07

    Insulin resistance has been associated with cardiovascular disease in peritoneal dialysis patients. Few studies have addressed the impact of fast transport status or dialysis prescription on insulin resistance. The aim of this study was to test whether insulin resistance is associated with obesity parameters, peritoneal transport rate, and glucose absorption. Insulin resistance was evaluated with homeostasis model assessment method (HOMA-IR), additionally corrected by adiponectin (HOMA-AD). Enrolled patients were prevalent nondiabetics attending at Santo António Hospital Peritoneal Dialysis Unit, who were free of hospitalization or infectious events in the previous 3 months (51 patients aged 50.4 ± 15.9 years, 59% women). Leptin, adiponectin, insulin-like growth factor-binding protein 1 (IGFBP-1), and daily glucose absorption were also measured. Lean tissue index, fat tissue index (FTI), and relative fat mass (rel.FM) were assessed using multifrequency bioimpedance. Patients were categorized according to dialysate to plasma creatinine ratio at 4 hours, 3.86% peritoneal equilibration test, and obesity parameters. Obesity was present in 49% of patients according to rel.FM. HOMA-IR correlated better with FTI than with body mass index. Significant correlations were found in obese, but not in nonobese patients, between HOMA-IR and leptin, leptin/adiponectin ratio (LAR), and IGFBP-1. HOMA-IR correlated with HOMA-AD, but did not correlate with glucose absorption or transport rate. There were no significant differences in insulin resistance indices, glucose absorption, and body composition parameters between fast and nonfast transporters. A total of 18 patients (35.3%) who had insulin resistance presented with higher LAR and rel.FM (7.3 [12.3, interquartile range] versus 0.7 [1.4, interquartile range], Pinsulin resistance. FTI and LAR were independent correlates of HOMA-IR in multivariate analysis adjusted for glucose absorption and small-solute transport (r=0

  3. Low-Dose Ethanol Preconditioning Protects Against Oxygen-Glucose Deprivation/Reoxygenation-Induced Neuronal Injury By Activating Large Conductance, Ca(2+)-Activated K(+) Channels In Vitro.

    Science.gov (United States)

    Su, Fang; Guo, An-Chen; Li, Wei-Wei; Zhao, Yi-Long; Qu, Zheng-Yi; Wang, Yong-Jun; Wang, Qun; Zhu, Yu-Lan

    2017-02-01

    Increasing evidence suggests that low to moderate ethanol ingestion protects against the deleterious effects of subsequent ischemia/reperfusion; however, the underlying mechanism has not been elucidated. In the present study, we showed that expression of the neuronal large-conductance, Ca(2+)-activated K(+) channel (BKCa) α-subunit was upregulated in cultured neurons exposed to oxygen-glucose deprivation/reoxygenation (OGD/R) compared with controls. Preconditioning with low-dose ethanol (10 mmol/L) increased cell survival rate in neurons subjected to OGD/R, attenuated the OGD/R-induced elevation of cytosolic Ca(2+) levels, and reduced the number of apoptotic neurons. Western blots revealed that ethanol preconditioning upregulated expression of the anti-apoptotic protein Bcl-2 and downregulated the pro-apoptotic protein Bax. The protective effect of ethanol preconditioning was antagonized by a BKCa channel inhibitor, paxilline. Inside-out patches in primary neurons also demonstrated the direct activation of the BKCa channel by 10 mmol/L ethanol. The above results indicated that low-dose ethanol preconditioning exerts its neuroprotective effects by attenuating the elevation of cytosolic Ca(2+) and preventing neuronal apoptosis, and this is mediated by BKCa channel activation.

  4. Hyperbaric oxygen preconditioning induces tolerance against oxidative injury and oxygen-glucose deprivation by up-regulating heat shock protein 32 in rat spinal neurons.

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    Guoyang Huang

    Full Text Available OBJECTIVE: Hyperbaric oxygen (HBO preconditioning (HBO-PC has been testified to have protective effects on spinal cord injury (SCI. However, the mechanisms remain enigmatic. The present study aimed to explore the effects of HBO-PC on primary rat spinal neurons against oxidative injury and oxygen-glucose deprivation (OGD and the relationship with heat shock proteins (HSPs. METHODS: Primary rat spinal neurons after 7 days of culture were used in this study. HSPs were detected in rat spinal neurons following a single exposure to HBO at different time points by Western blot. Using lactate dehydrogenase release assay and cell counting kit-8 assay, the injuries induced by hydrogen peroxide (H2O2 insult or OGD were determined and compared among neurons treated with HBO-PC with or without HSP inhibitors. RESULTS: The results of Western blot showed that HSP27, HSP70 and HSP90 have a slight but not significant increase in primary neurons following HBO exposure. However, HSP32 expression significantly increased and reached highest at 12 h following HBO exposure. HBO-PC significantly increased the cell viability and decreased the medium lactate dehydrogenase content in cultures treated with H2O2 or OGD. Pretreatment with zinc protoporphyrin IX, a specific inhibitor of HSP32, significantly blocked the protective effects of HBO-PC. CONCLUSIONS: These results suggest that HBO-PC could protect rat spinal neurons in vitro against oxidative injury and OGD mostly by up-regulating of HSP32 expression.

  5. Effect of training and detraining on skeletal muscle glucose transporter (GLUT4) content in rats.

    Science.gov (United States)

    Neufer, P D; Shinebarger, M H; Dohm, G L

    1992-09-01

    The aim of the present study was to examine the effects of treadmill exercise training and detraining on the skeletal muscle fiber type specific expression of the insulin-regulated glucose transporter protein (GLUT4) in rats. GLUT4 protein content was determined by Western and dot-blot analysis, using a polyclonal antibody raised against the carboxy-terminal peptide. Rats were sacrificed 24 h after the last training session. There were no significant changes in muscle GLUT4 after 1 day or 1 week of training. Six weeks of training increased GLUT4 protein content 1.4- to 1.7-fold (p < 0.05) over controls in the soleus and red vastus lateralis, whereas no significant change was evident in the white vastus lateralis muscle. GLUT4 protein content in both soleus and red vastus lateralis muscle returned to near control values after 7 days of detraining. Similar to GLUT4, citrate synthase activity showed no change after 1 day or 1 week of training, increased 1.8-fold over controls after 6 weeks of training, but returned to control values after 7 days detraining. These findings demonstrate that muscle GLUT4 protein is increased in rats with as little as 6 weeks of treadmill exercise training but that the adaptation is lost within 1 week of detraining. It is suggested that expression of the GLUT4 protein is coordinated with the well-documented adaptations in oxidative enzyme activity with endurance training and detraining.

  6. Apigenin inhibits the proliferation of adenoid cystic carcinoma via suppression of glucose transporter-1

    Science.gov (United States)

    FANG, JIN; BAO, YANG-YANG; ZHOU, SHUI-HONG; FAN, JUN

    2015-01-01

    Apigenin is a natural phyto-oestrogen flavonoid, which exerts various biological effects, including anti-oxidative, anti-inflammatory and anticancer activities. In addition, apigenin has recently been reported to target hypoxic markers; however, there are currently no studies regarding the association between apigenin and glucose transporter-1 (GLUT-1) in adenoid cystic carcinoma (ACC). The present study investigated whether apigenin inhibits the proliferation of ACC cells or suppresses the expression of GLUT-1 in ACC cells. The results of the present study demonstrated that apigenin inhibits ACC-2 cell growth in a dose- and time-dependent manner. Treatment with apigenin also induced apoptosis and G2/M-phase arrest in a dose- and time-dependent manner. Corresponding with the above results, the expression levels of GLUT-1 were significantly decreased following treatment in a dose- and time-dependent manner. These results suggest that the inhibition of ACC-2 cell growth by apigenin may be due to the decreased expression of GLUT-1. PMID:26300442

  7. Impairment of tight junctions and glucose transport in endothelial cells of human cerebral cavernous malformations.

    Science.gov (United States)

    Schneider, Hannah; Errede, Mariella; Ulrich, Nils H; Virgintino, Daniela; Frei, Karl; Bertalanffy, Helmut

    2011-06-01

    Cerebral cavernous malformations (CCMs) often cause hemorrhages that can result in severe clinical manifestations, including hemiparesis and seizures. The underlying mechanisms of the aggressive behavior of CCMs are undetermined to date, but alterations of vascular matrix components may be involved. We compared the localization of the tight junction proteins (TJPs) in 12 CCM specimens and the expression of glucose transporter 1 (GLUT-1), which is sensitive to alterations in TJP levels, in 5 CCM specimens with those in 5 control temporal lobectomy specimens without CCM by immunofluorescence microscopy. The TJPs occludin, claudin-5, and zonula occludens ZO-1 were downregulated at intercellular contact sites and partly redistributed within the surrounding tissue in the CCM samples; there was also a marked reduction of GLUT-1 immunoreactivity compared with that in control specimens. Corresponding analysis using quantitative real-time reverse transcription polymerase chain reaction on 8 CCM and 8 control specimens revealed significant downregulation of mRNA expression of occludin, claudin-5, ZO-1, and GLUT-1. The altered expression and localization of the TJPs at interendothelial contact sites accompanied by a reduction of GLUT-1 expression in dilated CCM microvessels likely affect vascular matrix stability and may contribute to hemorrhages of CCMs.

