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Equilibrative Nucleoside Transporter (equilibrative + nucleoside_transporter)
Selected AbstractsENT1 Regulates Ethanol-Sensitive EAAT2 Expression and Function in AstrocytesALCOHOLISM, Issue 6 2010Jinhua Wu Background:, Equilibrative nucleoside transporter 1 (ENT1) and excitatory amino acid transporter 2 (EAAT2) are predominantly expressed in astrocytes where they are thought to regulate synaptic adenosine and glutamate levels. Because mice lacking ENT1 display increased glutamate levels in the ventral striatum, we investigated whether ENT1 regulates the expression and function of EAAT2 in astrocytes, which could contribute to altered glutamate levels in the striatum. Methods:, We examined the effect of ENT1 inhibition and overexpression on the expression of EAAT2 using quantitative real-time PCR and measured glutamate uptake activity in cultured astrocytes. We also examined the effect of 0 to 200 mM ethanol doses for 0 to 24 hours of ethanol exposure on EAAT2 expression and glutamate uptake activity. We further examined the effect of ENT1 knockdown by a specific siRNA on ethanol-induced EAAT2 expression. Results:, An ENT1-specific antagonist and siRNA treatments significantly reduced both EAAT2 expression and glutamate uptake activity while ENT1 overexpression up-regulated EAAT2 mRNA expression. Interestingly, 100 or 200 mM ethanol exposure increased EAAT2 mRNA expression as well as glutamate uptake activity. Moreover, we found that ENT1 knockdown inhibited the ethanol-induced EAAT2 up-regulation. Conclusions:, Our results suggest that ENT1 regulates glutamate uptake activity by altering EAAT2 expression and function, which might be implicated in ethanol intoxication and preference. [source] The type 1 equilibrative nucleoside transporter regulates anxiety-like behavior in miceGENES, BRAIN AND BEHAVIOR, Issue 8 2007J. Chen Activation of adenosine receptors in the brain reduces anxiety-like behavior in animals and humans. Because nucleoside transporters regulate adenosine levels, we used mice lacking the type 1 equilibrative nucleoside transporter (ENT1) to investigate whether ENT1 contributes to anxiety-like behavior. The ENT1 null mice spent more time in the center of an open field compared with wild-type littermates. In the elevated plus maze, ENT1 null mice entered more frequently into and spent more time exploring the open arms. The ENT1 null mice also spent more time exploring the light side of a light,dark box compared with wild-type mice. Microinjection of an ENT1-specific antagonist, nitrobenzylthioinosine (nitrobenzylmercaptopurine riboside), into the amygdala of C57BL/6J mice reduced anxiety-like behavior in the open field and elevated plus maze. These findings show that amygdala ENT1 modulates anxiety-like behavior. The ENT1 may be a drug target for the treatment of anxiety disorders. [source] Insulin restores glucose inhibition of adenosine transport by increasing the expression and activity of the equilibrative nucleoside transporter 2 in human umbilical vein endotheliumJOURNAL OF CELLULAR PHYSIOLOGY, Issue 3 2006Gonzalo Muñoz L -Arginine transport and nitric oxide (NO) synthesis (L -arginine/NO pathway) are stimulated by insulin, adenosine or elevated extracellular D -glucose in human umbilical vein endothelial cells (HUVEC). Adenosine uptake via the human equilibrative nucleoside transporters 1 (hENT1) and 2 (hENT2) has been proposed as a mechanism regulating adenosine plasma concentration, and therefore its vascular effects in human umbilical veins. Thus, altered expression and/or activity of hENT1 or hENT2 could lead to abnormal physiological plasma adenosine level. We have characterized insulin effect on adenosine transport in HUVEC cultured in normal (5 mM) or high (25 mM) D -glucose. Insulin (1 nM) increased overall adenosine transport associated with higher hENT2-, but lower hENT1-mediated transport in normal D -glucose. Insulin increased hENT2 protein abundance in normal or high D -glucose, but reduced hENT1 protein abundance in normal D -glucose. Insulin did not alter the reduced hENT1 protein abundance, but blocked the reduced hENT1 and hENT2 mRNA expression induced by high D -glucose. Insulin effect on hENT1 mRNA expression in normal D -glucose was blocked by NG -nitro- L -arginine methyl ester (L-NAME, NO synthase inhibitor) and mimicked by S -nitroso- N -acetyl- L,D -penicillamine (SNAP, NO donor). L-NAME did not block insulin effect on hENT2 expression. In conclusion, insulin stimulation of overall adenosine transport results from increased hENT2 expression and activity via a NO-independent mechanism. These findings could be important in hyperglycemia-associated pathological pregnancies, such as gestational diabetes, where plasma adenosine removal by the endothelium is reduced, a condition that could alter the blood flow from the placenta to the fetus affecting fetus growth and development. J. Cell. Physiol. 