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Sample records for adenosine triphosphatase activity

  1. Hydrogen potassium adenosine triphosphatase activity inhibition and downregulation of its expression by bioactive fraction DLBS2411 from Cinnamomum burmannii in gastric parietal cells

    Tjandrawinata RR; Nailufar F; Arifin PF

    2013-01-01

    Raymond R Tjandrawinata1,2, Florensia Nailufar2, Poppy F Arifin11Section of Molecular Pharmacology, 2Section of Animal Pharmacology, Dexa Medica, Cikarang, IndonesiaAbstract: This study assessed the gastric acid antisecretory effect of DLBS2411 fractionated from Cinnamomum burmannii. Hydrogen potassium adenosine triphosphatase (H+/K+ ATPase) activity and its gene expression were observed, and the antioxidant activity of DLBS2411 was also investigated. Treatment of DLBS2411 decreased the level...

  2. Ultracytochemical localization of H+—adenosine triphosphatase activity in autophagic vacuoles induced by vinblastine in rat liver

    LUOSHENQIU; MASAHIROSAKAI; 等

    1990-01-01

    H-adenosine triphosphatase (H+-ATPase) activity was demonstrated cytochemically in autophagic vacuoles(AVs) of rat hepatocytes using a modification of the method for the demonstration of neutral p-nitrophenyl phosphatase(p-NPPase) activity[1].When an inhibitor of H+-ATPase,N-ethylmaleimide (NEM) or 4,4'-diisothiocyanostilbene-2,2'disulfonic acid,disodium salt (DIDS) was included in the incubation medium the enyzme activity was abolished indicating that p-NPPase demonstrated in this study represents H+-ATPase.Autophagy was induced by a single intraperitoneal injection of vinblastine sulfate(VBL).The number of AVs increased remarkably in hepatocytes from 40 min after VBL treatment.H+-ATPase activity was observed mainly on the membranes of lysosomes and AVs.However,early forms of AVs containing only incompletely digested material showed no H+-ATPase activity.Most AVs revealing a positive reaction seemed to be in advanced stages of development.Acid phosphatase acticity was demonstrable in mature but not in early forms of AVs.The present investigation showed that membranes of advanced stage AVs possess an H+-ATPase which may be derived from lysosomal membranes.

  3. Functional proteomics of adenosine triphosphatase system in the rat striatum during aging

    Roberto Federico Villa; Federica Ferrari; Antonella Gorini

    2012-01-01

    The maximum rates of adenosine triphosphatase (ATPase) systems related to energy consumption were systematically evaluated in synaptic plasma membranes isolated from the striata of male Wistar rats aged 2, 6, 12, 18, and 24 months, because of their key role in presynaptic nerve ending homeostasis. The following enzyme activities were evaluated: sodium-potassium-magnesium adenosine triphosphatase (Na+, K+, Mg2+-ATPase); ouabain-insensitive magnesium adenosine triphosphatase (Mg2+-ATPase); sodium-potassium adenosine triphosphatase (Na+, K+-ATPase); direct magnesium adenosine triphosphatase (Mg2+-ATPase); calcium-magnesium adenosine triphosphatase (Ca2+, Mg2+-ATPase); and acetylcholinesterase. The results showed that Na+, K+-ATPase decreased at 18 and 24 months, Ca2+, Mg2+-ATPase and acetylcholinesterase decreased from 6 months, while Mg2+-ATPase was unmodified. Therefore, ATPases vary independently during aging, suggesting that the ATPase enzyme systems are of neuropathological and pharmacological importance. This could be considered as an experimental model to study regeneration processes, because of the age-dependent modifications of specific synaptic plasma membranes. ATPases cause selective changes in some cerebral functions, especially bioenergetic systems. This could be of physiopathological significance, particularly in many central nervous system diseases, where, during regenerative processes, energy availability is essential.

  4. Adenosine triphosphatase, acetylcholinesterase and sulfhydryl groups of rat skin at local x-ray irradiation

    The activity of acetylcholinesterase (ACE), adenosine triphosphatase (ATPase), and sulfhydryl groups(SH-groups) in different structural elements of skin in the course of radiation epidermitis was studied. The posterior extremities of Wistar rats were exposed to single x-ray irradiation at a dose of 30 Gy

  5. Oxidative stress parameters and erythrocyte membrane adenosine triphosphatase activities in streptozotocin-induced diabetic rats administered aqueous preparation of Kalanchoe Pinnata leaves

    Nikhil Menon

    2016-01-01

    Full Text Available Background: Diabetes mellitus is a chronic metabolic disease that according to the World Health Organization affects more than 382 million people. The rise in diabetes mellitus coupled with the lack of an effective treatment has led many to investigate medicinal plants to identify a viable alternative. Objective: To evaluate red blood cell (RBC membrane adenosine triphosphatase (ATPase activities and antioxidant levels in streptozotocin-induced diabetic rats administered aqueous preparation of Kalanchoe pinnata leaves. Materials and Methods: Diabetes mellitus was induced in rats by a single administration of streptozotocin (60 mg/kg. Diabetic rats were then treated with aqueous K. pinnata preparation (three mature leaves ~ 9.96 g/70 kg body weight or about 0.14 g/kg body weight/day for 30 days. Serum glucose, RBC membrane ATPase activities, and antioxidant levels were determined. Results: We noted weight loss and reduced food consumption in the treated diabetic group. Serum glucose levels were reduced in the treated diabetic group compared to the other groups. Superoxide dismutase activity and glutathione levels were not significantly elevated in the treated group compared to the diabetic group. However, serum catalase activity was significantly (P < 0.05 increased in the treated diabetic group compared to the other groups. Serum thiobarbituric acid reactive substances were not significantly altered among the groups. There was a significant (P < 0.05 increase in Mg2+ ATPase activity and a nonsignificant increase in Na+/K+ ATPase activity in the RBC membrane of the treated diabetic group compared to the diabetic group. Conclusion: The consumption of aqueous preparation of K. pinnata may accrue benefits in the management of diabetes by lowering oxidative stress often associated with the disease and improving the availability of cellular magnesium through an increase in the magnesium ATPase pump in the RBC membrane for increased cellular metabolism

  6. Effect of an antioxidant combination on the distribution of acetylcholinesterase and adenosine triphosphatase activities in the cerebellum of in lindane-intoxicated mice

    Devendra Kumar Bhatt

    2013-04-01

    Full Text Available Objective: The present investigation ascertains a protective potential of a combination of antioxidants against lindane-induced toxicity in cerebellum of mice. Methods: For the study, animals are divided into four groups. First group is control and it is given only vehicle. Second group is treated with lindane and analysed if there are any lesions in the brain. Third group is purely antioxidants treated group; four antioxidants, i.e. resveratrol, ascorbic acid, alpha lipoic acid and vitamin E, are subcutaneously administered in a suitable combination to the animals of this group. Fourth group is treated with both lindane and antioxidants. Acetylcholinesterase (AChE and adenosine triphosphatase (ATPase activities are used as histochemical markers for manifestation of lindane-induced acute toxicity. Biochemical levels of glutathione (GSH and thiobarbituric acid reactive substances (TBARS were also evaluated for different groups to confirm the toxicity of lindane in cerebellum. Results: Treatment with lindane caused decrease in AChE and ATPase activities, and GSH levels in cerebellum whereas a significant increase was recorded in TBARS. Antioxidants treatment increased the enzymatic activities. A significant rise in GSH level was recorded in the antioxidant treatment group as compared to group I and group II whereas TBARS levels were significantly reduced. GSH and TBARS levels altered significantly in group IV as compared to control group and lindane-treated group. In group III, AChE and ATPase activities increased in layers and nuclei of cerebellum as compared to control group. Conclusions: Since the use of antioxidants prevents the inhibition of AChE and ATPase, functions which are altered due to lindane-toxicity may be protected. [J Exp Integr Med 2013; 3(2.000: 103-112

  7. Retinoic acid inhibits calmodulin binding to human erythrocyte membranes and reduces membrane Ca2(+)-adenosine triphosphatase activity.

    Davis, F B; Smith, T. J.; Deziel, M R; Davis, P J; Blas, S D

    1990-01-01

    Ca2(+)-ATPase activity in human red cell membranes is dependent on the presence of calmodulin. All trans-retinoic acid inhibited human red cell membrane Ca2(+)-ATPase activity in vitro in a concentration-dependent manner (10(-8) to 10(-4) M). In contrast, retinol, retinal, 13-cis-retinoic acid and the benzene ring analogue of retinoic acid did not alter enzyme activity. Purified calmodulin (up to 500 ng/ml, 3 X 10(-8) M) added to red cell membranes, in the presence of inhibitory concentration...

  8. Magnetic resonance and kinetic studies of the mechanism of membrane-bound sodium and potassium ion- activated adenosine triphosphatase.

    Grisham, C M; Mildvan, A S

    1975-01-01

    EPR and water proton relaxation rate (1/T1) studies of partially (40%) and "fully" (90%) purified preparations of membrane-bound (Na+ + K+) activated ATPase from sheep kidney indicate one tight binding site for Mn2+ per enzyme dimer, with a dissociation constant (KD = 0.88 muM) in agreement with the kinetically determined activator constant, identifying this Mn2+-binding site as the active site of the ATPase. Competition studies indicate that Mg2+ binds at this site with a dissociation constant of 1 mM in agreement with its activator constant. Inorganic phosphate and methylphosphonate bind to the enzyme-Mn2+ complex with similar high affinities and decrease 1/T1 of water protons due to a decrease from four to three in the number of rapidly exchanging water protons in the coordination sphere of enzyme-bound Mn2+. The relative effectiveness of Na+ and K+ in facilitating ternary complex formation with HPO2-4 and CH3PO2-3 as a function of pH indicates that Na+ induces the phosphate monoanion to interact with enzyme-bound Mn2+. Thus protonation of an enzyme-bound phosphoryl group would convert a K+-binding site to a Na+-binding site. Dissociation constants for K+ and Na+, estimated from NMR titrations, agreed with kinetically determined activator constants of these ions consistent with binding to the active site. Parallel 32Pi-binding studies show negligible formation (less than 7%) of a covalent E-P complex under these conditions, indicating that the NMR method has detected an additional noncovalent intermediate in ion transport. Ouabain, which increases the extent of phosphorylation of the enzyme to 24% at pH 7.8 and to 106% at pH 6.1, produced further decreases in 1/T1 of water protons. Preliminary 31P- relaxation studies of CH3PO2-3 in the presence of ATPase and Mn2+ yield an Mn to P distance (6.9 +/- 0.5 A) suggesting a second sphere enzyme-Mn-ligand-CH3PO2-3 complex. Previous kinetic studies have shown that T1+ substitutes for K+ in the activation of the enzyme

  9. Pre-steady-state studies of the adenosine triphosphatase activity of coupled submitochondrial particles. Regulation by ADP.

    Martins, O B; Tuena de Gómez-Puyou, M; Gómez-Puyou, A

    1988-09-20

    ATPase activities were measured in 10 mM MgCl2, 5 mM ATP, 1 mM ADP, and 1 microM FCCP with submitochondrial particles from bovine heart that had been stimulated by delta mu H+-forming substrates and with particles whose natural inhibitor protein was partially removed by heating. The activities were not linear with time. With both particles, the rate of ATP hydrolysis in the 7-fold greater than that in the steady state. Pre-steady-state and steady-state kinetic studies showed that the decrease of ATPase activity was due to the binding of ADP in a high-affinity site of the enzyme (K0.5 of 10 microM). Inhibition of ATP hydrolysis was accompanied by the binding of approximately 1 mol of ADP/mol of particulate F1; 10 microM ADP gave half-maximal binding. ADP could be replaced by IDP, but with an affinity 50-fold lower (K0.5 of 0.5 mM). Maximal inhibition by ADP and IDP was achieved in less than 5 s. Inhibition was enhanced by uncouplers. Even in the presence of pyruvate kinase and phosphoenolpyruvate, the rates of hydrolysis were about 2.5-fold higher in the first seconds of reaction than in the steady state. This decrease of ATPase activity also correlated with the binding of nearly 1 mol of ADP/mol of F1. This inhibitory ADP remained bound to the enzyme after several thousand turnovers. Apparently, it is possible to observe maximal rates of hydrolysis only in the first few catalytic cycles of the enzyme. PMID:2974725

  10. Mapping the active site of vaccinia virus RNA triphosphatase

    The RNA triphosphatase component of vaccinia virus mRNA capping enzyme (the product of the viral D1 gene) belongs to a family of metal-dependent phosphohydrolases that includes the RNA triphosphatases of fungi, protozoa, Chlorella virus, and baculoviruses. The family is defined by two glutamate-containing motifs (A and C) that form the metal-binding site. Most of the family members resemble the fungal and Chlorella virus enzymes, which have a complex active site located within the hydrophilic interior of a topologically closed eight-stranded β barrel (the so-called ''triphosphate tunnel''). Here we queried whether vaccinia virus capping enzyme is a member of the tunnel subfamily, via mutational mapping of amino acids required for vaccinia triphosphatase activity. We identified four new essential side chains in vaccinia D1 via alanine scanning and illuminated structure-activity relationships by conservative substitutions. Our results, together with previous mutational data, highlight a constellation of six acidic and three basic amino acids that likely compose the vaccinia triphosphatase active site (Glu37, Glu39, Arg77, Lys107, Glu126, Asp159, Lys161, Glu192, and Glu194). These nine essential residues are conserved in all vertebrate and invertebrate poxvirus RNA capping enzymes. We discerned no pattern of clustering of the catalytic residues of the poxvirus triphosphatase that would suggest structural similarity to the tunnel proteins (exclusive of motifs A and C). We infer that the poxvirus triphosphatases are a distinct lineage within the metal-dependent RNA triphosphatase family. Their unique active site, which is completely different from that of the host cell's capping enzyme, recommends the poxvirus RNA triphosphatase as a molecular target for antipoxviral drug discovery

  11. Possible mechanism for the inhibition of gastric (H+ + K+)-adenosine triphosphatase by the proton pump inhibitor AG-1749

    Mechanisms related to the inhibition of (H+ + K+)-adenosine triphosphatase (ATPase) by 2-[[[3-methyl-4-(2,2,2-trifluoroethoxy)-2-pyridyl]methyl] sulfinyl]-1H-benzimidazole (AG-1749) were studied using canine gastric microsomes. AG-1749 (1-100 microM) inhibited the K+-stimulated ATP hydrolysis and vesicular accumulation of H+. AG-1749 bound to the microsomes concentration-dependently and decreased the number of free SH groups; the binding correlating with the enzyme inhibition. Both the binding and inhibition were antagonized by dithiothreitol. N-ethylmaleimide inhibited the (H+ + K+)-ATPase and decreased the binding of [14C]AG-1749 to the microsomes. The inhibitory effect of AG-1749 gradually increased with incubation time, and was enhanced by lowering the pH. AG-2000 and AG-1812, acid-induced rearrangement products of AG-1749, inhibited (H+ + K+)-ATPase potently, rapidly and independently of pH; the inhibition was antagonized by dithiothreitol. We propose that AG-1749 is transformed into its active forms within the acidic compartment of the parietal cells and that the active compounds inhibit (H+ + K+)-ATPase activity by reacting with the SH groups of the enzyme

  12. A kinetic description for sodium and potassium effects on (Na+ plus K+)-adenosine triphosphatase: a model for a two-nonequivalent site potassium activation and an analysis of multiequivalent site models for sodium activation.

    Lindenmayer, G E; Schwartz, A; Thompson, H K

    1974-01-01

    1. Dissociation constants for sodium and potassium of a site that modulates the rate of ouabain-(Na(+)+K(+))-ATPase interaction were applied to models for potassium activation of (Na(+)+K(+))-ATPase. The constants for potassium (0.213 mM) and for sodium (13.7 mM) were defined, respectively, as activation constant, K(a) and inhibitory constant, K(i).2. Tests of the one- and the two-equivalent site models, that describe sodium and potassium competition, revealed that neither model adequately predicts the activation effects of potassium in the presence of 100 or 200 mM sodium.3. The potassium-activation data, obtained at low potassium and high sodium, were explained by a two-nonequivalent site model where the dissociation constants of the first site are 0.213 mM for potassium and 13.7 mM for sodium. The second site was characterized by dissociation constants of 0.091 mM for potassium and 74.1 mM for sodium.4. The two-nonequivalent site model adequately predicted the responses to concentrations of potassium between 0.25 and 5 mM in the presence of 100-500 mM sodium. At lower sodium concentrations the predicted responses formed an upper limit for the function of observed activities. This limit was reached at lower concentrations of potassium and higher concentrations of sodium, which inferred saturation of the sodium-activation sites with sodium.5. Sodium-activation data were corrected for sodium interaction with potassium-activation sites by use of the two-nonequivalent site model for potassium activation. Tests of equivalent site models suggested that the corrected data for sodium activation may be most consistent with a model that has three-equivalent sites. Other multiequivalent site models (n = 2, 4, 5 or 6), however, cannot be statistically eliminated as possibilities. The three-equivalent site activation model was characterized by dissociation constants of 1.39 mM for sodium and 11.7 mM for potassium. The system theoretically would be half-maximally activated by

  13. Insecticidal properties of essential oils against Tribolium castaneum (Herbst) and their inhibitory effects on acetylcholinesterase and adenosine triphosphatases.

    Abou-Taleb, Hamdy K; Mohamed, Magdy I E; Shawir, Mohamed S; Abdelgaleil, Samir A M

    2016-01-01

    Essential oils from 20 Egyptian plants were obtained by using hydrodistillation. The chemical composition of the isolated oils was identified by gas chromatograph/mass spectrometer. Fumigant and contact toxicities of the essential oils were evaluated against the adults of Tribolium castaneum. In fumigation assays, the oil of Origanum vulgare (LC50 = 9.97 mg/L air) displayed the highest toxicity towards the adults of T. castaneum. In contact assays, the oils of Artemisia monosperma (LC50 = 0.07 mg/cm(2)) and O. vulgare (LC50 = 0.07 mg/cm(2)) were the most potent toxicants against the adults of T. castaneum. Biochemical studies showed that the tested oils caused pronounced inhibition of acetylcholinesterase (AChE) and adenosine triphosphatases (ATPases) isolated from the larvae of T. castaneum. The oil Cupressus macrocarpa (IC50 = 12.3 mg/L) was the most potent inhibitor of AChE, while the oil of Calistemon viminals (IC50 = 4.4 mg/L) was the most potent inhibitor of ATPases. PMID:25978134

  14. Membrane bound pyrophosphatase and P-type adenosine triphosphatase of Leishmania donovani as possible chemotherapeutic targets: similarities and differences in inhibitor sensitivities.

    Sen, S S; Bhuyan, N R; Lakshman, K; Roy, A K; Chakraborty, B; Bera, T

    2009-12-01

    The activities of inorganic pyrophosphatase (PPase) and adenosine triphosphatase (ATPase) were studied in the plasma membrane of Leishmania donovani promastigotes and amastigotes. It was shown that the specific activity of PPase was greater than that of ATPase in the promastigote plasma membrane. We characterized H+-PPase present in the plasma membrane of L. donovani and investigated its possible role in the survival of promastigote and amastigote. PPase activity was stimulated by K+ and sodium orthovanadate and inhibited by pyrophosphate analogs (imidodiphosphate and alendronate), KF, N,N'-dicyclohexylcarbodiimide (DCCD), thiol reagents (p-chloromercuribenzenesulfonate (PCMBS), N-ethylmaleimide (NEM), and phenylarsine oxide (PAO)), the ABC superfamily transport modulator verapamil, and also by the F(1)F(o)-ATPase inhibitor quercetin. ATPase activity was stimulated by K+ and verapamil, inhibited by DCCD, PCMBS, NEM, sodium azide, sodium orthovanadate, and quercetin, and was unaffected by PAO. We conclude that there are significant differences within promastigote, amastigote, and mammalian host in cytosolic pH homeostasis to merit the inclusion of PPase transporter as a putative target for rational drug design. PMID:19961421

  15. Effects of basic fibroblast growth factor on adenosine triphosphatase activity and antioxidation of alcoholism rat models%碱性成纤维细胞生长因子对乙醇中毒大鼠ATP酶及抗氧化作用的影响

    黄俊杰; 王彩冰; 何显教; 黄丽娟; 黄彦峰; 梁祚仁; 阳秀英; 黎昀; 赵善民

    2012-01-01

    BACKGROUND: Basic fibroblast growth factor(bFGF) has multiple biological activities, and has positive effect on the recovery oftissue trauma. But the effect of bFGF against alcoholism has not been reported.OBJECTIVE: To study the effect of bFGF on the activities of adenosine triphosphatase (ATPase) and superoxide dismutase(SOD) and malondialdehyde (MDA)in cerebral cortex and liver tissue of alcoholism rat model.METHODS: The alcoholism rat models were established by perfusing stomach with alcohol. Thirty Wistar rats were randomlydivided into three groups. Normal saline group and bFGF treatment group were injected with normal saline and bFGF,respectively, at 60 days after modeling; Alcoholism model group was treated with no intervention. Another 10 rats without alcoholserved as control group.RESULTS AND CONCLUSION: The activities of ATPase and SOD in the cerebral cortex and livertissue of alcoholism modelgroup were significantly decreased than those in the control group, but the content of M DA was significantly higher than that inthe control group (P<0.05). After bFGF intervention, the activities of ATPase and SOD in the cerebral cortex and livertissuewere significantly increased compared with the normal saline group and alcoholism model group, but the content of MDA wassignificantly decreased (P<0.05-0.01). bFGF can increase the activities of ATPase and the antioxidative ability of brain tissue inalcoholism model rats. The bFGF can protect the brain and liver of rats against alcoholism.%背景:碱性成纤维细胞生长因子具有多种生物活性,对组织创伤具有修复作用,但其对乙醇中毒的保护作用至今未见报道.目的:观察碱性成纤维细胞生长因子对乙醇中毒大鼠脑和肝组织中ATP 酶、超氧化物歧化酶活力和丙二醛水平的影响.方法:选择成年Wistar 雄性大鼠,采用白酒灌胃建立乙醇中毒模型,30 只造模成功的大鼠分为3 组,生理盐水组和碱性成纤维细胞生长

  16. Effect of Ethanol on Branchial Adenosine Triphosphatases in Oreochromis mossambicus (Peters)

    Bhanu, Smitha V.; Philip, Babu

    2011-01-01

    The aim of this work was to determine the toxicity of ethanol in an aquatic system by means of bioassays with Oreochromis mossambicus (Peters) as a test organism. The study revealed changes in the gill ATPase activities. The results obtained indicated that ethanol brought about a decrease in the body weight, followed by significant inhibition on total ATPase, Na+/K+ ATPase, Ca2+ ATPase and Mg2+ ATPase activities. The studies also indicated that these can be employed as suitable biomarkers in ...

  17. Effect of nitration and D2O on the kinetics of beef heart mitochondrial adenosine triphosphatase

    The role of tyrosine in the catalytic mechanism of nucleoside triphosphate hydrolysis by beef heart mitochondrial ATPase is explored. Compared are the rates of the ATPase reaction by both nitrated and native F1 at both pH 8 and pH 6. The pH-activity profile of nitrated F1 is compared to the pH-activity profile of the unmodified enzyme. These data indicate that the phenolic group of an active-site tyrosine must be protonated during the hydrolysis reaction. Deuterium oxide is used in the reaction buffer to explore the role of protons in the ATPase reaction. Kinetic constants of the nucleoside triphosphates are obtained at various levels of D20 using both the nitrated and native forms of F1. Several nucleoside diphosphates are used as inhibitors of F1-catalyzed ITP hydrolysis. Dissociation constants of these inhibitors are obtained at both low and high concentrations of D20 for both the nitrated and native F1. We explore the possibility that a tyrosine and an arginine lie in close proximity in the F1 active site by studying the effects of sequential modification of arginine and tyrosine. These results are interpreted in terms of possible ATP hydrolysis mechanisms. Two possible roles for tyrosine in the hydrolysis of nucleoside triphosphates by F1 are suggested

  18. Influence of the α-, β- and γ-subunits of the energy-transducing adenosine triphosphatase from Micrococcus lysodeikticus in the immunochemical properties of the protein and in their reconstitution studied by a radioimmunoassay method

    A sensitive radioimmunoassay was developed for the energy-transducing adenosine triphosphatase (F1-ATPase, EC 3.6.1.3) of Micrococcus lysodeikticus and the assay was extended to the α-, β- and γ-subunits of the enzyme. These subunits were isolated and cross-reactions studied. (author)

  19. Adenosine-induced activation of esophageal nociceptors.

    Ru, F; Surdenikova, L; Brozmanova, M; Kollarik, M

    2011-03-01

    Clinical studies implicate adenosine acting on esophageal nociceptive pathways in the pathogenesis of noncardiac chest pain originating from the esophagus. However, the effect of adenosine on esophageal afferent nerve subtypes is incompletely understood. We addressed the hypothesis that adenosine selectively activates esophageal nociceptors. Whole cell perforated patch-clamp recordings and single-cell RT-PCR analysis were performed on the primary afferent neurons retrogradely labeled from the esophagus in the guinea pig. Extracellular recordings were made from the isolated innervated esophagus. In patch-clamp studies, adenosine evoked activation (inward current) in a majority of putative nociceptive (capsaicin-sensitive) vagal nodose, vagal jugular, and spinal dorsal root ganglia (DRG) neurons innervating the esophagus. Single-cell RT-PCR analysis indicated that the majority of the putative nociceptive (transient receptor potential V1-positive) neurons innervating the esophagus express the adenosine receptors. The neural crest-derived (spinal DRG and vagal jugular) esophageal nociceptors expressed predominantly the adenosine A(1) receptor while the placodes-derived vagal nodose nociceptors expressed the adenosine A(1) and/or A(2A) receptors. Consistent with the studies in the cell bodies, adenosine evoked activation (overt action potential discharge) in esophageal nociceptive nerve terminals. Furthermore, the neural crest-derived jugular nociceptors were activated by the selective A(1) receptor agonist CCPA, and the placodes-derived nodose nociceptors were activated by CCPA and/or the selective adenosine A(2A) receptor CGS-21680. In contrast to esophageal nociceptors, adenosine failed to stimulate the vagal esophageal low-threshold (tension) mechanosensors. We conclude that adenosine selectively activates esophageal nociceptors. Our data indicate that the esophageal neural crest-derived nociceptors can be activated via the adenosine A(1) receptor while the placodes

  20. Effect of nitration and D/sub 2/O on the kinetics of beef heart mitochondrial adenosine triphosphatase

    Dorgan, L.J.; Schuster, S.M.

    1981-04-25

    The role of tyrosine in the catalytic mechanism of nucleoside triphosphate hydrolysis by beef heart mitochondrial ATPase is explored. Compared are the rates of the ATPase reaction by both nitrated and native F1 at both pH 8 and pH 6. The pH-activity profile of nitrated F1 is compared to the pH-activity profile of the unmodified enzyme. These data indicate that the phenolic group of an active-site tyrosine must be protonated during the hydrolysis reaction. Deuterium oxide is used in the reaction buffer to explore the role of protons in the ATPase reaction. Kinetic constants of the nucleoside triphosphates are obtained at various levels of D20 using both the nitrated and native forms of F1. Several nucleoside diphosphates are used as inhibitors of F1-catalyzed ITP hydrolysis. Dissociation constants of these inhibitors are obtained at both low and high concentrations of D20 for both the nitrated and native F1. We explore the possibility that a tyrosine and an arginine lie in close proximity in the F1 active site by studying the effects of sequential modification of arginine and tyrosine. These results are interpreted in terms of possible ATP hydrolysis mechanisms. Two possible roles for tyrosine in the hydrolysis of nucleoside triphosphates by F1 are suggested.

  1. High inorganic triphosphatase activities in bacteria and mammalian cells: identification of the enzymes involved.

    Gregory Kohn

    Full Text Available BACKGROUND: We recently characterized a specific inorganic triphosphatase (PPPase from Nitrosomonas europaea. This enzyme belongs to the CYTH superfamily of proteins. Many bacterial members of this family are annotated as predicted adenylate cyclases, because one of the founding members is CyaB adenylate cyclase from A. hydrophila. The aim of the present study is to determine whether other members of the CYTH protein family also have a PPPase activity, if there are PPPase activities in animal tissues and what enzymes are responsible for these activities. METHODOLOGY/PRINCIPAL FINDINGS: Recombinant enzymes were expressed and purified as GST- or His-tagged fusion proteins and the enzyme activities were determined by measuring the release of inorganic phosphate. We show that the hitherto uncharacterized E. coli CYTH protein ygiF is a specific PPPase, but it contributes only marginally to the total PPPase activity in this organism, where the main enzyme responsible for hydrolysis of inorganic triphosphate (PPP(i is inorganic pyrophosphatase. We further show that CyaB hydrolyzes PPP(i but this activity is low compared to its adenylate cyclase activity. Finally we demonstrate a high PPPase activity in mammalian and quail tissue, particularly in the brain. We show that this activity is mainly due to Prune, an exopolyphosphatase overexpressed in metastatic tumors where it promotes cell motility. CONCLUSIONS AND GENERAL SIGNIFICANCE: We show for the first time that PPPase activities are widespread in bacteria and animals. We identified the enzymes responsible for these activities but we were unable to detect significant amounts of PPP(i in E. coli or brain extracts using ion chromatography and capillary electrophoresis. The role of these enzymes may be to hydrolyze PPP(i, which could be cytotoxic because of its high affinity for Ca(2+, thereby interfering with Ca(2+ signaling.

  2. New member of the guanosine triphosphatase activating protein family in the human epididymis

    Xiangqi Li; Qiang Liu; Shigui Liu; Jinsong Zhang; Yonglian Zhang

    2008-01-01

    The effect of the guanosine triphosphatase activating proteins (GAPs) on spermatogenesis has been studied for years,though no GAPs have been explored in epididymis, an essential organ for sperm maturation. In this study, a new GAP member, designated as MacGAP, was cloned in human epididymis. The MacGAP gene encodes a protein of 618 amino acids with a putative size of 70 kDa and harbors the conservedRhoGAPdomain. The N-terminal and C-terminal peptides of MacGAP were expressed and their corresponding antiserawere prepared. The antisera against N-terminal peptide could detect antigen as low as 0.3 ng, and its specificity was alsoconfirmed. However, the antisera against C-terminal peptide failed to detect its antigen because of its low sensitivity.Immunohistochemistry showed that the MacGAP protein was dependent on epididymis and had a region-specific expression pattern, with high expression in the epithelial cells' basal section in the caput region. The results have created a foundation for further interpretation of the biological effects of GAPs in sperm maturation.

  3. Nucleoside triphosphatase and RNA helicase activities associated with GB virus B nonstructural protein 3.

    Zhong, W; Ingravallo, P; Wright-Minogue, J; Skelton, A; Uss, A S; Chase, R; Yao, N; Lau, J Y; Hong, Z

    1999-09-01

    GB virus B (GBV-B) is a positive-stranded RNA virus that belongs to the Flaviviridae family. This virus is closely related to hepatitis C virus (HCV) and causes acute hepatitis in tamarins (Saguinus species). Nonstructural protein 3 (NS3) of GBV-B contains sequence motifs predictive of three enzymatic activities: serine protease, nucleoside triphosphatase (NTPase), and RNA helicase. The N-terminal serine protease has been characterized and shown to share similar substrate specificity with the HCV NS3 protease. In this report, a full-length GBV-B NS3 protein was expressed in Escherichia coli and purified to homogeneity. This recombinant protein was shown to possess polynucleotide-stimulated NTPase and double-stranded RNA (dsRNA) unwinding activities. Both activities were abolished by a single amino acid substitution, from the Lys (K) residue in the conserved walker motif A (or Ia) "AXXXXGK(210)S" to an Ala (A), confirming that they are intrinsic to GBV-B NS3. Kinetic parameters (K(m) and k(cat)) for hydrolysis of various NTPs or dNTPs were obtained. The dsRNA unwinding activity depends on the presence of divalent metal ions and ATP and requires an RNA duplex substrate with 3' unpaired regions (RNAs with 5' unpaired regions only or with blunt ends are not suitable substrates for this enzyme). This indicates that GBV-B NS3 RNA helicase unwinds dsRNA in the 3' to 5' direction. Direct interaction of the GBV-B NS3 protein with a single-stranded RNA was established using a gel-based RNA bandshift assay. Finally, a homology model of GBV-B NS3 RNA helicase domain based on the 3-dimensional structure of the HCV NS3 helicase that shows a great similarity in overall structure and surface charge distribution between the two proteins was proposed. PMID:10497107

  4. Neurofibromatosis: The role of guanosine triphosphatase activating proteins in sensory neuron function

    Cynthia M. Hingtgen

    2008-01-01

    Neurofibromatosis type 1 (NF1) is a common autosomal dominant disease characterized by formation of multiple benign and malignant tumors. People with this disorder also experience chronic pain, which can be disabling. Neurofibromin, the protein product of the Nfl gene, is a gnanosine triphosphatase activating protein (GAP) for p21Ras (Ras). Loss of Nfl results in an increase in activity of the Ras transduction cascade. Because of the growing evidence suggesting involvement of downstream components of the Ras transduction cascade in the sensitization of nociceptive sensory neurons, we examined the stimulus-evoked release of the neuropeptides, substance P (SP) and calcitonin gene-related peptide (CGRP), from primary sensory neurons of mice with a mutation of the Nfl gene (NfI+1-). Measuring the levels of SP and CGRP by radioimmunoassay, we demonstrated that capsaicin-stimulated release of neuropep-tides is 3-5 folds higher in spinal cord slices from Nfl+1-mice than that from wildtype mouse tissue. In addition, the potassium- and capsaicin-stimulated release of CGRP from the culture of sensory neurons isolated from Nfl+1- mice was more than double that from the culture of wildtype neurons. Using patch-clamp electrophysiological techniques, we also examined the excitability of capsaicin-sensitive sensory neurons. It was found that the number of action potentials generated by the neurons from Nfl+1- mice, responsing to a ramp of depolarizing current, was more than three times of that generated by wildtype neurons. Consistent with that observation, neurons from Nfl+1- mice had lower firing thresholds, lower rheobase currents and shorter firing latencies compared with wildtype neurons. These data clearly demonstrate that GAPs, such as neurofihromin, can alter the excitability of nociceptive sensory neurons. The augmented response of sensory neurons with altered Ras signaling may explain the abnormal pain sensations experienced by people with NFI and suggests an important

  5. Transcriptional activation of the nitrogenase promoter in vitro: adenosine nucleotides are required for inhibition of NIFA activity by NIFL.