  8. Relation Between Fluorodeoxyglucose Uptake and Glucose Transporter 1 Expression in Gastric signet Ring Cell Carcinoma

    Energy Technology Data Exchange (ETDEWEB)

    Choi, Bong Hoi; Song, Hee Sung; An, Young Sil; Han, Sang Uk; Kim, Jang Hee; Yoon, Joon kee [Ajou Univ. School of medicine, Suwon (Korea, Republic of)

    2011-03-15

    Gastic signet ring cell carcinoma (GSRC) is known to have low fluorodeoxyglucose (FDG) uptake. The aim of the study was to investigate the relation between FDG uptake and glucose transporter (GLUT) 1 expression and clinicopathologic parameters in cases of GSRC. Forty patients (28 men, mean age 54{+-}12 years) with histologically confirmed GSRC who underwent pre operative [{sup 18}F]FDG PET/CT were enrolled. Maximum standardized uptake values (SUVmax) were compared with clinicopathologic parameters and GLUT 1 expression. Cases were divided based on GLUT 1 expression in tumor tissues into a membranous group (n=17) and a cytoplasmic group (n=23). Mean SUVmax was significantly higher in the membranous group than in the cytoplasmic group (6.06{+-}2.79 vs, 3.67{+-}1.54, P=0.03). Gastric wall invasion, depth of invasion, extent of LN metastasis, overall stage, and tumor size were found to be related to SUVmax. On the other hand, age, sex, and the presence of distant metastasis were not related to SUVmax. Multiveriate analysis revealed that membranous GLCT 1 expression and the extent of LN metastasis independently predicted high FDG uptake. This study demonstrates that high FDG uptake is mediated by membranous GLUT 1 expression in GSRC.

  9. Glucose transporter 1-positive endothelial cells in infantile hemangioma exhibit features of facultative stem cells

    Science.gov (United States)

    Huang, Lan; Nakayama, Hironao; Klagsbrun, Michael; Mulliken, John B.; Bischoff, Joyce

    2014-01-01

    Endothelial glucose transporter 1 (GLUT1) is a definitive and diagnostic marker for infantile hemangioma (IH), a vascular tumor of infancy. To date, GLUT1-positive endothelial cells in IH have not been quantified nor directly isolated and studied. We isolated GLUT1-positive and GLUT1-negative endothelial cells from IH specimens and characterized their proliferation, differentiation and response to propranolol, a first-line therapy for IH, and to rapamycin, an mTOR pathway inhibitor used to treat an increasingly wide array of proliferative disorders. Although freshly isolated GLUT1-positive cells, selected using anti-GLUT1 magnetic beads, expressed endothelial markers CD31, VE-Cadherin and VEGFR2, they converted to a mesenchymal phenotype after three weeks in culture. In contrast, GLUT1-negative endothelial cells exhibited a stable endothelial phenotype in vitro. GLUT1-selected cells were clonogenic when plated as single cells and could be induced to re-differentiate into endothelial cells, or into pericyte/smooth muscle cells or into adipocytes, indicating a stem cell-like phenotype. These data demonstrate that, although they appear and function in the tumor as bona fide endothelial cells, the GLUT1-positive endothelial cells display properties of facultative stem cells. Pretreatment with rapamycin for 4 days significantly slowed proliferation of GLUT1-selected cells, whereas propranolol pretreatment had no effect. These results reveal for the first time the facultative nature of GLUT1-positive endothelial cells in infantile hemangioma. PMID:25187207

  10. Neuronal calcium sensor synaptotagmin-9 is not involved in the regulation of glucose homeostasis or insulin secretion

    DEFF Research Database (Denmark)

    Gustavsson, Natalia; Wang, Xiaorui; Wang, Yue

    2010-01-01

    /PRINCIPAL FINDINGS: In this study we tested whether synaptotagmin-9 participates in the regulation of glucose-stimulated insulin release by using pancreas-specific synaptotagmin-9 knockout (p-S9X) mice. Deletion of synaptotagmin-9 in the pancreas resulted in no changes in glucose homeostasis or body weight. Glucose...

  11. Glutamate transporter activity promotes enhanced Na+/K+-ATPase-mediated extracellular K+ management during neuronal activity

    DEFF Research Database (Denmark)

    Larsen, Brian Roland; Holm, Rikke; Vilsen, Bente;

    2016-01-01

    , in addition, Na+/K+-ATPase-mediated K+ clearance could be governed by astrocytic [Na+]i. During most neuronal activity, glutamate is released in the synaptic cleft and is re-absorbed by astrocytic Na+-coupled glutamate transporters, thereby elevating [Na+]i. It thus remains unresolved whether the different Na...... constellations in Xenopus oocytes and determined their apparent Na+ affinity in intact oocytes and isolated membranes. The Na+/K+-ATPase was not fully saturated at basal astrocytic [Na+]i, irrespective of isoform constellation, although the β1 subunit conferred lower apparent Na+ affinity to the α1 and α2...

  12. Genetic changes during a laboratory adaptive evolution process that allowed fast growth in glucose to an Escherichia coli strain lacking the major glucose transport system

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    Aguilar César

    2012-08-01

    Full Text Available Abstract Background Escherichia coli strains lacking the phosphoenolpyruvate: carbohydrate phosphotransferase system (PTS, which is the major bacterial component involved in glucose transport and its phosphorylation, accumulate high amounts of phosphoenolpyruvate that can be diverted to the synthesis of commercially relevant products. However, these strains grow slowly in glucose as sole carbon source due to its inefficient transport and metabolism. Strain PB12, with 400% increased growth rate, was isolated after a 120 hours adaptive laboratory evolution process for the selection of faster growing derivatives in glucose. Analysis of the genetic changes that occurred in the PB12 strain that lacks PTS will allow a better understanding of the basis of its growth adaptation and, therefore, in the design of improved metabolic engineering strategies for enhancing carbon diversion into the aromatic pathways. Results Whole genome analyses using two different sequencing methodologies: the Roche NimbleGen Inc. comparative genome sequencing technique, and high throughput sequencing with Illumina Inc. GAIIx, allowed the identification of the genetic changes that occurred in the PB12 strain. Both methods detected 23 non-synonymous and 22 synonymous point mutations. Several non-synonymous mutations mapped in regulatory genes (arcB, barA, rpoD, rna and in other putative regulatory loci (yjjU, rssA and ypdA. In addition, a chromosomal deletion of 10,328 bp was detected that removed 12 genes, among them, the rppH, mutH and galR genes. Characterization of some of these mutated and deleted genes with their functions and possible functions, are presented. Conclusions The deletion of the contiguous rppH, mutH and galR genes that occurred simultaneously, is apparently the main reason for the faster growth of the evolved PB12 strain. In support of this interpretation is the fact that inactivation of the rppH gene in the parental PB11 strain substantially increased

  13. Role of the water extract from Coccinia indica stem on the stimulation of glucose transport in L8 myotubes

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    Chaweewan Jansakul

    2006-11-01

    Full Text Available Hypoglycemic effect of Coccinia indica used for treatment of diabetes in traditional remedies has known to relate with increased transport of glucose into peripheral tissues. However, the cellular mechanisms for this effect remain unclear. This present study reports that the water extract (WE of C. indica stem exhibited a dose-dependent induction of 2-deoxyglucose (2-DG uptake in rat L8 myotubes. Maximal uptake was observed with approximately 3-fold increase in 2-DG transport in 16 h treatment compared with the control. Effect of WE was stronger than that of 1 mM metformin. The effects of insulin and WE were additive. WE-induced glucose uptake was significantly inhibited by cycloheximide and partially reversed by SB203580. GLUT1 protein was markedly increased in response to WE. Conversely, WE had no effect on GLUT4 protein level. Redistribution of GLUT4 to the plasma membrane was demonstrated. Triterpenoids and carbohydrates were detected in WE. In conclusion, new GLUT1 protein synthesis is necessary for WEstimulated glucose transport while p38-MAPK-dependent activation of transporter intrinsic activity partly contributes to WE action. These results may explain and support the use of C. indica for the prevention and treatment of diabetes.

  14. Suppressed intrinsic catalytic activity of GLUT1 glucose transporters in insulin-sensitive 3T3-L1 adipocytes

    Energy Technology Data Exchange (ETDEWEB)

    Harrison, S.A.; Buxton, J.M.; Czech, M.P. (Univ. of Massachusetts Medical Center, Worcester (United States))

    1991-09-01

    Previous studies indicated that the erythroid-type (GLUT1) glucose transporter isoform contributes to basal but not insulin-stimulated hexose transport in mouse 3T3-L1 adipocytes. In the present studies it was found that basal hexose uptake in 3T3-L1 adipocytes was about 50% lower than that in 3T3-L1 or CHO-K1 fibroblasts. Intrinsic catalytic activities of GLUT1 transporters in CHO-K1 and 3T3-L1 cells were compared by normalizing these hexose transport rates to GLUT1 content on the cell surface, as measured by two independent methods. Cell surface GLUT1 levels in 3T3-L1 fibroblasts and adipocytes were about 10- and 25-fold higher, respectively, than in CHO-K1 fibroblasts, as assessed with an anti-GLUT1 exofacial domain antiserum, delta. The large excess of cell surface GLUT1 transporters in 3T3-L1 adipocytes relative to CHO-K1 fibroblasts was confirmed by GLUT1 protein immunoblot analysis and by photoaffinity labeling (with 3-({sup 125}I)iodo-4-azidophenethylamido-7-O-succinyldeacetylforskolin) of glucose transporters in isolated plasma membranes. Thus, GLUT1 intrinsic activity is markedly reduced in 3T3-L1 fibroblasts compared with the CHO-K1 fibroblasts, and further reduction occurs upon differentiation to adipocytes. The authors conclude that a mechanism that markedly suppresses basal hexose transport catalyzed by GLUT1 is a major contributor to the dramatic insulin sensitivity of glucose uptake in 3T3-L1 adipocytes.