209: 826,835, 2006. © 2006 Wiley-Liss, Inc. [source] Role of Wake-Promoting Basal Forebrain and Adenosinergic Mechanisms in Sleep-Promoting Effects of EthanolALCOHOLISM, Issue 6 2010Mahesh M. Thakkar Background:, Ethanol intake has significant impact on sleep. However, the cellular substrates responsible for sleep promotion following ethanol intake are unknown. The purine nucleoside, adenosine, is responsible for mediating many neuronal and behavioral responses to ethanol. Studies performed in cell cultures suggest that ethanol inhibits equilibrative nucleoside transporter 1 to block the reuptake of adenosine resulting in increased extracellular adenosine. Adenosine also has a pivotal role in sleep regulation. Adenosine acts via A1 receptor to inhibit the wake-promoting neurons of the basal forebrain (BF) resulting in the promotion of sleep. Is ethanol-induced sleep associated with the inhibition of the BF wake-promoting neurons? Do adenosinergic mechanisms in the BF have a role in sleep-promoting effects of ethanol? Methods:, To address these questions, we performed 3 experiments in Sprague,Dawley rats. First, we verified the effect of ethanol on sleep promotion. Second, we evaluated the effect of ethanol on c-Fos expression (a marker of neuronal activation) in the BF wake-promoting neurons and third we monitored the effects of A1 receptor blockade in the BF on ethanol-induced sleep. Results:, Significant increase in non-rapid eye movement (NREM) sleep with a concomitant decrease in wakefulness was observed during the first 12 hours postethanol. REM sleep remained unaffected. Ethanol administration caused a significant decrease in the number of BF wake-promoting neurons with c-Fos immunoreactivity. Bilateral microinjections of a selective A1R receptor antagonist 8-cyclopentyl-1, 3-dipropylxanthine into the BF significantly attenuated sleep-promoting effects of ethanol. Conclusion:, These results suggest that the inhibition of BF wake-promoting neurons by adenosinergic mechanism may be responsible for the sleep promoting effects of ethanol. We believe our study is the first to investigate the cellular mechanisms responsible for the somnogenic effects of ethanol. [source] Determinants of sensitivity and resistance to gemcitabine: The roles of human equilibrative nucleoside transporter 1 and deoxycytidine kinase in non-small cell lung cancerCANCER SCIENCE, Issue 9 2004Hiroyuki Achiwa Gemcitabine is one of the most commonly used agents for lung cancer chemotherapy, but the determinants of sensitivity and/or resistance to this agent are not yet fully understood. In this study we used quantitative RT-PCR to examine the expression levels of human equilibrative nucleoside transporter 1 (hENT1) and deoxycytidine kinase (dCK) genes in non-small cell lung cancer (NSCLC) cell lines in relation to sensitivity and resistance to gemcitabine. The basal expression levels of hENT1 were significantly correlated with the IC50 values for gemcitabine (r=-0.6769, P=0.0005), whereas dCK expression levels were not. In a highly gemcitabine-sensitive cell line, NCI-H23, the sensitivity to gemcitabine was inhibited by nitrobenzylmercaptopurine ribonucleoside (NBMPR), an inhibitor of hENT1, without significant modulation of hENT1 expression. These data suggest that hENT1 is associated with gemcitabine sensitivity in lung cancer. We also continuously exposed NCI-H23 cells to gemcitabine and subsequently established the drug-resistant clone H23/GEM-R, which showed a significant decrease of dCK expression; however, hENT1 expression was not altered in the continuously exposed sublines or in the resistant clone. We conclude that increased hENT1 expression is a determinant of gemcitabine sensitivity, while decreased dCK expression is