    Eydmann, T; Söderbäck, E; Jones, T; Hill, S; Austin, S; Dixon, R

    1995-03-01

    The enhancer-binding protein NIFA is required for transcriptional activation of nif promoters by the alternative holoenzyme form of RNA polymerase, which contains the sigma factor sigma 54 (sigma N). NIFA hydrolyzes nucleoside triphosphates to catalyze the isomerization of closed promoter complexes to transcriptionally competent open complexes. The activity of NIFA is antagonized by the regulatory protein NIFL in response to oxygen and fixed nitrogen in vivo. We have investigated the requirement for nucleotides in the formation and stability of open promoter complexes by NIFA and inhibition of its activity by NIFL at the Klebsiella pneumoniae nifH promoter. Open complexes formed by sigma 54-containing RNA polymerase are considerably more stable to heparin challenge in the presence of GTP than in the presence of ATP. This differential stability is most probably a consequence of GTP being the initiating nucleotide at this promoter. Adenosine nucleosides are specifically required for Azotobacter vinelandii NIFL to inhibit open complex formation by native NIFA, and the nucleoside triphosphatase activity of NIFA is strongly inhibited by NIFL under these conditions. We propose a model in which NIFL modulates the activity of NIFA via an adenosine nucleotide switch. PMID:7868590

  6. Anxiolytic activity of adenosine receptor activation in mice.

    Jain, N; Kemp, N; Adeyemo, O; Buchanan, P; Stone, T W

    1995-10-01

    1. Purine analogues have been examined for anxiolytic- and anxiogenic-like activity in mice, by use of the elevated plus-maze. 2. The selective A1 receptor agonist, N6-cyclopentyladenosine (CPA) had marked anxiolytic-like activity at 10 and 50 microg kg(-1), with no effect on locomotor performance at these doses. 3. The A1 selective adenosine receptor antagonist, 1,3-dipropyl-8-cyclopentylxanthine (CPX) had no significant effect on anxiety-related measures or locomotor behaviour, but blocked the anxiolytic-like activity of CPA. The hydrophilic xanthine, 8-(p-sulphophenyl) theophylline did not prevent anxiolysis by CPA. 4. Caffeine had anxiogenic-like activity at 30 mg kg(-1) which was prevented by CPA at 50 micro kg(-1). 5. The A2 receptor agonist, N6-[2-(3,5-dimethoxyphenyl)-2(2-methylphenyl)-ethyl]adenosine (DPMA) had no effect on anxiety behaviour but depressed locomotor activity at the highest dose tested of 1 mg kg(-1). The A2 receptor antagonist, 1,3-dimethyl-l-propargylxanthine (DMPX) had no effect on anxiety-related measures or locomotion and did not modify the anxiolytic-like activity of CPA. 6. Administration of DPMA in combination with anxiolytic doses of CPA prevented the anxiolytic-like activity of the latter. 7. The results suggest that the selective activation of central A1 adenosine receptors induces anxiolytic-like behaviour, while the activation of A2 sites causes locomotor depression and reduces the effects of A1 receptor activation. The absence of any effect of CPX alone suggests that the receptors involved in modulating behaviour in the elevated plus-maze in mice are not activated tonically by endogenous adenosine. PMID:8640355

  7. [Properties of the enzymes of energy supply in the common frog under various physiological states. II. Changes in the heat resistance of adenosine triphosphatases and succinate dehydrogenase].

    Glushankova, M A; Ivanova, G I; Pisareva, L N

    1986-05-01

    The heat resistance of SDG, Na, K-ATPase and Mg-ATPase of the grass frog was determined in January, March and May, the number of animals examined being 30-40 in either experiment. It was found that the average level of the heat resistance of the enzymes studied shows significant, often differently directed changes, which depend on the physiological state of an organism. Negligible correlation between the thermal sensitivity of different enzymes of an organism during hibernation, completely disappear during the activity state. PMID:3016954

  8. Anxiolytic activity of adenosine receptor activation in mice.

    Jain, N; Kemp, N; Adeyemo, O; Buchanan, P.; Stone, T W

    1995-01-01

    1. Purine analogues have been examined for anxiolytic- and anxiogenic-like activity in mice, by use of the elevated plus-maze. 2. The selective A1 receptor agonist, N6-cyclopentyladenosine (CPA) had marked anxiolytic-like activity at 10 and 50 microg kg(-1), with no effect on locomotor performance at these doses. 3. The A1 selective adenosine receptor antagonist, 1,3-dipropyl-8-cyclopentylxanthine (CPX) had no significant effect on anxiety-related measures or locomotor behaviour, but blocked ...

  9. Investigating real-time activation of adenosine receptors by bioluminescence resonance energy transfer technique

    Huang, Yimei; Yang, Hongqin; Zheng, Liqin; Chen, Jiangxu; Wang, Yuhua; Li, Hui; Xie, Shusen

    2013-02-01

    Adenosine receptors play important roles in many physiological and pathological processes, for example regulating myocardial oxygen consumption and the release of neurotransmitters. The activations of adenosine receptors have been studied by some kinds of techniques, such as western blot, immunohistochemistry, etc. However, these techniques cannot reveal the dynamical response of adenosine receptors under stimulation. In this paper, bioluminescence resonance energy transfer technique was introduced to study the real-time activation of adenosine receptors by monitoring the dynamics of cyclic adenosine monophosphate (cAMP) level. The results showed that there were significant differences between adenosine receptors on real-time responses under stimulation. Moreover, the dynamics of cAMP level demonstrated that competition between adenosine receptors existed. Taken together, our study indicates that monitoring the dynamics of cAMP level using bioluminescence resonance energy transfer technique could be one potential approach to investigate the mechanism of competitions between adenosine receptors.

  10. Mechanism of A2 adenosine receptor activation. I. Blockade of A2 adenosine receptors by photoaffinity labeling

    Lohse, M.J.; Klotz, K.N.; Schwabe, U.

    1991-04-01

    It has previously been shown that covalent incorporation of the photoreactive adenosine derivative (R)-2-azido-N6-p-hydroxy-phenylisopropyladenosine ((R)-AHPIA) into the A1 adenosine receptor of intact fat cells leads to a persistent activation of this receptor, resulting in a reduction of cellular cAMP levels. In contrast, covalent incorporation of (R)-AHPIA into human platelet membranes, which contain only stimulatory A2 adenosine receptors, reduces adenylate cyclase stimulation via these receptors. This effect of (R)-AHPIA is specific for the A2 receptor and can be prevented by the adenosine receptor antagonist theophylline. Binding studies indicate that up to 90% of A2 receptors can be blocked by photoincorporation of (R)-AHPIA. However, the remaining 10-20% of A2 receptors are sufficient to mediate an adenylate cyclase stimulation of up to 50% of the control value. Similarly, the activation via these 10-20% of receptors occurs with a half-life that is only 2 times longer than that in control membranes. This indicates the presence of a receptor reserve, with respect to both the extent and the rate of adenylate cyclase stimulation. These observations require a modification of the models of receptor-adenylate cyclase coupling.

  11. Role of adenosine in the sympathetic activation produced by isometric exercise in humans.

    Costa, F.; Biaggioni, I

    1994-01-01

    Isometric exercise increases sympathetic nerve activity and blood pressure. This exercise pressor reflex is partly mediated by metabolic products activating muscle afferents (metaboreceptors). Whereas adenosine is a known inhibitory neuromodulator, there is increasing evidence that it activates afferent nerves. We, therefore, examined the hypothesis that adenosine stimulates muscle afferents and participates in the exercise pressor reflex in healthy volunteers. Intraarterial administration of...

  12. Adenosine, ketogenic diet and epilepsy: the emerging therapeutic relationship between metabolism and brain activity.

    Masino, S A; Kawamura, M; Wasser, C D; Wasser, C A; Pomeroy, L T; Ruskin, D N

    2009-09-01

    For many years the neuromodulator adenosine has been recognized as an endogenous anticonvulsant molecule and termed a "retaliatory metabolite." As the core molecule of ATP, adenosine forms a unique link between cell energy and neuronal excitability. In parallel, a ketogenic (high-fat, low-carbohydrate) diet is a metabolic therapy that influences neuronal activity significantly, and ketogenic diets have been used successfully to treat medically-refractory epilepsy, particularly in children, for decades. To date the key neural mechanisms underlying the success of dietary therapy are unclear, hindering development of analogous pharmacological solutions. Similarly, adenosine receptor-based therapies for epilepsy and myriad other disorders remain elusive. In this review we explore the physiological regulation of adenosine as an anticonvulsant strategy and suggest a critical role for adenosine in the success of ketogenic diet therapy for epilepsy. While the current focus is on the regulation of adenosine, ketogenic metabolism and epilepsy, the therapeutic implications extend to acute and chronic neurological disorders as diverse as brain injury, inflammatory and neuropathic pain, autism and hyperdopaminergic disorders. Emerging evidence for broad clinical relevance of the metabolic regulation of adenosine will be discussed. PMID:20190967

  13. Reduced striatal ecto-nucleotidase activity in schizophrenia patients supports the “adenosine hypothesis”

    Aliagas, Elisabet; Villar-Menéndez, Izaskun; Sévigny, Jean; Roca, Mercedes; Romeu, Miriam; Ferrer, Isidre; Martín-Satué, Mireia; Barrachina, Marta

    2013-01-01

    Schizophrenia (SZ) is a major chronic neuropsychiatric disorder characterized by a hyperdopaminergic state. The hypoadenosinergic hypothesis proposes that reduced extracellular adenosine levels contribute to dopamine D2 receptor hyperactivity. ATP, through the action of ecto-nucleotidases, constitutes a main source of extracellular adenosine. In the present study, we examined the activity of ecto-nucleotidases (NTPDases, ecto-5′-nucleotidase, and alkaline phosphatase) in the postmortem putame...

  14. Suppression of adenosine-activated chloride transport by ethanol in airway epithelia.

    Sammeta V Raju

    Full Text Available Alcohol abuse is associated with increased lung infections. Molecular understanding of the underlying mechanisms is not complete. Airway epithelial ion transport regulates the homeostasis of airway surface liquid, essential for airway mucosal immunity and lung host defense. Here, air-liquid interface cultures of Calu-3 epithelial cells were basolaterally exposed to physiologically relevant concentrations of ethanol (0, 25, 50 and 100 mM for 24 hours and adenosine-stimulated ion transport was measured by Ussing chamber. The ethanol exposure reduced the epithelial short-circuit currents (I(SC in a dose-dependent manner. The ion currents activated by adenosine were chloride conductance mediated by cystic fibrosis transmembrane conductance regulator (CFTR, a cAMP-activated chloride channel. Alloxazine, a specific inhibitor for A(2B adenosine receptor (A(2BAR, largely abolished the adenosine-stimulated chloride transport, suggesting that A(2BAR is a major receptor responsible for regulating the chloride transport of the cells. Ethanol significantly reduced intracellular cAMP production upon adenosine stimulation. Moreover, ethanol-suppression of the chloride secretion was able to be restored by cAMP analogs or by inhibitors to block cAMP degradation. These results imply that ethanol exposure dysregulates CFTR-mediated chloride transport in airways by suppression of adenosine-A(2BAR-cAMP signaling pathway, which might contribute to alcohol-associated lung infections.

  15. Alteration in the expression of Na+-K+-adenosine triphosphatase in denervated rat skeletal muscle%去除神经支配大鼠骨骼肌中钠钾三磷酸腺苷酶表达的变化

    赵海燕; Hundal HS

    2006-01-01

    苷酶亚基的表达机制被破坏.%BACKGROUND: Insulin-induced alteration in ion transport has impact on metabolic regulation and reduced Na+-K+-adenosine triphosphatase(ATPase) activity in insulin resistauce results in hypertension and decrease in glycolysis, glucose oxidation and production of ATP.OBJECTIVE: To investigate the regulation of expression of Na+-K+-AT-Pase in insulin resistance model system.DESIGN: Controlled experiment.SETTING: Division of Molecular Physiology, Faculty of Life Sciences, University of Dundee, UK.MATERIALS: The study was performed from October 1999 to February2000,in which, 15 adult male Sprague Dawley rats (200-250 g) of clean grade were employed. The experiment was performed with 5 rats each time and repeated for 3 times.METHODS:Unilateral hindlimbs innervation of rats were severed. Four days later, skeletal muscle was dissected from denervated and contralateral control hindlimbs. Grude muscle membranes were prepared from red muscle (soleus and red gastrocnemius) and white gastrocnemius. Western Blotting and Northern Blotting analysis were employed to examine protein and mRNA expression of Na+-K+-ATPase subunit isoforms respectively.The intensity of protein and mRNA signals was quantitated with a BioRad GS-670 imaging densitometer.MAIN OUTCOME MEASURES: Levels of protein and mRNA of Na+-K+-ATPase subunit isoforms in control and denervated hindlimb skeletal muscles.RESULTS: Fifteen rats all entered the stage of results analysis. Compared with control, in denervated hindlimb ① the protein levels of α2-and β1-subunit in red muscle were reduced markedly by 46% and 77% respectively. The abundance of α1 protein was increased in red and white muscle by 20% and 15% respectively. There was no significant change in protein expression of β2-subunit in red and white gastrocnemius (WG). ② The mRNA level of α2-subunit decreased by 29% and 39% in red gastrocnemius (RG) and soleus, respectively, and that of β1-subunit decreased 80%and

  16. Protective effects of inhibition of adenosine monophosphate activated protein kinase activity against cerebral ischemia-reperfusion injury in mice

    补娟

    2013-01-01

    Objective To observe the effect of inhibition of adenosine monophosphate activated protein kinase (AMPK) on shape,function and inflammatory factor of microglia for mice after cerebral ischemia-reperfusion

  17. Adenosine inhibits neutrophil vascular endothelial growth factor release and transendothelial migration via A2B receptor activation.

    Wakai, A

    2012-02-03

    The effects of adenosine on neutrophil (polymorphonuclear neutrophils; PMN)-directed changes in vascular permeability are poorly characterized. This study investigated whether adenosine modulates activated PMN vascular endothelial growth factor (vascular permeability factor; VEGF) release and transendothelial migration. PMN activated with tumour necrosis factor-alpha (TNF-alpha, 10 ng\\/mL) were incubated with adenosine and its receptor-specific analogues. Culture supernatants were assayed for VEGF. PMN transendothelial migration across human umbilical vein endothelial cell (HUVEC) monolayers was assessed in vitro. Adhesion molecule receptor expression was assessed flow cytometrically. Adenosine and some of its receptor-specific analogues dose-dependently inhibited activated PMN VEGF release. The rank order of potency was consistent with the affinity profile of human A2B receptors. The inhibitory effect of adenosine was reversed by 3,7-dimethyl-1-propargylxanthine, an A2 receptor antagonist. Adenosine (100 microM) or the A2B receptor agonist 5\\'-N-ethylcarboxamidoadenosine (NECA, 100 microM) significantly reduced PMN transendothelial migration. However, expression of activated PMN beta2 integrins and HUVEC ICAM-1 were not significantly altered by adenosine or NECA. Adenosine attenuates human PMN VEGF release and transendothelial migration via the A2B receptor. This provides a novel target for the modulation of PMN-directed vascular hyperpermeability in conditions such as the capillary leak syndrome.

  18. Adenosine activates brown adipose tissue and recruits beige adipocytes via A2A receptors

    Gnad, Thorsten; Scheibler, Saskia; von Kügelgen, Ivar;

    2014-01-01

    Brown adipose tissue (BAT) is specialized in energy expenditure, making it a potential target for anti-obesity therapies. Following exposure to cold, BAT is activated by the sympathetic nervous system with concomitant release of catecholamines and activation of β-adrenergic receptors. Because BAT...... that adenosine-A2A signalling plays an unexpected physiological role in sympathetic BAT activation and protects mice from diet-induced obesity. Those findings reveal new possibilities for developing novel obesity therapies....

  19. Adenosine induces vasoconstriction through Gi-dependent activation of phospholipase C in isolated perfused afferent arterioles of mice

    Hansen, Pernille B; Castrop, Hayo; Briggs, Josie; Schnermann, Jurgen

    2003-01-01

    Adenosine induces vasoconstriction of renal afferent arterioles through activation of A1 adenosine receptors (A1AR). A1AR are directly coupled to Gi/Go, resulting in inhibition of adenylate cyclase, but the contribution of this signaling pathway to smooth muscle cell activation is unclear. In......-induced vasoconstriction was stable for up to 30 min and was most pronounced in the most distal part of the afferent arterioles. Adenosine did not cause vasoconstriction in arterioles from A1AR-/- mice. Pretreatment with pertussis toxin (PTX) (400 ng/ml) for 2 h blocked the vasoconstricting action of adenosine or N(6......) blocked the constriction responses to both adenosine and angiotensin II. In contrast, the adenylate cyclase inhibitor SQ22536 (10 micro M) and the protein kinase A antagonist KT5720 (0.1 and 1 micro M) did not induce significant vasoconstriction of afferent arterioles. It is concluded that the...

  20. Alteration of membrane phospholipid methylation by adenosine analogs does not affect T lymphocyte activation

    Membrane phospholipid methylation has been described during activation of various immune cells. Moreover recent data indicated modulation of immune cells functions by adenosine. As S-adenosyl-methionine and S-adenosyl-homocysteine are adenosine analogs and modulators of transmethylation reactions, the effects of SAH and SAM were investigated on membrane phospholipid methylation and lymphocyte activation. SAM was shown to induce the membrane phospholipid methylation as assessed by the 3Hmethyl-incorporation in membrane extract. This effect was inhibited by SAH. In contrast SAM and SAH did not affect the phytohemagglutinin-induced proliferative response of peripheral blood mononuclear cells. SAH neither modified the early internalization of membrane CD3 antigens nor did it prevent the late expression of HLA-DR antigens on lymphocytes activated by phytohemagglutinin. These results indicate that in vitro alteration of phospholipid methylation does not affect subsequent steps of human T lymphocyte activation and proliferation

  1. Platelet aggregation and serum adenosine deaminase (ADA) activity in pregnancy associated with diabetes, hypertension and HIV.

    Leal, Claudio A M; Leal, Daniela B R; Adefegha, Stephen A; Morsch, Vera M; da Silva, José E P; Rezer, João F P; Schrekker, Clarissa M L; Abdalla, Faida H; Schetinger, Maria R C

    2016-07-01

    Platelet aggregation and adenosine deaminase (ADA) activity were evaluated in pregnant women living with some disease conditions including hypertension, diabetes mellitus and human immunodeficiency virus infection. The subject population is consisted of 15 non-pregnant healthy women [control group (CG)], 15 women with normal pregnancy (NP), 7 women with hypertensive pregnancy (HP), 10 women with gestational diabetes mellitus (GDM) and 12 women with human immunodeficiency virus-infected pregnancy (HIP) groups. The aggregation of platelets was checked using an optical aggregometer, and serum ADA activity was determined using the colorimetric method. After the addition of 5 µM of agonist adenosine diphosphate, the percentage of platelet aggregation was significantly (p pregnancy and pregnancy-associated diseases suggest that platelet aggregation and ADA activity could serve as peripheral markers for the development of effective therapy in the maintenance of homeostasis and some inflammatory process in these pathophysiological conditions. Copyright © 2016 John Wiley & Sons, Ltd. PMID:27273565

  2. Reduced striatal ecto-nucleotidase activity in schizophrenia patients supports the "adenosine hypothesis".

    Aliagas, Elisabet; Villar-Menéndez, Izaskun; Sévigny, Jean; Roca, Mercedes; Romeu, Miriam; Ferrer, Isidre; Martín-Satué, Mireia; Barrachina, Marta

    2013-12-01

    Schizophrenia (SZ) is a major chronic neuropsychiatric disorder characterized by a hyperdopaminergic state. The hypoadenosinergic hypothesis proposes that reduced extracellular adenosine levels contribute to dopamine D2 receptor hyperactivity. ATP, through the action of ecto-nucleotidases, constitutes a main source of extracellular adenosine. In the present study, we examined the activity of ecto-nucleotidases (NTPDases, ecto-5'-nucleotidase, and alkaline phosphatase) in the postmortem putamen of SZ patients (n = 13) compared with aged-matched controls (n = 10). We firstly demonstrated, by means of artificial postmortem delay experiments, that ecto-nucleotidase activity in human brains was stable up to 24 h, indicating the reliability of this tissue for these enzyme determinations. Remarkably, NTPDase-attributable activity (both ATPase and ADPase) was found to be reduced in SZ patients, while ecto-5'-nucleotidase and alkaline phosphatase activity remained unchanged. In the present study, we also describe the localization of these ecto-enzymes in human putamen control samples, showing differential expression in blood vessels, neurons, and glial cells. In conclusion, reduced striatal NTPDase activity may contribute to the pathophysiology of SZ, and it represents a potential mechanism of adenosine signalling impairment in this illness. PMID:23771238

  3. Respiratory activity in medulla oblongata and its modulation by adenosine and opioids

    Herlenius, Eric

    1998-01-01

    From the moment of birth the complex neuronal networks generating breathing has to function continuously and adapt to the new postnatal environmental demands. This thesis aims at studying the perinatal development of respiratory control and its modulation by adenosine and opioids. Respiratory activity was studied in vitro using brainstem spinal cord preparations and in vivo with a barometric plethysmograph. In vitro whole-cell patch clamp recordings of respiratory related ne...

  4. Biosynthesis of bacterial glycogen: activator specificity of the adenosine diphosphate glucose pyrophosphorylases from the genus Rhodospirillum.

    Preiss, J; Greenberg, E

    1981-01-01

    The adenosine diphosphate (ADP) glucose pyrophosphorylases from Rhodospirillum fulvum, Rhodospirillum molischianum, and Rhodospirillum tenue were partially purified, and their kinetic properties were studied. The enzyme from the three organisms was found to be activated by pyruvate and thus was similar to the Rhodospirillum rubrum enzyme that had been previously studied (C. E. Furlong, and J. Preiss, J. Biol. Chem. 244:2539-2548, 1979). The enzymes from R. fulvum, R. molischianum, and R. tenu...

  5. Modulation of bladder function by luminal adenosine turnover and A1 receptor activation

    Prakasam, H. Sandeep; Herrington, Heather; Roppolo, James R.; Jackson, Edwin K.; Apodaca, Gerard

    2012-01-01

    The bladder uroepithelium transmits information to the underlying nervous and musculature systems, is under constant cyclical strain, expresses all four adenosine receptors (A1, A2A, A2B, and A3), and is a site of adenosine production. Although adenosine has a well-described protective effect in several organs, there is a lack of information about adenosine turnover in the uroepithelium or whether altering luminal adenosine concentrations impacts bladder function or overactivity. We observed ...

  6. Serum adenosine deaminase and its isoenzyme activities in pregnancy

    Göksel Bahadır

    2011-12-01

    Full Text Available Objective: ADA is widely distributed in human tissues, which may contribute to the maturation of the immunological system, especially the proliferation and differentiation of lymphoid cells, and seems to be critical at different stages of the maturation process. The activity of ADA changes in diseases characterized by the alteration of cell-mediated immunity. In this study we examined changes in serum total ADA activity and the patterns of two ADA isoenzymes, ADA-1 and ADA-2, in healthy pregnant women, and evaluated the possible role of the alteration of cell-mediated immunity during pregnancy as causes of changes in ADA activity. Materials and Methods: We measured serum activities of total ADA, ADA-1 and ADA-2 in healthy pregnant women (n=129 and age-matched healthy nonpregnant women (n=42. We divided the study group into three different subgroups: first trimester, second trimester and third trimester. Results: Serum ADA, ADA-1 and ADA-2 activities in healthy pregnant women were significantly lower than in nonpregnant women (p<0.001, p<0.001 and p<0.01 respectively. ADA (p<0.001 and ADA-2 (p<0.001 activities in the first trimester were significantly lower than in the control group. However, there were no significant differences between the first trimester and control group according to their ADA-1 activities (p=0.016. ADA (p<0.001, ADA-1 (p<0.001 and ADA-2 (p<0.008 activities in the second trimester were significantly lower than in the control group. Combined trisomy 21 risk, biochemical trisomy 21 risk, age risk and trisomy 18 + Nuchal translucency (NT risk were calculated using a first trimester screening test in 63 pregnant women. Furthermore, trisomy 21 risk, age risk and trisomy 18 risk were calculated by triple test in 52 pregnant women. ADA, ADA-1 and ADA-2 activities were not significantly correlated with risks in the first trimester screening test. ADA-1 activity was slightly significantly negative correlated with age risk (r= -0

  7. Activation of adenosine receptors and inhibition of cyclooxygenases: two recent pharmacological approaches to modulation of radiation suppressed hematopoiesis

    Searching for drugs conforming to requirements for protection and/or treatment of radiation-induced damage belongs to the most important tasks of current radiobiology. In the Laboratory of Experimental Hematology, Institute of Biophysics, v.v.i., Academy of Sciences of the Czech Republic, Brno, Czech Republic, two original approaches for stimulation of radiation-suppressed hematopoiesis have been tested in recent years, namely activation of adenosine receptors and inhibition of cyclooxygenases. Non-selective activation of adenosine receptors, induced by combined administration of dipyridamole, a drug preventing adenosine uptake and supporting thus its extracellular receptor-mediated action, and adenosine monophosphate, an adenosine prodrug, has been found to stimulate hematopoiesis when the drugs were given either pre- or post-irradiation. When synthetic adenosine receptor agonists selective for individual adenosine receptor subtypes were tested, stimulatory effects in myelosuppressed mice have been found after administration of IB-MECA, a selective adenosine A3 receptor agonist. Non-selective cyclooxygenase inhibitors, inhibiting both cyclooxygenase-1 (COX-1) and cyclooxygenase-2 (COX-2), indomethacin, diclofenac, or flurbiprofen, have been observed to act positively on radiation-perturbed hematopoiesis in sublethally irradiated mice. However, their undesirable gastrointestinal side effects have been found to negatively influence survival of lethally irradiated animals. Recently tested selective COX-2 inhibitor meloxicam, preserving protective action of COX-1-synthesized prostaglandins in the gastrointestinal tissues, has been observed to retain the hematopoiesis-stimulating effects of non-selective cyclooxygenase inhibitors and to improve the survival of animals exposed to lethal radiation doses. These findings bear evidence for the possibility to use selective adenosine A3 receptor agonists and selective COX-2 inhibitors in human practice for treatment of

  8. Adenosine triphosphate stimulates Aquifex aeolicus MutL endonuclease activity.

    Jerome Mauris

    Full Text Available BACKGROUND: Human PMS2 (hPMS2 homologues act to nick 5' and 3' to misincorporated nucleotides during mismatch repair in organisms that lack MutH. Mn(++ was previously found to stimulate the endonuclease activity of these homologues. ATP was required for the nicking activity of hPMS2 and yPMS1, but was reported to inhibit bacterial MutL proteins from Thermus thermophilus and Aquifex aeolicus that displayed homology to hPMS2. Mutational analysis has identified the DQHA(X(2E(X(4E motif present in the C-terminus of PMS2 homologues as important for endonuclease activity. METHODOLOGIES/PRINCIPAL FINDINGS: We examined the effect ATP had on the Mn(++ induced nicking of supercoiled pBR322 by full-length and mutant A. aeolicus MutL (Aae MutL proteins. Assays were single time point, enzyme titration experiments or reaction time courses. The maximum velocity for MutL nicking was determined to be 1.6+/-0.08x10(-5 s(-1 and 4.2+/-0.3x10(-5 s(-1 in the absence and presence of ATP, respectively. AMPPNP stimulated the nicking activity to a similar extent as ATP. A truncated Aae MutL protein composed of only the C-terminal 123 amino acid residues was found to nick supercoiled DNA. Furthermore, mutations in the conserved C-terminal DQHA(X(2E(X(4E and CPHGRP motifs were shown to abolish Aae MutL endonuclease activity. CONCLUSIONS: ATP stimulated the Mn(++ induced endonuclease activity of Aae MutL. Experiments utilizing AMPPNP implied that the stimulation did not require ATP hydrolysis. A mutation in the DQHA(X(2E(X(4E motif of Aae MutL further supported the role of this region in endonclease activity. For the first time, to our knowledge, we demonstrate that changing the histidine residue in the conserved CPHGRP motif abolishes endonucleolytic activity of a hPMS2 homologue. Finally, the C-terminal 123 amino acid residues of Aae MutL were sufficient to display Mn(++ induced nicking activity.

  9. Adenosine Triphosphate Stimulates Aquifex aeolicus MutL Endonuclease Activity

    Jerome Mauris; Thomas C Evans

    2009-01-01

    BACKGROUND: Human PMS2 (hPMS2) homologues act to nick 5' and 3' to misincorporated nucleotides during mismatch repair in organisms that lack MutH. Mn(++) was previously found to stimulate the endonuclease activity of these homologues. ATP was required for the nicking activity of hPMS2 and yPMS1, but was reported to inhibit bacterial MutL proteins from Thermus thermophilus and Aquifex aeolicus that displayed homology to hPMS2. Mutational analysis has identified the DQHA(X)(2)E(X)(4)E motif pre...

  10. Staurosporine-induced apoptosis in astrocytes is prevented by A1 adenosine receptor activation.

    D'Alimonte, Iolanda; Ballerini, Patrizia; Nargi, Eleonora; Buccella, Silvana; Giuliani, Patricia; Di Iorio, Patrizia; Caciagli, Francesco; Ciccarelli, Renata

    2007-05-11

    Astrocyte apoptosis occurs in acute and chronic pathological processes at the central nervous system and the prevention of astrocyte death may represent an efficacious intervention in protecting neurons against degeneration. Our research shows that rat astrocyte exposure to 100 nM staurosporine for 3h caused apoptotic death accompanied by caspase-3, p38 mitogen-ed protein kinase (MAPK) and glycogen synthase kinase-3beta (GSK3beta) activation. N(6)-chlorocyclopentyladenosine (CCPA, 2.5-75 nM), a selective agonist of A(1) adenosine receptors, added to the cultures 1h prior to staurosporine, induced a dose-dependent anti-apoptotic effect, which was inhibited by the A(1) receptor antagonist, 8-cyclopentyl-1,3-dipropylxanthine. CCPA also caused a dose- and time-dependent phosphorylation/activation of Akt, a downstream effector of cell survival promoting phosphatidylinositol 3-kinase (PI3K) pathway, which in turn led to inhibition of staurosporine-induced GSK3beta and p38 MAPK activity. Accordingly, the anti-apoptotic effect of CCPA was abolished by culture pre-treatment with LY294002, a selective PI3K inhibitor, pointing out the prevailing role played by PI3K pathway in the protective effect exerted by A(1) receptor activation. Since an abnormal p38 and GSK3beta activity is implicated in acute (stroke) and chronic (Alzheimer's disease) neurodegenerative diseases, the results of the present study provide a hint to better understand adenosine relevance in these disorders. PMID:17400382

  11. Activation of NTS A1 adenosine receptors inhibits regional sympathetic responses evoked by activation of cardiopulmonary chemoreflex

    Ichinose, Tomoko K.; Minic, Zeljka; Li, Cailian; O'Leary, Donal S.; Scislo, Tadeusz J.

    2012-01-01

    Previously we have shown that adenosine operating via the A1 receptor subtype may inhibit glutamatergic transmission in the baroreflex arc within the nucleus of the solitary tract (NTS) and differentially increase renal (RSNA), preganglionic adrenal (pre-ASNA), and lumbar (LSNA) sympathetic nerve activity (ASNA>RSNA≥LSNA). Since the cardiopulmonary chemoreflex and the arterial baroreflex are mediated via similar medullary pathways, and glutamate is a primary transmitter in both pathways, it i...

  12. Non-linear quantitative structure-activity relationship for adenine derivatives as competitive inhibitors of adenosine deaminase

    Logistic regression and artificial neural networks have been developed as two non-linear models to establish quantitative structure-activity relationships between structural descriptors and biochemical activity of adenosine based competitive inhibitors, toward adenosine deaminase. The training set included 24 compounds with known k i values. The models were trained to solve two-class problems. Unlike the previous work in which multiple linear regression was used, the highest of positive charge on the molecules was recognized to be in close relation with their inhibition activity, while the electric charge on atom N1 of adenosine was found to be a poor descriptor. Consequently, the previously developed equation was improved and the newly formed one could predict the class of 91.66% of compounds correctly. Also optimized 2-3-1 and 3-4-1 neural networks could increase this rate to 95.83%

  13. Serum adenosine deaminase activity and its isoenzyme in patients treated for tuberculosis

    Objective: Increased serum adenosine deaminase (ADA) activity, mainly associated with tuberculosis can also occur in a number of other diseases thus negatively affecting the diagnostic utility of ADA measurements in tuberculosis. The aim of the study was to determine whether or not the combined use of the activity of ADA, its isoenzymes and differential cell counts would provide a more efficient means of diagnosing tuberculosis than the use of ADA levels alone. Results: Data suggested significant (p0.75) of ADA/sub 2/ADA was found to be better indicator of tuberculosis. Lymphocyte neutrophil ratio (L/N)> 0.69 gave additional benefit to increase the sensitivity and specificity for the use of ADA as marker in diagnosing tuberculosis. Conclusion: The combined use of activity of ADA, its isoenzymes and total and differential cell counts is a better indicator and gives better understanding to diagnose and evaluate tuberculosis and response to therapy. (author)

  14. Cerebrospinal fluid adenosine deaminase activity: A complimentary tool in the early diagnosis of tuberculous meningitis

    Taori Girdhar M

    2006-03-01

    Full Text Available Abstract Background Tuberculous meningitis (TBM is the commonest form of neurotuberculosis caused by Mycobacterium tuberculosis bacilli (MTB. The diagnosis of TBM is often difficult. A reliable, cost-effective and rapid diagnostic test, which can be performed in any standard pathology laboratory, could be of help in the diagnosis of TBM. In the present study we measured the adenosine deaminase (ADA activity in cerebrospinal fluid (CSF of TBM and non-TBM patients. Method ADA activity in CSF was determined according to a method based on the Berthlot reaction, which is the formation of a colored indophenol complex from ammonia liberated from adenosine, and quantified spectrophotometrically. Results The CSF ADA activity from TBM patients was compared with CSF ADA from non-TBM infectious meningitis patients, and from patients with non-infectious neurological disorders. The mean CSF ADA activity was found to be significantly higher in CSF of TBM patients, 14.31 ± 3.87 (2.99–26.94, mean ± SD with range, than in the CSF from non-TBM infectious meningitis, 9.25 ± 2.14 (4.99–13.96 and from the non-infectious neurological disorders group, 2.71 ± 1.96 (0.00–7.68, P Conclusion This study demonstrated that ADA activity in the CSF of TBM patients, using a cut-off value 11.39 U/L/min, can be useful for the early differential diagnosis of TBM. This test can be performed in any pathology laboratory where more sophisticated methods are not available.

  15. Serum Adenosine deaminase activity and C-reactive protein levels in unstable angina

    Rani Surekha

    2003-01-01

    Full Text Available In unstable angina (USA patients, immunological responses contributing to inflammation play a vital role in plaque rupture and thrombosis causing stroke. In the present study an attempt is made to estimate the levels of adenosine deaminase activity, an immunoenzyme marker and C-reactive protein, a marker of inflammation in USA patients. 45 patients presenting USA and 50 age and sex matched healthy controls were included in the study. Serum ADA activity was measured spectrophotometrically at 630nm and serum C-reactive protein was detected using Avitex CRP kit, which is a rapid latex agglutination test. The Mean ADA levels were 41.15 ± 11.04 in patients and 20.71±5.63 in controls and 66.6% of patients and none of the controls were positive to CRP. The present study observed the importance of ADA as a serum marker in addition to CRP for assessing the immune response in USA patients.