  15. Tamoxifen mediated estrogen receptor activation protects against early impairment of hippocampal neuron excitability in an oxygen/glucose deprivation brain slice ischemia model.

    Science.gov (United States)

    Zhang, Huaqiu; Xie, Minjie; Schools, Gary P; Feustel, Paul F; Wang, Wei; Lei, Ting; Kimelberg, Harold K; Zhou, Min

    2009-01-09

    Pretreatment of ovarectomized rats with estrogen shows long-term protection via activation of the estrogen receptor (ER). However, it remains unknown whether activation of the ER can provide protection against early neuronal damage when given acutely. We simulated ischemic conditions by applying oxygen and glucose deprived (OGD) solution to acute male rat hippocampal slices and examined the neuronal electrophysiological changes. Pyramidal neurons and interneurons showed a time-dependent membrane potential depolarization and reduction in evoked action potential frequency and amplitude over a 10 to 15 min OGD exposure. These changes were largely suppressed by 10 microM TAM. The TAM effect was neuron-specific as the OGD-induced astrocytic membrane potential depolarization was not altered. The TAM effect was mediated through ER activation because it could be simulated by 17beta-estradiol and was completely inhibited by the ER inhibitor ICI 182, 780, and is therefore an example of TAM's selective estrogen receptor modulator (SERM) action. We further show that TAM's effects on OGD-induced impairment of neuronal excitability was largely due to activation of neuroprotective BK channels, as the TAM effect was markedly attenuated by the BK channel inhibitor paxilline at 10 microM. TAM also significantly reduced the frequency and amplitude of AMPA receptor mediated spontaneous excitatory postsynaptic currents (sEPSCs) in pyramidal neurons which is an early consequence of OGD. Altogether, this study demonstrates that both 17beta-estradiol and TAM attenuate neuronal excitability impairment early on in a simulated ischemia model via ER activation mediated potentiation of BK K(+) channels and reduction in enhanced neuronal AMPA/NMDA receptor-mediated excitotoxicity.

  16. A possible role of the non-GAT1 GABA transporters in transfer of GABA from GABAergic to glutamatergic neurons in mouse cerebellar neuronal cultures.

    Science.gov (United States)

    Suñol, C; Babot, Z; Cristòfol, R; Sonnewald, U; Waagepetersen, H S; Schousboe, A

    2010-09-01

    Cultures of dissociated cerebellum from 7-day-old mice were used to investigate the mechanism involved in synthesis and cellular redistribution of GABA in these cultures consisting primarily of glutamatergic granule neurons and a smaller population of GABAergic Golgi and stellate neurons. The distribution of GAD, GABA and the vesicular glutamate transporter VGlut-1 was assessed using specific antibodies combined with immunofluorescence microscopy. Additionally, tiagabine, SKF 89976-A, betaine, beta-alanine, nipecotic acid and guvacine were used to inhibit the GAT1, betaine/GABA (BGT1), GAT2 and GAT3 transporters. Only a small population of cells were immuno-stained for GAD while many cells exhibited VGlut-1 like immuno-reactivity which, however, never co-localized with GAD positive neurons. This likely reflects the small number of GABAergic neurons compared to the glutamatergic granule neurons constituting the majority of the cells. GABA uptake exhibited the kinetics of high affinity transport and could be partly (20%) inhibited by betaine (IC(50) 142 microM), beta-alanine (30%) and almost fully (90%) inhibited by SKF 89976-A (IC(50) 0.8 microM) or nipecotic acid and guvacine at 1 mM concentrations (95%). Essentially all neurons showed GABA like immunostaining albeit with differences in intensity. The results indicate that GABA which is synthesized in a small population of GAD-positive neurons is redistributed to essentially all neurons including the glutamatergic granule cells. GAT1 is not likely involved in this redistribution since addition of 15 microM tiagabine (GAT1 inhibitor) to the culture medium had no effect on the overall GABA content of the cells. Likewise the BGT1 transporter cannot alone account for the redistribution since inclusion of 3 mM betaine in the culture medium had no effect on the overall GABA content. The inhibitory action of beta-alanine and high concentrations of nipecotic acid and guvacine on GABA transport strongly suggests that also

  17. A new Michaelis-Menten-based kinetic model for transport and phosphorylation of glucose and its analogs in skeletal muscle.

    Science.gov (United States)

    Huang, Hsuan-Ming; Ismail-Beigi, Faramarz; Muzic, Raymond F

    2011-08-01

    A new model is introduced that individually resolves the delivery, transport, and phosphorylation steps of metabolism of glucose and its analogs in skeletal muscle by interpreting dynamic positron emission tomography (PET) data. The model uniquely utilizes information obtained from the competition between glucose and its radiolabeled analogs. Importantly, the model avoids use of a lumped constant which may depend on physiological state. Four basic physiologic quantities constitute our model parameters, including the fraction of total tissue space occupied by interstitial space (f(IS)), a flow-extraction product and interstitial (IS(g)) and intracellular (IC(g)) glucose concentrations. Using the values of these parameters, cellular influx (CI) and efflux (CE) of glucose, glucose phosphorylation rate (PR), and maximal transport (V(G)) and phosphorylation capacities (V(H)) can all be determined. Herein, the theoretical derivation of our model is addressed and characterizes its properties via simulation. Specifically, the model performance is evaluated by simulation of basal and euglycemic hyperinsulinemic (EH) conditions. In fitting the model-generated, synthetic data (including noise), mean estimates of all but IC(g) of the parameter values are within 5% of their values for both conditions. In addition, mean errors of CI, PR, and V(G) are less than 5% whereas those of VH and CE are not. It is concluded that under the conditions tested, the novel model can provide accurate parameter estimates and physiological quantities, except IC(g) and two quantities that are dependent on IC(g), namely CE and VH. However, the ability to estimate IC(g) seems to improve with increases in intracellular glucose concentrations as evidenced by comparing IC(g) estimates under basal vs EH conditions.

  18. Tissue Specific Expression of Cre in Rat Tyrosine Hydroxylase and Dopamine Active Transporter-Positive Neurons.

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    Zhenyi Liu

    Full Text Available The rat is a preferred model system over the mouse for neurological studies, and cell type-specific Cre expression in the rat enables precise ablation of gene function in neurons of interest, which is especially valuable for neurodegenerative disease modeling and optogenetics. Yet, few such Cre rats are available. Here we report the characterization of two Cre rats, tyrosine hydroxylase (TH-Cre and dopamine active transporter (DAT or Slc6a3-Cre, by using a combination of immunohistochemistry (IHC and mRNA fluorescence in situ hybridization (FISH as well as a fluorescent reporter for Cre activity. We detected Cre expression in expected neurons in both Cre lines. Interestingly, we also found that in Th-Cre rats, but not DAT-Cre rats, Cre is expressed in female germ cells, allowing germline excision of the floxed allele and hence the generation of whole-body knockout rats. In summary, our data demonstrate that targeted integration of Cre cassette lead to faithful recapitulation of expression pattern of the endogenous promoter, and mRNA FISH, in addition to IHC, is an effective method for the analysis of the spatiotemporal gene expression patterns in the rat brain, alleviating the dependence on high quality antibodies that are often not available against rat proteins. The Th-Cre and the DAT-Cre rat lines express Cre in selective subsets of dopaminergic neurons and should be particularly useful for researches on Parkinson's disease.

  19. Tissue Specific Expression of Cre in Rat Tyrosine Hydroxylase and Dopamine Active Transporter-Positive Neurons.

    Science.gov (United States)

    Liu, Zhenyi; Brown, Andrew; Fisher, Dan; Wu, Yumei; Warren, Joe; Cui, Xiaoxia

    2016-01-01

    The rat is a preferred model system over the mouse for neurological studies, and cell type-specific Cre expression in the rat enables precise ablation of gene function in neurons of interest, which is especially valuable for neurodegenerative disease modeling and optogenetics. Yet, few such Cre rats are available. Here we report the characterization of two Cre rats, tyrosine hydroxylase (TH)-Cre and dopamine active transporter (DAT or Slc6a3)-Cre, by using a combination of immunohistochemistry (IHC) and mRNA fluorescence in situ hybridization (FISH) as well as a fluorescent reporter for Cre activity. We detected Cre expression in expected neurons in both Cre lines. Interestingly, we also found that in Th-Cre rats, but not DAT-Cre rats, Cre is expressed in female germ cells, allowing germline excision of the floxed allele and hence the generation of whole-body knockout rats. In summary, our data demonstrate that targeted integration of Cre cassette lead to faithful recapitulation of expression pattern of the endogenous promoter, and mRNA FISH, in addition to IHC, is an effective method for the analysis of the spatiotemporal gene expression patterns in the rat brain, alleviating the dependence on high quality antibodies that are often not available against rat proteins. The Th-Cre and the DAT-Cre rat lines express Cre in selective subsets of dopaminergic neurons and should be particularly useful for researches on Parkinson's disease.

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

    Directory of Open Access Journals (Sweden)

    Zahra Maria

    Full Text Available 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.

  1. Post-Translational Regulation of the Glucose-6-Phosphatase Complex by Cyclic Adenosine Monophosphate Is a Crucial Determinant of Endogenous Glucose Production and Is Controlled by the Glucose-6-Phosphate Transporter.

