Quantification of Extracellular and Intracellular Adenosine Production

Deußen, Andreas; Stappert, Margit; Schäfer, Stefan; Kelm, Malte

doi:10.1161/01.cir.99.15.2041

Cited by 99 publications

(105 citation statements)

References 27 publications

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“…However, it is not clear whether the source of adenosine is intracellular or extracellular under stress conditions. Deussen et al (9) showed that, in guinea pig hearts under well-oxygenated conditions, addition of NBMPR increased interstitial adenosine concentrations and the source of adenosine was extracellular.…”

Section: Discussionmentioning

confidence: 99%

d-Glucose upregulates adenosine transport in cultured human aortic smooth muscle cells

Leung

Man

Tse³

2005

American Journal of Physiology-Heart and Circulatory Physiology

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nucleoside transporter; diabetes ADENOSINE MODULATES CELLULAR functions via G protein-coupled receptors to protect cells and organs and is released in response to cell injury and stress. For instance, adenosine levels in blood and interstitial fluid increase during hypoxia and ischemia. Increased extracellular adenosine causes vasodilation by acting through A 2 adenosine receptors on vascular smooth muscle cells (SMCs) (5, 31) and, thus, increases blood flow and oxygenation. Adenosine inhibits platelet aggregation (33), decreases heart rate (14), and antagonizes the stimulatory effects of catecholamines (37). Because of these important functions of adenosine as a negative-feedback modulator of cell and organ energy demand and consumption and as a celland organ-protective agent, it is known as a "retaliatory metabolite" (25). Recent reports also showed that adenosine released during preconditioning by short periods of ischemia followed by reperfusion induces cardioprotection to a subsequent sustained ischemia (17). This effect is mediated in part by activation of A 1 and A 3 adenosine receptors in cardiomyocytes and involves PKC and mitochondrial ATP-sensitive K ϩ channels (24).Nucleoside transporters fine-tune the local concentrations of adenosine in the vicinity of adenosine receptors. Two major classes of nucleoside transporters in mammalian cells have been characterized by Na ϩ dependence (16). The equilibrative nucleoside transporters (ENTs) are facilitated-diffusion systems and are Na ϩ independent. They are further subdivided into two types on the basis of their sensitivities to inhibition by nitrobenzylmercaptopurine riboside (NBMPR) (4, 43). ENT-1 is potently inhibited by nanomolar concentrations of NBMPR, but ENT-2 is resistant to NBMPR up to 1 M. ENT-1 and ENT-2 are broadly selective, accepting purine and pyrimidine nucleosides, but ENT-2 also transports nucleobases such as hypoxanthine (6,26,46). In contrast, the concentrative nucleoside transporters (CNTs) are Na ϩ dependent. CNT-1 is pyrimidine selective (36), CNT-2 is purine selective (42), and CNT-3 is broadly selective (35,41).Diabetes mellitus is a major risk factor for cardiovascular diseases. Because adenosine inhibits growth of vascular SMCs (10,11,12,13) and nucleoside transporters are involved in adenosine homeostasis, in the present study, we characterized the nucleoside transport systems in cultured human aortic SMCs (HASMCs). We also determined the effect on adenosine transport of long-term exposure of cells to 25 mM D-glucose, a condition that mimics hyperglycemia of diabetes. MATERIALS AND METHODSCulture of HASMCs. HASMCs were obtained from American Type Culture Collection (Manassas, VA) and cultured in DMEM (containing 5 mM D-glucose) supplemented with 10% (vol/vol) fetal bovine serum, 100 U/ml penicillin, and 100 g/ml streptomycin at 37°C in 95% air-5% CO 2. All experiments were carried out with cells from passages 6 to 14. At 48 h before uptake study and mRNA and protein isolation, cells were incubated in serum-free DMEM. To study t...

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Section: Discussionmentioning

confidence: 99%

d-Glucose upregulates adenosine transport in cultured human aortic smooth muscle cells

Leung

Man

Tse³

2005

American Journal of Physiology-Heart and Circulatory Physiology

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“…22,23 Dipyridamole inhibits this facilitated diffusion in humans in vivo and thereby increases extracellular adenosine concentration, which subsequently activates adenosine receptors at sites of adenosine formation. 13,14,24 -26 These observations justify the use of dipyridamole-induced vasodilation as a read-out of extracellular adenosine formation.…”