  16. Activation of β1-adrenoceptor triggers oxidative stress mediated myocardial membrane destabilization in isoproterenol induced myocardial infarcted rats: 7-hydroxycoumarin and its counter action.

    Jagadeesh, Govindan Sangaran; Nagoor Meeran, Mohamed Fizur; Selvaraj, Palanisamy

    2016-04-15

    Activation of β1-adrenoceptor stimulates myocardial membrane destabilization in isoproterenol induced rats. Male albino Wistar rats were pre and co-treated with 7-hydroxycoumarin (16mg/kg body weight) daily for 8 days. Myocardial infarction was induced into rats by the subcutaneous administration of isoproterenol (100mg/kg body weight) at an interval of 24h daily for a period of two days (7th and 8th day). The levels/activities of serum cardiac troponin-T, lactate dehydrogenase and the concentrations of heart lipid peroxidation products were significantly increased and the antioxidant status was significantly decreased in isoproterenol induced rats. Furthermore, the activity of sodium/potassium-dependent adenosine triphosphatase was significantly decreased and the activities of calcium and magnesium-dependent adenosine triphosphatases were significantly increased in the heart of isoproterenol induced myocardial infarcted rats. Isoproterenol induced rats also revealed increased concentrations of sodium and calcium and decreased concentrations of potassium in the heart. 7-hydroxycoumarin pre- and co-treatment showed considerable impact on all biochemical parameters assessed. Also, 7-HC greatly reduced the infarct size of the myocardium. The in vitro study confirmed its potent free radical scavenging activity. Thus, the present study revealed that 7-HC attenuates myocardial membrane destabilization by reinstating the activities/levels of adenosine triphosphatases and minerals in isoproterenol induced rats by inhibiting oxidative stress. These effects are attributed to the membrane stabilizing and free radical scavenging properties of 7-hydroxycoumarin. PMID:26930228

  17. Endogenous activation of adenosine A1 receptors promotes post-ischemic electrocortical burst suppression

    Ilie, A; Ciocan, D; Constantinescu, A O; Zagrean, A-M; Nita, D A; Zagrean, L; Moldovan, M

    2009-01-01

    -vessel occlusion" model under chloral hydrate anesthesia. Quantification of BS recovery was carried out using BS ratio. During GCI full electrocortical suppression was attained (BS ratio reached 100%). During the following reperfusion the BS ratio returned to 0. The time course of the decay was exponential after 1...... and 5-min GCI and bi-exponential after 10-min GCI. The BS recovery was progressively delayed with the duration of ischemia. Administration of the A1R antagonist 1,3-dipropyl-8-cyclopentylxanthine (DPCPX, 1.25 mg/kg i.p.) accelerated the post-ischemic BS recovery for all GCI durations. Following the 10......-min GCI the effect of DPCPX was only apparent on the initial fast decay of the BS ratio. These data suggest that endogenous adenosine release promotes BS patterns during reperfusion following transient cerebral ischemia. Furthermore, the endogenous A1R activation may be the primary underlying cause of...

  18. Detergent inhibited, heat labile nucleoside triphosphatase in cores of avian myeloblastosis virus

    Jensen, Kaj Frank

    1978-01-01

    Endogenous DNA synthesis was studied in isolated core particles of avian myeloblastosis virus. It was found that cores contained an enzymatic activity which rapidly converted the added nucleoside triphosphates to diphosphates (but not further) at 0 degrees C, thus inhibiting DNA synthesis. This t...... triphosphatase probably originates from the viral membranes. In the cores the enzyme is completely inactivated by low concentrations (0.02%) of Nonident P-40. Also, the enzyme is very thermolabile and denatures rapidly at 38 degrees C....

  19. Activation of the A2A adenosine G-protein-coupled receptor by conformational selection.

    Ye, Libin; Van Eps, Ned; Zimmer, Marco; Ernst, Oliver P; Prosser, R Scott

    2016-05-12

    Conformational selection and induced fit are two prevailing mechanisms to explain the molecular basis for ligand-based activation of receptors. G-protein-coupled receptors are the largest class of cell surface receptors and are important drug targets. A molecular understanding of their activation mechanism is critical for drug discovery and design. However, direct evidence that addresses how agonist binding leads to the formation of an active receptor state is scarce. Here we use (19)F nuclear magnetic resonance to quantify the conformational landscape occupied by the adenosine A2A receptor (A2AR), a prototypical class A G-protein-coupled receptor. We find an ensemble of four states in equilibrium: (1) two inactive states in millisecond exchange, consistent with a formed (state S1) and a broken (state S2) salt bridge (known as 'ionic lock') between transmembrane helices 3 and 6; and (2) two active states, S3 and S3', as identified by binding of a G-protein-derived peptide. In contrast to a recent study of the β2-adrenergic receptor, the present approach allowed identification of a second active state for A2AR. Addition of inverse agonist (ZM241385) increases the population of the inactive states, while full agonists (UK432097 or NECA) stabilize the active state, S3', in a manner consistent with conformational selection. In contrast, partial agonist (LUF5834) and an allosteric modulator (HMA) exclusively increase the population of the S3 state. Thus, partial agonism is achieved here by conformational selection of a distinct active state which we predict will have compromised coupling to the G protein. Direct observation of the conformational equilibria of ligand-dependent G-protein-coupled receptor and deduction of the underlying mechanisms of receptor activation will have wide-reaching implications for our understanding of the function of G-protein-coupled receptor in health and disease. PMID:27144352

  20. Influence of metabotropic glutamate receptor agonists on the inhibitory effects of adenosine A1 receptor activation in the rat hippocampus

    de Mendonça, Alexandre; Ribeiro, J. A.

    1997-01-01

    Glutamate and other amino acids are the main excitatory neurotransmitters in many brain regions, including the hippocampus, by activating ion channel-coupled glutamate receptors, as well as metabotropic receptors linked to G proteins and second messenger systems. Several conditions which promote the release of glutamate, like frequency stimulation and hypoxia, also lead to an increase in the extracellular levels of the important neuromodulator, adenosine. We studied whether the activation of ...

  1. Adenosine can thwart antitumor immune responses elicited by radiotherapy. Therapeutic strategies alleviating protumor ADO activities

    By studying the bioenergetic status we could show that the development of tumor hypoxia is accompanied, apart from myriad other biologically relevant effects, by a substantial accumulation of adenosine (ADO). ADO has been shown to act as a strong immunosuppressive agent in tumors by modulating the innate and adaptive immune system. In contrast to ADO, standard radiotherapy (RT) can either stimulate or abrogate antitumor immune responses. Herein, we present ADO-mediated mechanisms that may thwart antitumor immune responses elicited by RT. An overview of the generation, accumulation, and ADO-related multifaceted inhibition of immune functions, contrasted with the antitumor immune effects of RT, is provided. Upon hypoxic stress, cancer cells release ATP into the extracellular space where nucleotides are converted into ADO by hypoxia-sensitive, membrane-bound ectoenzymes (CD39/CD73). ADO actions are mediated upon binding to surface receptors, mainly A2A receptors on tumor and immune cells. Receptor activation leads to a broad spectrum of strong immunosuppressive properties facilitating tumor escape from immune control. Mechanisms include (1) impaired activity of CD4 + T and CD8 + T, NK cells and dendritic cells (DC), decreased production of immuno-stimulatory lymphokines, and (2) activation of Treg cells, expansion of MDSCs, promotion of M2 macrophages, and increased activity of major immunosuppressive cytokines. In addition, ADO can directly stimulate tumor proliferation and angiogenesis. ADO mechanisms described can thwart antitumor immune responses elicited by RT. Therapeutic strategies alleviating tumor-promoting activities of ADO include respiratory hyperoxia or mild hyperthermia, inhibition of CD39/CD73 ectoenzymes or blockade of A2A receptors, and inhibition of ATP-release channels or ADO transporters. (orig.)

  2. Increased orbitofrontal brain activation after administration of a selective adenosine A2A antagonist in cocaine dependent subjects

    F. Gerard eMoeller

    2012-05-01

    Full Text Available Background: Positron Emission Tomography imaging studies provide evidence of reduced dopamine function in cocaine dependent subjects in the striatum, which is correlated with prefrontal cortical glucose metabolism, particularly in the orbitofrontal cortex. However, whether enhancement of dopamine in the striatum in cocaine dependent subjects would be associated with changes in prefrontal cortical brain activation is unknown. One novel class of medications that enhance dopamine function via heteromer formation with dopamine receptors in the striatum is the selective adenosine A2A receptor antagonists. This study sought to determine the effects administration of the selective adenosine A2A receptor antagonist SYN115 on brain function in cocaine dependent subjects. Methodology/Principle Findings: Twelve cocaine dependent subjects underwent two fMRI scans (one after a dose of placebo and one after a dose of 100 mg of SYN115 while performing a working memory task with 3 levels of difficulty (3, 5, and 7 digits. fMRI results showed that for 7-digit working memory activation there was significantly greater activation from SYN115 compared to placebo in portions of left (L lateral orbitofrontal cortex, L insula, and L superior and middle temporal pole. Conclusion/Significance: These findings are consistent with enhanced dopamine function in the striatum in cocaine dependent subjects via blockade of adenosine A2A receptors producing increased brain activation in the orbitofrontal cortex and other cortical regions. This suggests that at least some of the changes in brain activation in prefrontal cortical regions in cocaine dependent subjects may be related to altered striatal dopamine function, and that enhancement of dopamine function via adenosine A2A receptor blockade could be explored further for amelioration of neurobehavioral deficits associated with chronic cocaine use.

  3. Adenosine Monophosphate-Activated Protein Kinase (AMPK) as a Diverse Therapeutic Target: A Computational Perspective.

    Ramesh, M; Vepuri, Suresh B; Oosthuizen, Frasia; Soliman, Mahmoud E

    2016-02-01

    Adenosine monophosphate-activated protein kinase (AMPK) is viewed as a privileged therapeutic target for several diseases such as cancer, diabetes, inflammation, obesity, etc. In addition, AMPK has entered the limelight of current drug discovery with its evolution as a key metabolic regulator. AMPK also plays a key role in the maintenance of cellular energy homeostasis. Structurally, AMPK is a heterotrimeric protein, which consists of three protein subunits (α, β, and γ). The crystal structure of AMPK was solved, and several computational studies including homology modeling, molecular docking, molecular dynamics, and QSAR have been reported in order to explore the structure and function of this diverse therapeutic target. In this review, we present a comprehensive up-to-date overview on the computational and molecular modeling approaches that have been carried out on AMPK in order to understand its structure, function, dynamics, and its drug binding landscape. Information provided in this review would be of great interest to a wide pool of researchers involved in the design of new molecules against various diseases where AMPK plays a predominant role. Graphical Abstract ᅟ. PMID:26541160

  4. ADENOSINE DEAMINASE ACTIVITY AND SERUM C-REACTIVE PROTEIN AS PROGNOSTIC MARKERS OF CHAGAS DISEASE SEVERITY

    Iván Darío BRAVO-TOBAR

    2015-10-01

    Full Text Available SUMMARY Chagas disease is a public health problem worldwide. The availability of diagnostic tools to predict the development of chronic Chagas cardiomyopathy is crucial to reduce morbidity and mortality. Here we analyze the prognostic value of adenosine deaminase serum activity (ADA and C-reactive protein serum levels (CRP in chagasic individuals. One hundred and ten individuals, 28 healthy and 82 chagasic patients were divided according to disease severity in phase I (n = 35, II (n = 29, and III (n = 18. A complete medical history, 12-lead electrocardiogram, chest X-ray, and M-mode echocardiogram were performed on each individual. Diagnosis of Chagas disease was confirmed by ELISA and MABA using recombinant antigens; ADA was determined spectrophotometrically and CRP by ELISA. The results have shown that CRP and ADA increased linearly in relation to disease phase, CRP being significantly higher in phase III and ADA at all phases. Also, CRP and ADA were positively correlated with echocardiographic parameters of cardiac remodeling and with electrocardiographic abnormalities, and negatively with ejection fraction. CRP and ADA were higher in patients with cardiothoracic index ≥ 50%, while ADA was higher in patients with ventricular repolarization disturbances. Finally, CRP was positively correlated with ADA. In conclusion, ADA and CRP are prognostic markers of cardiac dysfunction and remodeling in Chagas disease.

  5. Purine metabolic enzymes in lymphocytes. III. Effects of immunosuppressants on adenosine deaminase and purine nucleoside phosphorylase activities

    Kurashige, Satonori; Akuzawa, Yuki; Yoshida, Toshiharu; Kodama, Kazue

    1983-01-01

    Mice were treated with a single injection of 6-mercaptopurine riboside (6MP-R), predonine or cyclophosphamide (CY), and the effects of these immunosuppressants on blastogenic responses to phytohemagglutinin P (PHA-P) or bacterial lipopoly saccharide (LPS) and on adenosine deaminase (ADA) or purine nucleoside phosphorylase (PNP) activities were studied with spleen lymphocytes. The retardation of blastogenic responses to both PHA-P and LPS were associated with the retardation of both intracellu...

  6. Drugs elevating extracellular adenosine administered in vivo induce serum colony-stimulating activity and interleukin-6 in mice

    Weiterová, Lenka; Hofer, Michal; Pospíšil, Milan; Znojil, V.; Štreitová, Denisa

    2007-01-01

    Roč. 56, č. 4 (2007), s. 463-473. ISSN 0862-8408 R&D Projects: GA ČR(CZ) GP305/03/D050 Institutional research plan: CEZ:AV0Z50040507; CEZ:AV0Z50040702 Keywords : extracellular adenosine * serum colony-stimulating activity * interleukin -6 Subject RIV: BO - Biophysics Impact factor: 1.505, year: 2007

  7. Recombinant Mouse PAP Has pH-Dependent Ectonucleotidase Activity and Acts through A1-Adenosine Receptors to Mediate Antinociception

    Sowa, Nathaniel A.; Kunjumon I. Vadakkan; Zylka, Mark J.

    2009-01-01

    Prostatic acid phosphatase (PAP) is expressed in nociceptive neurons and functions as an ectonucleotidase. When injected intraspinally, the secretory isoforms of human and bovine PAP protein have potent and long-lasting antinociceptive effects that are dependent on A1-adenosine receptor (A1R) activation. In this study, we purified the secretory isoform of mouse (m)PAP using the baculovirus expression system to determine if recombinant mPAP also had antinociceptive properties. We found that mP...

  8. Cardiovascular protection and antioxidant activity of the extracts from the mycelia of Cordyceps sinensis act partially via adenosine receptors.

    Yan, Xiao-Feng; Zhang, Zhong-Miao; Yao, Hong-Yi; Guan, Yan; Zhu, Jian-Ping; Zhang, Lin-Hui; Jia, Yong-Liang; Wang, Ru-Wei

    2013-11-01

    Mycelia of cultured Cordyceps sinensis (CS) is one of the most common substitutes for natural CS and was approved for arrhythmia in China. However, the role of CS in ameliorating injury during ischemia-reperfusion (I/R) is still unclear. We examined effects of extracts from CS on I/R and investigated the possible mechanisms. Post-ischemic coronary perfusion pressure, ventricular function, and coronary flow were measured using the Langendorff mouse heart model. Oxidative stress of cardiac homogenates was performed using an ELISA. Our results indicate that CS affords cardioprotection possibly through enhanced adenosine receptor activation. Cardioprotection was demonstrated by reduced post-ischemic diastolic dysfunction and improved recovery of pressure development and coronary flow. Treatment with CS largely abrogates oxidative stress and damage in glucose- or pyruvate-perfused hearts. Importantly, observed reductions in oxidative stress [glutathione disulfide (GSSG)]/[GSSG + glutathione] and [malondialdehyde (MDA)]/[superoxide dismutase + MDA] ratios as well as the resultant damage upon CS treatment correlate with functional markers of post-ischemic myocardial outcome. These effects of CS were partially blocked by 8-ρ-sulfophenyltheophylline, an adenosine receptor antagonist. Our results demonstrate a suppressive role of CS in ischemic contracture. Meanwhile, the results also suggest pre-ischemic adenosine receptor activation may be involved in reducing contracture in hearts pretreated with CS. PMID:23192916

  9. Evaluation of adenosine deaminase seric activity in the diagnosis of bovine tuberculosis

    Márcio Roberto Silva

    2006-06-01

    Full Text Available Determination of seric levels of adenosine deaminase (ADA, an enzyme produced by monocytes/macrophages and lymphocytes, has been used in the diagnosis of human tuberculosis (TB. In the present study, ADA seric activity was evaluated comparatively to the comparative tuberculin test in the diagnosis of bovine tuberculosis. Two hundred fifty-six cattle were classified by origin and by the comparative tuberculin test as TB-positive animals (n = 52, from herds where the Mycobacterium bovis had previously been isolated, and TB-negative animals (n = 204, TB-free herds. The mean ADA seric value from the TB-positive group (4.45 ± 2.33 U/L was significantly lower (p = 0.008 than that observed in sera from the TB-negative group (6.12 ± 4.47 U/L. When animals from a herd with clinical cases of enzootic bovine leukosis of TB-negative group were withdrawn from analysis, the mean ADA seric values of TB-negative group (5.12 ± 3.75 U/L was not significantly different anymore from that of the TB-positive group (p = 0.28. There was no agreement in the diagnosis of bovine TB between comparative tuberculin test and determination of ADA seric values, using two different cutoff points, being 6.12 U/L and 15.0 U/L, (kappa = -0.086 and kappa = -0.082, respectively. In conclusion, the determination of ADA seric activity was not a good auxiliary test for bovine TB, because it was not able to distinguish between TB-positive and TB-negative animals.

  10. A New Activity of Anti-HIV and Anti-tumor Protein GAP31: DNA Adenosine Glycosidase – Structural and Modeling Insight into its Functions

    Li, H.; Huang, P; Zhang, D; Sun, Y; Chen, H; Zhang, J; Huang, P; Kong, X; Lee-Huang, S

    2010-01-01

    We report here the high-resolution atomic structures of GAP31 crystallized in the presence of HIV-LTR DNA oligonucleotides systematically designed to examine the adenosine glycosidase activity of this anti-HIV and anti-tumor plant protein. Structural analysis and molecular modeling lead to several novel findings. First, adenine is bound at the active site in the crystal structures of GAP31 to HIV-LTR duplex DNA with 5' overhanging adenosine ends, such as the 3'-processed HIV-LTR DNA but not to DNA duplex with blunt ends. Second, the active site pocket of GAP31 is ideally suited to accommodate the 5' overhanging adenosine of the 3'-processed HIV-LTR DNA and the active site residues are positioned to perform the adenosine glycosidase activity. Third, GAP31 also removes the 5'-end adenine from single-stranded HIV-LTR DNA oligonucleotide as well as any exposed adenosine, including that of single nucleotide dAMP but not from AMP. Fourth, GAP31 does not de-purinate guanosine from di-nucleotide GT. These results suggest that GAP31 has DNA adenosine glycosidase activity against accessible adenosine. This activity is distinct from the generally known RNA N-glycosidase activity toward the 28S rRNA. It may be an alternative function that contributes to the antiviral and anti-tumor activities of GAP31. These results provide molecular insights consistent with the anti-HIV mechanisms of GAP31 in its inhibition on the integration of viral DNA into the host genome by HIV-integrase as well as irreversible topological relaxation of the supercoiled viral DNA.

  11. A new activity of anti-HIV and anti-tumor protein GAP31: DNA adenosine glycosidase - Structural and modeling insight into its functions

    Li, Hui-Guang [Department of Biochemistry, New York University School of Medicine, New York, NY 10016 (United States); Huang, Philip L. [American Biosciences, Boston, MA 02114 (United States); Zhang, Dawei; Sun, Yongtao [Department of Biochemistry, New York University School of Medicine, New York, NY 10016 (United States); Chen, Hao-Chia [Endocrinology and Reproduction Research Branch, National Institute of Child Health and Human Development, NIH, Bethesda, MD 20892 (United States); Zhang, John [Department of Chemistry, New York University, New York, NY 10003 (United States); Huang, Paul L. [Department of Medicine, Harvard Medical School and Massachusetts General Hospital, Boston, MA 02114 (United States); Kong, Xiang-Peng, E-mail: xiangpeng.kong@med.nyu.edu [Department of Biochemistry, New York University School of Medicine, New York, NY 10016 (United States); Lee-Huang, Sylvia, E-mail: sylvia.lee-huang@med.nyu.edu [Department of Biochemistry, New York University School of Medicine, New York, NY 10016 (United States)

    2010-01-01

    We report here the high-resolution atomic structures of GAP31 crystallized in the presence of HIV-LTR DNA oligonucleotides systematically designed to examine the adenosine glycosidase activity of this anti-HIV and anti-tumor plant protein. Structural analysis and molecular modeling lead to several novel findings. First, adenine is bound at the active site in the crystal structures of GAP31 to HIV-LTR duplex DNA with 5' overhanging adenosine ends, such as the 3'-processed HIV-LTR DNA but not to DNA duplex with blunt ends. Second, the active site pocket of GAP31 is ideally suited to accommodate the 5' overhanging adenosine of the 3'-processed HIV-LTR DNA and the active site residues are positioned to perform the adenosine glycosidase activity. Third, GAP31 also removes the 5'-end adenine from single-stranded HIV-LTR DNA oligonucleotide as well as any exposed adenosine, including that of single nucleotide dAMP but not from AMP. Fourth, GAP31 does not de-purinate guanosine from di-nucleotide GT. These results suggest that GAP31 has DNA adenosine glycosidase activity against accessible adenosine. This activity is distinct from the generally known RNA N-glycosidase activity toward the 28S rRNA. It may be an alternative function that contributes to the antiviral and anti-tumor activities of GAP31. These results provide molecular insights consistent with the anti-HIV mechanisms of GAP31 in its inhibition on the integration of viral DNA into the host genome by HIV-integrase as well as irreversible topological relaxation of the supercoiled viral DNA.

  12. A new activity of anti-HIV and anti-tumor protein GAP31: DNA adenosine glycosidase - Structural and modeling insight into its functions

    We report here the high-resolution atomic structures of GAP31 crystallized in the presence of HIV-LTR DNA oligonucleotides systematically designed to examine the adenosine glycosidase activity of this anti-HIV and anti-tumor plant protein. Structural analysis and molecular modeling lead to several novel findings. First, adenine is bound at the active site in the crystal structures of GAP31 to HIV-LTR duplex DNA with 5' overhanging adenosine ends, such as the 3'-processed HIV-LTR DNA but not to DNA duplex with blunt ends. Second, the active site pocket of GAP31 is ideally suited to accommodate the 5' overhanging adenosine of the 3'-processed HIV-LTR DNA and the active site residues are positioned to perform the adenosine glycosidase activity. Third, GAP31 also removes the 5'-end adenine from single-stranded HIV-LTR DNA oligonucleotide as well as any exposed adenosine, including that of single nucleotide dAMP but not from AMP. Fourth, GAP31 does not de-purinate guanosine from di-nucleotide GT. These results suggest that GAP31 has DNA adenosine glycosidase activity against accessible adenosine. This activity is distinct from the generally known RNA N-glycosidase activity toward the 28S rRNA. It may be an alternative function that contributes to the antiviral and anti-tumor activities of GAP31. These results provide molecular insights consistent with the anti-HIV mechanisms of GAP31 in its inhibition on the integration of viral DNA into the host genome by HIV-integrase as well as irreversible topological relaxation of the supercoiled viral DNA.

  13. Aqueous extract from taxus baccata inhibits adenosine deaminase activity significantly in cancerous and noncancerous human gastric and colon tissues

    Zahide Esra Durak

    2014-01-01

    Full Text Available Objective: To investigate possible effects of aqueous taxus baccata extract on adenosine deaminase (ADA activity in cancerous and noncancerous human tissues and to clarify mechanism(s of its anticancer potential. Materials and Methods: Cancerous and noncancerous human gastric and colon tissues were used in the study. The extracts were prepared in distilled water. Before and after treatment with the extracts, ADA activities in the tissue homogenates were measured. Results: ADA activity was found to be higher in gastric tissue compared with colon tissue, but no differences were found between ADA activities of cancerous and noncancerous tissues for both as well. In the plant extract studies, it was found that taxus extract significantly inhibited ADA activity both in cancerous and noncancerous gastric and colon tissues. Conclusion: Our results suggest that aqueous extract from taxus baccata inhibits ADA activities in both gastric and colon tissues significantly. It is suggested that in addition to other proposed mechanisms, accumulated adenosine due to the inhibition of ADA enzyme might also play part in the anticancer properties of taxus species.

  14. Hybrid integrated biological-solid-state system powered with adenosine triphosphate

    Roseman, Jared M.; Lin, Jianxun; Ramakrishnan, Siddharth; Rosenstein, Jacob K.; Shepard, Kenneth L.

    2015-12-01

    There is enormous potential in combining the capabilities of the biological and the solid state to create hybrid engineered systems. While there have been recent efforts to harness power from naturally occurring potentials in living systems in plants and animals to power complementary metal-oxide-semiconductor integrated circuits, here we report the first successful effort to isolate the energetics of an electrogenic ion pump in an engineered in vitro environment to power such an artificial system. An integrated circuit is powered by adenosine triphosphate through the action of Na+/K+ adenosine triphosphatases in an integrated in vitro lipid bilayer membrane. The ion pumps (active in the membrane at numbers exceeding 2 × 106 mm-2) are able to sustain a short-circuit current of 32.6 pA mm-2 and an open-circuit voltage of 78 mV, providing for a maximum power transfer of 1.27 pW mm-2 from a single bilayer. Two series-stacked bilayers provide a voltage sufficient to operate an integrated circuit with a conversion efficiency of chemical to electrical energy of 14.9%.

  15. Adenosine: front and center in linking nutrition and metabolism to neuronal activity

    Greene, Robert W

    2011-01-01

    Many individuals with epilepsy benefit from consuming a ketogenic diet, which is similar to the more commonly known Atkins diet. The underlying molecular reason for this has not been determined. However, in this issue of the JCI, Masino et al. have elucidated the mechanism responsible for the antiepileptic effects of the ketogenic diet in mice. The diet is shown to decrease expression of the enzyme adenosine kinase (Adk), which is responsible for clearing the endogenous antiepileptic agent ad...

  16. Adenosine can thwart antitumor immune responses elicited by radiotherapy. Therapeutic strategies alleviating protumor ADO activities

    Vaupel, Peter [Klinikum rechts der Isar, Technische Universitaet Muenchen (TUM), Department of Radiation Oncology, Munich (Germany); Multhoff, Gabriele [Klinikum rechts der Isar, Technische Universitaet Muenchen (TUM), Department of Radiation Oncology, Munich (Germany); Helmholtz Zentrum Muenchen, Institute for innovative Radiotherapy (iRT), Experimental Immune Biology, Neuherberg (Germany)

    2016-05-15

    By studying the bioenergetic status we could show that the development of tumor hypoxia is accompanied, apart from myriad other biologically relevant effects, by a substantial accumulation of adenosine (ADO). ADO has been shown to act as a strong immunosuppressive agent in tumors by modulating the innate and adaptive immune system. In contrast to ADO, standard radiotherapy (RT) can either stimulate or abrogate antitumor immune responses. Herein, we present ADO-mediated mechanisms that may thwart antitumor immune responses elicited by RT. An overview of the generation, accumulation, and ADO-related multifaceted inhibition of immune functions, contrasted with the antitumor immune effects of RT, is provided. Upon hypoxic stress, cancer cells release ATP into the extracellular space where nucleotides are converted into ADO by hypoxia-sensitive, membrane-bound ectoenzymes (CD39/CD73). ADO actions are mediated upon binding to surface receptors, mainly A2A receptors on tumor and immune cells. Receptor activation leads to a broad spectrum of strong immunosuppressive properties facilitating tumor escape from immune control. Mechanisms include (1) impaired activity of CD4 + T and CD8 + T, NK cells and dendritic cells (DC), decreased production of immuno-stimulatory lymphokines, and (2) activation of Treg cells, expansion of MDSCs, promotion of M2 macrophages, and increased activity of major immunosuppressive cytokines. In addition, ADO can directly stimulate tumor proliferation and angiogenesis. ADO mechanisms described can thwart antitumor immune responses elicited by RT. Therapeutic strategies alleviating tumor-promoting activities of ADO include respiratory hyperoxia or mild hyperthermia, inhibition of CD39/CD73 ectoenzymes or blockade of A2A receptors, and inhibition of ATP-release channels or ADO transporters. (orig.) [German] Untersuchungen des bioenergetischen Status ergaben, dass Tumorhypoxie neben vielen anderen bedeutsamen biologischen Effekten zu einem starken

  17. Hyperglycemia alters E-NTPDases, ecto-5'-nucleotidase, and ectosolic and cytosolic adenosine deaminase activities and expression from encephala of adult zebrafish (Danio rerio).

    Capiotti, Katiucia Marques; Siebel, Anna Maria; Kist, Luiza Wilges; Bogo, Maurício Reis; Bonan, Carla Denise; Da Silva, Rosane Souza

    2016-06-01

    Hyperglycemia is the main feature for the diagnosis of diabetes mellitus (DM). Some studies have demonstrated the relationship between DM and dysfunction on neurotransmission systems, such as the purinergic system. In this study, we evaluated the extracellular nucleotide hydrolysis and adenosine deamination activities from encephalic membranes of hyperglycemic zebrafish. A significant decrease in ATP, ADP, and AMP hydrolyses was observed at 111-mM glucose-treated group, which returned to normal levels after 7 days of glucose withdrawal. A significant increase in ecto-adenosine deaminase activity was observed in 111-mM glucose group, which remain elevated after 7 days of glucose withdrawal. The soluble-adenosine deaminase activity was significantly increased just after 7 days of glucose withdrawal. We also evaluated the gene expressions of ecto-nucleoside triphosphate diphosphohydrolases (E-NTPDases), ecto-5'-nucleotidase, ADA, and adenosine receptors from encephala of adult zebrafish. The entpd 2a.1, entpd 2a.2, entpd 3, and entpd 8 mRNA levels from encephala of adult zebrafish were decreased in 111-mM glucose-treated and glucose withdrawal groups. The gene expressions of adenosine receptors (adora 1 , adora 2aa , adora 2ab , and adora 2b ) were decreased in 111-mM glucose-treated and glucose withdrawal groups. The gene expression of ADA (ada 2a.1) was decreased in glucose withdrawal group. Maltodextrin, used as a control, did not affect the expression of adenosine receptors, ADA and E-NTPDases 2, 3, and 8, while the expression of ecto-5'-nucleotidase was slightly increased and the E-NTPDases 1 decreased. These findings demonstrated that hyperglycemia might affect the ecto-nucleotidase and adenosine deaminase activities and gene expression in zebrafish, probably through a mechanism involving the osmotic effect, suggesting that the modifications caused on purinergic system may also contribute to the diabetes-induced progressive cognitive impairment. PMID:26769247

  18. Peripheral Adenosine A3 Receptor Activation Causes Regulated Hypothermia in Mice That Is Dependent on Central Histamine H1 Receptors.

    Carlin, Jesse Lea; Tosh, Dilip K; Xiao, Cuiying; Piñol, Ramón A; Chen, Zhoumou; Salvemini, Daniela; Gavrilova, Oksana; Jacobson, Kenneth A; Reitman, Marc L

    2016-02-01

    Adenosine can induce hypothermia, as previously demonstrated for adenosine A1 receptor (A1AR) agonists. Here we use the potent, specific A3AR agonists MRS5698, MRS5841, and MRS5980 to show that adenosine also induces hypothermia via the A3AR. The hypothermic effect of A3AR agonists is independent of A1AR activation, as the effect was fully intact in mice lacking A1AR but abolished in mice lacking A3AR. A3AR agonist-induced hypothermia was attenuated by mast cell granule depletion, demonstrating that the A3AR hypothermia is mediated, at least in part, via mast cells. Central agonist dosing had no clear hypothermic effect, whereas peripheral dosing of a non-brain-penetrant agonist caused hypothermia, suggesting that peripheral A3AR-expressing cells drive the hypothermia. Mast cells release histamine, and blocking central histamine H1 (but not H2 or H4) receptors prevented the hypothermia. The hypothermia was preceded by hypometabolism and mice with hypothermia preferred a cooler environmental temperature, demonstrating that the hypothermic state is a coordinated physiologic response with a reduced body temperature set point. Importantly, hypothermia is not required for the analgesic effects of A3AR agonists, which occur with lower agonist doses. These results support a mechanistic model for hypothermia in which A3AR agonists act on peripheral mast cells, causing histamine release, which stimulates central histamine H1 receptors to induce hypothermia. This mechanism suggests that A3AR agonists will probably not be useful for clinical induction of hypothermia. PMID:26606937

  19. 钙离子ATP酶2a基因修饰骨髓间充质干细胞移植改善慢性心力衰竭大鼠的心功能%Enhancement of cardiac function of chronic heart failure rats by marrow stromal cell-based sarcoplasmic reticulum Ca2+ adenosine triphosphatase gene therapy

    郭豫涛; 李小鹰; 鲁小春; 吴迪; 姚克群; 陈平; 马康涛; 周春燕

    2008-01-01

    were given the interventions of SERCA2a gene, MSC transplantation, MSC+Ad/SERCa2a and empty adenoviral vector, respectively. MSCs were separated and cultured, and then Ad-SERCA2a-GFP was used to transfer MSC in the 3rd and 8th generations.MAIN OUTCOME MEASURES: Ad-SERCA2a-GFP transfection rate of MSC was measured by using flow cytometer. Before and at 14 and 21 days after treatment, cardiac function was evaluated by ultrasonic echocardiogram. Expression of cytokine Ⅷ was tested by immunohistochemical staining. SERCA2a gene and protein expression were evaluated by RT-PCR and Western blot respectively, as well as SERCA2a enzyme activity. RESULTS: ① Transfection rate: The infection efficiency of adenovirus-medicated gene into different passages of MSC was over 80%, and there was no difference between passage three (P3) MSC and P8 MSC (P > 0.05). ② Heart function: Left ventricle wall was thickened obviously in group MSC and group MSC+Ad/SERCa2a on the 21st day after treatment, while volume was shortened and gradually rounded. Compared to control group, ejection fraction (EF) and shortening fraction (FS) of group Ad-SERCa2a, group MSC and group MSC+Ad/SERCa2a were elevated significantly on the 14th day after therapy (P 0.05).②大鼠心功能:治疗后14 d,与腺病毒空载体对照组相比,其余3组左室射血分数均明显升高(P < 0.01).治疗后 21 d,与腺病毒空载体对照组相比,干细胞移植治疗组和基因修饰的干细胞移植组大鼠室壁增厚;干细胞移植治疗组和基因修饰的干细胞移植组左室射血分数和左室短轴缩短率改善率持续升高(P < 0.01),基因治疗组两指标改善率较治疗14 d时下降.与腺病毒空载体对照组相比,基因修饰的干细胞移植组左室前壁和室间隔收缩期纵向峰值速度显著升高(P < 0.01),左室前壁和室间隔舒张期纵向峰值速度亦呈现相同改善(P < 0.01).③心肌SERCA2a的基因、蛋白水平表达和功能活性,

  20. A1 adenosine receptor-induced phosphorylation and modulation of transglutaminase 2 activity in H9c2 cells: A role in cell survival.