    Science.gov (United States)

    Soty, Maud; Chilloux, Julien; Delalande, François; Zitoun, Carine; Bertile, Fabrice; Mithieux, Gilles; Gautier-Stein, Amandine

    2016-04-01

    The excessive endogenous glucose production (EGP) induced by glucagon participates in the development of type 2 diabetes. To further understand this hormonal control, we studied the short-term regulation by cyclic adenosine monophosphate (cAMP) of the glucose-6-phosphatase (G6Pase) enzyme, which catalyzes the last reaction of EGP. In gluconeogenic cell models, a 1-h treatment by the adenylate cyclase activator forskolin increased G6Pase activity and glucose production independently of any change in enzyme protein amount or G6P content. Using specific inhibitors or protein overexpression, we showed that the stimulation of G6Pase activity involved the protein kinase A (PKA). Results of site-directed mutagenesis, mass spectrometry analyses, and in vitro phosphorylation experiments suggested that the PKA stimulation of G6Pase activity did not depend on a direct phosphorylation of the enzyme. However, the temperature-dependent induction of both G6Pase activity and glucose release suggested a membrane-based mechanism. G6Pase is composed of a G6P transporter (G6PT) and a catalytic unit (G6PC). Surprisingly, we demonstrated that the increase in G6PT activity was required for the stimulation of G6Pase activity by forskolin. Our data demonstrate the existence of a post-translational mechanism that regulates G6Pase activity and reveal the key role of G6PT in the hormonal regulation of G6Pase activity and of EGP.

  2. High glucose decreases the expression of ATP-binding cassette transporter G1 in human vascular smooth muscle cells

    Institute of Scientific and Technical Information of China (English)

    Jiahong Xue; Zuyi Yuan; Yue Wu; Yan Zhao; Zhaofei Wan

    2008-01-01

    Objective:ATP-binding cassette transporters(ABC) A1 and G1 play an important role in mediating cholesterol efflux and preventing macrophage foam cell formation. In this study, we examined the regulation of ABC transporters by high glucose in human vascular smooth muscle cells(VSMCs), the other precursor of foam cells. Methods:Incubation of human VSMCs with D-ghicose(5 to 30 mM) for 1 to 7 days in the presence or absence of antioxidant and nuclear factor(NF)-kB inhibitors, the expressions of ABCA1 and ABCG1 were analyzed by real time PCR and Western blotting. Results:High glucose decreased ABCG1 mRNA and protein expression in cultured VSMCs, whereas the expression of ABCA1 was not significantly decreased. Down-regulation of ABCG1 mRNA expression by high glucose was abolished by antioxidant N-acetyl-L-cysteine(NAC) and NF-kB inhibitors, BAY 11-7085 and tosyl-phenylalanine chloromethyl-ketone(TPCK). Conclusion:High glucose suppresses the expression of ABCG1 in VSMCs, which is the possible mechanism of VSMC derived foam cell transformation.

  3. Cannabinoid receptor 1 signalling dampens activity and mitochondrial transport in networks of enteric neurones.

    Science.gov (United States)

    Boesmans, W; Ameloot, K; van den Abbeel, V; Tack, J; Vanden Berghe, P

    2009-09-01

    Cannabinoid (CB) receptors are expressed in the enteric nervous system (ENS) and CB(1) receptor activity slows down motility and delays gastric emptying. This receptor system has become an important target for GI-related drug development such as in obesity treatment. The aim of the study was to investigate how CB(1) ligands and antagonists affect ongoing activity in enteric neurone networks, modulate synaptic vesicle cycling and influence mitochondrial transport in nerve processes. Primary cultures of guinea-pig myenteric neurones were loaded with different fluorescent markers: Fluo-4 to measure network activity, FM1-43 to image synaptic vesicles and Mitotracker green to label mitochondria. Synaptic vesicle cluster density was assessed by immunohistochemistry and expression of CB(1) receptors was confirmed by RT-PCR. Spontaneous network activity, displayed by both excitatory and inhibitory neurones, was significantly increased by CB(1) receptor antagonists (AM-251 and SR141716), abolished by CB(1) activation (methanandamide, mAEA) and reduced by two different inhibitors (arachidonylamide serotonin, AA-5HT and URB597) of fatty acid amide hydrolase. Antagonists reduced the number of synaptic vesicles that were recycled during an electrical stimulus. CB(1) agonists (mAEA and WIN55,212) reduced and antagonists enhanced the fraction of transported mitochondria in enteric nerve fibres. We found immunohistochemical evidence for an enhancement of synaptophysin-positive release sites with SR141716, while WIN55,212 caused a reduction. The opposite effects of agonists and antagonists suggest that enteric nerve signalling is under the permanent control of CB(1) receptor activity. Using inhibitors of the endocannabinoid degrading enzyme, we were able to show there is endogenous production of a CB ligand in the ENS.

  4. Neuronal exosomal miRNA-dependent translational regulation of astroglial glutamate transporter GLT1.

    Science.gov (United States)

    Morel, Lydie; Regan, Melissa; Higashimori, Haruki; Ng, Seng Kah; Esau, Christine; Vidensky, Svetlana; Rothstein, Jeffrey; Yang, Yongjie

    2013-03-08

    Perisynaptic astrocytes express important glutamate transporters, especially excitatory amino acid transporter 2 (EAAT2, rodent analog GLT1) to regulate extracellular glutamate levels and modulate synaptic activation. In this study, we investigated an exciting new pathway, the exosome-mediated transfer of microRNA (in particular, miR-124a), in neuron-to-astrocyte signaling. Exosomes isolated from neuron-conditioned medium contain abundant microRNAs and small RNAs. These exosomes can be directly internalized into astrocytes and increase astrocyte miR-124a and GLT1 protein levels. Direct miR-124a transfection also significantly and selectively increases protein (but not mRNA) expression levels of GLT1 in cultured astrocytes. Consistent with our in vitro findings, intrastriatal injection of specific antisense against miR-124a into adult mice dramatically reduces GLT1 protein expression and glutamate uptake levels in striatum without reducing GLT1 mRNA levels. MiR-124a-mediated regulation of GLT1 expression appears to be indirect and is not mediated by its suppression of the putative GLT1 inhibitory ligand ephrinA3. Moreover, miR-124a is selectively reduced in the spinal cord tissue of end-stage SOD1 G93A mice, the mouse model of ALS. Subsequent exogenous delivery of miR-124a in vivo through stereotaxic injection significantly prevents further pathological loss of GLT1 proteins, as determined by GLT1 immunoreactivity in SOD1 G93A mice. Together, our study characterized a new neuron-to-astrocyte communication pathway and identified miRNAs that modulate GLT1 protein expression in astrocytes in vitro and in vivo.

  5. Neuronal Exosomal miRNA-dependent Translational Regulation of Astroglial Glutamate Transporter GLT1*

    Science.gov (United States)

    Morel, Lydie; Regan, Melissa; Higashimori, Haruki; Ng, Seng Kah; Esau, Christine; Vidensky, Svetlana; Rothstein, Jeffrey; Yang, Yongjie

    2013-01-01

    Perisynaptic astrocytes express important glutamate transporters, especially excitatory amino acid transporter 2 (EAAT2, rodent analog GLT1) to regulate extracellular glutamate levels and modulate synaptic activation. In this study, we investigated an exciting new pathway, the exosome-mediated transfer of microRNA (in particular, miR-124a), in neuron-to-astrocyte signaling. Exosomes isolated from neuron-conditioned medium contain abundant microRNAs and small RNAs. These exosomes can be directly internalized into astrocytes and increase astrocyte miR-124a and GLT1 protein levels. Direct miR-124a transfection also significantly and selectively increases protein (but not mRNA) expression levels of GLT1 in cultured astrocytes. Consistent with our in vitro findings, intrastriatal injection of specific antisense against miR-124a into adult mice dramatically reduces GLT1 protein expression and glutamate uptake levels in striatum without reducing GLT1 mRNA levels. MiR-124a-mediated regulation of GLT1 expression appears to be indirect and is not mediated by its suppression of the putative GLT1 inhibitory ligand ephrinA3. Moreover, miR-124a is selectively reduced in the spinal cord tissue of end-stage SOD1 G93A mice, the mouse model of ALS. Subsequent exogenous delivery of miR-124a in vivo through stereotaxic injection significantly prevents further pathological loss of GLT1 proteins, as determined by GLT1 immunoreactivity in SOD1 G93A mice. Together, our study characterized a new neuron-to-astrocyte communication pathway and identified miRNAs that modulate GLT1 protein expression in astrocytes in vitro and in vivo. PMID:23364798

  6. Combination of mild hypothermia with neuroprotectants has greater neuroprotective effects during oxygen-glucose deprivation and reoxygenation-mediated neuronal injury.