Section: Rosuvastatin Increases Dipyridamole-induced Vasodilationmentioning

confidence: 99%

Rosuvastatin Increases Extracellular Adenosine Formation in Humans In Vivo

Meijer

Oyen

Dekker

et al. 2009

ATVB

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Objective-Statins may increase extracellular adenosine formation from adenosine monophosphate by enhancing ecto-5Ј-nucleotidase activity. This theory was tested in humans using dipyridamole-induced vasodilation as a read-out for local adenosine formation. Dipyridamole inhibits the transport of extracellular adenosine into the cytosol resulting in increased extracellular adenosine and subsequent vasodilation. In addition, we studied the effect of statin therapy in a forearm model of ischemia-reperfusion injury. Methods and Results-Volunteers randomly received rosuvastatin or placebo in a double-blind parallel design (nϭ21).The forearm vasodilator response to intraarterial dipyridamole was determined in the absence and presence of the adenosine antagonist caffeine. During a separate visit the vasodilator response to nitroprusside and adenosine was established. In addition, healthy men were randomly divided in 3 groups to receive either placebo (nϭ10), rosuvastatin (nϭ22), or rosuvastatin combined with intravenous caffeine (nϭ12). Subsequently, volunteers performed forearm ischemic exercise. At reperfusion, Tc-99m-labeled annexin A5 was infused intravenously and scintigraphic images were acquired, providing an early marker of cell injury. Rosuvastatin treatment significantly increased the vasodilator response to dipyridamole, which was prevented by caffeine. Rosuvastatin did not influence the response to either sodium nitroprusside or adenosine indicating a specific interaction between rosuvastatin and dipyridamole, which does not result from an effect of rosuvastatin on adenosine clearance nor adenosine-receptor affinity or efficacy. Rosuvastatin increased tolerance to ischemia-reperfusion injury, which was attenuated by caffeine. Conclusions-Rosuvastatin increases extracellular adenosine formation, which provides protection against ischemiareperfusion injury in humans in vivo. [1][2][3] This benefit of statins has been attributed to the lowering of plasma cholesterol. However, preclinical research indicates that HMG-CoA reductase inhibition has additional effects, including the activation of ecto-5Ј-nucleotidase which converses extracellular adenosine monophosphate into adenosine. 4 -6 Increased adenosine formation favorably influences cardiovascular disease by reducing platelet aggregation, 7 atherosclerosis formation, 8 and ischemia-reperfusion injury. 9 As statins possess similar properties, adenosine is a possible candidate to mediate these effects. 3,10 -12 To our knowledge, the role of extracellular adenosine formation in the benefit of statins has not been explored in humans.Here, we report on the results of 3 studies which addressed the effect of rosuvastatin on adenosine formation and its potential relevance in humans in vivo. For this purpose, we used the adenosine receptor antagonist caffeine and the nucleoside transport inhibitor dipyridamole, as pharmacological tools to assess the involvement of endogenous adenosine in statin-induced effects. Dipyridamole reduces clearance of extracellular adenosi...

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“…Figure 1A illustrates that a physiological concentration of adenosine (1 nmol/L) [17,18] decreased I h from 16.1±0.3 pA (I h (Con) ) to 9.9±0.7 pA (I h (Ado) ) by 39% (n=8, P<0.05, upper panel). The decrease in I p was not due to pump "rundown" because the adenosine effect was reversible upon washout (lower panel of Figure 1A).…”

Section: Resultsmentioning

confidence: 99%

Isoform-specific regulation of the Na+-K+ pump by adenosine in guinea pig ventricular myocytes

et al. 2009

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Aim: The present study investigated the effect of adenosine on Na + -K + pumps in acutely isolated guinea pig (Cavia sp.) ventricular myocytes. Methods: The whole-cell, patch-clamp technique was used to record the Na + -K + pump current (I p ) in acutely isolated guinea pig ventricular myocytes. Results: Adenosine inhibited the high DHO-affinity pump current (I h ) in a concentration-dependent manner, which was blocked by the selective adenosine A 1 receptor antagonist DPCPX and the general protein kinase C (PKC) antagonists staurosporine, GF 109203X or the specific δ isoform antagonist rottlerin. In addition, the inhibitory action of adenosine was mimicked by a selective A 1 receptor agonist CCPA and a specific activator peptide of PKC-δ, PP114. In contrast, the selective A 2A receptor agonist CGS21680 and A 3 receptor agonist Cl-IB-MECA did not affect I h . Application of the selective A 2A receptor antagonist SCH58261 and A 3 receptor antagonist MRS1191 also failed to block the effect of adenosine. Furthermore, H89, a selective protein kinase A (PKA) antagonist, did not exert any effect on adenosine-induced I h inhibition. Conclusion:The present study provides the electrophysiological evidence that adenosine can induce significant inhibition of I h via adenosine A 1 receptors and the PKC-δ isoform.

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Quantification of Extracellular and Intracellular Adenosine Production

Cited by 99 publications

References 27 publications

d-Glucose upregulates adenosine transport in cultured human aortic smooth muscle cells

d-Glucose upregulates adenosine transport in cultured human aortic smooth muscle cells

Rosuvastatin Increases Extracellular Adenosine Formation in Humans In Vivo

Isoform-specific regulation of the Na+-K+ pump by adenosine in guinea pig ventricular myocytes

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