    Vyas, Falguni S; Hargreaves, Alan J; Bonner, Philip L R; Boocock, David J; Coveney, Clare; Dickenson, John M

    2016-05-01

    The regulation of tissue transglutaminase (TG2) activity by the GPCR family is poorly understood. In this study, we investigated the modulation of TG2 activity by the A1 adenosine receptor in cardiomyocyte-like H9c2 cells. H9c2 cells were lysed following stimulation with the A1 adenosine receptor agonist N(6)-cyclopentyladenosine (CPA). Transglutaminase activity was determined using an amine incorporating and a protein cross linking assay. TG2 phosphorylation was assessed via immunoprecipitation and Western blotting. The role of TG2 in A1 adenosine receptor-induced cytoprotection was investigated by monitoring hypoxia-induced cell death. CPA induced time and concentration-dependent increases in amine incorporating and protein crosslinking activity of TG2. CPA-induced increases in TG2 activity were attenuated by the TG2 inhibitors Z-DON and R283. Responses to CPA were blocked by PKC (Ro 31-8220), MEK1/2 (PD 98059), p38 MAPK (SB 203580) and JNK1/2 (SP 600125) inhibitors and by removal of extracellular Ca(2+). CPA triggered robust increases in the levels of TG2-associated phosphoserine and phosphothreonine, which were attenuated by PKC, MEK1/2 and JNK1/2 inhibitors. Fluorescence microscopy revealed TG2-mediated biotin-X-cadaverine incorporation into proteins and proteomic analysis identified known (Histone H4) and novel (Hexokinase 1) protein substrates for TG2. CPA pre-treatment reversed hypoxia-induced LDH release and decreases in MTT reduction. TG2 inhibitors R283 and Z-DON attenuated A1 adenosine receptor-induced cytoprotection. TG2 activity was stimulated by the A1 adenosine receptor in H9c2 cells via a multi protein kinase dependent pathway. These results suggest a role for TG2 in A1 adenosine receptor-induced cytoprotection. PMID:27005940

  1. Endogenous activation of adenosine A(1) receptors accelerates ischemic suppression of spontaneous electrocortical activity

    Ilie, Andrei; Ciocan, Dragos; Zagrean, Ana-Maria;

    2006-01-01

    either 1.25 mg/kg DPCPX dissolved in 2 ml/kg dimethyl sulfoxide (DMSO) or the same volume of DMSO alone, 15 min before the third ischemic episode. Time to electrocortical suppression was estimated based on the decay of the root mean square of two-channel electrocorticographic recordings. During the first...... two ischemic episodes, electrocortical suppression appeared after approximately 12 s in both groups. After DMSO administration, ischemic suppression remained unchanged. After DPCPX administration, the time to electrocortical suppression was increased by approximately 10 s, and bursts of activity were...

  2. Comparative study of adenosine deaminase activity, insulin resistance and lipoprotein(a) among smokers and healthy non-smokers

    Ramesh Ramasamy; Sathish Babu Murugaiyan; Arulkumaran U.; Sathiya R.; Kuzhandai Velu V.; Niranjan Gopal

    2016-01-01

    Background: Adenosine deaminase also known as adenosine aminohydrolase involved in purine metabolism. Its primary function is development and maintenance of immune system. The main objective of the study was to estimate adenosine deaminase (ADA) enzyme and find its correlation with lipoprotein(a) and insulin resistance among smokers and healthy non-smokers. Methods: Fifty smokers and fifty healthy non-smokers were selected based on WHO definition. ADA, lipid profile and glucose was estimat...

  3. A2A adenosine receptor antagonism enhances synaptic and motor effects of cocaine via CB1 cannabinoid receptor activation.

    Alessandro Tozzi

    Full Text Available BACKGROUND: Cocaine increases the level of endogenous dopamine (DA in the striatum by blocking the DA transporter. Endogenous DA modulates glutamatergic inputs to striatal neurons and this modulation influences motor activity. Since D2 DA and A2A-adenosine receptors (A2A-Rs have antagonistic effects on striatal neurons, drugs targeting adenosine receptors such as caffeine-like compounds, could enhance psychomotor stimulant effects of cocaine. In this study, we analyzed the electrophysiological effects of cocaine and A2A-Rs antagonists in striatal slices and the motor effects produced by this pharmacological modulation in rodents. PRINCIPAL FINDINGS: Concomitant administration of cocaine and A2A-Rs antagonists reduced glutamatergic synaptic transmission in striatal spiny neurons while these drugs failed to produce this effect when given in isolation. This inhibitory effect was dependent on the activation of D2-like receptors and the release of endocannabinoids since it was prevented by L-sulpiride and reduced by a CB1 receptor antagonist. Combined application of cocaine and A2A-R antagonists also reduced the firing frequency of striatal cholinergic interneurons suggesting that changes in cholinergic tone might contribute to this synaptic modulation. Finally, A2A-Rs antagonists, in the presence of a sub-threshold dose of cocaine, enhanced locomotion and, in line with the electrophysiological experiments, this enhanced activity required activation of D2-like and CB1 receptors. CONCLUSIONS: The present study provides a possible synaptic mechanism explaining how caffeine-like compounds could enhance psychomotor stimulant effects of cocaine.

  4. Absence epileptic activity changing effects of non-adenosine nucleoside inosine, guanosine and uridine in Wistar Albino Glaxo Rijswijk rats.

    Kovács, Z; Kékesi, K A; Dobolyi, Á; Lakatos, R; Juhász, G

    2015-08-01

    Adenosine (Ado) and non-adenosine (non-Ado) nucleosides such as inosine (Ino), guanosine (Guo) and uridine (Urd) may have regionally different roles in the regulation of physiological and pathophysiological processes in the central nervous system (CNS) such as epilepsy. It was demonstrated previously that Ino and Guo decreased quinolinic acid (QA)-induced seizures and Urd reduced penicillin-, bicuculline- and pentylenetetrazole (PTZ)-induced seizures. It has also been demonstrated that Ino and Urd may exert their effects through GABAergic system by altering the function of GABA(A) type of gamma-aminobutyric acid receptors (GABAA receptors) whereas Guo decreases glutamate-induced excitability through glutamatergic system, which systems (GABAergic and glutamatergic) are involved in pathomechanisms of absence epilepsy. Thus, we hypothesized that Ino and Guo, similarly to the previously described effect of Urd, might also decrease absence epileptic activity. We investigated in the present study whether intraperitoneal (i.p.) application of Ino (500 and 1000mg/kg), Guo (20 and 50mg/kg), Urd (500 and 1000mg/kg), GABA(A) receptor agonist muscimol (1 and 3mg/kg), GABA(A) receptor antagonist bicuculline (2 and 4mg/kg), non-selective Ado receptor antagonist theophylline (5 and 10mg/kg) and non-competitive N-methyl-d-aspartate (NMDA) receptor antagonist (+)-5-methyl-10,11-dihydro-5H-dibenzo (a,d) cyclohepten-5,10-imine maleate (MK-801, 0.0625 and 0.1250mg/kg) alone and in combination have modulatory effects on absence epileptic activity in Wistar Albino Glaxo Rijswijk (WAG/Rij) rats. We found that Guo decreased the number of spike-wave discharges (SWDs) whereas Ino increased it dose-dependently. We strengthened that Urd can decrease absence epileptic activity. Our results suggest that Guo, Urd and their analogs could be potentially effective drugs for treatment of human absence epilepsy. PMID:26037802

  5. Adenosine and Sleep

    Bjorness, Theresa E.; Greene, Robert W.

    2009-01-01

    Over the last several decades the idea that adenosine (Ado) plays a role in sleep control was postulated due in large part to pharmacological studies that showed the ability of Ado agonists to induce sleep and Ado antagonists to decrease sleep. A second wave of research involving in vitro cellular analytic approaches and subsequently, the use of neurochemical tools such as microdialysis, identified a population of cells within the brainstem and basal forebrain arousal centers, with activity t...

  6. Adenosine-Associated Delivery Systems

    Kazemzadeh-Narbat, Mehdi; Annabi, Nasim; Tamayol, Ali; Oklu, Rahmi; Ghanem, Amyl; Khademhosseini, Ali

    2016-01-01

    Adenosine is a naturally occurring purine nucleoside in every cell. Many critical treatments such as modulating irregular heartbeat (arrhythmias), regulation of central nervous system (CNS) activity, and inhibiting seizural episodes can be carried out using adenosine. Despite the significant potential therapeutic impact of adenosine and its derivatives, the severe side effects caused by their systemic administration have significantly limited their clinical use. In addition, due to adenosine’s extremely short half-life in human blood (less than 10 s), there is an unmet need for sustained delivery systems to enhance efficacy and reduce side effects. In this paper, various adenosine delivery techniques, including encapsulation into biodegradable polymers, cell-based delivery, implantable biomaterials, and mechanical-based delivery systems, are critically reviewed and the existing challenges are highlighted. PMID:26453156

  7. Structure-Activity Analysis of Biased Agonism at the Human Adenosine A3 Receptor.

    Baltos, Jo-Anne; Paoletta, Silvia; Nguyen, Anh T N; Gregory, Karen J; Tosh, Dilip K; Christopoulos, Arthur; Jacobson, Kenneth A; May, Lauren T

    2016-07-01

    Biased agonism at G protein-coupled receptors (GPCRs) has significant implications for current drug discovery, but molecular determinants that govern ligand bias remain largely unknown. The adenosine A3 GPCR (A3AR) is a potential therapeutic target for various conditions, including cancer, inflammation, and ischemia, but for which biased agonism remains largely unexplored. We now report the generation of bias "fingerprints" for prototypical ribose containing A3AR agonists and rigidified (N)-methanocarba 5'-N-methyluronamide nucleoside derivatives with regard to their ability to mediate different signaling pathways. Relative to the reference prototypical agonist IB-MECA, (N)-methanocarba 5'-N-methyluronamide nucleoside derivatives with significant N(6) or C2 modifications, including elongated aryl-ethynyl groups, exhibited biased agonism. Significant positive correlation was observed between the C2 substituent length (in Å) and bias toward cell survival. Molecular modeling suggests that extended C2 substituents on (N)-methanocarba 5'-N-methyluronamide nucleosides promote a progressive outward shift of the A3AR transmembrane domain 2, which may contribute to the subset of A3AR conformations stabilized on biased agonist binding. PMID:27136943

  8. Activation of Adenosine Receptor A2A Increases HSC Proliferation and Inhibits Death and Senescence by Down-regulation of p53 and Rb

    Md. Kaimul eAhsan

    2014-04-01

    Full Text Available Background & Aims: During fibrosis hepatic stellate cells (HSC undergo activation, proliferation and senescence but the regulation of these important processes is poorly understood. The adenosine A2A receptor (A2A is known to be present on HSC, and its activation results in liver fibrosis. In this study, we tested if A2A has a role in the regulation of HSC proliferation, apoptosis, senescence, and the relevant molecular mechanism.Methods: The ability of adenosine to regulate p53 and Rb protein levels, proliferation, apoptosis and senescence was tested in the human HSC cell line LX-2 and rat primary HSC.Results: Adenosine receptor activation down-regulates p53 and Rb protein levels, increases BrdU incorporation and increases cell survival in LX-2 cells and in primary rat HSC. These effects of NECA were reproduced by an adenosine A2A receptor specific agonist (CGS21680 and blocked by a specific antagonist (ZM241385. By day twenty-one of culture primary rat HSC entered senescence and expressed -gal which was significantly inhibited by NECA. Furthermore, NECA induced down regulation of p53 and Rb and Rac1, and decreased phosphorylation of p44-42 MAP Kinase in LX-2 cells and primary rat HSC. These effects were reproduced by the cAMP analog 8-Bromo-cAMP, and the adenylyl cyclase activator forskolin, and were blocked by PKA inhibitors.Conclusions: These results demonstrate that A2A receptor regulates a number of HSC fate decisions and induces greater HSC proliferation, reduces apoptosis and senescence by decreasing p53 and Rb through cAMP-PKA/Rac1/p38 MAPK pathway. This provides a mechanism for adenosine induced HSC regulation and liver fibrosis.

  9. Influence of metabotropic glutamate receptor agonists on the inhibitory effects of adenosine A1 receptor activation in the rat hippocampus.

    de Mendonça, A; Ribeiro, J A

    1997-08-01

    1. Glutamate and other amino acids are the main excitatory neurotransmitters in many brain regions, including the hippocampus, by activating ion channel-coupled glutamate receptors, as well as metabotropic receptors linked to G proteins and second messenger systems. Several conditions which promote the release of glutamate, like frequency stimulation and hypoxia, also lead to an increase in the extracellular levels of the important neuromodulator, adenosine. We studied whether the activation of different subgroups of metabotropic glutamate receptors (mGluR) could modify the known inhibitory effects of a selective adenosine A1 receptor agonist on synaptic transmission in the hippocampus. The experiments were performed on hippocampal slices taken from young (12-14 days old) rats. Stimulation was delivered to the Schaffer collateral/commissural fibres, and evoked field excitatory postsynaptic potentials (fe. p.s.p.) recorded extracellularly from the stratum radiatum in the CAI area. 2. The concentration-response curve for the inhibitory effects of the selective adenosine A1 receptor agonist, N6-cyclopentyladenosine (CPA; 2-50 nM), on the fe.p.s.p. slope (EC50 = 12.5 (9.2-17.3; 95% confidence intervals)) was displaced to the right by the group I mGluR selective agonist, (R,S)-3,5-dihydroxyphenylglycine (DPHG; 10 microM) (EC50 = 27.2 (21.4-34.5) nM, n = 4). The attenuation of the inhibitory effect of CPA (10 nM) on the fe.p.s.p. slope by DHPG (10 microM) was blocked in the presence of the mGluR antagonist (which blocks group I and II mGluR), (R,S)-alpha-methyl-4-carboxyphenylglycine (MCPG; 500 microM). DHPG (10 microM) itself had an inhibitory effect of 20.1 +/- 1.9% (n = 4) on the fe.p.s.p. slope. 3. The concentration-response curves for the inhibitory effects of CPA (2-20 nM) on the fe.p.s.p. slope were not modified either in the presence of the group II mGluR selective agonist, (2S,3S,4S)-alpha-(carboxycyclopropyl)glycine (L-CCG-I; 1 microM), or in the presence of

  10. Dietary Supplementation of Ginger and Turmeric Rhizomes Modulates Platelets Ectonucleotidase and Adenosine Deaminase Activities in Normotensive and Hypertensive Rats.

    Akinyemi, Ayodele Jacob; Thomé, Gustavo Roberto; Morsch, Vera Maria; Bottari, Nathieli B; Baldissarelli, Jucimara; de Oliveira, Lizielle Souza; Goularte, Jeferson Ferraz; Belló-Klein, Adriane; Oboh, Ganiyu; Schetinger, Maria Rosa Chitolina

    2016-07-01

    Hypertension is associated with platelet alterations that could contribute to the development of cardiovascular complications. Several studies have reported antiplatelet aggregation properties of ginger (Zingiber officinale) and turmeric (Curcuma longa) with limited scientific basis. Hence, this study assessed the effect of dietary supplementation of these rhizomes on platelet ectonucleotidase and adenosine deaminase (ADA) activities in Nω-nitro-l-arginine methyl ester hydrochloride (l-NAME) induced hypertensive rats. Animals were divided into seven groups (n = 10): normotensive control rats; induced (l-NAME hypertensive) rats; hypertensive rats treated with atenolol (10 mg/kg/day); normotensive and hypertensive rats treated with 4% supplementation of turmeric or ginger, respectively. After 14 days of pre-treatment, the animals were induced with hypertension by oral administration of l-NAME (40 mg/kg/day). The results revealed a significant (p < 0.05) increase in platelet ADA activity and ATP hydrolysis with a concomitant decrease in ADP and AMP hydrolysis of l-NAME hypertensive rats when compared with the control. However, dietary supplementation with turmeric or ginger efficiently prevented these alterations by modulating the hydrolysis of ATP, ADP and AMP with a concomitant decrease in ADA activity. Thus, these activities could suggest some possible mechanism of the rhizomes against hypertension-derived complications associated to platelet hyperactivity. Copyright © 2016 John Wiley & Sons, Ltd. PMID:27151061

  11. Immobilized polyphosphate kinase: preparation, properties, and potential for use in adenosine 5'-triphosphate regeneration

    Polyphosphate kinase partially purified from Escherichia coli, has been immobilized on glutaraldehyde-activated aminoethyl cellulose with a 10% retention of enzymatic activity. The immobilized enzyme can carry out the synthesis of ATP from ADP, using long-chain inorganic polyphosphate as a phosphoryl donor. Chromatographic analyses of the product mixture produced from ADP and [32P]polyphosphate demonstrated that 98% of the 32P was incorporated into ATP, indicating that the immobilized polyphosphate kinase is substantially free from contaminating polyphosphate phosphohydrolase, adenosine triphosphatase, and adenylate kinase. Immobilized polyphosphate kinase loses no activity when stored in an aqueous suspension for 2 months at 5 degrees C or for 1-2 weeks at 25 degrees C. It may be stored indefinitely as a lyophilized powder at -10 degrees C. Michaelis constants for ADP and polyphosphate were determined to be 160 and 120 microM, respectively, for the immobilized enzyme. A small-batch reactor was found to produce ATP linearly with time up to 65% conversion of polyphosphate into ATP and to attain greater than 85% conversion to ATP at equilibrium. The ease of purification and immobilization of E. coli polyphosphate kinase, its storage stability, the purity and yield of its ATP product, and the low values of the Michaelis constants for its substrates make it a highly promising enzyme for ATP regeneration

  12. Plasmalemma- and tonoplast-ATPase activity in mesophyll protoplasts, vacuoles and microsomes of the Crassulacean-acid-metabolism plant Kalanchoe daigremontiana.

    Balsamo, R A; Uribe, E G

    1988-02-01

    Adenosine-triphosphatase activity on the plasmalemma and tonoplast of isolated mesophyll protoplasts, isolated vacuoles and tonoplast-derived microsomes of the Crassulacean-acid-metabolism plant Kalanchoe daigremontiana Hamet et Perr., was localized by a cytochemical procedure using lead citrate. Enzyme activity was detected on the cytoplasmic surfaces of the plasmalemma and tonoplast. The identity of the enzymes was confirmed by various treatments differentiating the enzymes by their sensitivity to inhibitors of plasmalemma and tonoplast H(+)-ATPase. Isolated vacuoles and microsomes prepared from isolated vacuoles clearly exhibited single-sided deposition on membrane surfaces. PMID:24226399

  13. Lower frequency of the low activity adenosine deaminase allelic variant (ADA1*2) in schizophrenic patients Diminuição da frequência da variante alélica de baixa atividade da adenosina desaminase (ADA1*2) em pacientes esquizofrênicos

    Gustavo Pimentel Dutra; Gustavo L. Ottoni; Lara, Diogo R; Maurício Reis Bogo

    2010-01-01

    OBJECTIVE: Adenosine may play a role in the pathophysiology of schizophrenia, since it modulates the release of several neurotransmitters such as glutamate, dopamine, serotonin and acetylcholine, decreases neuronal activity by pos-synaptic hyperpolarization and inhibits dopaminergic activity. Adenosine deaminase participates in purine metabolism by converting adenosine into inosine. The most frequent functional polymorphism of adenosine deaminase (22G→A) (ADA1*2) exhibits 20-30% lower e...

  14. Triiodothyronine causes rapid reversal of alpha 1/cyclic adenosine monophosphate synergism on brown adipocyte respiration and type II deiodinase activity.

    Noronha, M; Raasmaja, A; Moolten, N; Larsen, P R

    1991-12-01

    Previous studies have shown that thyroid status affects the response of brown adipose tissue (BAT) to the sympathetic nervous system. For example, hypothyroidism is associated with the development of a marked synergism between alpha 1- and beta-adrenergic pathways to stimulate type II iodothyronine 5'-deiodinase activity. Hypothyroidism also attenuates the respiratory response (thermogenesis) of isolated brown adipocytes to norepinephrine. To explore the interactions of the sympathetic nervous system and thyroid status in these cells, we compared the thermogenic and 5'-deiodinase responses to adrenergic agonists in isolated brown adipocytes from hypothyroid rats during treatment with 3,5,3'-triiodothyronine (T3). The fivefold synergism of alpha 1- and beta-adrenergic catecholamines to increase the deiodinase activity was progressively reduced, reaching a control euthyroid value of unity after 5 days of T3 treatment. Hypothyroidism reduced both the O2max (twofold to threefold) and increased the concentration of agonist required for 50% stimulation (10-fold) for both norepinephrine and forskolin. In hypothyroid cells, there was a twofold synergism between the alpha 1-agonist cirazoline and forskolin to increase respiration, which was blocked by prazosin and reproduced by the calcium ionophore, A23187. This synergistic effect of the alpha 1-agonist was lost within 2 days of T3 administration. These studies identify a second Ca(2+)-dependent intra-adrenergic synergism, which functions to ameliorate the reduced cyclic adenosine monophosphate (cAMP) responsiveness of the hypothyroid brown adipocyte. PMID:1683679

  15. Calcium and adenosine triphosphate control of cellular pathology: asparaginase-induced pancreatitis elicited via protease-activated receptor 2.

    Peng, Shuang; Gerasimenko, Julia V; Tsugorka, Tatiana; Gryshchenko, Oleksiy; Samarasinghe, Sujith; Petersen, Ole H; Gerasimenko, Oleg V

    2016-08-01

    Exocytotic secretion of digestive enzymes from pancreatic acinar cells is elicited by physiological cytosolic Ca(2+) signals, occurring as repetitive short-lasting spikes largely confined to the secretory granule region, that stimulate mitochondrial adenosine triphosphate (ATP) production. By contrast, sustained global cytosolic Ca(2+) elevations decrease ATP levels and cause necrosis, leading to the disease acute pancreatitis (AP). Toxic Ca(2+) signals can be evoked by products of alcohol and fatty acids as well as bile acids. Here, we have investigated the mechanism by which l-asparaginase evokes AP. Asparaginase is an essential element in the successful treatment of acute lymphoblastic leukaemia, the most common type of cancer affecting children, but AP is a side-effect occurring in about 5-10% of cases. Like other pancreatitis-inducing agents, asparaginase evoked intracellular Ca(2+) release followed by Ca(2+) entry and also substantially reduced Ca(2+) extrusion because of decreased intracellular ATP levels. The toxic Ca(2+) signals caused extensive necrosis. The asparaginase-induced pathology depended on protease-activated receptor 2 and its inhibition prevented the toxic Ca(2+) signals and necrosis. We tested the effects of inhibiting the Ca(2+) release-activated Ca(2+) entry by the Ca(2+) channel inhibitor GSK-7975A. This markedly reduced asparaginase-induced Ca(2+) entry and also protected effectively against the development of necrosis.This article is part of the themed issue 'Evolution brings Ca(2+) and ATP together to control life and death'. PMID:27377732

  16. Activation of the adenosine A2A receptor attenuates experimental autoimmune encephalomyelitis and is associated with increased intracellular calcium levels.

    Liu, Yumei; Zou, Haifeng; Zhao, Ping; Sun, Bo; Wang, Jinghua; Kong, Qingfei; Mu, Lili; Zhao, Sihan; Wang, Guangyou; Wang, Dandan; Zhang, Yao; Zhao, Jiaying; Yin, Pengqi; Liu, Lei; Zhao, Xiuli; Li, Hulun

    2016-08-25

    Multiple sclerosis (MS) is a common autoimmune disease that inevitably causes inflammatory nerve demyelination. However, an effective approach to prevent its course is still lacking and urgently needed. Recently, the adenosine A2A receptor (A2AR) has emerged as a novel inflammation regulator. Manipulation of A2AR activity may suppress the MS process and protect against nerve damage. To test this hypothesis, we treated murine experimental autoimmune encephalomyelitis (EAE), a model for MS, with the selective A2AR agonist, CGS21680 (CGS). We evaluated the effects of CGS on the pathological features of EAE progression, including CNS cellular infiltration, inflammatory cytokine expression, lymphocyte proliferation, and cell surface markers. Treatment with CGS significantly suppressed specific lymphocyte proliferation, reduced infiltration of CD4(+) T lymphocytes, and attenuated the expression of inflammatory cytokines, which in turn inhibited the EAE progression. For the first time, we demonstrate that CGS can increase the intracellular calcium concentration ([Ca(2+)]i) in murine lymphocytes, which may be the mechanism underlying the suppressive effects of CGS-induced A2AR activation on EAE progression. Our findings strongly suggest that A2AR is a potential therapeutic target for MS and provide insight into the mechanism of action of A2AR agonists, which may offer a therapeutic option for this disease. PMID:27217214

  17. Calcium and adenosine triphosphate control of cellular pathology: asparaginase-induced pancreatitis elicited via protease-activated receptor 2

    Peng, Shuang; Gerasimenko, Julia V.; Tsugorka, Tatiana; Gryshchenko, Oleksiy; Samarasinghe, Sujith; Gerasimenko, Oleg V.

    2016-01-01

    Exocytotic secretion of digestive enzymes from pancreatic acinar cells is elicited by physiological cytosolic Ca2+ signals, occurring as repetitive short-lasting spikes largely confined to the secretory granule region, that stimulate mitochondrial adenosine triphosphate (ATP) production. By contrast, sustained global cytosolic Ca2+ elevations decrease ATP levels and cause necrosis, leading to the disease acute pancreatitis (AP). Toxic Ca2+ signals can be evoked by products of alcohol and fatty acids as well as bile acids. Here, we have investigated the mechanism by which l-asparaginase evokes AP. Asparaginase is an essential element in the successful treatment of acute lymphoblastic leukaemia, the most common type of cancer affecting children, but AP is a side-effect occurring in about 5–10% of cases. Like other pancreatitis-inducing agents, asparaginase evoked intracellular Ca2+ release followed by Ca2+ entry and also substantially reduced Ca2+ extrusion because of decreased intracellular ATP levels. The toxic Ca2+ signals caused extensive necrosis. The asparaginase-induced pathology depended on protease-activated receptor 2 and its inhibition prevented the toxic Ca2+ signals and necrosis. We tested the effects of inhibiting the Ca2+ release-activated Ca2+ entry by the Ca2+ channel inhibitor GSK-7975A. This markedly reduced asparaginase-induced Ca2+ entry and also protected effectively against the development of necrosis. This article is part of the themed issue ‘Evolution brings Ca2+ and ATP together to control life and death’. PMID:27377732

  18. Enzymatic activity of granulations tissues under low doses of radiation. Biochemical analysis in rats

    This paper was designed to investigate in the rat subcutaneous sponge-induced granulation tissue under low doses of X-ray, the activity of alkaline phosphatase, 5'nucleotide phosphodiesterase and adenosine triphosphatase (ATPase) enzymes. One hundred and fourteen Wistar rats were divided into three groups, as follows: Group I as control, Group II that received single 7,14 R in split-dosis immediately after sponge-implantation at the third and fifth days postoperatively. Biopsies were taken after 7, 11, 14, 21 and 28 days and the activity of the three enzymes was determined. The results have shown that in Group II alkaline phosphatase had higher activity in the 14th day of tissue evolution when compared to Groups I and III . The 5'nucleotide phosphodiesterase activity in Group I was similar in all days checked, although in Group II the enzyme showed higher activity in 7th day and lower in 21st. In Group III the activity was higher after 14 and 7 days and lower after 28 and 21 days. There was no observation of changing in adenosine triphosphatase (ATPase) activity when the three groups were compared. (author)

  19. Optogenetic Activation of Adenosine A2A Receptor Signaling in the Dorsomedial Striatopallidal Neurons Suppresses Goal-Directed Behavior.

    Li, Yan; He, Yan; Chen, Mozi; Pu, Zhilan; Chen, Li; Li, Ping; Li, Bo; Li, Haiyan; Huang, Zhi-Li; Li, Zhihui; Chen, Jiang-Fan

    2016-03-01

    The striatum has an essential role in neural control of instrumental behaviors by reinforcement learning. Adenosine A(2A) receptors (A(2A)Rs) are highly enriched in the striatopallidal neurons and are implicated in instrumental behavior control. However, the temporal importance of the A(2A)R signaling in relation to the reward and specific contributions of the striatopallidal A(2A)Rs in the dorsolateral striatum (DLS) and the dorsomedial striatum (DMS) to the control of instrumental learning are not defined. Here, we addressed temporal relationship and sufficiency of transient activation of optoA(2A)R signaling precisely at the time of the reward to the control of instrumental learning, using our newly developed rhodopsin-A2AR chimeras (optoA(2A)R). We demonstrated that transient light activation of optoA(2A)R signaling in the striatopallidal neurons in 'time-locked' manner with the reward delivery (but not random optoA(2A)R activation) was sufficient to change the animal's sensitivity to outcome devaluation without affecting the acquisition or extinction phases of instrumental learning. We further demonstrated that optogenetic activation of striatopallidal A(2A)R signaling in the DMS suppressed goal-directed behaviors, as focally genetic knockdown of striatopallidal A(2A)Rs in the DMS enhanced goal-directed behavior by the devaluation test. By contrast, optogenetic activation or focal AAV-Cre-mediated knockdown of striatopallidal A(2A)R in the DLS had relatively limited effects on instrumental learning. Thus, the striatopallidal A(2A)R signaling in the DMS exerts inhibitory and predominant control of goal-directed behavior by acting precisely at the time of reward, and may represent a therapeutic target to reverse abnormal habit formation that is associated with compulsive obsessive disorder and drug addiction. PMID:26216520

  20. Amplification of neuromuscular transmission by methylprednisolone involves activation of presynaptic facilitatory adenosine A2A receptors and redistribution of synaptic vesicles.

    Oliveira, L; Costa, A C; Noronha-Matos, J B; Silva, I; Cavalcante, W L G; Timóteo, M A; Corrado, A P; Dal Belo, C A; Ambiel, C R; Alves-do-Prado, W; Correia-de-Sá, P

    2015-02-01

    The mechanisms underlying improvement of neuromuscular transmission deficits by glucocorticoids are still a matter of debate despite these compounds have been used for decades in the treatment of autoimmune myasthenic syndromes. Besides their immunosuppressive action, corticosteroids may directly facilitate transmitter release during high-frequency motor nerve activity. This effect coincides with the predominant adenosine A2A receptor tonus, which coordinates the interplay with other receptors (e.g. muscarinic) on motor nerve endings to sustain acetylcholine (ACh) release that is required to overcome tetanic neuromuscular depression in myasthenics. Using myographic recordings, measurements of evoked [(3)H]ACh release and real-time video microscopy with the FM4-64 fluorescent dye, results show that tonic activation of facilitatory A2A receptors by endogenous adenosine accumulated during 50 Hz bursts delivered to the rat phrenic nerve is essential for methylprednisolone (0.3 mM)-induced transmitter release facilitation, because its effect was prevented by the A2A receptor antagonist, ZM 241385 (10 nM). Concurrent activation of the positive feedback loop operated by pirenzepine-sensitive muscarinic M1 autoreceptors may also play a role, whereas the corticosteroid action is restrained by the activation of co-expressed inhibitory M2 and A1 receptors blocked by methoctramine (0.1 μM) and DPCPX (2.5 nM), respectively. Inhibition of FM4-64 loading (endocytosis) by methylprednisolone following a brief tetanic stimulus (50 Hz for 5 s) suggests that it may negatively modulate synaptic vesicle turnover, thus increasing the release probability of newly recycled vesicles. Interestingly, bulk endocytosis was rehabilitated when methylprednisolone was co-applied with ZM241385. Data suggest that amplification of neuromuscular transmission by methylprednisolone may involve activation of presynaptic facilitatory adenosine A2A receptors by endogenous adenosine leading to synaptic

  1. Campylobacter jejuni adenosine triphosphate phosphoribosyltransferase is an active hexamer that is allosterically controlled by the twisting of a regulatory tail.

    Mittelstädt, Gerd; Moggré, Gert-Jan; Panjikar, Santosh; Nazmi, Ali Reza; Parker, Emily J

    2016-08-01

    Adenosine triphosphate phosphoribosyltransferase (ATP-PRT) catalyzes the first committed step of the histidine biosynthesis in plants and microorganisms. Here, we present the functional and structural characterization of the ATP-PRT from the pathogenic ε-proteobacteria Campylobacter jejuni (CjeATP-PRT). This enzyme is a member of the long form (HisGL ) ATP-PRT and is allosterically inhibited by histidine, which binds to a remote regulatory domain, and competitively inhibited by AMP. In the crystalline form, CjeATP-PRT was found to adopt two distinctly different hexameric conformations, with an open homohexameric structure observed in the presence of substrate ATP, and a more compact closed form present when inhibitor histidine is bound. CjeATP-PRT was observed to adopt only a hexameric quaternary structure in solution, contradicting previous hypotheses favoring an allosteric mechanism driven by an oligomer equilibrium. Instead, this study supports the conclusion that the ATP-PRT long form hexamer is the active species; the tightening of this structure in response to remote histidine binding results in an inhibited enzyme. PMID:27191057

  2. Impact of aspirin dose on adenosine diphosphate-mediated platelet activities. Results of an in vitro pilot investigation.