associated with acquired resistance to gemcitabine in NSCLC cells. Thus, hENT1 and dCK might be useful as predictive markers for efficacy of gemcitabine therapy in NSCLC. [source] Nucleoside transporter expression and function in cultured mouse astrocytesGLIA, Issue 1 2005Liang Peng Abstract Uptake of purine and pyrimidine nucleosides in astrocytes is important for several reasons: (1) uptake of nucleosides contributes to nucleic acid synthesis; (2) astrocytes synthesize AMP, ADP, and ATP from adenosine and GTP from guanosine; and (3) adenosine and guanosine function as neuromodulators, whose effects are partly terminated by cellular uptake. It has previously been shown that adenosine is rapidly accumulated by active uptake in astrocytes (Hertz and Matz, Neurochem Res 14:755,760, 1989), but the ratio between active uptake and metabolism-driven uptake of adenosine is unknown, as are uptake characteristics for guanosine. The present study therefore aims at providing detailed information of nucleoside transport and transporters in primary cultures of mouse astrocytes. Reverse transcription-polymerase chain reaction identified the two equilibrative nucleoside transporters, ENT1 and ENT2, together with the concentrative nucleoside transporter CNT2, whereas CNT3 was absent, and CNT1 expression could not be investigated. Uptake studies of tritiated thymidine, formycin B, guanosine, and adenosine (3-s uptakes at 1,4°C to study diffusional uptake and 1,60-min uptakes at 37°C to study concentrative uptake) demonstrated a fast diffusional uptake of all four nucleosides, a small, Na+ -independent and probably metabolism-driven uptake of thymidine (consistent with DNA synthesis), larger metabolism-driven uptakes of guanosine (consistent with synthesis of DNA, RNA, and GTP) and especially of adenosine (consistent with rapid nucleotide synthesis), and Na+ -dependent uptakes of adenosine (consistent with its concentrative uptake) and guanosine, rendering neuromodulator uptake independent of nucleoside metabolism. Astrocytes are accordingly well suited for both intense nucleoside metabolism and metabolism-independent uptake to terminate neuromodulator effects of adenosine and guanosine. © 2005 Wiley-Liss, Inc. [source] Insulin restores glucose inhibition of adenosine transport by increasing the expression and activity of the equilibrative nucleoside transporter 2 in human umbilical vein endotheliumJOURNAL OF CELLULAR PHYSIOLOGY, Issue 3 2006Gonzalo Muñoz L -Arginine transport and nitric oxide (NO) synthesis (L -arginine/NO pathway) are stimulated by insulin, adenosine or elevated extracellular D -glucose in human umbilical vein endothelial cells (HUVEC). Adenosine uptake via the human equilibrative nucleoside transporters 1 (hENT1) and 2 (hENT2) has been proposed as a mechanism regulating adenosine plasma concentration, and therefore its vascular effects in human umbilical veins. Thus, altered expression and/or activity of hENT1 or hENT2 could lead to abnormal physiological plasma adenosine level. We have characterized insulin effect on adenosine transport in HUVEC cultured in normal (5 mM) or high (25 mM) D -glucose. Insulin (1 nM) increased overall adenosine transport associated with higher hENT2-, but lower hENT1-mediated transport in normal D -glucose. Insulin increased hENT2 protein abundance in normal or high D -glucose, but reduced hENT1 protein abundance in normal D -glucose. Insulin did not alter the reduced hENT1 protein abundance, but blocked the reduced hENT1 and hENT2 mRNA expression induced by high D -glucose. Insulin effect on hENT1 mRNA expression in normal D -glucose was blocked by NG -nitro- L -arginine methyl ester (L-NAME, NO synthase inhibitor) and mimicked by S -nitroso- N -acetyl- L,D -penicillamine (SNAP, NO donor). L-NAME did not block insulin effect on hENT2 expression. In conclusion, insulin stimulation of overall adenosine transport results from increased hENT2 expression and activity via a NO-independent mechanism. These findings could be important in hyperglycemia-associated pathological pregnancies, such as gestational diabetes, where plasma adenosine removal by the endothelium is reduced, a condition that could alter the blood flow from the placenta to the fetus affecting fetus growth and development. J. Cell. Physiol. 209: 826,835, 2006. © 2006 Wiley-Liss, Inc. [source] | |||||||||||||