    Science.gov (United States)

    Gao, Xiao-Ya; Huang, Jian-Ou; Hu, Ya-Fang; Gu, Yong; Zhu, Shu-Zhen; Huang, Kai-Bin; Chen, Jin-Yu; Pan, Su-Yue

    2014-11-18

    Co-treatment of neuroprotective reagents may improve the therapeutic efficacy of hypothermia in protecting neurons during ischemic stroke. This study aimed to find promising drugs that enhance the neuroprotective effect of mild hypothermia (MH). 26 candidate drugs were selected based on different targets. Primary cultured cortical neurons were exposed to oxygen-glucose deprivation and reoxygenation (OGD/R) to induce neuronal damage, followed by either single treatment (a drug or MH) or a combination of a drug and MH. Results showed that, compared with single treatment, combination of MH with brain derived neurotrophic factor, glibenclamide, dizocilpine, human urinary kallidinogenase or neuroglobin displayed higher proportion of neuronal cell viability. The latter three drugs also caused less apoptosis rate in combined treatment. Furthermore, co-treatment of those three drugs and MH decreased the level of reactive oxygen species (ROS) and intracellular calcium accumulation, as well as stabilized mitochondrial membrane potential (MMP), indicating the combined neuroprotective effects are probably via inhibiting mitochondrial apoptosis pathway. Taken together, the study suggests that combined treatment with hypothermia and certain neuroprotective reagents provide a better protection against OGD/R-induced neuronal injury.

  7. Near infrared radiation protects against oxygen-glucose deprivation-induced neurotoxicity by down-regulating neuronal nitric oxide synthase (nNOS) activity in vitro.

    Science.gov (United States)

    Yu, Zhanyang; Li, Zhaoyu; Liu, Ning; Jizhang, Yunneng; McCarthy, Thomas J; Tedford, Clark E; Lo, Eng H; Wang, Xiaoying

    2015-06-01

    Near infrared radiation (NIR) has been shown to be neuroprotective against neurological diseases including stroke and brain trauma, but the underlying mechanisms remain poorly understood. In the current study we aimed to investigate the hypothesis that NIR may protect neurons by attenuating oxygen-glucose deprivation (OGD)-induced nitric oxide (NO) production and modulating cell survival/death signaling. Primary mouse cortical neurons were subjected to 4 h OGD and NIR was applied at 2 h reoxygenation. OGD significantly increased NO level in primary neurons compared to normal control, which was significantly ameliorated by NIR at 5 and 30 min post-NIR. Neither OGD nor NIR significantly changed neuronal nitric oxide synthase (nNOS) mRNA or total protein levels compared to control groups. However, OGD significantly increased nNOS activity compared to normal control, and this effect was significantly diminished by NIR. Moreover, NIR significantly ameliorated the neuronal death induced by S-Nitroso-N-acetyl-DL-penicillamine (SNAP), a NO donor. Finally, NIR significantly rescued OGD-induced suppression of p-Akt and Bcl-2 expression, and attenuated OGD-induced upregulation of Bax, BAD and caspase-3 activation. These results suggest NIR may protect against OGD at least partially through reducing NO production by down-regulating nNOS activity, and modulating cell survival/death signaling.

  8. Activated microglia induce bone marrow mesenchymal stem cells to produce glial cell-derived neurotrophic factor and protect neurons against oxygen-glucose deprivation injury

    Directory of Open Access Journals (Sweden)

    Bingke Lv

    2016-12-01

    Full Text Available In this study, we investigated interactions among microglia (MG, bone marrow mesenchymal stem cells (BMSCs and neurons in cerebral ischemia and the potential mechanisms using an in vitro oxygen-glucose deprivation (OGD model. Rat BMSCs were incubated with conditioned medium (CM from in vitro cultures of OGD-activated rat MG and murine BV2 MG cells. Effects of glial cell-derived neurotrophic factor (GDNF on rat neuron viability, apoptosis, lactate dehydrogenase (LDH leakage and mitochondrial membrane potential (MMP were analyzed in this model. OGD-activated MG promoted GDNF production by BMSCs (P < 0.01. TNFα, but not IL6 or IL1β, promoted GDNF production by BMSCs (P < 0.001. GDNF or CM pre-treated BMSCs elevated neuronal viability and suppressed apoptosis (P < 0.05 or P < 0.01; these effects were inhibited by the RET antibody. GDNF activated MEK/ERK and PI3K/AKT signaling but not JNK/c-JUN. Furthermore, GDNF upregulated B cell lymphoma 2 (BCL2 and heat shock 60 kDa protein 1 (HSP60 levels, suppressed LDH leakage, and promoted MMP. Thus, activated MG produce TNFα to stimulate GDNF production by BMSCs, which prevents and repairs OGD-induced neuronal injury, possibly via regulating MEK/ERK and PI3K/AKT signaling. These findings will facilitate the prevention and treatment of neuronal injury by cerebral ischemia.

  9. Benefits and Harms of Sodium-Glucose Co-Transporter 2 Inhibitors in Patients with Type 2 Diabetes

    DEFF Research Database (Denmark)

    Storgaard, Heidi; Gluud, Lise L; Bennett, Cathy

    2016-01-01

    OBJECTIVE: Sodium-glucose co-transporter 2 inhibitors (SGLT2-i) are a novel drug class for the treatment of diabetes. We aimed at describing the maximal benefits and risks associated with SGLT2-i for patients with type 2 diabetes. DESIGN: Systematic review and meta-analysis. DATA SOURCES AND STUDY...... increased risk in non-serious adverse events. The analyses may overestimate the intervention benefit due bias....

  10. The role of GluN2A and GluN2B NMDA receptor subunits in AgRP and POMC neurons on body weight and glucose homeostasis

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    Aykut Üner

    2015-10-01

    Conclusions: GluN2B-containing NMDA receptors in AgRP neurons play a critical role in central control of body weight homeostasis and blood glucose balance via mechanisms that likely involve regulation of AgRP neuronal survival and structure, and modulation of hypothalamic leptin action.

  11. Expression Patterns of Glucose Transporter-1 Gene and Thyroid Specific Genes in Human Papillary Thyroid Carcinoma

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Sungeun; Chung, Junekey; Min Haesook and others

    2014-06-15

    The expression of glucose transporter-1 (Glut-1) gene and those of major thyroid-specific genes were examined in papillary carcinoma tissues, and the expressions of these genes were compared with cancer differentiation grades. Twenty-four human papillary carcinoma tissues were included in this study. The expressions of Glut-1- and thyroid-specific genes [sodium/iodide symporter (NIS), thyroid peroxidase, thyroglobulin, TSH receptor and pendrin] were analyzed by RT-PCR. Expression levels were expressed as ratios versus the expression of beta-actin. Pathologic differentiation of papillary carcinoma was classified into a relatively well-differentiated group (n=13) and relatively less differentiated group (n=11). Glut-1 gene expression was significantly higher in the less differentiated group (0.66±0.04) than in the well-differentiated group (0.59±0.07). The expression levels of the NIS, PD and TG genes were significantly higher in the well-differentiated group (NIS: 0.67±0.20, PD: 0.65±0.21, TG: 0.74±0.16) than in the less differentiated group (NIS: 0.36±0.05, PD: 0.49±0.08, TG: 0.60±0.11), respectively. A significant negative correlation was found between Glut-1 and NIS expression, and positive correlations were found between NIS and TG, and between NIS and PD. The NIS, PD and TG genes were highly expressed in well-differentiated thyroid carcinomas, whereas the Glut-1 gene was highly expressed in less differentiated thyroid carcinomas. These findings provide a molecular rationale for the management of papillary carcinoma, especially in the selection of FDG PET or radioiodine whole-body scan and I-131-based therapy.

  12. Expression of glucose transporter 4 in endometrium of polycystic ovary syndrome

    Institute of Scientific and Technical Information of China (English)

    Wang Wei; Li Xiao-dong; Hao Gui-min; Xu Su-xin; Cui Na; Cao Jin-feng

    2008-01-01

    Objective:The aim of this study was to detect the expression of glucose transporter 4(GLUT4),insulin(INS)and insulin receptor(INS-R)genes and proteins in endometrium of patients with polycystic ovarian syndrome(P-COS)and control group and investigate the relationship between GLUT4 and insulin resistance(IR).Methods:The patients with PCOS were divided into two groups:hypertestosteronemia(h-T)group and hyperin-sulinemia(h-I)group.The expression of GLUT4,INS and INS-R genes and proteins in endometrium of PCOS groups and control group was detected by immunohistochemistry and reverse transcription-polymerase chain reaction.Results:There was the expression of GLUT4,INS and INS-R proteins and genes in endometrium of PCOS groups and control group.Compared with the control group,PCOS groups were significantly lower in the expres-sion of GLUT4 protein and gene as well as INS-R protein and gene but much stronger in the expression of INS pro-tein and gene.The GLUT4 gene expression in h-I group was lower than that of h-T group but the expression of INS protein in h-I group was siginificantly higher than that of h-T group.Conclusion:The expression of GLUT4,INS and INS-R in endometrium of PCOS and control groups is con-firmed.Hyperinsulinemia and hyperandrogen can cause the expression of GLUT4 to decrease.The decreased ex-pression of GLUT4 in PCOS may be related with the IR of endometrium.

  13. Quantitative PCR for glucose transporter and tristetraprolin family gene expression in cultured mouse adipocytes and macrophages.