    Tello-Montoliu, Antonio; Thano, Estela; Rollini, Fabiana; Patel, Ronakkumar; Wilson, Ryan E; Muñiz-Lozano, Ana; Franchi, Francesco; Darlington, Andrew; Desai, Bhaloo; Guzman, Luis A; Bass, Theodore A; Angiolillo, Dominick J

    2013-10-01

    Different aspirin dosing regimens have been suggested to impact outcomes when used in combination with adenosine diphosphate (ADP) P2Y12 receptor antagonists. Prior investigations have shown that not only aspirin, but also potent ADP P2Y12 receptor blockade can inhibit thromboxane A2-mediated platelet activation. The impact of aspirin dosing on ADP mediated platelet activities is unknown and represents the aim of this in vitro pilot pharmacodynamic (PD) investigation. Twenty-six patients with stable coronary artery disease on aspirin 81 mg/day and P2Y12 naïve were enrolled. PD assessments were performed at baseline, while patients were on 81 mg/day aspirin and after switching to 325 mg/day for 7 ± 2 days with and without escalating concentrations (vehicle, 1, 3, and 10 μM) of prasugrel's active metabolite (P-AM). PD assays included flow cytometric assessment of VASP to define the platelet reactivity index (PRI) and the Multiplate Analyzer (MEA) using multiple agonists [ADP, ADP + prostaglandin (PGE1), arachidonic acid (AA), and collagen]. Escalating P-AM concentrations showed incremental platelet P2Y12 inhibition measured by VASP-PRI (paspirin dosing regimen at any P-AM concentration (vehicle: p=0.899; 1 μM: p=0.888; 3 μM: p=0.524; 10 μM: p=0.548). Similar findings were observed in purinergic markers assessed by MEA (ADP and ADP+PGE1). P-AM addition significantly reduced AA and collagen induced platelet aggregation (paspirin dose. In conclusion, aspirin dosing does not appear to affect PD measures of ADP-mediated platelet reactivity irrespective of the degree of P2Y12 receptor blockade. P2Y12 receptor blockade modulates platelet reactivity mediated by alternative activators. PMID:23884248

  3. Comparison of the Effects of Adenosine A1 Receptors Activity in CA1 Region of the Hippocampus on Entorhinal Cortex and Amygdala Kindled Seizures in Rats

    A. Heidarianpour

    2008-10-01

    Full Text Available Introduction & Objective: In the CNS, adenosine is known to suppress repetitive neuronal Firing, suggesting a role as an endogenous modifier of seizures. Indeed, intracerebral adenosine concentrations rise acutely during seizure activity and are thought to be responsible for terminating seizures and establishing a period of post-ictal refractoriness. However, it is unclear whether this suppression results from a general depression of brain excitability or through action on particular sites critical for the control of after discharge generation and/or seizure development and propagation. In this regard, comparison of the effects of adenosine A1 receptors of CA1 (region of the ‎hippocampus on entorhinal cortex and amygdala kindled seizures was ‎investigated in this study. Materials & Methods: In this experimental study, Animals were kindled by daily electrical stimulation of amygdale (group A or entorhinal cortex (group B. In the fully kindled animals, N6-‎cyclohexyladenosine (CHA;1 and 10 M; a selective adenosine A1 receptor ‎agonist and 1,3-dimethyl-8-cyclohexylxanthine(CPT;1 ‎µ‎M; a selective ‎adenosine A1 receptors antagonist were microinfused bilaterally into the CA1 ‎region of hippocampus (1l/2min and animals were stimulated at 5 and 15 minutes after drug ‎injection. All animals were received artificial cerebrospinal fluid, 24 h before ‎each drug injection and this result were used as control. Results: The seizure parameters were measured at 5 and 15min post injection. Obtained data showed that CHA at concentrations of 10 ‎µ‎M reduced ‎entorhinal cortex and amygdala after discharge and stage5 seizure durations and ‎increased stage4 latency. CHA at concentration 1‎µ‎M significantly alters ‎seizure parameters of group A but not effect on group B. Intrahippocampal (CA1 region pretreatment of CPT (1 ‎µ‎M before CHA abolished the effects of CHA on seizure parameters.Conclusion: It ‎may be

  4. Effect of adenosine and adenosine analogs on [14C]aminopyrine accumulation by rabbit parietal cells

    Adenosine receptors that modulate adenylate cyclase activity have been identified recently in a number of tissues. Adenosine A2 receptor is stimulatory to adenylate cyclase, whereas adenosine A1 receptor is inhibitory to adenylate cyclase. We investigated the effect of adenosine and its analogs on [14C]aminopyrine accumulation by rabbit parietal cells. Rabbit gastric mucosal cells were isolated by enzyme digestion. Parietal cells were enriched by nonlinear percoll gradients. [14C]Aminopyrine accumulation was used as an indicator of acid secretion. The effect of 2-chloroadenosine on histamine-stimulated [14C]aminopyrine accumulation was studied. The effects of N-ethylcarboxamideadenosine, 2-chloroadenosine, stable analogs of adenosine, and adenosine on [14C]aminopyrine accumulation were assessed. Cyclic AMP content of parietal cells was determined by radioimmunoassay. Histamine and carbachol, known secretagogues, stimulated [14C]aminopyrine accumulation. 2-Chloroadenosine did not suppress histamine-stimulated [14C]aminopyrine accumulation. 2-Chloroadenosine, N-ethylcarboxamideadenosine, and adenosine dose dependently increased [14C]aminopyrine accumulation. The order of potency was N-ethylcarboxamideadenosine greater than 2-chloroadenosine greater than adenosine. 8-Phenyltheophylline and theophylline, adenosine-receptor antagonists, or cimetidine did not have significant effects on the increase of AP uptake induced by 2-chloroadenosine. Coadministration of dipyridamole, and adenosine uptake inhibitor, augmented the effect of adenosine on [14C]aminopyrine accumulation. 2-Chloroadenosine, N-ethylcarboxamideadenosine, and adenosine each induced a significant increase in cellular cyclic AMP. We conclude that there may be adenosine A2 receptors on rabbit parietal cells which modulate gastric acid secretion

  5. Adenosine A2A receptor activation reduces recurrence and mortality from Clostridium difficile infection in mice following vancomycin treatment

    Li Yuesheng

    2012-12-01

    Full Text Available Abstract Background Activation of the A2A adenosine receptor (A2AAR decreases production of inflammatory cytokines, prevents C. difficile toxin A-induced enteritis and, in combination with antibiotics, increases survival from sepsis in mice. We investigated whether A2AAR activation improves and A2AAR deletion worsens outcomes in a murine model of C. difficile (strain VPI10463 infection (CDI. Methods C57BL/6 mice were pretreated with an antibiotic cocktail prior to infection and then treated with vancomycin with or without an A2AAR agonist. A2AAR-/- and littermate wild-type (WT mice were similarly infected, and IFNγ and TNFα were measured at peak of and recovery from infection. Results Infected, untreated mice rapidly lost weight, developed diarrhea, and had mortality rates of 50-60%. Infected mice treated with vancomycin had less weight loss and diarrhea during antibiotic treatment but mortality increased to near 100% after discontinuation of antibiotics. Infected mice treated with both vancomycin and an A2AAR agonist, either ATL370 or ATL1222, had minimal weight loss and better long-term survival than mice treated with vancomycin alone. A2AAR KO mice were more susceptible than WT mice to death from CDI. Increases in cecal IFNγ and blood TNFα were pronounced in the absence of A2AARs. Conclusion In a murine model of CDI, vancomycin treatment resulted in reduced weight loss and diarrhea during acute infection, but high recurrence and late-onset death, with overall mortality being worse than untreated infected controls. The administration of vancomycin plus an A2AAR agonist reduced inflammation and improved survival rates, suggesting a possible benefit of A2AAR agonists in the management of CDI to prevent recurrent disease.

  6. Adenosine regulation of alveolar fluid clearance

    Factor, Phillip; Mutlu, Göskhan M.; Chen, Lan; Mohameed, Jameel; Akhmedov, Alexander T.; Meng, Fan Jing; Jilling, Tamas; Lewis, Erin Rachel; Johnson, Meshell D.; Xu, Anna; Kass, Daniel; Martino, Janice M.; Bellmeyer, Amy; Albazi, John S.; Emala, Charles

    2007-01-01

    Adenosine is a purine nucleoside that regulates cell function through G protein-coupled receptors that activate or inhibit adenylyl cyclase. Based on the understanding that cAMP regulates alveolar epithelial active Na+ transport, we hypothesized that adenosine and its receptors have the potential to regulate alveolar ion transport and airspace fluid content. Herein, we report that type 1 (A1R), 2a (A2aR), 2b (A2bR), and 3 (A3R) adenosine receptors are present in rat and mouse lungs and alveol...

  7. Adenosine: An immune modulator of inflammatory bowel diseases

    Jeff Huaqing Ye; Vazhaikkurichi M Rajendran

    2009-01-01

    Inflammatory bowel disease (IBD) is a common and lifelong disabling gastrointestinal disease. Emerging treatments are being developed to target inflammatory cytokines which initiate and perpetuate the immune response. Adenosine is an important modulator of inflammation and its anti-inflammatory effects have been well established in humans as well as in animal models. High extracellular adenosine suppresses and resolves chronic inflammation in IBD models. High extracellular adenosine levels could be achieved by enhanced adenosine absorption and increased de novo synthesis. Increased adenosine concentration leads to activation of the A2a receptor on the cell surface of immune and epithelial cells that would be a potential therapeutic target for chronic intestinal inflammation. Adenosine is transported via concentrative nucleoside transporter and equilibrative nucleoside transporter transporters that are localized in apical and basolateral membranes of intestinal epithelial cells, respectively. Increased extracellular adenosine levels activate the A2a receptor, which would reduce cytokines responsible for chronic inflammation.

  8. Targeting energy metabolic and oncogenic signaling pathways in triple-negative breast cancer by a novel adenosine monophosphate-activated protein kinase (AMPK) activator.

    Lee, Kuen-Haur; Hsu, En-Chi; Guh, Jih-Hwa; Yang, Hsiao-Ching; Wang, Dasheng; Kulp, Samuel K; Shapiro, Charles L; Chen, Ching-Shih

    2011-11-11

    The antitumor activities of the novel adenosine monophosphate-activated protein kinase (AMPK) activator, OSU-53, were assessed in in vitro and in vivo models of triple-negative breast cancer. OSU-53 directly stimulated recombinant AMPK kinase activity (EC(50), 0.3 μM) and inhibited the viability and clonogenic growth of MDA-MB-231 and MDA-MB-468 cells with equal potency (IC(50), 5 and 2 μM, respectively) despite lack of LKB1 expression in MDA-MB-231 cells. Nonmalignant MCF-10A cells, however, were unaffected. Beyond AMPK-mediated effects on mammalian target of rapamycin signaling and lipogenesis, OSU-53 also targeted multiple AMPK downstream pathways. Among these, the protein phosphatase 2A-dependent dephosphorylation of Akt is noteworthy because it circumvents the feedback activation of Akt that results from mammalian target of rapamycin inhibition. OSU-53 also modulated energy homeostasis by suppressing fatty acid biosynthesis and shifting the metabolism to oxidation by up-regulating the expression of key regulators of mitochondrial biogenesis, such as a peroxisome proliferator-activated receptor γ coactivator 1α and the transcription factor nuclear respiratory factor 1. Moreover, OSU-53 suppressed LPS-induced IL-6 production, thereby blocking subsequent Stat3 activation, and inhibited hypoxia-induced epithelial-mesenchymal transition in association with the silencing of hypoxia-inducible factor 1a and the E-cadherin repressor Snail. In MDA-MB-231 tumor-bearing mice, daily oral administration of OSU-53 (50 and 100 mg/kg) suppressed tumor growth by 47-49% and modulated relevant intratumoral biomarkers of drug activity. However, OSU-53 also induced protective autophagy that attenuated its antiproliferative potency. Accordingly, cotreatment with the autophagy inhibitor chloroquine increased the in vivo tumor-suppressive activity of OSU-53. OSU-53 is a potent, orally bioavailable AMPK activator that acts through a broad spectrum of antitumor activities. PMID

  9. Adenosine in exercise adaptation.

    Simpson, R E; Phillis, J. W.

    1992-01-01

    By influencing the regulation of the mechanisms of angiogenesis, erythropoietin production, blood flow, myocardial glucose uptake, glycogenolysis, systolic blood pressure, respiration, plasma norepinephrine and epinephrine levels, adenosine may exert a significant effect on the body's adaptation response to exercise. However, adenosine's possible influence over the vasodilatory response to exercise in skeletal muscle is controversial and more research is required to resolve this issue. Variou...

  10. Role of adenosine receptors in caffeine tolerance

    Caffeine is a competitive antagonist at adenosine receptors. Receptor up-regulation during chronic drug treatment has been proposed to be the mechanism of tolerance to the behavioral stimulant effects of caffeine. This study reassessed the role of adenosine receptors in caffeine tolerance. Separate groups of rats were given scheduled access to drinking bottles containing plain tap water or a 0.1% solution of caffeine. Daily drug intake averaged 60-75 mg/kg and resulted in complete tolerance to caffeine-induced stimulation of locomotor activity, which could not be surmounted by increasing the dose of caffeine. 5'-N-ethylcarboxamidoadenosine (0.001-1.0 mg/kg) dose dependently decreased the locomotor activity of caffeine-tolerant rats and their water-treated controls but was 8-fold more potent in the latter group. Caffeine (1.0-10 mg/kg) injected concurrently with 5-N-ethylcarboxamidoadenosine antagonized the decreases in locomotor activity comparably in both groups. Apparent pA2 values for tolerant and control rats also were comparable: 5.05 and 5.11. Thus, the adenosine-antagonist activity of caffeine was undiminished in tolerant rats. The effects of chronic caffeine administration on parameters of adenosine receptor binding and function were measured in cerebral cortex. There were no differences between brain tissue from control and caffeine-treated rats in number and affinity of adenosine binding sites or in receptor-mediated increases (A2 adenosine receptor) and decreases (A1 adenosine receptor) in cAMP accumulation. These results are consistent with theoretical arguments that changes in receptor density should not affect the potency of a competitive antagonist. Experimental evidence and theoretical considerations indicate that up-regulation of adenosine receptors is not the mechanism of tolerance to caffeine-induced stimulation of locomotor activity

  11. Effect of the growth stage and cultivar on policosanol profiles of barley sprouts and their adenosine 5'-monophosphate-activated protein kinase activation.

    Seo, Woo Duck; Yuk, Heung Joo; Curtis-Long, Marcus J; Jang, Ki Chang; Lee, Jin Hwan; Han, Sang-Ik; Kang, Hang Won; Nam, Min Hee; Lee, Sung-Joon; Lee, Ji Hae; Park, Ki Hun

    2013-02-01

    Adenosine 5'-monophosphate-activated protein kinase (AMPK) is an intracellular sensor that can regulate glucose levels within the cell. For this reason, it is well-known to be a target for drugs against diabetes and obesity. AMPK was activated significantly by the hexane extract of barley sprouts. This AMPK activation emerges across the growth stages of the sprout, becoming most significant (3 times above the initial stages) 10 days after sprouting. After this time, the activation decreased between 13 and 20 days post-sprouting. Analysis of the hexane extracts by gas chromatography-mass spectrometry showed that the amounts of policosanols (PCs, which are linear, primary aliphatic alcohols with 20-30 carbons) in the plant dramatically increased between 5 days (109.7 mg/100 g) and 10 days (343.7 mg/100 g) post-sprouting and then levels fell back down, reaching 76.4 mg/100 g at 20 days post-sprouting. This trend is consistent with PCs being the active ingredient in the barley plants. We validate this by showing that hexacosanol is an activator of AMPK. The richest cultivar for PCs was found to be the Daejin cultivar. Cultivars had a significant effect on the total PC content (113.2-183.5 mg/100 g) within the plant up to 5 days post-sprouting. However this dependence upon the cultivar was not so apparent at peak stages of PC production (10 days post-sprouting). The most abundant PC in barley sprout, hexacosanol, contributed 62-80% of the total PC content at every stage. These results are valuable to determine the optimal times of harvest to obtain the highest yield of PCs. PMID:23301834

  12. Aldehyde dehydrogenase type 2 activation by adenosine and histamine inhibits ischemic norepinephrine release in cardiac sympathetic neurons: mediation by protein kinase Cε.

    Robador, Pablo A; Seyedi, Nahid; Chan, Noel Yan-Ki; Koda, Kenichiro; Levi, Roberto

    2012-10-01

    During myocardial ischemia/reperfusion, lipid peroxidation leads to the formation of toxic aldehydes that contribute to ischemic dysfunction. Mitochondrial aldehyde dehydrogenase type 2 (ALDH2) alleviates ischemic heart damage and reperfusion arrhythmias via aldehyde detoxification. Because excessive norepinephrine release in the heart is a pivotal arrhythmogenic mechanism, we hypothesized that neuronal ALDH2 activation might diminish ischemic norepinephrine release. Incubation of cardiac sympathetic nerve endings with acetaldehyde, at concentrations achieved in myocardial ischemia, caused a concentration-dependent increase in norepinephrine release. A major increase in norepinephrine release also occurred when sympathetic nerve endings were incubated in hypoxic conditions. ALDH2 activation substantially reduced acetaldehyde- and hypoxia-induced norepinephrine release, an action prevented by inhibition of ALDH2 or protein kinase Cε (PKCε). Selective activation of G(i/o)-coupled adenosine A(1), A(3), or histamine H(3) receptors markedly inhibited both acetaldehyde- and hypoxia-induced norepinephrine release. These effects were also abolished by PKCε and/or ALDH2 inhibition. Moreover, A(1)-, A(3)-, or H(3)-receptor activation increased ALDH2 activity in a sympathetic neuron model (differentiated PC12 cells stably transfected with H(3) receptors). This action was prevented by the inhibition of PKCε and ALDH2. Our findings suggest the existence in sympathetic neurons of a protective pathway initiated by A(1)-, A(3)-, and H(3)-receptor activation by adenosine and histamine released in close proximity of these terminals. This pathway comprises the sequential activation of PKCε and ALDH2, culminating in aldehyde detoxification and inhibition of hypoxic norepinephrine release. Thus, pharmacological activation of PKCε and ALDH2 in cardiac sympathetic nerves may have significant protective effects by alleviating norepinephrine-induced life-threatening arrhythmias that

  13. A heterodimer of human 3'-phospho-adenosine-5'-phosphosulphate (PAPS) synthases is a new sulphate activating complex

    3'-Phospho-adenosine-5'-phosphosulphate (PAPS) synthases are fundamental to mammalian sulphate metabolism. These enzymes have recently been linked to a rising number of human diseases. Despite many studies, it is not yet understood how the mammalian PAPS synthases 1 and 2 interact with each other. We provide first evidence for heterodimerisation of these two enzymes by pull-down assays and Foerster resonance energy transfer (FRET) measurements. Kinetics of dimer dissociation/association indicates that these heterodimers form as soon as PAPSS1 and -S2 encounter each other in solution. Affinity of the homo- and heterodimers were found to be in the low nanomolar range using anisotropy measurements employing proteins labelled with the fluorescent dye IAEDANS that - in spite of its low quantum yield - is well suited for anisotropy due to its large Stokes shift. Within its kinase domain, the PAPS synthase heterodimer displays similar substrate inhibition by adenosine-5'-phosphosulphate (APS) as the homodimers. Due to divergent catalytic efficacies of PAPSS1 and -S2, the heterodimer might be a way of regulating PAPS synthase function within mammalian cells.

  14. Adenosine Receptors and Asthma

    Wilson, Constance N; Nadeem, Ahmed; Spina, Domenico; Brown, Rachel; Page, Clive P.; Jamal Mustafa, S.

    2009-01-01

    The pathophysiological processes underlying respiratory diseases like asthma are complex, resulting in an overwhelming choice of potential targets for the novel treatment of this disease. Despite this complexity, asthmatic subjects are uniquely sensitive to a range of substances like adenosine, thought to act indirectly to evoke changes in respiratory mechanics and in the underlying pathology, and thereby to offer novel insights into the pathophysiology of this disease. Adenosine is of partic...

  15. A CORRELATIVE STUDY OF ADENOSINE DEAMINASE ACTIVITY & T.B. IgG IN SERUM IN CASES OF TUBERCULOSIS

    Ajay

    2012-11-01

    Full Text Available ABSTRACT: INTRODUCTION: Tuberculosis is major cause of morbidity and mort ality in India as well in other parts of world. It is caused by myc obacterium tuberculosis which primarily affects lung and cause pulmonary tuberculosis. Diag nosis of tuberculosis rests upon a positive history of contact, clinical symptoms, x-ray chest, sputum positivity and AFB culture. Adenosine deaminase (ADA is an enzyme which catalyzes the de amination of adenosine into inosine and ammonia. ADA level is found to be elevated in tuber culosis and typhoid fever where cell mediated immunity is elevated. The ADA level is sig nificantly elevated in tuberculosis and helps to differentiate between tubercular and non tubercu lar diseases. The ADA level is also found to be elevated in serum and pleural fluid in patients of tubercular pleural effusion, tubercular ascitis and tubercular pericardial effusion. METHODS : Routine hemogram, Montoux test, X-ray chest, FNAC of lymph nodes, biopsy of lymph node whene ver required, estimation of serum ADA level and T.B.IgG studies were performed in each cas e. RESULTS: In the present study a total of 45 cases were selected for the study. There are 30 cases of pulmonary tuberculosis and 15 controls. The values of serum ADA and tubercular Ig G in pulmonary tubercular group are significantly higher as compared to those of control s. None of the control for ADA showed significant ratio of positivity (≥1.7. One of the 1 5 cases showed remarkable ratio of positivity (>1.2-1.6 and 14 (93.3% cases showed insignifican t ratio of positivity. Only 2 (13.33% of the 15 cases showed positivity for TB IgG and rest 13 (8 6.66% were regarded negative. CONCLUSIONS: Thus it can be concluded that determination of ser um adenosine deaminase levels can effectively diagnose tuberculosis with s ensitivity of 96.66% and specificity of 93.33% as compared to TBIgG showing sensitivity of 90% and specificity of 86.6%. Also cost of ADA estimation is remarkably

  16. Mixed Inhibition of Adenosine Deaminase Activity by 1,3-Dinitrobenzene: A Model for Understanding Cell-Selective Neurotoxicity in Chemically-Induced Energy Deprivation Syndromes in Brain

    Wang, Yipei; Liu, Xin; Schneider, Brandon; Zverina, Elaina A.; Russ, Kristen; Wijeyesakere, Sanjeeva J.; Fierke, Carol A.; Richardson., Rudy J.; Philbert, Martin A.

    2011-01-01

    Astrocytes are acutely sensitive to 1,3-dinitrobenzene (1,3-DNB) while adjacent neurons are relatively unaffected, consistent with other chemically-induced energy deprivation syndromes. Previous studies have investigated the role of astrocytes in protecting neurons from hypoxia and chemical injury via adenosine release. Adenosine is considered neuroprotective, but it is rapidly removed by extracellular deaminases such as adenosine deaminase (ADA). The present study tested the hypothesis that ...

  17. Running out of time: the decline of channel activity and nucleotide activation in adenosine triphosphate-sensitive K-channels.

    Proks, Peter; Puljung, Michael C; Vedovato, Natascia; Sachse, Gregor; Mulvaney, Rachel; Ashcroft, Frances M

    2016-08-01

    KATP channels act as key regulators of electrical excitability by coupling metabolic cues-mainly intracellular adenine nucleotide concentrations-to cellular potassium ion efflux. However, their study has been hindered by their rapid loss of activity in excised membrane patches (rundown), and by a second phenomenon, the decline of activation by Mg-nucleotides (DAMN). Degradation of PI(4,5)P2 and other phosphoinositides is the strongest candidate for the molecular cause of rundown. Broad evidence indicates that most other determinants of rundown (e.g. phosphorylation, intracellular calcium, channel mutations that affect rundown) also act by influencing KATP channel regulation by phosphoinositides. Unfortunately, experimental conditions that reproducibly prevent rundown have remained elusive, necessitating post hoc data compensation. Rundown is clearly distinct from DAMN. While the former is associated with pore-forming Kir6.2 subunits, DAMN is generally a slower process involving the regulatory sulfonylurea receptor (SUR) subunits. We speculate that it arises when SUR subunits enter non-physiological conformational states associated with the loss of SUR nucleotide-binding domain dimerization following prolonged exposure to nucleotide-free conditions. This review presents new information on both rundown and DAMN, summarizes our current understanding of these processes and considers their physiological roles.This article is part of the themed issue 'Evolution brings Ca(2+) and ATP together to control life and death'. PMID:27377720

  18. Attenuation of gastric mucosal inflammation induced by aspirin through activation of A2A adenosine receptor in rats

    Masaru Odashima; Reina Ohba; Sumio Watanabe; Joel Linden; Michiro Otaka; Mario Jin; Koga Komatsu; Isao Wada; Youhei Horikawa; Tamotsu Matsuhashi; Natsumi Hatakeyama; Jinko Oyake

    2006-01-01

    AIM: To determine whether a specific adenosine A2A receptor agonist (ATL-146e) can ameliorate aspirin-induced gastric mucosal lesions in rats, and reduce neutrophil accumulation and production of pro-inflammatory cytokines.METHODS: Gastric lesions were produced by oral gavage of aspirin (200 mg/kg) and HCl (0.15 mol/L,8.0 mL/kg). 4-{3-[6-Amino-9-(5-ethylcarbamoyl-3,4-dihydroxy-tetrahydro-furan-2-yl)-9H-purin-2-yl]-prop-2-ynyl}-cyclohexanecarboxylic acid methyl ester (ATL-146e,2.5-5 μg/kg, IP) was injected 30 min before the administration of aspirin. Tissue myeloperoxidase (MPO) concentration in gastric mucosa was measured as an index of neutrophil infiltration. Gastric mucosal concentrations of tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β) were determined by ELISA. Also, we examined the effect of ATL-146e on tissue prostaglandin E2 (PGE2) production and gastric secretion.RESULTS: Intragastric administration of aspirin induced multiple hemorrhagic erosions in rat gastric mucosa. The total length of gastric erosions (ulcer index) in control rats was 29.8±7.75 mm and was reduced to 3.8±1.42 mm after pretreatment with 5.0 g/kg ATL-146e (P< 0.01).The gastric contents of MPO and pro-inflammatory cytokines were all increased after the administration of aspirin and reduced to nearly normal levels by ATL-146e.Gastric mucosal PGE2 concentration was not affected by intraperitoneal injection of ATL-146e.CONCLUSION: The specific adenosine A2A receptor agohist, ATL-146e, has potent anti-ulcer effects presumably mediated by its anti-inflammatory properties.

  19. Effect of adenosine and adenosine analogs on ( sup 14 C)aminopyrine accumulation by rabbit parietal cells

    Ota, S.; Hiraishi, H.; Terano, A.; Mutoh, H.; Kurachi, Y.; Shimada, T.; Ivey, K.J.; Sugimoto, T. (Univ. of Tokyo (Japan))

    1989-12-01

    Adenosine receptors that modulate adenylate cyclase activity have been identified recently in a number of tissues. Adenosine A2 receptor is stimulatory to adenylate cyclase, whereas adenosine A1 receptor is inhibitory to adenylate cyclase. We investigated the effect of adenosine and its analogs on (14C)aminopyrine accumulation by rabbit parietal cells. Rabbit gastric mucosal cells were isolated by enzyme digestion. Parietal cells were enriched by nonlinear percoll gradients. (14C)Aminopyrine accumulation was used as an indicator of acid secretion. The effect of 2-chloroadenosine on histamine-stimulated (14C)aminopyrine accumulation was studied. The effects of N-ethylcarboxamideadenosine, 2-chloroadenosine, stable analogs of adenosine, and adenosine on (14C)aminopyrine accumulation were assessed. Cyclic AMP content of parietal cells was determined by radioimmunoassay. Histamine and carbachol, known secretagogues, stimulated (14C)aminopyrine accumulation. 2-Chloroadenosine did not suppress histamine-stimulated (14C)aminopyrine accumulation. 2-Chloroadenosine, N-ethylcarboxamideadenosine, and adenosine dose dependently increased (14C)aminopyrine accumulation. The order of potency was N-ethylcarboxamideadenosine greater than 2-chloroadenosine greater than adenosine. 8-Phenyltheophylline and theophylline, adenosine-receptor antagonists, or cimetidine did not have significant effects on the increase of AP uptake induced by 2-chloroadenosine. Coadministration of dipyridamole, and adenosine uptake inhibitor, augmented the effect of adenosine on (14C)aminopyrine accumulation. 2-Chloroadenosine, N-ethylcarboxamideadenosine, and adenosine each induced a significant increase in cellular cyclic AMP. We conclude that there may be adenosine A2 receptors on rabbit parietal cells which modulate gastric acid secretion.

  20. Selective detection of adenosine A1 receptor-dependent G-protein activity in basal and stimulated conditions of rat brain [35S]guanosine 5minutes or feet-(γ-thio)triphosphate autoradiography

    [35S]Guanosine 5minutes or feet-(γ-thio)triphosphate autoradiography is a novel technique to detect receptor-dependent activation of G-proteins in brain tissue sections. While an increasing number of reports using this approach are beginning to appear, little effort has been directed to the identification of factors responsible for the heterogeneously distributed [35S]guanosine 5minutes or feet-(γ-thio)triphosphate signal in basal conditions. The present study demonstrates that endogenously formed adenosine generates a widespread and prominent adenosine A1 receptor-dependent signal in basal conditions using this technique. Treatment of rat brain tissue sections with the A1-selective antagonist 8-cyclopentyl-1,3-dipropylxanthine dose-dependently (ec5035S]guanosine 5minutes or feet-(γ-thio)triphosphate binding in a region-specific manner, an effect fully mimicked by the adenosine-depleting enzyme adenosine deaminase, and less so by the A1 antagonist cirsimarin and by caffeine. That adenosine was continuously formed during the incubation is supported by the constant requirements of adenosine deaminase in order to suppress basal radioligand binding and further by the fact that low micromolar concentrations of adenine nucleotides evoked only adenosine-mimicking and fully 8-cyclopentyl-1,3-dipropylxanthine-sensitive binding responses. In the presence of adenosine deaminase, all responses to adenine nucleotides were abolished, indicating that prior conversion to adenosine was required. Upon stimulation, this technique selectively detected A1 receptor-activated G-proteins, as the non-selective agonists adenosine and 2-chloroadenosine and the A1-selective agonist N6-p-sulfophenyladenosine all evoked only 8-cyclopentyl-1,3-dipropylxanthine-sensitive responses in identical gray matter areas, and also in several white matter areas such as the corpus callosum, anterior commissure, optic tract and cerebellar white matter. Dose-response studies revealed region

  1. Pretreatment with adenosine and adenosine A1 receptor agonist protects against intestinal ischemia-reperfusion injury in rat

    V Haktan Ozacmak; Hale Sayan

    2007-01-01

    AIM: To examine the effects of adenosine and A1 receptor activation on reperfusion-induced small intestinal injury.METHODS: Rats were randomized into groups with sham operation, ischemia and reperfusion, and systemic treatments with either adenosine or 2-chloro-N6-cyclopentyladenosine, A1 receptor agonist or 8-cyclopentyl-1,3-dipropylxanthine, A1 receptor antagonist, plus adenosine before ischemia. Following reperfusion, contractions of ileum segments in response to KCl, carbachol and substance P were recorded. Tissue myeloperoxidase,malondialdehyde, and reduced glutathione levels were measured.RESULTS: Ischemia significantly decreased both contraction and reduced glutathione level which were ameliorated by adenosine and agonist administration. Treatment also decreased neutrophil infiltration and membrane lipid peroxidation. Beneficial effects of adenosine were abolished by pretreatment with A1 receptor antagonist.CONCLUSION: The data suggest that adenosine and A1 receptor stimulation attenuate ischemic intestinal injury via decreasing oxidative stress, lowering neutrophil infiltration, and increasing reduced glutathione content.

  2. A Metabolic Immune Checkpoint: Adenosine in Tumor Microenvironment

    Ohta, Akio

    2016-01-01

    Within tumors, some areas are less oxygenated than others. Since their home ground is under chronic hypoxia, tumor cells adapt to this condition by activating aerobic glycolysis; however, this hypoxic environment is very harsh for incoming immune cells. Deprivation of oxygen limits availability of energy sources and induces accumulation of extracellular adenosine in tumors. Extracellular adenosine, upon binding with adenosine receptors on the surface of various immune cells, suppresses pro-inflammatory activities. In addition, signaling through adenosine receptors upregulates a number of anti-inflammatory molecules and immunoregulatory cells, leading to the establishment of a long-lasting immunosuppressive environment. Thus, due to hypoxia and adenosine, tumors can discourage antitumor immune responses no matter how the response was induced, whether it was spontaneous or artificially introduced with a therapeutic intention. Preclinical studies have shown the significance of adenosine in tumor survival strategy by demonstrating tumor regression after inactivation of adenosine receptors, inhibition of adenosine-producing enzymes, or reversal of tissue hypoxia. These promising results indicate a potential use of the inhibitors of the hypoxia–adenosine pathway for cancer immunotherapy. PMID:27066002

  3. Adenosine stress protocols for myocardial perfusion imaging

    Baškot Branislav

    2008-01-01

    Full Text Available Background/Aim. Treadmill test combined with myocardial perfusion scintigraphy (MPS is a commonly used technique in the assessment of coronary artery disease. There are many patients, however, who may not be able to undergo treadmill test. Such patients would benefit from pharmacological stress procedures combined with MPS. The most commonly used pharmacological agents for cardiac stress are coronary vasodilatators (adenosine, dipyridamol and catecholamines. Concomitant low-level treadmill exercise with adenosine pharmacologic stress (AdenoEX during MPS has become commonly used in recent years. A number of studies have demonstrated a beneficial impact of AdenoEX protocol. The aim of the study was, besides introducing into practice the two types of protocols of pharmatological stress test with adenosine, as a preparation for MPS, to compare and monitor the frequency of their side effects to quality, acquisition, as well as to standardize the onset time of acquisition (diagnostic imaging for both protocols. Methods. A total of 130 patients underwent pharmacological stress test with adenosine (vasodilatator. In 108 of the patients we performed concomitant exercise (AdenoEX of low level (50W by a bicycle ergometar. In 28 of the patients we performed Adenosine abbreviated protocol (AdenoSCAN. Side effects of adenosine were followed and compared between the two kinds of protocols AdenoEX and AdenoSCAN. Also compared were image quality and suggested time of acquisition after the stress test. Results. Numerous side effects were found, but being short-lived they did not require any active interventions. The benefit of AdenoEX versus AdenoSCAN included decreased side effects (62% vs 87%, improved safety and patients tolerance, improved target-to-background ratios because of less subdiaphragmatic activity, earlier acquisition, and improved sensitivity. Conclusion. The safety and efficacy of adenosine pharmacological stress is even better with concomitant

  4. Turnover of adenosine in plasma of human and dog blood

    To determine half-life and turnover of plasma adenosine, heparinized blood from healthy volunteers was incubated with radiolabeled adenosine in the physiological concentration range of 0.1-1 microM. Plasma levels of adenosine in vitro were 82 +/- 14 nM and were similar to those determined immediately after blood collection with a ''stopping solution.'' Dipyridamole (83 microM) and erythro-9(2-hydroxynon-3yl)-adenine (EHNA) (8 microM) did not measurably alter basal adenosine levels but completely blocked the uptake of added adenosine. Inhibition of ecto-5'-nucleotidase with 100 microM alpha, beta-methyleneadenosine 5'-diphosphate (AOPCP) reduced plasma adenosine to 22 +/- 6 nM. For the determination of adenosine turnover, the decrease in specific radioactivity of added [3H]adenosine was measured using a dipyridamole-containing stopping solution. Without altering basal adenosine levels, the half-life was estimated to be 0.6 s. Similar experiments were carried out with washed erythrocytes or in the presence of AOPCP, yielding half-lives of 0.7 and 0.9 s, respectively. When the initial adenosine concentration was 1 microM, its specific activity decreased by only 11% within 5 s, whereas total plasma adenosine exponentially decreased with a half-life of 1.5 s. Venous plasma concentrations were measured after relief of a 3-min forearm ischemia. Changes in plasma adenosine did not correlate well with changes in blood flow but were augmented in the presence of dipyridamole

  5. Optogenetic activation of intracellular adenosine A2A receptor signaling in the hippocampus is sufficient to trigger CREB phosphorylation and impair memory.