    Science.gov (United States)

    Cao, Heping; Cao, Fangping; Roussel, Anne-Marie; Anderson, Richard A

    2013-12-01

    Quantitative real-time PCR (qPCR) such as TaqMan and SYBR Green qPCR are widely used for gene expression analysis. The drawbacks of SYBR Green assay are that the dye binds to any double-stranded DNA which can generate false-positive signals and that the length of the amplicon affects the intensity of the amplification. Previous results demonstrate that TaqMan assay is more sensitive but generates lower calculated expression levels than SYBR Green assay in quantifying seven mRNAs in tung tree tissues. The objective of this study is to expand the analysis using animal cells. We compared both qPCR assays for quantifying 24 mRNAs including those coding for glucose transporter (Glut) and mRNA-binding protein tristetraprolin (TTP) in mouse 3T3-L1 adipocytes and RAW264.7 macrophages. The results showed that SYBR Green and TaqMan qPCR were reliable for quantitative gene expression in animal cells. This result was supported by validation analysis of Glut and TTP family gene expression. However, SYBR Green qPCR overestimated the expression levels in most of the genes tested. Finally, both qPCR instruments (Bio-Rad's CFX96 real-time system and Applied Biosystems' Prism 7700 real-time PCR instrument) generated similar gene expression profiles in the mouse cells. These results support the conclusion that both qPCR assays (TaqMan and SYBR Green qPCR) and both qPCR instruments (Bio-Rad's CFX96 real-time system and Applied Biosystems' Prism 7700 real-time PCR instrument) are reliable for quantitative gene expression analyses in animal cells but SYBR Green qPCR generally overestimates gene expression levels than TaqMan qPCR.

  14. Maltose and maltodextrin utilization by Listeria monocytogenes depend on an inducible ABC transporter which is repressed by glucose.

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    Shubha Gopal

    Full Text Available BACKGROUND: In the environment as well as in the vertebrate intestine, Listeriae have access to complex carbohydrates like maltodextrins. Bacterial exploitation of such compounds requires specific uptake and utilization systems. METHODOLOGY/PRINCIPAL FINDINGS: We could show that Listeria monocytogenes and other Listeria species contain genes/gene products with high homology to the maltodextrin ABC transporter and utilization system of B. subtilis. Mutant construction and growth tests revealed that the L. monocytogenes gene cluster was required for the efficient utilization of maltodextrins as well as maltose. The gene for the ATP binding protein of the transporter was located distant from the cluster. Transcription analyses demonstrated that the system was induced by maltose/maltodextrins and repressed by glucose. Its induction was dependent on a LacI type transcriptional regulator. Repression by glucose was independent of the catabolite control protein CcpA, but was relieved in a mutant defective for Hpr kinase/phosphorylase. CONCLUSIONS/SIGNIFICANCE: The data obtained show that in L. monocytogenes the uptake of maltodextrin and, in contrast to B. subtilis, also maltose is exclusively mediated by an ABC transporter. Furthermore, the results suggest that glucose repression of the uptake system possibly is by inducer exclusion, a mechanism not described so far in this organism.

  15. A three-year-old boy with glucose transporter type 1 deficiency syndrome presenting with episodic ataxia.

    Science.gov (United States)

    Ohshiro-Sasaki, Akiko; Shimbo, Hiroko; Takano, Kyoko; Wada, Takahito; Osaka, Hitoshi

    2014-01-01

    Glucose transporter type 1 deficiency syndrome is a metabolic encephalopathy that results from impaired glucose transport into the brain as the result of a mutation of the SLC2A1 gene. It has been recognized recently that these patients can present with a much broader clinical spectrum than previously thought. We describe a 3-year-old boy presenting with episodic ataxia. Our patient exhibited periodic abnormal eye movements, including opsoclonus, since he was 4 months of age. At 2 years of age, he experienced acute cerebellar ataxia after a vaccination. Since then, he has had periodic attacks of ataxic gait, repeated vomiting, and abnormal eye movement. He was diagnosed as having episodic ataxia type 2 because the administration of acetazolamide seemed effective. By 3 years and 10 months of age, he exhibited mild mental retardation and mild trunk ataxia. The attacks were more likely to occur when he was hungry. Molecular analysis revealed that the SLC2A1 gene had a de novo mutation of heterozygous seven nucleotide insertion within exon 7, resulting in a frameshift. He has recently begun a modified Atkins diet; the frequency of attacks has been reduced, and his psychomotor and language skills have begun to develop. Glucose transporter type 1 deficiency syndrome should be considered in the differential diagnosis in children with episodic ataxia, even if acetazolamide is effective. Copyright © 2014 Elsevier Inc. All rights reserved.

  16. Botulinum neurotoxins A and E undergo retrograde axonal transport in primary motor neurons.

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    Laura Restani

    2012-12-01

    Full Text Available The striking differences between the clinical symptoms of tetanus and botulism have been ascribed to the different fate of the parental neurotoxins once internalised in motor neurons. Tetanus toxin (TeNT is known to undergo transcytosis into inhibitory interneurons and block the release of inhibitory neurotransmitters in the spinal cord, causing a spastic paralysis. In contrast, botulinum neurotoxins (BoNTs block acetylcholine release at the neuromuscular junction, therefore inducing a flaccid paralysis. Whilst overt experimental evidence supports the sorting of TeNT to the axonal retrograde transport pathway, recent findings challenge the established view that BoNT trafficking is restricted to the neuromuscular junction by highlighting central effects caused by these neurotoxins. These results suggest a more complex scenario whereby BoNTs also engage long-range trafficking mechanisms. However, the intracellular pathways underlying this process remain unclear. We sought to fill this gap by using primary motor neurons either in mass culture or differentiated in microfluidic devices to directly monitor the endocytosis and axonal transport of full length BoNT/A and BoNT/E and their recombinant binding fragments. We show that BoNT/A and BoNT/E are internalised by spinal cord motor neurons and undergo fast axonal retrograde transport. BoNT/A and BoNT/E are internalised in non-acidic axonal carriers that partially overlap with those containing TeNT, following a process that is largely independent of stimulated synaptic vesicle endo-exocytosis. Following intramuscular injection in vivo, BoNT/A and TeNT displayed central effects with a similar time course. Central actions paralleled the peripheral spastic paralysis for TeNT, but lagged behind the onset of flaccid paralysis for BoNT/A. These results suggest that the fast axonal retrograde transport compartment is composed of multifunctional trafficking organelles orchestrating the simultaneous transfer

  17. Decreased [{sup 18}F]fluoro-2-deoxy-D-glucose incorporation and increased glucose transport are associated with resistance to 5FU in MCF7 cells in vitro

    Energy Technology Data Exchange (ETDEWEB)

    Smith, Tim A.D. [PET Unit, Department of Biomedical Physics, University of Aberdeen, Foresterhill, Aberdeen AB25 2ZD (United Kingdom)], E-mail: t.smith@biomed.abdn.ac.uk; Sharma, Rituka I. [PET Unit, Department of Biomedical Physics, University of Aberdeen, Foresterhill, Aberdeen AB25 2ZD (United Kingdom); Wang, Weiguang G. [Department of Medicine and Therapeutics, University of Aberdeen, Foresterhill, Aberdeen AB25 2ZD (United Kingdom); School of Applied Sciences, University of Wolverhampton, City Campus-South, Wolverhampton WV1 1SB (United Kingdom); Welch, Andy E.; Schweiger, Lutz F. [PET Unit, Department of Biomedical Physics, University of Aberdeen, Foresterhill, Aberdeen AB25 2ZD (United Kingdom); Collie-Duguid, Elaina S.R. [Department of Medicine and Therapeutics, University of Aberdeen, Foresterhill, Aberdeen AB25 2ZD (United Kingdom)

    2007-11-15

    Introduction: Tumor refractoriness to chemotherapy is frequently due to the acquisition of resistance. Resistant cells selected by exposure to chemotherapy agents may exhibit differences in [{sup 18}F]fluoro-2-deoxy-D-glucose (FDG) incorporation, as compared with sensitive cells. Methods: FDG incorporation, hexokinase (HK) activity, glucose transport and ATP content were determined in clones of 5-fluorouracil (5FU)-resistant MCF7 cells, established by long-term exposure to increasing 5FU concentrations, and in parental MCF7 cells. Results: FDG incorporation was decreased in MCF7 cells resistant to 5FU; HK activity was similar in the resistant and sensitive cells, while glucose transport was increased, as compared with sensitive cells. Treatment of cells with the glucose efflux inhibitor phloretin increased FDG incorporation to similar levels in the resistant and sensitive cells. Analysis of microarray data demonstrated the expression of GLUT1, 8 and 10 transporters in MCF7 cells. GLUT8 and 10 expression was decreased in the resistant cells, while GLUT1 was only increased in cells resistant to the lowest 5FU concentration. Conclusion: FDG incorporation in 5FU-resistant MCF7 cells is decreased, as compared with sensitive cells. Our findings also suggest that this may be due to high rates of membrane glucose transport in the resistant cells resulting in enhanced efflux of FDG. We believe that this is the first demonstration that facilitative glucose transporters can actually decrease the incorporation of FDG.

  18. Application of the Co-culture Membrane System Pointed to a Protective Role of Catestatin on Hippocampal Plus Hypothalamic Neurons Exposed to Oxygen and Glucose Deprivation.