    Li, P; Rial, D; Canas, P M; Yoo, J-H; Li, W; Zhou, X; Wang, Y; van Westen, G J P; Payen, M-P; Augusto, E; Gonçalves, N; Tomé, A R; Li, Z; Wu, Z; Hou, X; Zhou, Y; IJzerman, A P; PIJzerman, Ad; Boyden, E S; Cunha, R A; Qu, J; Chen, J-F

    2015-11-01

    Human and animal studies have converged to suggest that caffeine consumption prevents memory deficits in aging and Alzheimer's disease through the antagonism of adenosine A2A receptors (A2ARs). To test if A2AR activation in the hippocampus is actually sufficient to impair memory function and to begin elucidating the intracellular pathways operated by A2AR, we have developed a chimeric rhodopsin-A2AR protein (optoA2AR), which retains the extracellular and transmembrane domains of rhodopsin (conferring light responsiveness and eliminating adenosine-binding pockets) fused to the intracellular loop of A2AR to confer specific A2AR signaling. The specificity of the optoA2AR signaling was confirmed by light-induced selective enhancement of cAMP and phospho-mitogen-activated protein kinase (p-MAPK) (but not cGMP) levels in human embryonic kidney 293 (HEK293) cells, which was abolished by a point mutation at the C terminal of A2AR. Supporting its physiological relevance, optoA2AR activation and the A2AR agonist CGS21680 produced similar activation of cAMP and p-MAPK signaling in HEK293 cells, of p-MAPK in the nucleus accumbens and of c-Fos/phosphorylated-CREB (p-CREB) in the hippocampus, and similarly enhanced long-term potentiation in the hippocampus. Remarkably, optoA2AR activation triggered a preferential p-CREB signaling in the hippocampus and impaired spatial memory performance, while optoA2AR activation in the nucleus accumbens triggered MAPK signaling and modulated locomotor activity. This shows that the recruitment of intracellular A2AR signaling in the hippocampus is sufficient to trigger memory dysfunction. Furthermore, the demonstration that the biased A2AR signaling and functions depend on intracellular A2AR loops prompts the possibility of targeting the intracellular A2AR-interacting partners to selectively control different neuropsychiatric behaviors. PMID:25687775

  6. Mechanism of protection of adenosine from sulphate radical anion and repair of adenosine radicals by caffeic acid in aqueous solution

    M Sudha Swaraga; L Charitha; M Adinarayana

    2005-07-01

    The photooxidation of adenosine in presence of peroxydisulphate (PDS) has been studied by spectrophotometrically measuring the absorbance of adenosine at 260 nm. The rates of oxidation of adenosine by sulphate radical anion have been determined in the presence of different concentrations of caffeic acid. Increase in [caffeic acid] is found to decrease the rate of oxidation of adenosine suggesting that caffeic acid acts as an efficient scavenger of $SO_{4}^{\\bullet-}$ and protects adenosine from it. Sulphate radical anion competes for adenosine as well as for caffeic acid. The quantum yields of photooxidation of adenosine have been calculated from the rates of oxidation of adenosine and the light intensity absorbed by PDS at 254 nm, the wavelength at which PDS is activated to sulphate radical anion. From the results of experimentally determined quantum yields (exptl) and the quantum yields calculated (cal) assuming caffeic acid acting only as a scavenger of $SO_{4}^{\\bullet-}$ show that exptl values are lower than cal values. The ' values, which are experimentally found quantum yield values at each caffeic acid concentration and corrected for $SO_{4}^{\\bullet-}$ scavenging by caffeic acid, are also found to be greater than exptl values. These observations suggest that the transient adenosine radicals are repaired by caffeic acid in addition to scavenging of sulphate radical anions.

  7. Activity of cholinesterases, pyruvate kinase and adenosine deaminase in rats experimentally infected by Fasciola hepatica: Influences of these enzymes on inflammatory response and pathological findings.

    Baldissera, Matheus D; Bottari, Nathieli B; Mendes, Ricardo E; Schwertz, Claiton I; Lucca, Neuber J; Dalenogare, Diessica; Bochi, Guilherme V; Moresco, Rafael N; Morsch, Vera M; Schetinger, Maria R C; Rech, Virginia C; Jaques, Jeandre A; Da Silva, Aleksandro S

    2015-11-01

    The aim of this study was to investigate acetylcholinesterase (AChE) in total blood and liver tissue; butyrylcholinesterase (BChE) in serum and liver tissue; adenosine deaminase (ADA) in serum and liver tissue; and pyruvate kinase (PK) in liver tissue of rats experimentally infected by Fasciola hepatica. Animals were divided into two groups with 12 animals each, as follows: group A (uninfected) and group B (infected). Samples were collected at 20 (A1 and B1;n=6 each) and 150 (A2 and B2; n=6 each) days post-infection (PI). Infected animals showed an increase in AChE activity in whole blood and a decrease in AChE activity in liver homogenates (P<0.05) at 20 and 150 days PI. BChE and PK activities were decreased (P<0.05) in serum and liver homogenates of infected animals at 150 days PI. ADA activity was decreased in serum at 20 and 150 days PI, while in liver homogenates it was only decreased at 150 days PI (P<0.05). Aspartate aminotransferase and alanine aminotransferase activities in serum were increased (P<0.05), while concentrations of total protein and albumin were decreased (P<0.05) when compared to control. The histological analysis revealed fibrous perihepatitis and necrosis. Therefore, we conclude that the liver fluke is associated with cholinergic and purinergic dysfunctions, which in turn may influence the pathogenesis of the disease. PMID:26452485

  8. Effect of theophylline on adenosine production in the canine myocardium

    Adenosine is thought to participate in local regulation of coronary blood flow. However, competitive antagonists of adenosine fail to block myocardial active hyperemia. The authors examined the effect of locally administered theophylline on active hyperemia and myocardial adenosine production during intracoronary isoproterenol infusion in the dog heart. Isoproterenol decreased coronary resistance and increased myocardial adenosine production. Infusion of theophylline at a rate that attenuated the vasodilator response to exogenously administered adenosine failed to attenuate the increase in coronary blood flow produced by isoproterenol. However, theophylline plus isoproterenol production greater increases in myocardial adensine production than isoproterenol alone. The curves relating resistance and adenosine in the presence of theophylline fell to the right of those in the absence of theophylline. These findings suggest that the failure of theophylline to attenuate isoproterenol hyperemia in the dog heart results at least in part from an increase in adenosine concentration at the arteriole to a level beyond that blocked by this competitive antagonist and that adenosine may in fact play a role in isoproterenol-induced active hyperemia

  9. Adenosine and sleep

    Behavioral and biochemical approaches have been used to determine the relative contribution of endogenous adenosine and adenosine receptors to the sleep-wake cycle in the rat. Adenosine concentrations in specific areas of the rat brain were not affected by 24 hours of total sleep deprivation, or by 24 or 48 hours of REM sleep deprivation. In order to assess the effect of REM sleep deprivation on adenosine A1 receptors, 3H-L-PIA binding was measured. The Bmax values for 3H-L-PIA binding to membrane preparations of the cortices and corpus striata from 48 hour REM sleep-deprived animals were increased 14.8% and 23%, respectively. These increases were not maintained following the cessation of sleep deprivation and recovered within 2 hours. The results of a 96 hour REM deprivation experiment were similar to those of the 48 hour REM sleep deprivation experiment. However, these increases were not evident in similar structures taken from stress control animals, and conclusively demonstrated that the changes in 3H-L-PIA binding resulted from REM sleep deprivation and not from stress

  10. Adenosine and sleep

    Yanik, G.M. Jr.

    1987-01-01

    Behavioral and biochemical approaches have been used to determine the relative contribution of endogenous adenosine and adenosine receptors to the sleep-wake cycle in the rat. Adenosine concentrations in specific areas of the rat brain were not affected by 24 hours of total sleep deprivation, or by 24 or 48 hours of REM sleep deprivation. In order to assess the effect of REM sleep deprivation on adenosine A/sub 1/ receptors, /sup 3/H-L-PIA binding was measured. The Bmax values for /sup 3/H-L-PIA binding to membrane preparations of the cortices and corpus striata from 48 hour REM sleep-deprived animals were increased 14.8% and 23%, respectively. These increases were not maintained following the cessation of sleep deprivation and recovered within 2 hours. The results of a 96 hour REM deprivation experiment were similar to those of the 48 hour REM sleep deprivation experiment. However, these increases were not evident in similar structures taken from stress control animals, and conclusively demonstrated that the changes in /sup 3/H-L-PIA binding resulted from REM sleep deprivation and not from stress.

  11. Ectonucleotidases and adenosine deaminase activity in laying hens naturally infected by Salmonella Gallinarum and their effects on the pathogenesis of the disease.

    Boiago, Marcel M; Baldissera, Matheus D; Doleski, Pedro H; Bottari, Nathieli B; do Carmo, Guilherme M; Araujo, Denise N; Giuriatti, Jessica; Baggio, Vanessa; Leal, Daniela B R; Casagrande, Renata A; Wisser, Cláudia S; Stefani, Lenita M; da Silva, Aleksandro S

    2016-04-01

    Salmonella Gallinarum is the etiologic agent of fowl typhoid that affects chickens and turkeys causing egg production drops, infertility, lower hatchability, high mortality, and as a consequence severe economic losses to the poultry industry. The alterations in NTPDase and adenosine deaminase (ADA) activities have been demonstrated in several inflammatory conditions; however, there are no data in the literature associated with this infection. Thus, the aim of this study was to evaluate the activities of NTPDase, 5'nucleotidase, and ADA in serum and hepatic tissue of laying hens naturally infected by Salmonella Gallinarum. Liver and serum samples were collected of 27 laying hens (20 S. Gallinarum infected and 7 uninfected). NTPDase and 5'-nucleotidase activities in serum were increased (P laying hens naturally infected by S. Gallinarum; as well as increased (P laying hens. Histopathological analyses revealed that S. Gallinarum caused fibrinoid necrosis in liver and spleen associated with infiltrates of heterophils, macrophages, lymphocytes, and plasma cells. Considering that NTPDase and ADA are involved in the cell-mediated immunity, this study suggests that activities of these enzymes could be important biomarkers to determine the severity of inflammatory and immune responses in salmonellosis, contributing to clarify the pathogenesis of the disease. PMID:26911648

  12. Cytotoxic purine nucleoside analogues bind to A1, A2A and A3 adenosine receptors

    Jensen, Kyle; Johnson, L’Aurelle A.; Jacobson, Pamala A.; Kachler, Sonja; Kirstein, Mark N.; Lamba, Jatinder; Klotz, Karl-Norbert

    2012-01-01

    Fludarabine, clofarabine and cladribine are anti-cancer agents which are analogues of the purine nucleoside adenosine. These agents have been associated with cardiac and neurological toxicities. Because these agents are analogues of adenosine, they may act through adenosine receptors to elicit their toxic effects. The objective of this study was to evaluate the ability of cytotoxic nucleoside analogues to bind and activate adenosine receptor subtypes (A1, A2A, A2B, and A3). Radioligand bindin...

  13. Adenosine as a signaling molecule in the retina: biochemical and developmental aspects

    ROBERTO PAES-DE-CARVALHO

    2002-01-01

    The nucleoside adenosine plays an important role as a neurotransmitter or neuromodulator in the central nervous system, including the retina. In the present paper we review compelling evidence showing that adenosine is a signaling molecule in the developing retina. In the chick retina, adenosine transporters are present since early stages of development before the appearance of adenosine A1 receptors modulating dopamine-dependent adenylate cyclase activity or A2 receptors that directly activa...

  14. AMP-activated protein kinase mediates mitochondrial fission in response to energy stress

    Courchet, Julien; Lewis, Tommy L.; Losón, Oliver C.; Hellberg, Kristina; Young, Nathan P.; Chen, Hsiuchen; Polleux, Franck; Chan, David C.; Shaw, Reuben J.

    2016-01-01

    Mitochondria undergo fragmentation in response to electron transport chain (ETC) poisons and mitochondrial DNA–linked disease mutations, yet how these stimuli mechanistically connect to the mitochondrial fission and fusion machinery is poorly understood. We found that the energy-sensing adenosine monophosphate (AMP)–activated protein kinase (AMPK) is genetically required for cells to undergo rapid mitochondrial fragmentation after treatment with ETC inhibitors. Moreover, direct pharmacological activation of AMPK was sufficient to rapidly promote mitochondrial fragmentation even in the absence of mitochondrial stress. A screen for substrates of AMPK identified mitochondrial fission factor (MFF), a mitochondrial outer-membrane receptor for DRP1, the cytoplasmic guanosine triphosphatase that catalyzes mitochondrial fission. Nonphosphorylatable and phosphomimetic alleles of the AMPK sites in MFF revealed that it is a key effector of AMPK-mediated mitochondrial fission. PMID:26816379

  15. Adenosine Diphosphate Ribosylation Factor-GTPaseActivating Protein Stimulates the Transport of AUX1Endosome, Which Relies on Actin Cytoskeletal Organization in Rice Root DevelopmentF

    Cheng Du; Yunyuan XU; Yingdian Wang; Kang Chong

    2011-01-01

    Polar auxin transport,which depends on polarized subcellular distribution of AUXIN RESISTANT 1/LIKE AUX1 (AUX1/LAX) influx carriers and PIN-FORMED (PIN) efflux carriers,mediates various processes of plant growth and development.Endosomal recycling of PIN1 is mediated by an adenosine diphosphate (ADP)ribosylation factor (ARF)-GTPase exchange factor protein,GNOM.However,the mediation of auxin influx carrier recycling is poorly understood.Here,we report that overexpression of OsAGAP,an ARF-GTPase-activating protein in rice,stimulates vesicle transport from the plasma membrane to the Golgi apparatus in protoplasts and transgenic plants and induces the accumulation of early endosomes and AUX1.AUX1 endosomes could partially colocalize with FM4-64 labeled early endosome after actin disruption.Furthermore,OsAGAP is involved in actin cytoskeletal organization,and its overexpression tends to reduce the thickness and bundling of actin filaments.Fluorescence recovery after photobleaching analysis revealed exocytosis of the AUX1 recycling endosome was not affected in the OsAGAP overexpression cells,and was only slightly promoted when the actin filaments were completely disrupted by Lat B.Thus,we propose that AUX1 accumulation in the OsAGAP overexpression and actin disrupted cells may be due to the fact that endocytosis of the auxin influx carrier AUX1 early endosome was greatly promoted by actin cytoskeleton disruption.

  16. Role of hypothalamic adenosine 5'-monophosphate-activated protein kinase in the impaired counterregulatory response induced by repetitive neuroglucopenia.

    Alquier, Thierry; Kawashima, Junji; Tsuji, Youki; Kahn, Barbara B

    2007-03-01

    Antecedent hypoglycemia blunts counterregulatory responses that normally restore glycemia, a phenomenon known as hypoglycemia-associated autonomic failure (HAAF). The mechanisms leading to impaired counterregulatory responses are largely unknown. Hypothalamic AMP-activated protein kinase (AMPK) acts as a glucose sensor. To determine whether failure to activate AMPK could be involved in the etiology of HAAF, we developed a model of HAAF using repetitive intracerebroventricular (icv) injection of 2-deoxy-D-glucose (2DG) resulting in transient neuroglucopenia in normal rats. Ten minutes after a single icv injection of 2DG, both alpha1- and alpha2-AMPK activities were increased 30-50% in arcuate and ventromedial/dorsomedial hypothalamus but not in other hypothalamic regions, hindbrain, or cortex. Increased AMPK activity persisted in arcuate hypothalamus at 60 min after 2DG injection when serum glucagon and corticosterone levels were increased 2.5- to 3.4-fold. When 2DG was injected icv daily for 4 d, hypothalamic alpha1- and alpha2-AMPK responses were markedly blunted in arcuate hypothalamus, and alpha1-AMPK was also blunted in mediobasal hypothalamus 10 min after 2DG on d 4. Both AMPK isoforms were activated normally in arcuate hypothalamus at 60 min. Counterregulatory hormone responses were impaired by recurrent neuroglucopenia and were partially restored by icv injection of 5-aminoimidazole-4-carboxamide 1-beta-D-ribofuranoside, an AMPK activator, before 2DG. Glycogen content increased 2-fold in hypothalamus after recurrent neuroglucopenia, suggesting that glycogen supercompensation could be involved in down-regulating the AMPK glucose-sensing pathway in HAAF. Thus, activation of hypothalamic AMPK may be important for the full counterregulatory hormone response to neuroglucopenia. Furthermore, impaired or delayed AMPK activation in specific hypothalamic regions may play a critical role in the etiology of HAAF. PMID:17185376

  17. Activation of A2b adenosine receptor regulates ovarian cancer cell growth: involvement of Bax/Bcl-2 and caspase-3.

    Hajiahmadi, Sima; Panjehpour, Mojtaba; Aghaei, Mahmoud; Shabani, Mahdi

    2015-08-01

    A2b adenosine receptor (A2bAR) acts as a potent regulator of cell growth in various cell lines. The present study was designed to understand the controlling mechanism of A2bAR agonist (NECA)-induced apoptosis in ovarian cancer cells. Real-time PCR and western blotting assays were used to evaluate the gene and protein expression profiles of A2bAR, respectively. MTT assay was used to study the cell proliferation effect of A2bAR agonist (NECA). Detection of apoptosis was conducted using annexin V-FITC/PI staining, caspase-3 activation assay, and the expression of Bax and Bcl-2 proteins analysis. The mitochondrial membrane potential (ΔΨM) was analyzed by employing JC-1 prob. The mRNA and protein expression levels of A2bAR in ovarian cancer cells were detected. NECA significantly reduced cell viability in a dose-dependent manner in OVCAR-3 and Caov-4 cell lines. The growth inhibition effect of NECA was related to the induction of cell apoptosis, which was manifested by annexin V-FITC staining, activation of caspase-3, and loss of mitochondrial membrane potentials (ΔΨm). In addition, downregulation of the regulatory protein Bcl-2 and upregulation of Bax protein by NECA were also observed. These findings demonstrated that NECA induces apoptosis via the mitochondrial signaling pathway. Thus, A2bAR agonists may be a potential agent for induction of apoptosis in ovarian cancer cells. PMID:25877700

  18. Distinct Roles for the A2B Adenosine Receptor in Acute and Chronic Stages of Bleomycin-Induced Lung Injury

    Yang ZHOU; Schneider, Daniel J.; Morschl, Eva; Song, Ling; Pedroza, Mesias; Karmouty-Quintana, Harry; Le, Thuy.; Sun, Chun-Xiao; Blackburn, Michael R.

    2010-01-01

    Adenosine is an extracellular signaling molecule that is generated in response to cell injury where it orchestrates tissue protection and repair. Whereas adenosine is best known for promoting anti-inflammatory activities during acute injury responses, prolonged elevations can enhance destructive tissue remodeling processes associated with chronic disease states. The generation of adenosine and the subsequent activation of the adenosine 2B receptor (A2BR) is an important processes in the regul...

  19. WISP3 and RhoC guanosine triphosphatase cooperate in the development of inflammatory breast cancer

    Kleer, Celina G.; Zhang, Yanhong; Pan, Quintin; Gallagher, Gary; Wu, Mei; Wu, Zhi-fen; Merajver, Sofia D

    2003-01-01

    Background Inflammatory breast cancer (IBC) is the most lethal form of locally advanced breast cancer. We found concordant and consistent alterations of two genes in 90% of IBC tumors when compared with stage-matched non-IBC tumors: overexpression of RhoC guanosine triphosphatase and loss of WNT-1 induced secreted protein 3 (WISP3). Further work revealed that RhoC is a transforming oncogene for human mammary epithelial (HME) cells. Despite the aggressiveness of the RhoC-driven phenotype, it d...

  20. Characterization of a Trifunctional Mimivirus mRNA Capping Enzyme and Crystal Structure of the RNA Triphosphatase Domain

    Benarroch,D.; Smith, P.; Shuman, S.

    2008-01-01

    The RNA triphosphatase (RTPase) components of the mRNA capping apparatus are a bellwether of eukaryal taxonomy. Fungal and protozoal RTPases belong to the triphosphate tunnel metalloenzyme (TTM) family, exemplified by yeast Cet1. Several large DNA viruses encode metal-dependent RTPases unrelated to the cysteinyl-phosphatase RTPases of their metazoan host organisms. The origins of DNA virus RTPases are unclear because they are structurally uncharacterized. Mimivirus, a giant virus of amoeba, resembles poxviruses in having a trifunctional capping enzyme composed of a metal-dependent RTPase module fused to guanylyltransferase (GTase) and guanine-N7 methyltransferase domains. The crystal structure of mimivirus RTPase reveals a minimized tunnel fold and an active site strikingly similar to that of Cet1. Unlike homodimeric fungal RTPases, mimivirus RTPase is a monomer. The mimivirus TTM-type RTPase-GTase fusion resembles the capping enzymes of amoebae, providing evidence that the ancestral large DNA virus acquired its capping enzyme from a unicellular host.

  1. Role of adenosine 5'-monophosphate-activated protein kinase in interleukin-6 release from isolated mouse skeletal muscle

    Glund, Stephan; Treebak, Jonas Thue; Long, Yun Chau;

    2009-01-01

    IL-6 is released from skeletal muscle during exercise and has consequently been implicated to mediate beneficial effects on whole-body metabolism. Using 5-aminoimidazole-4-carboxamide-1-beta-4-ribofuranoside (AICAR), a pharmacological activator of 5'-AMP-activated protein kinase (AMPK), we tested...... the hypothesis that AMPK modulates IL-6 release from isolated muscle. Skeletal muscle from AMPKalpha2 kinase-dead transgenic, AMPKalpha1 knockout (KO) and AMPKgamma3 KO mice and respective wild-type littermates was incubated in vitro, in the absence or presence of 2 mmol/liter AICAR. Skeletal muscle...... from wild-type mice was also incubated with the AMPK activator A-769662. Incubation of mouse glycolytic extensor digitorum longus and oxidative soleus muscle for 2 h was associated with profound IL-6 mRNA production and protein release, which was suppressed by AICAR (P < 0.001). Basal IL-6 release from...

  2. Apoptotic effects of extract from Cnidium monnieri (L.) Cusson by adenosine monosphosphate-activated protein kinase-independent pathway in HCT116 colon cancer cells.

    Lim, Eun Gyeong; Kim, Guen Tae; Lee, Se Hee; Kim, Sang-Yong; Kim, Young Min

    2016-06-01

    Colon cancer, a common malignancy, can occur due to poor eating habits and increasing age. Consequently, careful regulation of eating habits may serve as a possible method for preventing the occurrence or progression of colon cancer. Extracts of the fruit of Cnidium monnieri (L.) Cusson are well‑known as an effective herbal medicine for the treatment of pain in female genitalia and carbuncle. However, there have been no studies on the apoptotic effects of Cnidium monnieri (L.) Cusson (CME). Adenosine monophosphate‑activated protein kinase (AMPK), the major regulator of energy metabolism, is activated by metabolic stress, including hypoxia and glucose deprivation. Activation of AMPK inhibits cell proliferation and induces apoptosis through the inhibition of phosphorylated (p)‑Akt and control of B‑cell lymphoma 2 (Bcl‑2) family members. The pro‑apoptotic proteins Bcl‑2‑associated X protein (Bax) and Bcl‑2‑homologous antagonist killer (Bak), are activated by their translocation to mitochondria from the cytosol. Translocation of Bax/Bak induces outer membrane permeabilization and is likely to lead to apoptosis through cytochrome C release and caspase activity. In the present study, the apoptotic effects and influence on mitochondria‑mediated apoptotic proteins of CME in HCT116 cells were assessed. We hypothesized that CME may have an effect on the inhibition of p‑Akt in an AMPK‑independent pathway. The present study demonstrated that CME induced the release of LDH and apoptosis through its inhibition of p‑Akt to control Bcl‑2 and activate Bax and Bak. Co‑treatment with CME and AMPK inhibitors showed that CME‑induced apoptosis does not occurr through a AMPK‑dependent pathway. Therefore, the present study determined, for the first time, that CME induced apoptosis as a result of causing metabolic stresses due to directly regulation of the de‑phosphorylation of Akt, whereas it did not control the AMPK-dependent pathway in HCT116

  3. New ribosome-inactivating proteins with polynucleotide:adenosine glycosidase and antiviral activities from Basella rubra L. and bougainvillea spectabilis Willd.

    Bolognesi, A; Polito, L; Olivieri, F; Valbonesi, P; Barbieri, L; Battelli, M G; Carusi, M V; Benvenuto, E; Del Vecchio Blanco, F; Di Maro, A; Parente, A; Di Loreto, M; Stirpe, F

    1997-12-01

    New single-chain (type 1) ribosome-inactivating proteins (RIPs) were isolated from the seeds of Basella rubra L. (two proteins) and from the leaves of Bougainvillea spectabilis Willd. (one protein). These RIPs inhibit protein synthesis both in a cell-free system, with an IC50 (concentration causing 50% inhibition) in the 10(-10) M range, and by various cell lines, with IC50S in the 10(-8)-10(-6) M range. All three RIPs released adenine not only from rat liver ribosomes but also from Escherichia coli rRNA, polyadenylic acid, herring sperm DNA, and artichoke mottled crinkle virus (AMCV) genomic RNA, thus being polynucleotide:adenosine glycosidases. The proteins from Basella rubra had toxicity to mice similar to that of most type 1 RIPs (Barbieri et al., 1993, Biochim Biophys Acta 1154: 237-282) with an LD50 (concentration that is 50% lethal) 32 mg.kg-1. The N-terminal sequence of the two RIPs from Basella rubra had 80-93% identity, whereas it differed from the sequence of the RIP from Bougainvillea spectabilis. When tested with antibodies against various RIPs, the RIPs from Basella gave some cross-reactivity with sera against dianthin 32, and weak cross-reactivity with momordin I and momorcochin-S, whilst the RIP from Bougainvillea did not cross-react with any antiserum tested. An RIP from Basella rubra and one from Bougainvillea spectabilis were tested for antiviral activity, and both inhibited infection of Nicotiana benthamiana by AMCV. PMID:9421927

  4. Role of adenosine 5'-monophosphate-activated protein kinase subunits in skeletal muscle mammalian target of rapamycin signaling

    Deshmukh, Atul S.; Treebak, Jonas Thue; Long, Yun Chau;

    2008-01-01

    AMP-activated protein kinase (AMPK) is an important energy-sensing protein in skeletal muscle. Mammalian target of rapamycin (mTOR) mediates translation initiation and protein synthesis through ribosomal S6 kinase 1 (S6K1) and eukaryotic initiation factor 4E-binding protein 1 (4E-BP1). AMPK...... activation reduces muscle protein synthesis by down-regulating mTOR signaling, whereas insulin mediates mTOR signaling via Akt activation. We hypothesized that AMPK-mediated inhibitory effects on mTOR signaling depend on catalytic alpha2 and regulatory gamma3 subunits. Extensor digitorum longus muscle from...... extensor digitorum longus muscle from either alpha2 or gamma3 AMPK KO mice, indicating functional alpha2 and gamma3 subunits of AMPK are required for the reduction in mTOR signaling. AICAR alone was without effect on basal phosphorylation of S6K1 (Thr389), ribosomal protein S6 (Ser235/236), and 4E-BP1 (Thr...

  5. Activation of adenosine A3 receptors supports hematopoiesis-stimulating effects of granulocyte colony-stimulating factor in sublethally irradiated mice

    Hofer, Michal; Pospíšil, Milan; Šefc, L.; Dušek, L.; Vacek, Antonín; Holá, Jiřina; Hoferová, Zuzana; Štreitová, Denisa

    2010-01-01

    Roč. 86, č. 8 (2010), s. 649-656. ISSN 0955-3002 R&D Projects: GA ČR(CZ) GA305/08/0158 Institutional research plan: CEZ:AV0Z50040507; CEZ:AV0Z50040702 Keywords : ionising radiation * hematopoiesis * adenosine A3 receptors Subject RIV: BO - Biophysics Impact factor: 1.861, year: 2010

  6. Vitality Improvement of the Mediterranean Fruit Fly, Ceratitis capitata Wied 2- Measured by using ME and At Pase Enzyme Activities and Total Protein Content

    The present investigation aims at producing sterile adult Mediterranean fruit fly, Ceratitis capitata Wied. Having the best possible vitality through the use of irradiation and /or a mutagenic substances to be used in a sterile insect technique program. Several types of mutagenic that were thought to cause mutations were used as IGR's, temperature, formaldehyde, colchicine, alcohols, serve ral types of larval rearing media and gamma-rays. In a common pathway, malic enzyme (ME) activity, adenosine triphosphatase (ATPase) enzyme activity and the total protein contents are studied as direct parameters for measuring vitality of the insect. It was found that there is an increment at levels of these parameters due to the treatment of egg stage by the previously mentioned treatments specially the usage of the rice hulls as a bulking component in the larval rearing media alone or followed by irradiation of the pupal stage with 90 Gy

  7. Effects of adenosine metabolism in astrocytes on central nervous system oxygen toxicity.

    Chen, Yu-liang; Zhang, Ya-nan; Wang, Zhong-zhuang; Xu, Wei-gang; Li, Run-ping; Zhang, Jun-dong

    2016-03-15

    Hyperbaric oxygen (HBO) is widely used in military operations, especially underwater missions. However, prolonged and continuous inhalation of HBO can cause central nervous system oxygen toxicity (CNS-OT), which greatly limits HBO's application. The regulation of astrocytes to the metabolism of adenosine is involved in epilepsy. In our study, we aimed to observe the effects of HBO exposure on the metabolism of adenosine in the brain. Furthermore, we aimed to confirm the possible mechanism underlying adenosine's mediation of the CNS-OT. Firstly, anesthetized rats exposed to 5 atm absolute HBO for 80 min. The concentrations of extracellular adenosine, ATP, ADP, and AMP were detected. Secondly, free-moving rats were exposed to HBO at the same pressure for 20 min, and the activities of 5'-nucleotidase and ADK in brain tissues were measured. For the mechanism studies, we observed the effects of a series of different doses of drugs related to adenosine metabolism on the latency of CNS-OT. Results showed HBO exposure could increase adenosine content by inhibiting ADK activity and improving 5'-nucleotidase activity. And adenosine metabolism during HBO exposure may be a protective response against HBO-induced CNS-OT. Moreover, the improvement of adenosine concentration, activation of adenosine A1R, or suppression of ADK and adenosine A2AR, which are involved in the prevention of HBO-induced CNS-OT. This is the first study to demonstrate HBO exposure regulated adenosine metabolism in the brain. Adenosine metabolism and adenosine receptors are related to HBO-induced CNS-OT development. These results will provide new potential targets for the termination or the attenuation of CNS-OT. PMID:26806404

  8. The pharmacological activation of adenosine A1 and A3 receptors does not modulate the long- or short-term repopulating ability of hematopoietic stem and multipotent progenitor cells in mice

    Hofer, Michal; Pospíšil, Milan; Hoferová, Zuzana; Komůrková, Denisa; Páral, Petr; Savvulidi, Filipp; Šefc, Luděk

    2012-01-01

    This study continues our earlier findings on the hematopoiesis-modulating effects of adenosine A1 and A3 receptor agonists that were performed on committed hematopoietic progenitor and precursor cell populations. In the earlier experiments, N6-cyclopentyladenosine (CPA), an adenosine A1 receptor agonist, was found to inhibit proliferation in the above-mentioned hematopoietic cell systems, whereas N6-(3-iodobenzyl)adenosine-5′-N-methyluronamide (IB-MECA), an adenosine A3 receptor agonist, was ...

  9. Adenosine A1 Receptor Antagonist Versus Montelukast on Airway Reactivity and Inflammation

    Nadeem, Ahmed; Obiefuna, Peter C.M.; Wilson, Constance N.; Mustafa, S. Jamal

    2006-01-01

    Adenosine produces bronchoconstriction in allergic rabbits, primates, and humans by activating adenosine A1 receptors. Previously, it is reported that a high dose of L-97-1, a water-soluble, small molecule adenosine A1 receptor antagonist, blocks early and late allergic responses, and bronchial hyper-responsiveness to histamine in a hyper-responsive rabbit model of allergic asthma. Effects of a lower dose of L-97-1 are compared to montelukast, a cysteinyl leukotriene-1 receptor antagonist on ...

  10. Protective effect of taurine on hypochlorous acid toxicity to nuclear nucleoside triphosphatase in isolated nuclei from rat liver

    Ju-Xiang Li; Yong-Zheng Pang; Chao-Shu Tang; Zai-Quan Li

    2004-01-01

    AIM: Taurine has been shown to be an effective scavenger of hypochlorous acid (HOCI). The role of HOCI is well established in tissue damage associated with inflammation and injury. In the present study, the effect of HOCI on nuclear nucleoside triphosphatase of hepatocytes and the ability of taurine to prevent this effect were investigated.METHODS: Isolated hepatic nuclei from rat liver were exposed to HOCI with or without taurine. The NTPase activity on nuclear envelope was assayed using ATP and GTP as substrates, respectively.RESULTS: The first series of experiments evaluated the toxicity of HOCl and the efficacy of taurine to protect NTPase.HOCI at 10-9-5×10-6 mol/L reduced nuclear NTPase activities in a concentration dependent manner (ATP and GTP as substrates) (P<0.01). HOCI at 10-6 mol/L reduced the NTPase activity by 65% (ATP as substrate) and 76% (GTP as substrate). Taurine (10-7 to 10-4 mol/L) was tested for protection against HOCI at 10-6 mol/L and the nuclei treated with 5x10-4 mol/L taurine exhibited only 20% and 12% reduction in NTPase activities compared to untreated controls. A second study was performed comparing taurine to glutathione (GSH). GSH and HOCI at 10-6 mol/L exhibited 46% and 67.4% reduction in NTPase activities compared with control. GSH (10-4 mol/L) which was incubated with the nuclei and HOCi still exhibited 44.2% and 44.8% reduction in NTPase activities of untreated control. Taurine with HOCI only exhibited 15.2% and 17.1% reduction in NTPase activities, which provided more powerful protection against HOCI than GSH. The third experiment was undertaken to evaluate the specificity of taurine against HOCI. Incubation of rat hepatic nuclei with Fe3+/H2O2 (1 m mol/L vS 5μ mol/L) resulted in a decrease in nuclear NTPase activities (P<0.01).When hepatic nuclei were incubated with Tau (10-4 mol/L) and Fe3+/H2O2 (1m mol/L vS 5μ mol/L), nuclear NTPase activities were only slightly increased as compared with that of incubation with Fe3+/H

  11. Intracellular signalling pathways in the vasoconstrictor response of mouse afferent arterioles to adenosine

    Hansen, Pernille B. Lærkegaard; Friis, Ulla Glenert; Uhrenholt, Torben Rene;

    2007-01-01

    calcium from the sarcoplasmic reticulum (SR), stimulated presumably by IP(3), is involved in the adenosine contraction mechanism of the afferent arteriole. In agreement with this notion is the observation that 2 aminoethoxydiphenyl borate (100 microM) blocked the adenosine-induced constriction whereas the...... protein kinase C inhibitor calphostin C had no effect. The calcium-activated chloride channel inhibitor IAA-94 (30 microM) inhibited the adenosine-mediated constriction. Patch clamp experiments showed that adenosine treatment induced a depolarizing current in preglomerular smooth muscle cells which was....... METHODS AND RESULTS: Adenosine (10(-7) M) significantly increased the intracellular calcium concentration in mouse isolated afferent arterioles measured by fura-2 fluorescence. Pre-treatment with thapsigargin (2 microM) blocked the vasoconstrictor action of adenosine (10(-7) M) indicating that release of...