    Science.gov (United States)

    Mele, Maria; Morelli, Sabrina; Fazzari, Gilda; Avolio, Ennio; Alò, Raffaella; Piscioneri, Antonella; De Bartolo, Loredana; Facciolo, Rosa Maria; Canonaco, Marcello

    2016-11-05

    Depletion of oxygen and glucose even for brief periods is sufficient to cause cerebral ischemia, which is a predominant worldwide cause of motor deficits with the reduction of life quality and subsequently death. Hence, more insights regarding protective measures against ischemic events are becoming a major research goal. Among the many neuronal factors, N-methyl-D-aspartate receptors (NMDAR), orexinergic neuroreceptors (ORXR), and sympatho-inhibitory neuropeptide catestatin (CST) are widely involved with ischemic episodes. In this study, it was possible to induce in vitro ischemic conditions of the hamster (Mesocricetus auratus) hippocampal and hypothalamic neuronal cultures, grown on a newly compartmentalized membrane system, via oxygen and glucose deprivation (OGD). These cultures displayed notably differentiated NMDARergic and ORXergic receptor expression activities along with evident brain-derived neurotrophic factor (BDNF) plus orexin A (ORX-A) secretion, especially under co-cultured conditions. Interestingly, addition of CST in OGD-insulted hippocampal cells accounted for upregulated GluN1 and ORX1R transcripts that in the case of the latter neuroreceptor was very strongly (p BDNF and ORX-A secretion in the presence of hippocampal cells. Overall, the preferential CST effects on BDNF plus ORX-A production together with altered NMDAR and ORXR levels, especially in co-cultured hypothalamic cells pointed to ORX-containing neurons as major protective constituents against ischemic damages thus opening new scenarios on the cross-talking roles of CST during ischemic disorders.

  19. α-Tubulin Tyrosination and CLIP-170 Phosphorylation Regulate the Initiation of Dynein-Driven Transport in Neurons

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    Jeffrey J. Nirschl

    2016-03-01

    Full Text Available Motor-cargo recruitment to microtubules is often the rate-limiting step of intracellular transport, and defects in this recruitment can cause neurodegenerative disease. Here, we use in vitro reconstitution assays with single-molecule resolution, live-cell transport assays in primary neurons, computational image analysis, and computer simulations to investigate the factors regulating retrograde transport initiation in the distal axon. We find that phosphorylation of the cytoskeletal-organelle linker protein CLIP-170 and post-translational modifications of the microtubule track combine to precisely control the initiation of retrograde transport. Computer simulations of organelle dynamics in the distal axon indicate that while CLIP-170 primarily regulates the time to microtubule encounter, the tyrosination state of the microtubule lattice regulates the likelihood of binding. These mechanisms interact to control transport initiation in the axon in a manner sensitive to the specialized cytoskeletal architecture of the neuron.

  20. In vivo neuronal synthesis and axonal transport of Kunitz protease inhibitor (KPI)-containing forms of the amyloid precursor protein.

    Science.gov (United States)

    Moya, K L; Confaloni, A M; Allinquant, B

    1994-11-01

    We have shown previously that the amyloid precursor protein (APP) is synthesized in retinal ganglion cells and is rapidly transported down the axons, and that different molecular weight forms of the precursor have different developmental time courses. Some APP isoforms contain a Kunitz protease inhibitor (KPI) domain, and APP that lacks the KPI domain is considered the predominant isoform in neurons. We now show that, among the various rapidly transported APPs, a 140-kDa isoform contains the KPI domain. This APP isoform is highly expressed in rapidly growing retinal axons, and it is also prominent in adult axon endings. This 140-kDa KPI-containing APP is highly sulfated compared with other axonally transported isoforms. These results show that APP with the KPI domain is a prominent isoform synthesized in neurons in vivo, and they suggest that the regulation of protease activity may be an important factor during the establishment of neuronal connections.

  1. BDNF regulates the expression and distribution of vesicular glutamate transporters in cultured hippocampal neurons.

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    Carlos V Melo

    Full Text Available BDNF is a pro-survival protein involved in neuronal development and synaptic plasticity. BDNF strengthens excitatory synapses and contributes to LTP, presynaptically, through enhancement of glutamate release, and postsynaptically, via phosphorylation of neurotransmitter receptors, modulation of receptor traffic and activation of the translation machinery. We examined whether BDNF upregulated vesicular glutamate receptor (VGLUT 1 and 2 expression, which would partly account for the increased glutamate release in LTP. Cultured rat hippocampal neurons were incubated with 100 ng/ml BDNF, for different periods of time, and VGLUT gene and protein expression were assessed by real-time PCR and immunoblotting, respectively. At DIV7, exogenous application of BDNF rapidly increased VGLUT2 mRNA and protein levels, in a dose-dependent manner. VGLUT1 expression also increased but only transiently. However, at DIV14, BDNF stably increased VGLUT1 expression, whilst VGLUT2 levels remained low. Transcription inhibition with actinomycin-D or α-amanitine, and translation inhibition with emetine or anisomycin, fully blocked BDNF-induced VGLUT upregulation. Fluorescence microscopy imaging showed that BDNF stimulation upregulates the number, integrated density and intensity of VGLUT1 and VGLUT2 puncta in neurites of cultured hippocampal neurons (DIV7, indicating that the neurotrophin also affects the subcellular distribution of the transporter in developing neurons. Increased VGLUT1 somatic signals were also found 3 h after stimulation with BDNF, further suggesting an increased de novo transcription and translation. BDNF regulation of VGLUT expression was specifically mediated by BDNF, as no effect was found upon application of IGF-1 or bFGF, which activate other receptor tyrosine kinases. Moreover, inhibition of TrkB receptors with K252a and PLCγ signaling with U-73122 precluded BDNF-induced VGLUT upregulation. Hippocampal neurons express both isoforms during

  2. Flozins, inhibitors of type 2 renal sodium-glucose co-transporter – not only antihyperglycemic drugs

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    Mizerski Grzegorz

    2015-09-01

    Full Text Available The kidneys play a crucial role in the regulation of the carbohydrate metabolism. In normal physiological conditions, the glucose that filters through the renal glomeruli is subsequently nearly totally reabsorbed in the proximal renal tubules. Two transporters are engaged in this process: sodium-glucose co-transporter type 1 (SGLT1, and sodium-glucose co-transporter type type 2 (SGLT2 - this being located in the luminal membrane of the renal tubular epithelial cells. It was found that the administration of dapagliflozin, a selective SGLT2 inhibitor, in patients with type 2 diabetes, is associated with the reduction of HbA1c concentration by 0.45-1.11%. Additional benefits from the treatment with dapagliflozin are the reduction of arterial blood pressure and a permanent reduction of body weight. This outcome is related to the effect of osmotic diuresis and to the considerable loss of the glucose load by way of urine excretion. Dapagliflozin may be successfully applied in type 2 diabetes monotherapy, as well as in combined therapy (including insulin, where it is equally effective as other oral anti-diabetic drugs. Of note: serious adverse effects of dapagliflozin administration are rarely observed. What is more, episodes of severe hypoglycaemia related with the treatment occur only sporadically, most often in the course of diabetes polytherapy. The most frequent effects of the SGLT2 inhibitors are inseparably associated with the mechanism of their action (the glucuretic effect, and cover urogenital infections with a mild clinical course. At present, clinical trials are being continued of the administration of several subsequent drugs from this group, the most advanced of these being the use of canagliflozin and empagliflozin.

  3. Skeletal muscle sodium glucose co-transporters in older adults with type 2 diabetes undergoing resistance training

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    Francisco Castaneda, Jennifer E. Layne, Carmen Castaneda

    2006-01-01

    Full Text Available We examined the expression of the sodium-dependent glucose co-transporter system (hSGLT3 in skeletal muscle of Hispanic older adults with type 2 diabetes. Subjects (65±8 yr were randomized to resistance training (3x/wk, n=13 or standard of care (controls, n=5 for 16 weeks. Skeletal muscle hSGLT3 and GLUT4 mRNA transcript levels were determined by real time RT-PCR. hSGLT3 transcripts increased by a factor of ten following resistance training compared to control subjects (0.10, P=0.03. There were no differences in GLUT4 mRNA expression levels between groups. Protein expression levels of these transporters were confirmed by immunohistochemistry and Western blotting. hSGLT3 after resistance exercise was found not to be co-localized with the nicotinic acetylcholine receptor. The change in hSGLT3 transcript levels in the vastus lateralis muscle was positively correlated with glucose uptake, as measured by the change in muscle glycogen stores (r=0.53, P=0.02; and with exercise intensity, as measured by the change in muscle strength (r=0.73, P=0.001. Group assignment was be the only independent predictor of hSGLT3 transcript levels, explaining 68% of its variability (P=0.01. Our data show that hSGLT3, but not GLTU4, expression was enhanced in skeletal muscle after 16 weeks of resistance training. This finding suggests that hSGLT3, an insulin-independent glucose transporter, is activated with exercise and it may play a significant role in glycemic control with muscle contraction. The hSGLT3 exact mechanism is not well understood and requires further investigation. However its functional significance regarding a reduction of glucose toxicity and improvement of insulin resistance is the subject of ongoing research.

  4. Empagliflozin: a new sodium-glucose co-transporter 2 (SGLT2 inhibitor for the treatment of type 2 diabetes

    Directory of Open Access Journals (Sweden)

    Joshua J Neumiller

    2014-06-01

    Full Text Available Type 2 diabetes is increasing in prevalence worldwide, and hyperglycemia is often poorly controlled despite a number of therapeutic options. Unlike previously available agents, sodium-glucose co-transporter 2 (SGLT2 inhibitors offer an insulin-independent mechanism for improving blood glucose levels, since they promote urinary glucose excretion (UGE by inhibiting glucose reabsorption in the kidney. In addition to glucose control, SGLT2 inhibitors are associated with weight loss and blood pressure reductions, and do not increase the risk of hypoglycemia. Empagliflozin is a selective inhibitor of SGLT2, providing dose-dependent UGE increases in healthy volunteers, with up to 90 g of glucose excreted per day. It can be administered orally, and studies of people with renal or hepatic impairment indicated empagliflozin needed no dose adjustment based on pharmacokinetics. In Phase II trials in patients with type 2 diabetes, empagliflozin provided improvements in glycosylated hemoglobin (HbA1c and other measures of glycemic control when given as monotherapy or add-on to metformin, as well as reductions in weight and systolic blood pressure. As add-on to basal insulin, empagliflozin not only improved HbA1c levels but also reduced insulin doses. Across studies, empagliflozin was generally well tolerated with a similar rate of hypoglycemia to placebo; however, patients had a slightly increased frequency of genital infections, but not urinary tract infections, versus placebo. Phase III studies have also reported a good safety profile along with significant improvements in HbA1c, weight and blood pressure, with no increased risk of hypoglycemia versus placebo. Based on available data, it appears that empagliflozin may be a useful option in a range of patients; however, clinical decisions will be better informed by the results of ongoing studies, in particular, a large cardiovascular outcome study (EMPA-REG OUTCOME™.

  5. Dynamic transport and localization of alpha-synuclein in primary hippocampal neurons

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    Woods Wendy S

    2010-02-01

    Full Text Available Abstract Background Alpha-synuclein is a presynaptic protein with a proposed role in neurotransmission and dopamine homeostasis. Abnormal accumulation of α-synuclein aggregates in dopaminergic neurons of the substantia nigra is diagnostic of sporadic Parkinson's disease, and mutations in the protein are linked to early onset forms of the disease. The folded conformation of the protein varies depending upon its environment and other factors that are poorly understood. When bound to phospholipid membranes, α-synuclein adopts a helical conformation that mediates specific interactions with other proteins. Results To investigate the role of the helical domain in transport and localization of α-synuclein, eGFP-tagged constructs were transfected into rat primary hippocampal neurons at 7 DIV. A series of constructs were analyzed in which each individual exon was deleted, for comparison to previous studies of lipid affinity and α-helix content. A53T and A30P substitutions, representing Parkinson's disease-associated variants, were analyzed as well. Single exon deletions within the lipid-binding N-terminal domain of α-synuclein (exons 2, 3, and 4 partially disrupted its presynaptic localization at 17-21 DIV, resulting in increased diffuse labeling of axons. Similar results were obtained for A30P, which exhibits decreased lipid binding, but not A53T. To examine whether differences in presynaptic enrichment were related to deficiencies in transport velocity, transport was visualized via live cell microscopy. Tagged α-synuclein migrated at a rate of 1.85 ± 0.09 μm/s, consistent with previous reports, and single exon deletion mutants migrated at similar rates, as did A30P. Deletion of the entire N-terminal lipid-binding domain (Δ234GFP did not significantly alter rates of particle movement, but decreased the number of moving particles. Only the A53TGFP mutant exhibited a significant decrease in transport velocity as compared to ASGFP. Conclusions

  6. JIP3 regulates neuronal radial migration by mediating TrkB axonal anterograde transport in the developing cerebral cortex.

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    Ma, Huixian; Yu, Hui; Li, Ting; Zhao, Yan; Hou, Ming; Chen, Zheyu; Wang, Yue; Sun, Tao

    2017-04-15

    Radial migration is essential for the precise lamination and the coordinated function of the cerebral cortex. However, the molecular mechanisms for neuronal radial migration are not clear. Here, we report that c-Jun NH2-terminal kinase (JNK)-interacting protein-3 (JIP3) is highly expressed in the brain of embryonic mice and essential for radial migration. Knocking down JIP3 by in utero electroporation specifically perturbs the radial migration of cortical neurons but has no effect on neurogenesis and neuronal differentiation. Furthermore, we illustrate that JIP3 knockdown delays but does not block the migration of cortical neurons by investigating the distribution of neurons with JIP3 knocked down in the embryo and postnatal mouse. Finally, we find that JIP3 regulates cortical neuronal migration by mediating TrkB axonal anterograde transport during brain development. These findings deepen our understanding of the regulation of neuronal development by JIP3 and provide us a novel view on the regulating mechanisms of neuronal radial migration. Copyright © 2017 Elsevier Inc. All rights reserved.

  7. Comparative Study of the Effect of Baicalin and Its Natural Analogs on Neurons with Oxygen and Glucose Deprivation Involving Innate Immune Reaction of TLR2/TNFα

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    Hui-Ying Li

    2012-01-01

    Full Text Available This work is to study the baicalin and its three analogs, baicalin, wogonoside, and wogonin, on the protective effect of neuron from oxygen-glucose deprivation (OGD and toll-like receptor 2 (TLR2 expression in OGD damage. The results showed that baicalin and its three analogs did protect neurons from OGD damage and downregulated protein level of TLR2. D-Glucopyranosiduronic acid on site 7 in the structure played a core of cytotoxicity of these flavonoid analogs. The methoxyl group on carbon 8 of the structure had the relation with TLR2 protein expression, as well as the anti-inflammation. In addition, we detected caspase3 and antioxidation capability, to investigate the effect of four analogs on cell apoptosis and total antioxidation competence in OGD model.

  8. Live imaging of mitochondrial dynamics in CNS dopaminergic neurons in vivo demonstrates early reversal of mitochondrial transport following MPP(+) exposure.

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    Dukes, April A; Bai, Qing; Van Laar, Victor S; Zhou, Yangzhong; Ilin, Vladimir; David, Christopher N; Agim, Zeynep S; Bonkowsky, Joshua L; Cannon, Jason R; Watkins, Simon C; Croix, Claudette M St; Burton, Edward A; Berman, Sarah B

    2016-11-01

    Extensive convergent evidence collectively suggests that mitochondrial dysfunction is central to the pathogenesis of Parkinson's disease (PD). Recently, changes in the dynamic properties of mitochondria have been increasingly implicated as a key proximate mechanism underlying neurodegeneration. However, studies have been limited by the lack of a model in which mitochondria can be imaged directly and dynamically in dopaminergic neurons of the intact vertebrate CNS. We generated transgenic zebrafish in which mitochondria of dopaminergic neurons are labeled with a fluorescent reporter, and optimized methods allowing direct intravital imaging of CNS dopaminergic axons and measurement of mitochondrial transport in vivo. The proportion of mitochondria undergoing axonal transport in dopaminergic neurons decreased overall during development between 2days post-fertilization (dpf) and 5dpf, at which point the major period of growth and synaptogenesis of the relevant axonal projections is complete. Exposure to 0.5-1.0mM MPP(+) between 4 and 5dpf did not compromise zebrafish viability or cause detectable changes in the number or morphology of dopaminergic neurons, motor function or monoaminergic neurochemistry. However, 0.5mM MPP(+) caused a 300% increase in retrograde mitochondrial transport and a 30% decrease in anterograde transport. In contrast, exposure to higher concentrations of MPP(+) caused an overall reduction in mitochondrial transport. This is the first time mitochondrial transport has been observed directly in CNS dopaminergic neurons of a living vertebrate and quantified in a PD model in vivo. Our findings are compatible with a model in which damage at presynaptic dopaminergic terminals causes an early compensatory increase in retrograde transport of compromised mitochondria for degradation in the cell body. These data are important because manipulation of early pathogenic mechanisms might be a valid therapeutic approach to PD. The novel transgenic lines and

  9. A synthetic peptide shows retro- and anterograde neuronal transport before disrupting the chemosensation of plant-pathogenic nematodes.

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    Dong Wang

    Full Text Available Cyst nematodes are a group of plant pathogens each with a defined host range that cause major losses to crops including potato, soybean and sugar beet. The infective mobile stage hatches from dormant eggs and moves a short distance through the soil to plant roots, which it then invades. A novel strategy for control has recently been proposed in which the plant is able to secrete a peptide which disorientates the infective stage and prevents invasion of the pathogen. This study provides indirect evidence to support the mechanism by which one such peptide disrupts chemosensory function in nematodes. The peptide is a disulphide-constrained 7-mer with the amino acid sequence CTTMHPRLC that binds to nicotinic acetylcholine receptors. A fluorescently tagged version of this peptide with both epifluorescent and confocal microscopy was used to demonstrate that retrograde transport occurs from an aqueous environment along bare-ending primary cilia of chemoreceptive sensilla. The peptide is transported to the cell bodies of these neurons and on to a limited number of other neurons to which they connect. It appears to be localised in both neuronal processes and organelles adjacent to nuclei of some neurons suggesting it could be transported through the Golgi apparatus. The peptide takes 2.5 h to reach the neuronal cell bodies. Comparative studies established that similar but less abundant uptake occurs for Caenorhabditis elegans along its well studied dye-filling chemoreceptive neurons. Incubation in peptide solution or root-exudate from transgenic plants that secrete the peptide disrupted normal orientation of infective cyst nematodes to host root diffusate. The peptide probably undergoes transport along the dye-filling non-cholinergic chemoreceptive neurons to their synapses where it is taken up by the interneurons to which they connect. Coordinated responses to chemoreception are disrupted when the sub-set of cholinergic interneurons secrete the peptide

  10. A synthetic peptide shows retro- and anterograde neuronal transport before disrupting the chemosensation of plant-pathogenic nematodes.

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    Wang, Dong; Jones, Laura M; Urwin, Peter E; Atkinson, Howard J

    2011-03-07

    Cyst nematodes are a group of plant pathogens each with a defined host range that cause major losses to crops including potato, soybean and sugar beet. The infective mobile stage hatches from dormant eggs and moves a short distance through the soil to plant roots, which it then invades. A novel strategy for control has recently been proposed in which the plant is able to secrete a peptide which disorientates the infective stage and prevents invasion of the pathog