  12. Latent N-methyl-D-aspartate receptors in the recurrent excitatory pathway between hippocampal CA1 pyramidal neurons: Ca(2+)-dependent activation by blocking A1 adenosine receptors.

    Klishin, A; Tsintsadze, T.; Lozovaya, N.; Krishtal, O

    1995-01-01

    When performed at increased external [Ca2+]/[Mg2+] ratio (2.5 mM/0.5 mM), temporary block of A1 adenosine receptors in hippocampus [by 8-cyclopentyltheophylline (CPT)] leads to a dramatic and irreversible change in the excitatory postsynaptic current (EPSC) evoked by Schaffer collateral/commissural (SCC) stimulation and recorded by in situ patch clamp in CA1 pyramidal neurons. The duration of the EPSC becomes stimulus dependent, increasing with increase in stimulus strength. The later occurri...

  13. Cyclic adenosine monophosphate phosphodiesterase in brain: effect on anxiety.

    Beer, B; Chasin, M; Clody, D E; Vogel, J R

    1972-04-28

    Drugs that reduce anxiety may be mediated by cyclic adenosine monophosphate in the brain because (i) potent anxiety-reducing drugs are also potent inhibitors of brain phosphodiesterase activity; (ii) dibutyryl cyclic adenosine monophosphate has the ability to reduce anxiety; (iii) the methylxanthines show significant anxiety-reducing effects; (iv) theophylline and chlordiazepoxide produce additive anxiety-reducing activity; and (v) there is a significant correlation between the anxiety-reducing property of drugs and their ability to inhibit phosphodiesterase activity in the brain. PMID:4402069

  14. Molecular signalling mediating the protective effect of A1 adenosine and mGlu3 metabotropic glutamate receptor activation against apoptosis by oxygen/glucose deprivation in cultured astrocytes.

    Ciccarelli, Renata; D'Alimonte, Iolanda; Ballerini, Patrizia; D'Auro, Mariagrazia; Nargi, Eleonora; Buccella, Silvana; Di Iorio, Patrizia; Bruno, Valeria; Nicoletti, Ferdinando; Caciagli, Francesco

    2007-05-01

    Astrocyte death may occur in neurodegenerative disorders and complicates the outcome of brain ischemia, a condition associated with high extracellular levels of adenosine and glutamate. We show that pharmacological activation of A(1) adenosine and mGlu3 metabotropic glutamate receptors with N(6)-chlorocyclopentyladenosine (CCPA) and (-)2-oxa-4-aminocyclo-[3.1.0]hexane-4,6-dicarboxylic acid (LY379268), respectively, protects cultured astrocytes against apoptosis induced by a 3-h exposure to oxygen/glucose deprivation (OGD). Protection by CCPA and LY379268 was less than additive and was abrogated by receptor blockade with selective competitive antagonists or pertussis toxin. Both in control astrocytes and in astrocytes exposed to OGD, CCPA and LY379268 induced a rapid activation of the phosphatidylinositol-3-kinase (PI3K) and extracellular signal-regulated kinases 1 and 2 (ERK1/2)/mitogen-activated protein kinase (MAPK) pathways, which are known to support cell survival. In cultures exposed to OGD, CCPA and LY379268 reduced the activation of c-Jun N-terminal kinase and p38/MAPK, reduced the levels of the proapoptotic protein Bad, increased the levels of the antiapoptotic protein Bcl-X(L), and were highly protective against apoptotic death, as shown by nuclear 4'-6-diamidino-2-phenylindole staining and measurements of caspase-3 activity. All of these effects were attenuated by treatment with 1,4-diamino-2,3-dicyano-1,4-bis(methylthio)butadiene (U0126) and 2-(4-morpholinyl)-8-phenyl-1(4H)-benzopyran-4-one hydrochloride (LY294002), which inhibit the MAPK and the PI3K pathways, respectively. These data suggest that pharmacological activation of A(1) and mGlu3 receptors protects astrocytes against hypoxic/ischemic damage by stimulating the PI3K and ERK1/2 MAPK pathways. PMID:17293559

  15. Berberine treatment prevents cardiac dysfunction and remodeling through activation of 5'-adenosine monophosphate-activated protein kinase in type 2 diabetic rats and in palmitate-induced hypertrophic H9c2 cells.

    Chang, Wenguang; Zhang, Ming; Meng, Zhaojie; Yu, Yang; Yao, Fan; Hatch, Grant M; Chen, Li

    2015-12-15

    Diabetic cardiomyopathy is the major cause of death in type 2 diabetic patients. Berberine is an isoquinoline alkaloid extract from traditional chinese herbs and its hypoglycemic and hypolipidemic effects make it a promising drug for treatment of type 2 diabetes. We examined if berberine improved cardiac function and attenuated cardiac hypertrophy and fibrosis in high fat diet and streptozotocin induced-type 2 diabetic rats in vivo and reduced expression of hypertrophy markers in palmitate-induced hypertrophic H9c2 cells in vitro. Treatment of diabetic animals with berberine partially improved cardiac function and restored fasting blood insulin, fasting blood glucose, total cholesterol, and triglyceride levels to that of control. In addition, berberine treatment of diabetic animals increased cardiac 5'-adenosine monophosphate-activated protein kinase (AMPK) and protein kinase B (AKT) activation and reduced glycogen synthase kinase 3 beta (GSK3β) activation compared to control. Palmitate incubation of H9c2 cells resulted in cellular hypertrophy and decreased expression of alpha-myosin heavy chain (α-MHC) and increased expression of beta-myosin heavy chain (β-MHC) compared to controls. Berberine treatment of palmitate-incubated H9c2 cells reduced hypertrophy, increased α-MHC expression and decreased β-MHC expression. In addition, berberine treatment of palmitate-incubated H9c2 cells increased AMPK and AKT activation and reduced GSK3β activation. The presence of the AMPK inhibitor Compound C attenuated the effects of berberine. The results strongly indicate that berberine treatment may be protective against the development of diabetic cardiomyopathy. PMID:26522928

  16. Single Nucleotide Polymorphisms That Increase Expression of the Guanosine Triphosphatase RAC1 Are Associated With Ulcerative Colitis

    Aleixo M Muise; Walters, Thomas; Xu, Wei; Shen-Tu, Grace; Guo, Cong-Hui; Fattouh, Ramzi; Lam, Grace Y; Wolters, Victorien M; Bennitz, Joshua; Van Limbergen, Johan; Renbaum, Paul; Kasirer, Yair; Ngan, Bo-yee; Turner, Dan; Denson, Lee A.

    2011-01-01

    Background & AimsRAC1 is a guanosine triphosphatase that has an evolutionarily conserved role in coordinating immune defenses, from plants to mammals. Chronic inflammatory bowel diseases are associated with dysregulation of immune defenses. We studied the role of RAC1 in inflammatory bowel diseases using human genetic and functional studies and animal models of colitis.MethodsWe used a candidate gene approach to HapMap-Tag single nucleotide polymorphisms in a discovery cohort; findings we...

  17. Possible therapeutic benefits of adenosine-potentiating drugs in reducing age-related degenerative disease in dogs and cats.

    Scaramuzzi, R J; Baker, D J

    2003-10-01

    Adenosine is a ubiquitous, biologically important molecule that is a precursor of other biologically active molecules. It also is a component of some co-factors and has distinct physiological actions in its own right. Levels are maintained by synthesis from dietary precursors and re-cycling. The daily turnover of adenosine is very high. Adenosine can act either as a hormone by binding to adenosine receptors, four adenosine receptor subtypes have been identified, and as an intracellular modulator, after transport into the cell by membrane transporter proteins. One of the principal intracellular actions of adenosine is inhibition of the enzyme phosphodiesterase. Extracellular adenosine also has specific neuromodulatory actions on dopamine and glutamate. Selective and nonselective agonists and antagonists of adenosine are available. The tasks of developing, evaluating and exploiting the therapeutic potential of these compounds is still in its infancy. Adenosine has actions in the central nervous system (CNS), heart and vascular system, skeletal muscle and the immune system and the presence of receptors suggests potential actions in the gonads and other organs. Adenosine agonists improve tissue perfusion through actions on vascular smooth muscle and erythrocyte fluidity and they can be used to improve the quality of life in aged dogs. This article reviews the therapeutic potential of adenosine-potentiating drugs in the treatment of age-related conditions in companion animals, some of which may be exacerbated by castration or spaying at an early age. PMID:14633184

  18. Dual activity of certain HIT-proteins: A. thaliana Hint4 and C. elegans DcpS act on adenosine 5'-phosphosulfate as hydrolases (forming AMP) and as phosphorylases (forming ADP).

    Guranowski, Andrzej; Wojdyła, Anna Maria; Zimny, Jarosław; Wypijewska, Anna; Kowalska, Joanna; Jemielity, Jacek; Davis, Richard E; Bieganowski, Paweł

    2010-01-01

    Histidine triad (HIT)-family proteins interact with different mono- and dinucleotides and catalyze their hydrolysis. During a study of the substrate specificity of seven HIT-family proteins, we have shown that each can act as a sulfohydrolase, catalyzing the liberation of AMP from adenosine 5'-phosphosulfate (APS or SO(4)-pA). However, in the presence of orthophosphate, Arabidopsis thaliana Hint4 and Caenorhabditis elegans DcpS also behaved as APS phosphorylases, forming ADP. Low pH promoted the phosphorolytic and high pH the hydrolytic activities. These proteins, and in particular Hint4, also catalyzed hydrolysis or phosphorolysis of some other adenylyl-derivatives but at lower rates than those for APS cleavage. A mechanism for these activities is proposed and the possible role of some HIT-proteins in APS metabolism is discussed. PMID:19896942

  19. An adenosine at position 27 in the human immunodeficiency virus type 1 trans-activation response element is not critical for transcriptional or translational activation by Tat.

    Blanchard, A. D.; Powell, R; Braddock, M; Kingsman, A J; Kingsman, S M

    1992-01-01

    Tat protein binds to the trans-activation response (TAR) element of human immunodeficiency virus type 1 RNAs and activates gene expression at the level of transcription in mammalian cell lines and translation in Xenopus oocytes. Certain residues within TAR are important for Tat binding in vitro, including residue A-27, which appears to be able to be modified in a Tat-dependent manner in Xenopus oocytes (L. Sharmeen, B. Bass, N. Sonenberg, H. Weintraub, and M. Groudine, Proc. Natl. Acad. Sci. ...

  20. Antagonism by theophylline of respiratory inhibition induced by adenosine.

    Eldridge, F L; Millhorn, D E; Kiley, J P

    1985-11-01

    The effects on respiration of an analogue of adenosine, L-2-N6-(phenylisopropyl)adenosine (PIA), and of the methylxanthine, theophylline, were determined in 19 vagotomized glomectomized cats whose end-tidal PCO2 was kept constant by means of a servo-controlled ventilator. Integrated phrenic nerve activity was used to represent respiratory output. Our results show that PIA, whether given systemically or into the third cerebral ventricle, depressed respiration. Systemically administered theophylline stimulated respiration. Theophylline given intravenously, or into the third ventricle not only reversed the depressive effects of previously administered PIA but caused further increases of respiration above the control level. Prior systemic administration of theophylline blocked both respiratory and hypotensive effects of subsequently administered PIA. Effects of either agent on medullary extracellular fluid pH did not explain the results. We conclude that the adenosine analogue PIA, acts to inhibit neurons in the brain that are involved in the control of respiration and that its effects are blocked by theophylline. We suggest that adenosine acts as a tonic modulator of respiration and that theophylline stimulates breathing by competitive antagonism of adenosine at neuronal receptor sites. PMID:4066573

  1. Adenosine A(1) Receptors in the Central Nervous System : Their Functions in Health and Disease, and Possible Elucidation by PET Imaging

    Paul, S.; Elsinga, P. H.; Ishiwata, K.; Dierckx, R. A. J. O.; van Waarde, A.

    2011-01-01

    Adenosine is a neuromodulator with several functions in the central nervous system (CNS), such as inhibition of neuronal activity in many signaling pathways. Most of the sedating, anxiolytic, seizure-inhibiting and protective actions of adenosine are mediated by adenosine A(1) receptors (A(1)R) on t

  2. 二甲双胍通过激活腺苷酸活化蛋白激酶(AMPK)的抗肿瘤机制%Antitumor Mechanism of Metformin via Adenosine Monophosphate-activated Protein Kinase (AMPK) Activation

    陈兆煜; 王连唐; 陈雁扬

    2013-01-01

    Metformin, as a traditional oral hypoglycemic agent, is commonly used in the clinical treatment for type 2 diabetes. Recently, a large number of epidemiological researches have shown that metformin could reduce the tumor morbidity of type 2 diabetes, moreover, it has also been indicated that metformin could inhibit the growth, proliferation and transformation of cancer cells in metabolic pathways, cell cycle, oxidative stress and cancer/tumor stem cells transformation via AMPK pathway activation. But the antitumor effect of metformin via AMPK activation still exists arguments, and the deifnite mechanism remains to be further investigated and conifrmed by extensive clinical trials.%二甲双胍是一种传统的口服降糖药,临床上普遍用于2型糖尿病的治疗。近年来大量流行病学研究报道二甲双胍能够降低2型糖尿病患者的肿瘤发病率,亦有研究发现二甲双胍能在代谢途径、细胞周期、氧化应激、肿瘤干细胞转化等方面通过激活腺苷酸活化蛋白激酶(adenosine monophosphate-activated protein kinase, AMPK)信号通路,从而抑制肿瘤细胞的生长、增殖以及转化。但二甲双胍通过激活AMPK的抗肿瘤机制仍存在着争议,其确切的作用机制有待进一步深入的研究,同时亟需大规模的临床试验来证实。

  3. The role of muscarinic receptors in the beneficial effects of adenosine against myocardial reperfusion injury in rats.

    Lei Sun

    Full Text Available Adenosine, a catabolite of ATP, displays a wide variety of effects in the heart including regulation of cardiac response to myocardial ischemia and reperfusion injury. Nonetheless, the precise mechanism of adenosine-induced cardioprotection is still elusive. Isolated Sprague-Dawley rat hearts underwent 30 min global ischemia and 120 min reperfusion using a Langendorff apparatus. Both adenosine and acetylcholine treatment recovered the post-reperfusion cardiac function associated with adenosine and muscarinic receptors activation. Simultaneous administration of adenosine and acetylcholine failed to exert any additive protective effect, suggesting a shared mechanism between the two. Our data further revealed a cross-talk between the adenosine and acetylcholine receptor signaling in reperfused rat hearts. Interestingly, the selective M(2 muscarinic acetylcholine receptor antagonist methoctramine significantly attenuated the cardioprotective effect of adenosine. In addition, treatment with adenosine upregulated the expression and the maximal binding capacity of muscarinic acetylcholine receptor, which were inhibited by the selective A(1 adenosine receptor antagonist 8-Cyclopentyl-1,3-dipropylxanthine (DPCPX and the nitric oxide synthase inhibitor N(ω-nitro-L-arginine methyl ester (L-NAME. These data suggested a possible functional coupling between the adenosine and muscarinic receptors behind the observed cardioprotection. Furthermore, nitric oxide was found involved in triggering the response to each of the two receptor agonist. In summary, there may be a cross-talk between the adenosine and muscarinic receptors in ischemic/reperfused myocardium with nitric oxide synthase might serve as the distal converging point. In addition, adenosine contributes to the invigorating effect of adenosine on muscarinic receptor thereby prompting to regulation of cardiac function. These findings argue for a potentially novel mechanism behind the adenosine

  4. Diversification of the RAB Guanosine Triphosphatase Family in Dicots and Monocots

    2007-01-01

    RAB guanosine triphosphatases (GTPases) are key regulators of vesicle trafficking and are essential to the growth and development of all eukaryotic cells. During evolution, the RAB family has expanded in different patterns to facilitate distinct cellular, developmental and physiological adaptations. Yeast has only 11 family members, whereas mammalian RABs have expanded to 18 RAB subfamilies. Plant RABs have diversified primarily by duplicating members within a single subfamily. Plant RABs are divided into eight subfamilies, corresponding to mammalian RAB1, RAB2, RAB5, RAB6,RAB7, RAB8, RAB11 and RAB18. Functional diversification of these is exemplified by the RAB11s, orthologs of which are partitioned into unique cell compartments in plants where they function to transport vesicles during localized tip growth.Similarly, the RAB2 family in grasses is likely involved in vesicle secretion associated with wall expansion, as determined by analysis of over-expression mutants. We propose that dicots and monocots have also diverged in their RAB profiles to accommodate unique cellular functions between the two groups. Here we present a bioinformatics analysis comparing the RAB sub-families of rice, maize and Arabidopsis. These results will guide future functional studies to test for the role of diversification of subfamilies unique to monocots compared to dicots.

  5. Characterization of spontaneous, transient adenosine release in the caudate-putamen and prefrontal cortex.

    Michael D Nguyen

    Full Text Available Adenosine is a neuroprotective agent that inhibits neuronal activity and modulates neurotransmission. Previous research has shown adenosine gradually accumulates during pathologies such as stroke and regulates neurotransmission on the minute-to-hour time scale. Our lab developed a method using carbon-fiber microelectrodes to directly measure adenosine changes on a sub-second time scale with fast-scan cyclic voltammetry (FSCV. Recently, adenosine release lasting a couple of seconds has been found in murine spinal cord slices. In this study, we characterized spontaneous, transient adenosine release in vivo, in the caudate-putamen and prefrontal cortex of anesthetized rats. The average concentration of adenosine release was 0.17±0.01 µM in the caudate and 0.19±0.01 µM in the prefrontal cortex, although the range was large, from 0.04 to 3.2 µM. The average duration of spontaneous adenosine release was 2.9±0.1 seconds and 2.8±0.1 seconds in the caudate and prefrontal cortex, respectively. The concentration and number of transients detected do not change over a four hour period, suggesting spontaneous events are not caused by electrode implantation. The frequency of adenosine transients was higher in the prefrontal cortex than the caudate-putamen and was modulated by A1 receptors. The A1 antagonist DPCPX (8-cyclopentyl-1,3-dipropylxanthine, 6 mg/kg i.p. increased the frequency of spontaneous adenosine release, while the A1 agonist CPA (N(6-cyclopentyladenosine, 1 mg/kg i.p. decreased the frequency. These findings are a paradigm shift for understanding the time course of adenosine signaling, demonstrating that there is a rapid mode of adenosine signaling that could cause transient, local neuromodulation.

  6. The adenosine metabolite inosine is a functional agonist of the adenosine A2A receptor with a unique signaling bias.

    Welihinda, Ajith A; Kaur, Manmeet; Greene, Kelly; Zhai, Yongjiao; Amento, Edward P

    2016-06-01

    Inosine is an endogenous purine nucleoside that is produced by catabolism of adenosine. Adenosine has a short half-life (approximately 10s) and is rapidly deaminated to inosine, a stable metabolite with a half-life of approximately 15h. Resembling adenosine, inosine acting through adenosine receptors (ARs) exerts a wide range of anti-inflammatory and immunomodulatory effects in vivo. The immunomodulatory effects of inosine in vivo, at least in part, are mediated via the adenosine A2A receptor (A2AR), an observation that cannot be explained fully by in vitro pharmacological characterization of inosine at the A2AR. It is unclear whether the in vivo effects of inosine are due to inosine or a metabolite of inosine engaging the A2AR. Here, utilizing a combination of label-free, cell-based, and membrane-based functional assays in conjunction with an equilibrium agonist-binding assay we provide evidence for inosine engagement at the A2AR and subsequent activation of downstream signaling events. Inosine-mediated A2AR activation leads to cAMP production with an EC50 of 300.7μM and to extracellular signal-regulated kinase-1 and -2 (ERK1/2) phosphorylation with an EC50 of 89.38μM. Our data demonstrate that inosine produces ERK1/2-biased signaling whereas adenosine produces cAMP-biased signaling at the A2AR, highlighting pharmacological differences between these two agonists. Given the in vivo stability of inosine, our data suggest an additional, previously unrecognized, mechanism that utilizes inosine to functionally amplify and prolong A2AR activation in vivo. PMID:26903141

  7. Overexpression, purification and crystallographic analysis of a unique adenosine kinase from Mycobacterium tuberculosis

    Adenosine kinase from M. tuberculosis has been overexpressed, purified and crystallized in the presence of adenosine. Structure determination using molecular replacement with diffraction data collected at 2.2 Å reveals a dimeric structure. Adenosine kinase from Mycobacterium tuberculosis is the only prokaryotic adenosine kinase that has been isolated and characterized. The enzyme catalyzes the phosphorylation of adenosine to adenosine monophosphate and is involved in the activation of 2-methyladenosine, a compound that has demonstrated selective activity against M. tuberculosis. The mechanism of action of 2-methyladenosine is likely to be different from those of current tuberculosis treatments and this compound (or other adenosine analogs) may prove to be a novel therapeutic intervention for this disease. The M. tuberculosis adenosine kinase was overexpressed in Escherichia coli and the enzyme was purified with activity comparable to that reported previously. The protein was crystallized in the presence of adenosine using the vapour-diffusion method. The crystals diffracted X-rays to high resolution and a complete data set was collected to 2.2 Å using synchrotron radiation. The crystal belonged to space group P3121, with unit-cell parameters a = 70.2, c = 111.6 Å, and contained a single protein molecule in the asymmetric unit. An initial structural model of the protein was obtained by the molecular-replacement method, which revealed a dimeric structure. The monomers of the dimer were related by twofold crystallographic symmetry. An understanding of how the M. tuberculosis adenosine kinase differs from the human homolog should aid in the design of more potent and selective antimycobacterial agents that are selectively activated by this enzyme

  8. Activation of microglial cells triggers a release of brain-derived neurotrophic factor (BDNF inducing their proliferation in an adenosine A2A receptor-dependent manner: A2A receptor blockade prevents BDNF release and proliferation of microglia

    Gomes Catarina

    2013-01-01

    Full Text Available Abstract Background Brain-derived neurotrophic factor (BDNF has been shown to control microglial responses in neuropathic pain. Since adenosine A2A receptors (A2ARs control neuroinflammation, as well as the production and function of BDNF, we tested to see if A2AR controls the microglia-dependent secretion of BDNF and the proliferation of microglial cells, a crucial event in neuroinflammation. Methods Murine N9 microglial cells were challenged with lipopolysaccharide (LPS, 100 ng/mL in the absence or in the presence of the A2AR antagonist, SCH58261 (50 nM, as well as other modulators of A2AR signaling. The BDNF cellular content and secretion were quantified by Western blotting and ELISA, A2AR density was probed by Western blotting and immunocytochemistry and cell proliferation was assessed by BrdU incorporation. Additionally, the A2AR modulation of LPS-driven cell proliferation was also tested in primary cultures of mouse microglia. Results LPS induced time-dependent changes of the intra- and extracellular levels of BDNF and increased microglial proliferation. The maximal LPS-induced BDNF release was time-coincident with an LPS-induced increase of the A2AR density. Notably, removing endogenous extracellular adenosine or blocking A2AR prevented the LPS-mediated increase of both BDNF secretion and proliferation, as well as exogenous BDNF-induced proliferation. Conclusions We conclude that A2AR activation plays a mandatory role controlling the release of BDNF from activated microglia, as well as the autocrine/paracrine proliferative role of BDNF.

  9. Why do premature newborn infants display elevated blood adenosine levels?

    Panfoli, Isabella; Cassanello, Michela; Bruschettini, Matteo; Colella, Marina; Cerone, Roberto; Ravera, Silvia; Calzia, Daniela; Candiano, Giovanni; Ramenghi, Luca

    2016-05-01

    Our preliminary data show high levels of adenosine in the blood of very low birth weight (VLBW) infants, positively correlating to their prematurity (i.e. body weight class). This prompted us to look for a mechanism promoting such impressive adenosine increase. We hypothesized a correlation with oxygen challenge. In fact, it is recognized that either oxygen lack or its excess contribute to the pathogenesis of the injuries of prematurity, such as retinopathy (ROP) and periventricular white matter lesions (PWMI). The optimal concentration of oxygen for resuscitation of VLBW infants is currently under revision. We propose that the elevated adenosine blood concentrations of VLBW infants recognizes two sources. The first could be its activity-dependent release from unmyelinated brain axons. Adenosine in this respect would be an end-product of the hypometabolic VLBW newborn unmyelinated axon intensely firing in response to the environmental stimuli consequent to premature birth. Adenosine would be eventually found in the blood due to blood-brain barrier immaturity. In fact, adenosine is the primary activity-dependent signal promoting differentiation of premyelinating oligodendrocyte progenitor cells (OPC) into myelinating cells in the Central Nervous System, while inhibiting their proliferation and inhibiting synaptic function. The second, would be the ecto-cellular ATP synthesized by the endothelial cell plasmalemma exposed to ambient oxygen concentrations due to premature breathing, especially in lung. ATP would be rapidly transformed into adenosine by the ectonucleotidase activities such as NTPDase I (CD39), and NT5E (CD73). An ectopic extra-mitochondrial aerobic ATP synthetic ability was reported in many cell plasma-membranes, among which endothelial cells. The potential implications of the cited hypotheses for the neonatology area would be great. The amount of oxygen administration for reviving of newborns would find a molecular basis for its assessment. VLBW

  10. Adenosine stimulates DNA fragmentation in human thymocytes by Ca(2+)-mediated mechanisms.

    Szondy, Z

    1994-12-15

    Incubation of human thymocytes with an optimum concentration of adenosine and its receptor site agonist, 2-chloroadenosine, induced increases in intracellular cyclic AMP (cAMP) (from a resting 0.6 +/- 0.1 to 4.1 +/- 0.2 pmol/10(7) cells within 5 min) and Ca2+ (from the resting 85 +/- 7 nM to a peak of 210 +/- 25 nM) levels and resulted in internucleosomal DNA fragmentation and cell death (apoptosis). Other adenosine analogues were also effective at inducing DNA fragmentation, the order of potency being 2-p-(carboxyethylphenylethylamino)-5'-carboxyamidoadenosine 13399-13402], at 60 ng/ml concentration also prevented adenosine-induced DNA fragmentation when added prior to adenosine. This suggested a complex cross-talk between the adenosine-triggered signal transduction cascade and the activation state of protein kinase C in regulating apoptosis of human thymocytes. PMID:7818494

  11. Adenosine in inflammatory joint diseases

    Chan, E. S. L.; Fernandez, P.; Cronstein, B. N.

    2007-01-01

    Inflammatory joint diseases are a group of heterogeneous disorders with a variety of different etiologies and disease manifestations. However, there are features that are common to all of them: first, the recruitment of various inflammatory cell types that are attracted to involved tissues over the course of the disease process. Second, the treatments used in many of these diseases are commonly medications that suppress or alter immune function. The demonstration that adenosine has endogenous...

  12. Cancer exosomes express CD39 and CD73, which suppress T cells through adenosine production.

    Clayton, Aled; Al-Taei, Saly; Webber, Jason; Mason, Malcolm D; Tabi, Zsuzsanna

    2011-07-15

    Extracellular adenosine is elevated in cancer tissue, and it negatively regulates local immune responses. Adenosine production from extracellular ATP has attracted attention as a mechanism of regulatory T cell-mediated immune regulation. In this study, we examined whether small vesicles secreted by cancer cells, called exosomes, contribute to extracellular adenosine production and hence modulate immune effector cells indirectly. We found exosomes from diverse cancer cell types exhibit potent ATP- and 5'AMP-phosphohydrolytic activity, partly attributed to exosomally expressed CD39 and CD73, respectively. Comparable levels of activity were seen with exosomes from pleural effusions of mesothelioma patients. In such fluids, exosomes accounted for 20% of the total ATP-hydrolytic activity. Exosomes can perform both hydrolytic steps sequentially to form adenosine from ATP. This exosome-generated adenosine can trigger a cAMP response in adenosine A(2A) receptor-positive but not A(2A) receptor-negative cells. Similarly, significantly elevated cAMP was also triggered in Jurkat cells by adding exosomes with ATP but not by adding exosomes or ATP alone. A proportion of healthy donor T cells constitutively express CD39 and/or CD73. Activation of T cells by CD3/CD28 cross-linking could be inhibited by exogenously added 5'AMP in a CD73-dependent manner. However, 5'AMP converted to adenosine by exosomes inhibits T cell activation independently of T cell CD73 expression. This T cell inhibition was mediated through the adenosine A(2A) receptor. In summary, the data highlight exosome enzymic activity in the production of extracellular adenosine, and this may play a contributory role in negative modulation of T cells in the tumor environment. PMID:21677139

  13. WISP3 and RhoC guanosine triphosphatase cooperate in the development of inflammatory breast cancer

    Inflammatory breast cancer (IBC) is the most lethal form of locally advanced breast cancer. We found concordant and consistent alterations of two genes in 90% of IBC tumors when compared with stage-matched non-IBC tumors: overexpression of RhoC guanosine triphosphatase and loss of WNT-1 induced secreted protein 3 (WISP3). Further work revealed that RhoC is a transforming oncogene for human mammary epithelial (HME) cells. Despite the aggressiveness of the RhoC-driven phenotype, it does not quantitatively reach that of the true IBC tumors. We have demonstrated that WISP3 has tumor growth and angiogenesis inhibitory functions in IBC. We proposed that RhoC and WISP3 cooperate in the development of IBC. Using an antisense approach, we blocked WISP3 expression in HME cells. Cellular proliferation and growth were determined using the 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl-tetrazolium bromide (MTT) assay and anchorage-independent growth in a soft agar assay. Vascular endothelial growth factor (VEGF) was measured in conditioned medium by enzyme-linked immunosorbent assay. Antisense inhibition of WISP3 in HME cells increased RhoC mRNA levels and resulted in an increase in cellular proliferation, anchorage-independent growth and VEGF levels in the conditioned medium. Conversely, restoration of WISP3 expression in the highly malignant IBC cell line SUM149 was able to decrease the expression of RhoC protein. WISP3 modulates RhoC expression in HME cells and in the IBC cell line SUM149. This provides further evidence that these two genes act in concert to give rise to the highly aggressive IBC phenotype. We propose a model of this interaction as a starting point for further investigations

  14. The role of adenosine A2A receptors on neuromuscular transmission upon ageing

    Pousinha, Paula Isabel Antunes, 1978-

    2012-01-01

    Tese de doutoramento, Ciências Biomédicas (Neurociências), Universidade de Lisboa, Faculdade de Medicina, 2012 Adenosine is a neuromodulator with important actions in the nervous system. The activation of adenosine A2A receptors has been shown to modulate the action of other receptors. Considering that it was observed an interaction between adenosine A2A receptors and TrkB receptors in hippocampus, I hypothesized that the activation of A2A receptors could also facilitate BDNF actions on ne...

  15. Role of extracellular adenosine in Drosophila

    FENCKOVÁ, Michaela

    2011-01-01

    This thesis describes several aspects of the role for extracellular adenosine in Drosophila. Reverse genetic, molecular and microscopic methods together with the most forefront Drosophila research techniques have been applied to elucidate the role of adenosine signaling in the regulation of development, physiology and metabolism of Drosophila larvae. The thesis helps to establish the model for extracellular adenosine as a stress-signal for the release of energy stores. It also describes the e...

  16. Specificity of synergistic coronary flow enhancement by adenosine and pulsatile perfusion in the dog.

    Pagliaro, P; Senzaki, H; Paolocci, N; Isoda, T; Sunagawa, G; Recchia, F A; Kass, D A

    1999-10-01

    1. Coronary flow elevation from enhanced perfusion pulsatility is synergistically amplified by adenosine. This study determined the specificity of this interaction and its potential mechanisms. 2. Mean and phasic coronary flow responses to increasing pulsatile perfusion were assessed in anaesthetized dogs, with the anterior descending coronary artery servoperfused to regulate real-time physiological flow pulsatility at constant mean pressure. Pulsatility was varied between 40 and 100 mmHg. Hearts ejected into the native aorta whilst maintaining stable loading. 3. Increasing pulsatility elevated mean coronary flow +11.5 +/- 1.7 % under basal conditions. Co-infusion of adenosine sufficient to raise baseline flow 66 % markedly amplified this pulsatile perfusion response (+82. 6 +/- 14.3 % increase in mean flow above adenosine baseline), due to a leftward shift of the adenosine-coronary flow response curve at higher pulsatility. Flow augmentation with pulsatility was not linked to higher regional oxygen consumption, supporting direct rather than metabolically driven mechanisms. 4. Neither bradykinin, acetylcholine nor verapamil reproduced the synergistic amplification of mean flow by adenosine and higher pulsatility, despite being administered at doses matching basal flow change with adenosine. 5. ATP-sensitive potassium (KATP) activation (pinacidil) amplified the pulse-flow response 3-fold, although this remained significantly less than with adenosine. Co-administration of the phospholipase A2 inhibitor quinacrine virtually eliminated adenosine-induced vasodilatation, yet synergistic interaction between adenosine and pulse perfusion persisted, albeit at a reduced level. 6. Thus, adenosine and perfusion pulsatility specifically interact to enhance coronary flow. This synergy is partially explained by KATP agonist action and additional non-flow-dependent mechanisms, and may be important for modulating flow reserve during exercise or other high output states where

  17. The major nucleoside triphosphatase in pea (Pisum sativum L.) nuclei and in rat liver nuclei share common epitopes also present in nuclear lamins

    Tong, C. G.; Dauwalder, M.; Clawson, G. A.; Hatem, C. L.; Roux, S. J.

    1993-01-01

    The major nucleoside triphosphatase (NTPase) activities in mammalian and pea (Pisum sativum L.) nuclei are associated with enzymes that are very similar both biochemically and immunochemically. The major NTPase from rat liver nuclei appears to be a 46-kD enzyme that represents the N-terminal portion of lamins A and C, two lamina proteins that apparently arise from the same gene by alternate splicing. Monoclonal antibody (MAb) G2, raised to human lamin C, both immunoprecipitates the major (47 kD) NTPase in pea nuclei and recognizes it in western blot analyses. A polyclonal antibody preparation raised to the 47-kD pea NTPase (pc480) reacts with the same lamin bands that are recognized by MAb G2 in mammalian nuclei. The pc480 antibodies also bind to the same lamin-like bands in pea nuclear envelope-matrix preparations that are recognized by G2 and three other MAbs known to bind to mammalian lamins. In immunofluorescence assays, pc480 and anti-lamin antibodies stain both cytoplasmic and nuclear antigens in plant cells, with slightly enhanced staining along the periphery of the nuclei. These results indicate that the pea and rat liver NTPases are structurally similar and that, in pea nuclei as in rat liver nuclei, the major NTPase is probably derived from a lamin precursor by proteolysis.

  18. Impairment of ATP hydrolysis decreases adenosine A1 receptor tonus favoring cholinergic nerve hyperactivity in the obstructed human urinary bladder.

    Silva-Ramos, M; Silva, I; Faria, M; Magalhães-Cardoso, M T; Correia, J; Ferreirinha, F; Correia-de-Sá, P

    2015-12-01

    This study was designed to investigate whether reduced adenosine formation linked to deficits in extracellular ATP hydrolysis by NTPDases contributes to detrusor neuromodulatory changes associated with bladder outlet obstruction in men with benign prostatic hyperplasia (BPH). The kinetics of ATP catabolism and adenosine formation as well as the role of P1 receptor agonists on muscle tension and nerve-evoked [(3)H]ACh release were evaluated in mucosal-denuded detrusor strips from BPH patients (n = 31) and control organ donors (n = 23). The neurogenic release of ATP and [(3)H]ACh was higher (P bladders. Relaxation of detrusor contractions induced by acetylcholine required 30-fold higher concentrations of adenosine. Despite VAChT-positive cholinergic nerves exhibiting higher A(1) immunoreactivity in BPH bladders, the endogenous adenosine tonus revealed by adenosine deaminase is missing. Restoration of A1 inhibition was achieved by favoring (1) ATP hydrolysis with apyrase (2 U mL(-1)) or (2) extracellular adenosine accumulation with dipyridamole or EHNA, as these drugs inhibit adenosine uptake and deamination, respectively. In conclusion, reduced ATP hydrolysis leads to deficient adenosine formation and A(1) receptor-mediated inhibition of cholinergic nerve activity in the obstructed human bladder. Thus, we propose that pharmacological manipulation of endogenous adenosine levels and/or A(1) receptor activation might be useful to control bladder overactivity in BPH patients. PMID:26521170

  19. Severe hemorrhage attenuates cardiopulmonary chemoreflex control of regional sympathetic outputs via NTS adenosine receptors.

    Minic, Zeljka; Li, Cailian; O'Leary, Donal S; Scislo, Tadeusz J

    2014-09-15

    Selective stimulation of inhibitory A1 and facilitatory A2a adenosine receptor subtypes located in the nucleus of the solitary tract (NTS) powerfully inhibits cardiopulmonary chemoreflex (CCR) control of regional sympathetic outputs via different mechanisms: direct inhibition of glutamate release and facilitation of an inhibitory neurotransmitter release, respectively. However, it remains unknown whether adenosine naturally released into the NTS has similar inhibitory effects on the CCR as the exogenous agonists do. Our previous study showed that adenosine is released into the NTS during severe hemorrhage and contributes to reciprocal changes of renal (decreases) and adrenal (increases) sympathetic nerve activity observed in this setting. Both A1 and A2a adenosine receptors are involved. Therefore, we tested the hypothesis that, during severe hemorrhage, CCR control of the two sympathetic outputs is attenuated by adenosine naturally released into the NTS. We compared renal and adrenal sympathoinhibitory responses evoked by right atrial injections of 5HT3 receptor agonist phenylbiguanide (2-8 μg/kg) under control conditions, during hemorrhage, and during hemorrhage preceded by blockade of NTS adenosine receptors with bilateral microinjections of 8-(p-sulfophenyl) theophylline (1 nmol/100 nl) in urethane/chloralose anesthetized rats. CCR-mediated inhibition of renal and adrenal sympathetic activity was significantly attenuated during severe hemorrhage despite reciprocal changes in the baseline activity levels, and this attenuation was removed by bilateral blockade of adenosine receptors in the caudal NTS. This confirmed that adenosine endogenously released into the NTS has a similar modulatory effect on integration of cardiovascular reflexes as stimulation of NTS adenosine receptors with exogenous agonists. PMID:25063794

  20. Radio-chromatographic determination of plasmatic adenosine deaminase (A.D.)

    We were able, by using a radio-chromatographic method, to measure an adenosine deaminase activity in normal human heparinized platelet-poor plasma, which can degrade 0.016 μM adenosine. This activity suppressed by heating 56 C for 30 minutes is inhibited by high concentrations of urea and is proportional to the amount of plasma, source of enzyme, in the systems. (authors)

  1. Receptor crosstalk: haloperidol treatment enhances A2A adenosine receptor functioning in a transfected cell model

    Trincavelli, Maria Letizia; Cuboni, Serena; Catena Dell’Osso, Mario; Maggio, Roberto; Klotz, Karl-Norbert; Novi, Francesca; Panighini, Anna; Daniele, Simona; Martini, Claudia

    2010-01-01

    A2A adenosine receptors are considered an excellent target for drug development in several neurological and psychiatric disorders. It is noteworthy that the responses evoked by A2A adenosine receptors are regulated by D2 dopamine receptor ligands. These two receptors are co-expressed at the level of the basal ganglia and interact to form functional heterodimers. In this context, possible changes in A2A adenosine receptor functional responses caused by the chronic blockade/activation of D2 dop...

  2. Anti-Inflammatory and Immunosuppressive Effects of the A2A Adenosine Receptor

    Gillian R. Milne; Palmer, Timothy M.

    2011-01-01

    The production of adenosine represents a critical endogenous mechanism for regulating immune and inflammatory responses during conditions of stress, injury, or infection. Adenosine exerts predominantly protective effects through activation of four 7-transmembrane receptor subtypes termed A1, A2A, A2B, and A3, of which the A2A adenosine receptor (A2AAR) is recognised as a major mediator of anti-inflammatory responses. The A2AAR is widely expressed on cells of the immune system and numerous in ...

  3. Diabetic complications within the context of aging: Nicotinamide adenine dinucleotide redox, insulin C-peptide, sirtuin 1-liver kinase B1-adenosine monophosphate-activated protein kinase positive feedback and forkhead box O3.

    Ido, Yasuo

    2016-07-01

    Recent research in nutritional control of aging suggests that cytosolic increases in the reduced form of nicotinamide adenine dinucleotide and decreasing nicotinamide adenine dinucleotide metabolism plays a central role in controlling the longevity gene products sirtuin 1 (SIRT1), adenosine monophosphate-activated protein kinase (AMPK) and forkhead box O3 (FOXO3). High nutrition conditions, such as the diabetic milieu, increase the ratio of reduced to oxidized forms of cytosolic nicotinamide adenine dinucleotide through cascades including the polyol pathway. This redox change is associated with insulin resistance and the development of diabetic complications, and might be counteracted by insulin C-peptide. My research and others' suggest that the SIRT1-liver kinase B1-AMPK cascade creates positive feedback through nicotinamide adenine dinucleotide synthesis to help cells cope with metabolic stress. SIRT1 and AMPK can upregulate liver kinase B1 and FOXO3, key factors that help residential stem cells cope with oxidative stress. FOXO3 directly changes epigenetics around transcription start sites, maintaining the health of stem cells. 'Diabetic memory' is likely a result of epigenetic changes caused by high nutritional conditions, which disturb the quiescent state of residential stem cells and impair tissue repair. This could be prevented by restoring SIRT1-AMPK positive feedback through activating FOXO3. PMID:27181414

  4. Local adenosine receptor blockade accentuates the sympathetic responses to fatiguing exercise

    Cui, Jian; Leuenberger, Urs A.; Blaha, Cheryl; Yoder, Jonathan; Gao, Zhaohui; Sinoway, Lawrence I.

    2010-01-01

    The role adenosine plays in evoking the exercise pressor reflex in humans remains controversial. We hypothesized that localized forearm adenosine receptor blockade would attenuate muscle sympathetic nerve activity (MSNA) responses to fatiguing handgrip exercise in humans. Blood pressure (Finometer), heart rate, and MSNA from the peroneal nerve were assessed in 11 healthy young volunteers during fatiguing isometric handgrip, postexercise circulatory occlusion (PECO), and passive muscle stretch...

  5. Identification of a specific assembly of the G protein Golf as a critical and regulated module of dopamine and adenosine-activated cAMP pathways in the striatum

    Denis eHervé

    2011-08-01

    Full Text Available In the principal neurons of striatum (medium spiny neurons, MSNs, cAMP pathway is primarily activated through the stimulation of dopamine D1 and adenosine A2A receptors, these receptors being mainly expressed in striatonigral and striatopallidal MSNs, respectively. Since cAMP signaling pathway could be altered in various physiological and pathological situations, including drug addiction and Parkinson’s disease, it is of crucial importance to identify the molecular components involved in the activation of this pathway. In MSNs, cAMP pathway activation is not dependent on the classical Gs GTP-binding protein but requires a specific G protein subunit heterotrimer containing Galpha-olf/beta2/gamma7 in particular association with adenylate cyclase type 5. This assembly forms an authentic functional signaling unit since loss of one of its members leads to defects of cAMP pathway activation in response to D1 or A2A receptor stimulation, inducing dramatic impairments of behavioral responses dependent on these receptors. Interestingly, D1 receptor-dependent cAMP signaling is modulated by the neuronal levels of Galpha-olf, indicating that Galpha-olf represents the rate-limiting step in this signaling cascade and could constitute a critical element for regulation of D1 receptor responses. In both Parkinsonian patients and several animal models of Parkinson’s disease, the lesion of dopamine neurons produces a prolonged elevation of Galpha-olf levels. This observation gives an explanation for the cAMP pathway hypersensitivity to D1 stimulation, occurring despite an unaltered D1 receptor density. In conclusion, alterations in the highly specialized assembly of Galpha-olf/beta2/gamma7 subunits can happen in pathological conditions, such as Parkinson’s disease, and it could have important functional consequences in relation to changes in D1 receptor signaling in the striatum.

  6. Optical Aptasensors for Adenosine Triphosphate

    Ng, Stella; Lim, Hui Si; Ma, Qian; Gao, Zhiqiang

    2016-01-01

    Nucleic acids are among the most researched and applied biomolecules. Their diverse two- and three-dimensional structures in conjunction with their robust chemistry and ease of manipulation provide a rare opportunity for sensor applications. Moreover, their high biocompatibility has seen them being used in the construction of in vivo assays. Various nucleic acid-based devices have been extensively studied as either the principal element in discrete molecule-like sensors or as the main component in the fabrication of sensing devices. The use of aptamers in sensors - aptasensors, in particular, has led to improvements in sensitivity, selectivity, and multiplexing capacity for a wide verity of analytes like proteins, nucleic acids, as well as small biomolecules such as glucose and adenosine triphosphate (ATP). This article reviews the progress in the use of aptamers as the principal component in sensors for optical detection of ATP with an emphasis on sensing mechanism, performance, and applications with some discussion on challenges and perspectives. PMID:27446501

  7. NTS adenosine A2a receptors inhibit the cardiopulmonary chemoreflex control of regional sympathetic outputs via a GABAergic mechanism.

    Minic, Zeljka; O'Leary, Donal S; Scislo, Tadeusz J

    2015-07-01

    Adenosine is a powerful central neuromodulator acting via opposing A1 (inhibitor) and A2a (activator) receptors. However, in the nucleus of the solitary tract (NTS), both adenosine receptor subtypes attenuate cardiopulmonary chemoreflex (CCR) sympathoinhibition of renal, adrenal, and lumbar sympathetic nerve activity and attenuate reflex decreases in arterial pressure and heart rate. Adenosine A1 receptors inhibit glutamatergic transmission in the CCR pathway, whereas adenosine A2a receptors most likely facilitate release of an unknown inhibitory neurotransmitter, which, in turn, inhibits the CCR. We hypothesized that adenosine A2a receptors inhibit the CCR via facilitation of GABA release in the NTS. In urethane-chloralose-anesthetized rats (n = 51), we compared regional sympathetic responses evoked by stimulation of the CCR with right atrial injections of the 5-HT3 receptor agonist phenylbiguanide (1-8 μg/kg) before and after selective stimulation of NTS adenosine A2a receptors [microinjections into the NTS of CGS-21680 (20 pmol/50 nl)] preceded by blockade of GABAA or GABAB receptors in the NTS [bicuculline (10 pmol/100 nl) or SCH-50911 (1 nmol/100 nl)]. Blockade of GABAA receptors virtually abolished adenosine A2a receptor-mediated inhibition of the CCR. GABAB receptors had much weaker but significant effects. These effects were similar for the different sympathetic outputs. We conclude that stimulation of NTS adenosine A2a receptors inhibits CCR-evoked hemodynamic and regional sympathetic reflex responses via a GABA-ergic mechanism. PMID:25910812

  8. The adenosine A2B receptor is involved in anion secretion in human pancreatic duct Capan-1 epithelial cells.

    Hayashi, M; Inagaki, A; Novak, I; Matsuda, H

    2016-07-01

    Adenosine modulates a wide variety of biological processes via adenosine receptors. In the exocrine pancreas, adenosine regulates transepithelial anion secretion in duct cells and is considered to play a role in acini-to-duct signaling. To identify the functional adenosine receptors and Cl(-) channels important for anion secretion, we herein performed experiments on Capan-1, a human pancreatic duct cell line, using open-circuit Ussing chamber and gramicidin-perforated patch-clamp techniques. The luminal addition of adenosine increased the negative transepithelial potential difference (V te) in Capan-1 monolayers with a half-maximal effective concentration value of approximately 10 μM, which corresponded to the value obtained on whole-cell Cl(-) currents in Capan-1 single cells. The effects of adenosine on V te, an equivalent short-circuit current (I sc), and whole-cell Cl(-) currents were inhibited by CFTRinh-172, a cystic fibrosis transmembrane conductance regulator (CFTR) Cl(-) channel inhibitor. The adenosine A2B receptor agonist, BAY 60-6583, increased I sc and whole-cell Cl(-) currents through CFTR Cl(-) channels, whereas the A2A receptor agonist, CGS 21680, had negligible effects. The A2B receptor antagonist, PSB 603, inhibited the response of I sc to adenosine. Immunohistochemical analysis showed that the A2A and A2B receptors colocalized with Ezrin in the luminal membranes of Capan-1 monolayers and in rat pancreatic ducts. Adenosine elicited the whole-cell Cl(-) currents in guinea pig duct cells. These results demonstrate that luminal adenosine regulates anion secretion by activating CFTR Cl(-) channels via adenosine A2B receptors on the luminal membranes of Capan-1 cells. The present study endorses that purinergic signaling is important in the regulation of pancreatic secretion. PMID:26965147

  9. Involvement of adenosine A2a receptor in intraocular pressure decrease induced by 2-(1-octyn-1-yl)adenosine or 2-(6-cyano-1-hexyn-1-yl)adenosine.

    Konno, Takashi; Murakami, Akira; Uchibori, Takehiro; Nagai, Akihiko; Kogi, Kentaro; Nakahata, Norimichi

    2005-04-01

    The aim of the present study is to clarify the mechanism for the decrease in intraocular pressure by 2-alkynyladenosine derivatives in rabbits. The receptor binding analysis revealed that 2-(1-octyn-1-yl)adenosine (2-O-Ado) and 2-(6-cyano-1-hexyn-1-yl)adenosine (2-CN-Ado) selectively bound to the A(2a) receptor with a high affinity. Ocular hypotensive responses to 2-O-Ado and 2-CN-Ado were inhibited by the adenosine A(2a)-receptor antagonist 1,3,7-trimethyl-8-(3-chlorostyryl)xanthine (CSC), but not by the adenosine A(1)-receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine (DPCPX) or the adenosine A(2b)-receptor antagonist alloxazine. In addition, 2-O-Ado and 2-CN-Ado caused an increase in outflow facility, which was inhibited by CSC, but not by DPCPX or alloxazine. Moreover, 2-O-Ado and 2-CN-Ado increased cAMP in the aqueous humor, and the 2-O-Ado-induced an increase in cAMP was inhibited by CSC. These results suggest that 2-O-Ado and 2-CN-Ado reduced intraocular pressure via an increase in outflow facility. The ocular hypotension may be mainly mediated through the activation of adenosine A(2a) receptor, although a possible involvement of adenosine A(1) receptor cannot be completely ruled out. 2-O-Ado and 2-CN-Ado are useful lead compounds for the treatment of glaucoma. PMID:15821340

  10. 5’-Phosphate and 5’-Phosphonate Ester Derivatives of (N)-Methanocarba Adenosine with in Vivo Cardioprotective Activity

    Kumar, T. Santhosh; Yang, Tiehong; Mishra, Shilpi; Cronin, Chunxia; Charkaborty, Saibal; Shen, Jian-Bing; Liang, Bruce T.; Jacobson, Kenneth A.

    2013-01-01

    Activation of a cardiac myocyte P2X4 receptor protects in heart failure. 5’-Phosphonate and 5’-phosphate analogues of AMP containing a (N)-methanocarba (bicyclo[3.1.0]hexane) system could protect from heart failure by potentially activating this cardioprotective channel. Phosphoesters and phosphonodiesters were synthesized and administered in vivo via a mini-osmotic pump in a mouse ischemic heart failure model; most significantly increased intact heart contractile function (echocardiography) ...

  11. Inosine triphosphatase allele frequency and association with ribavirin-induced anaemia in Brazilian patients receiving antiviral therapy for chronic hepatitis C

    Nathália Delvaux

    2015-08-01

    Full Text Available Inosine triphosphatase (ITPA single nucleotide polymorphisms (SNPs are strongly associated with protection against ribavirin (RBV-induced anaemia in European, American and Asian patients; however, there is a paucity of data for Brazilian patients. The aim of this study was to evaluate the ITPA SNP (rs7270101/rs1127354 frequency in healthy and hepatitis C virus (HCV-infected patients from Brazil and the association with the development of severe anaemia during antiviral therapy. ITPA SNPs were determined in 200 HCV infected patients and 100 healthy individuals by sequencing. Biochemical parameters and haemoglobin (Hb levels were analysed in 97 patients who underwent antiviral therapy. A combination of AArs7270101+CCrs1127354 (100% ITPase activity was observed in 236/300 individuals. Anaemia was observed in 87.5% and 86.2% of treated patients with AA (rs7270101 and CC genotypes (rs1127354, respectively. Men with AA (rs7270101 showed a considerable reduction in Hb at week 12 compared to those with AC/CC (p = 0.1475. In women, there was no influence of genotype (p = 0.5295. For rs1127354, men with the CC genotype also showed a sudden reduction in Hb compared to those with AC. Allelic distribution of rs7270101 and rs1127354 shows high rates of the genotypes AA and CC, respectively, suggesting that the study population had a great propensity for developing RBV-induced anaemia. A progressive Hb reduction during treatment was observed; however, this reduction was greater in men at week 12 than in women.

  12. Adenosine triphosphate inhibition of yeast trehalase.

    Panek, A D

    1969-09-01

    Yeast trehalase has been found to be inhibited non-competitively by adenosine triphosphate. Such a biological control could explain the accumulation of trehalose during the stationary phase of the growth curve. PMID:5370287

  13. Adenosine signaling in striatal circuits and alcohol use disorders.

    Nam, Hyung Wook; Bruner, Robert C; Choi, Doo-Sup

    2013-09-01

    Adenosine signaling has been implicated in the pathophysiology of alcohol use disorders and other psychiatric disorders such as anxiety and depression. Numerous studies have indicated a role for A1 receptors (A1R) in acute ethanol-induced motor incoordination, while A2A receptors (A2AR) mainly regulate the rewarding effect of ethanol in mice. Recent findings have demonstrated that dampened A2AR-mediated signaling in the dorsomedial striatum (DMS) promotes ethanol-seeking behaviors. Moreover, decreased A2AR function is associated with decreased CREB activity in the DMS, which enhances goal-oriented behaviors and contributes to excessive ethanol drinking in mice. Interestingly, caffeine, the most commonly used psychoactive substance, is known to inhibit both the A1R and A2AR. This dampened adenosine receptor function may mask some of the acute intoxicating effects of ethanol. Furthermore, based on the fact that A2AR activity plays a role in goal-directed behavior, caffeine may also promote ethanol-seeking behavior. The A2AR is enriched in the striatum and exclusively expressed in striatopallidal neurons, which may be responsible for the regulation of inhibitory behavioral control over drug rewarding processes through the indirect pathway of the basal ganglia circuit. Furthermore, the antagonistic interactions between adenosine and dopamine receptors in the striatum also play an integral role in alcoholism and addiction-related disorders. This review focuses on regulation of adenosine signaling in striatal circuits and the possible implication of caffeine in goal-directed behaviors and addiction. PMID:23912595

  14. Electrocardiographic profile of adenosine pharmacological stress testing

    Sun, Hao; TIAN, YUEQIN; ZHENG, LIHUI; Pan, Qingrong; XIE, BOQIA

    2015-01-01

    Adenosine stress testing in conjunction with radionuclide myocardial perfusion imaging has become a common approach for the detection of coronary artery diseases in patients who are unable to perform adequate levels of exercise. However, specific electrocardiographic alterations during the test have been rarely described. Using a Chinese population, the aim of the present study was to provide a detailed electrocardiographic profile of adenosine stress testing. The study population included 1,...

  15. Aldehyde Dehydrogenase Type 2 Activation by Adenosine and Histamine Inhibits Ischemic Norepinephrine Release in Cardiac Sympathetic Neurons: Mediation by Protein Kinase Cε

    Robador, Pablo A.; Seyedi, Nahid; Chan, Noel Yan-Ki; Koda, Kenichiro; Levi, Roberto

    2012-01-01

    During myocardial ischemia/reperfusion, lipid peroxidation leads to the formation of toxic aldehydes that contribute to ischemic dysfunction. Mitochondrial aldehyde dehydrogenase type 2 (ALDH2) alleviates ischemic heart damage and reperfusion arrhythmias via aldehyde detoxification. Because excessive norepinephrine release in the heart is a pivotal arrhythmogenic mechanism, we hypothesized that neuronal ALDH2 activation might diminish ischemic norepinephrine release. Incubation of cardiac sym...

  16. Increased Na+/K(+)-pump activity and adenosine triphosphate utilization after compound 48/80-induced histamine secretion from rat mast cells

    Johansen, Torben; Praetorius, Birger Hans

    1994-01-01

    -production were measured by the bioluminescence technique (firefly lantern) and by measurement of the lactate production under anaerobic conditions (antimycin A, oligomycin), respectively. There was an increased requirement for ATP after the secretory response associated with an increased activity of the Na...

  17. Nucleophilic behavior of lysine-501 of the alpha-polypeptide of sodium and potassium ion activated adenosinetriphosphatase consistent with a role in binding adenosine triphosphate

    An immunoadsorbent specific for the carboxy-terminal sequence -GAPER, which comprises residues 502-506 of the alpha-polypeptide of ovine sodium and potassium ion activated adenosinetriphosphatase [(Na+ + K+)-ATPase], was used to isolate the products of the reaction between the lysine immediately preceding this sequence in the intact protein and either [3H]acetic anhydride or fluorescein 5'-isothiocyanate. Changes in the apparent nucleophilicity of this lysine, Lys501, were observed with both reagents when ATP was bound by the intact, native enzyme poised in the E1 conformation or when the structure of the enzyme was changed from the E1 conformation into the E2-P conformation. With both reagents, a decrease of more than 4-fold in the yield of incorporation occurred during the former change, but a decrease of only 2-fold occurred during the latter. Because a much larger decrease occurred when ATP was bound in the absence of a conformational change than occurred when a major conformational change took place in the absence of the occupation of the active site, these changes in the incorporation of [3H]acetyl suggest that Lys501 from the alpha polypeptide is directly involved in binding ATP within the active site of (Na+ + K+)-ATPase. The immunochemical reactions between the specific polyclonal antibodies raised against the sequence-GAPER and denatured or enzymically active (Na+ + K+)-ATPase were also investigated. Western blots and the inhibition of enzymic activity caused by the antibody have shown that it can bind to both the denatured and the native form of the alpha-polypeptide, respectively

  18. Online cleanup of accelerated solvent extractions for determination of adenosine 5'-triphosphate (ATP), adenosine 5'-diphosphate (ADP), and adenosine 5'-monophosphate (AMP) in royal jelly using high-performance liquid chromatography.

    Xue, Xiaofeng; Wang, Feng; Zhou, Jinhui; Chen, Fang; Li, Yi; Zhao, Jing

    2009-06-10

    Determination of the levels of adenosine 5'-triphosphate (ATP), adenosine 5'-diphosphate (ADP), and adenosine 5'-monophosphate (AMP) in royal jelly is important for the study of its pharmacological activities, health benefits, and adenosine phosphate degradation. In this study was developed a novel method to determine ATP, ADP, and AMP levels in royal jelly using accelerated solvent extraction (ASE) followed by online cleanup and high-performance liquid chromatography (HPLC) with diode array detection (DAD). The optimum extraction conditions were obtained using an 11 mL ASE cell, ethanol/water (5:5 v/v) as the extraction solvent, 1500 psi, 80 degrees C, a 5 min static time, and a 60% flush volume. Optimum separation of the three compounds was achieved in extraction procedures developed here were compared with the classical adenosine phosphate extraction procedures (perchloric acid). The results indicate that the two techniques are similar in terms of recovery and reproducibility, but when other factors such as extraction time, environmental protection, and worker's health are considered, ASE is preferable to the classical extraction method. With this ASE-HPLC method, a minisurvey of ATP, ADP, and AMP levels in 15 samples of royal jelly of different origins was performed. Sample results indicated that the AMP concentration was 24.2-2214.4 mg kg(-1), whereas ATP and ADP were not detectable or present only at low levels. PMID:19435312

  19. KATP-channel activation: effects on myocardial recovery from ischaemia and role in the cardioprotective response to adenosine A1-receptor stimulation

    Ford, William R.; Lopaschuk, Gary D.; Schulz, Richard; Clanachan, Alexander S

    1998-01-01

    Optimization of myocardial energy substrate metabolism improves the recovery of mechanical function of the post-ischaemic heart. This study investigated the role of KATP-channels in the regulation of the metabolic and mechanical function of the aerobic and post-ischaemic heart by measuring the effects of the selective KATP-channel activator, cromakalim, and the effects of the KATP-channel antagonist, glibenclamide, in rat fatty acid perfused, working hearts in vitro. The role of KATP channels...

  20. Pretreatment of rats with increased bioavailable berberine attenuates cerebral ischemia-reperfusion injury via down regulation of adenosine-5'monophosphate kinase activity.

    Chen, Weijia; Wei, Shengnan; Yu, Yang; Xue, Huan; Yao, Fan; Zhang, Ming; Xiao, Jun; Hatch, Grant M; Chen, Li

    2016-05-15

    Berberine (BBR) exhibits multiple beneficial biological effects. However, poor bioavailability of BBR has limited its clinical application. We previously demonstrated that solid dispersion of BBR with sodium caprate (HGSD) remarkably improves its bioavailability. We examined whether this increased bioavailability of BBR could protect the brain from ischemia-reperfusion (IR) induced injury. Rats treated with HGSD, SC and saline for 7 days then subjected to cerebral ischemia reperfusion by middle cerebral artery occlusion for 2h followed 12h reperfusion. Neurological deficit scores, infarct size, SOD, MDA and NO levels were examined. P-AMPK, Bax, cleaved-Caspase-3 in brain was determined. To further probe for the mechanism of beneficial effect of HGSD, PC12 cells were incubated with serum from control or HGSD pretreated animals, incubated with 300μM H2O2 to induce apoptosis. Caspase-3 activity and cell apoptosis was evaluated. HGSD pretreatment significantly attenuated neurological deficit scores, reduced infarct size, increased SOD and decreased MDA and NO after cerebral IR injury compared to controls. Meanwhile, HGSD pretreatment significantly reduced expression of p-AMPK, Bax, cleaved-Caspase-3 after cerebral IR injury. Sodium caprate (100mg/kg/d) pretreatment alone did not exhibit any of these beneficial effects. PC12 cell apoptosis was attenuated when cells were cultured with HGSD serum compared to control. The presence of AMPK activator (AICAR) attenuated whereas AMPK inhibitor (Compound C) augmented the protective effect of HGSD serum on PC12 cell apoptosis.The results indicate that HGSD-pretreatment of rats protects the brain from ischemia-reperfusion injury and the mechanism is due to its anti-apoptotic effect mediated by decreased activation of AMPK. PMID:26957053

  1. Liver MicroRNA-291b-3p Promotes Hepatic Lipogenesis through Negative Regulation of Adenosine 5'-Monophosphate (AMP)-activated Protein Kinase α1.

    Meng, Xiangyu; Guo, Jun; Fang, Weiwei; Dou, Lin; Li, Meng; Huang, Xiuqing; Zhou, Shutong; Man, Yong; Tang, Weiqing; Yu, Liqing; Li, Jian

    2016-05-13

    In a microarray study, we found that hepatic miR-291b-3p was significantly increased in leptin-receptor-deficient type 2 mice (db/db), a mouse model of diabetes. The function of miR-291b-3p is unknown. The potential role of miR-291b-3p in regulating hepatic lipid metabolism was explored in this study. High-fat diet (HFD)- and chow-fed mice were injected with an adenovirus expressing a miR-291b-3p inhibitor and a miR-291b-3p mimic through the tail vein. Hepatic lipids and lipogenic gene expression were analyzed. Additionally, gain- and loss-of-function studies were performed in vitro to identify direct targets of miR-291b-3p. MiR-291b-3p expression and the protein levels of sterol regulatory element-binding protein 1 (SREBP1) and fatty acid synthase (FAS) were increased in the steatotic liver of db/db mice and HFD-fed mice versus their respective controls. Inhibition of hepatic miR-291b-3p expression prevented increases in hepatic lipogenesis and steatosis in HFD-fed mice. The opposite was observed when miR-291b-3p was overexpressed in the livers of chow-fed C57BL/6J wild-type mice. In vitro studies revealed that silencing of miR-291b-3p in NCTC1469 hepatic cells ameliorated oleic acid/palmitic acid mixture-induced elevation of cellular triglycerides. Importantly, we identified AMP-activated protein kinase (AMPK)-α1 as a direct target of miR-291b-3p. Using metformin, an activator of AMPK, we showed that AMPK activation-induced inhibition of hepatic lipid accumulation was accompanied by reduced expression of miR-291b-3p in the liver. Liver miR-291b-3p promoted hepatic lipogenesis and lipid accumulation in mice. AMPKα1 is a direct target of miR-291b-3p. In conclusion, our findings indicate that miR-291b-3p promotes hepatic lipogenesis by suppressing AMPKα1 expression and activity, indicating the therapeutic potential of miR-291b-3p inhibitors in fatty liver disease. PMID:27013659

  2. Urinary cyclic adenosine 3',5'-monophosphate response in McCune-Albright syndrome: clinical evidence for altered renal adenylate cyclase activity.

    Zung, A; Chalew, S A; Schwindinger, W F; Levine, M A; Phillip, M; Jara, A; Counts, D R; Kowarski, A A

    1995-12-01

    The recent finding of an activating mutation in the Gs alpha protein, the protein that couples receptors to stimulation of adenylate cyclase, from endocrine and nonendocrine tissues of patients with McCune-Albright syndrome (MAS) suggests that alterations in adenylate cyclase activity may account for the clinical abnormalities in these patients. Many patients with MAS have hypophosphatemia. This may result from the presence of the activating Gs alpha mutation in proximal renal tubules or the elaboration of a phosphaturic factor from fibrous dysplasia. We, therefore, sought to characterize renal cAMP generation and phosphate handling in MAS patients. Intravenous infusion of PTH is a classic clinical test used to evaluate hormonal responsiveness of renal proximal tubule adenylate cyclase and examine PTH-dependent phosphate clearance. We performed PTH infusion in 6 MAS patients, 10 normal subjects, and 6 patients with pseudohypoparathyroidism (PHP). The basal urinary cAMP (UcAMP) level in the MAS group [5.5 +/- 2.6 nmol/dL glomerular filtration (GF)] was elevated (P PHP (1.9 +/- 0.6 nmol/dL GF). However, PTH-stimulated peak UcAMP (15.0 +/- 7.0 nmol/dL GF) and the peak/basal UcAMP ratio (3.1 +/- 1.7) in MAS were significantly lower than the respective values in normal subjects (30.8 +/- 16.9 nmol/dL GF and 9.3 +/- 2.9; P PHP (respectively, 3.1 +/- 1.5 nmol/dL GF and 2.0 +/- 1.7). By contrast, the PTH-induced phosphaturic response in MAS patients was similar to that in the normal subjects. Our study provides clinical evidence that MAS patients have altered renal adenylate cyclase activity, manifested by an elevated basal UcAMP, but a blunted UcAMP response to PTH stimulation. These observations are presumably due to a mutation in the Gs alpha protein in the renal tubules. Despite the blunted UcAMP excretion, the phosphaturic response to PTH in MAS patients is intact. PMID:8530601

  3. Adenosine modulates hypoxia-induced responses in rat PC12 cells via the A2A receptor.

    Kobayashi, S; Conforti, L; Pun, R Y; Millhorn, D E

    1998-04-01

    1. The present study was undertaken to determine the role of adenosine in mediating the cellular responses to hypoxia in rat phaeochromocytoma (PC12) cells, an oxygen-sensitive clonal cell line. 2. Reverse transcriptase polymerase chain reaction studies revealed that PC12 cells express adenosine deaminase (the first catalysing enzyme of adenosine degradation) and the A2A and A2B adenosine receptors, but not the A1 or A3 adenosine receptors. 3. Whole-cell current- and voltage-clamp experiments showed that adenosine attenuated the hypoxia-induced membrane depolarization. The hypoxia-induced suppression of the voltage-sensitive potassium current (IK(V)) was markedly reduced by adenosine. Furthermore, extracellularly applied adenosine increased the peak amplitudes of IK(V) in a concentration-dependent manner. This increase was blocked by pretreatment not only with a non-specific adenosine receptor antagonist, 8-phenyltheophylline (8-PT), but also with a selective A2A receptor antagonist, ZM241385. 4. Ca2+ imaging studies using fura-2 acetoxymethyl ester (fura-2 AM) revealed that the increase in intracellular free Ca2+ during hypoxic exposure was attenuated significantly by adenosine. Voltage-clamp studies showed that adenosine inhibited the voltage-dependent Ca2+ currents (ICa) in a concentration-dependent fashion. This inhibition was also abolished by both 8-PT and ZM241385. 5. The modulation of both IK(V) and ICa by adenosine was prevented by intracellular application of an inhibitor of protein kinase A (PKA), PKA inhibitor fragment (6-22) amide. In addition, the effect of adenosine on either IK(V) or ICa was absent in PKA-deficient PC12 cells. 6. These results indicate that the modulatory effects of adenosine on the hypoxia-induced membrane responses of PC12 cells are likely to be mediated via activation of the A2A receptor, and that the PKA pathway is required for these modulatory actions. We propose that this modulation serves to regulate membrane excitability in

  4. Genistein suppresses tumor necrosis factor α-induced inflammation via modulating reactive oxygen species/Akt/nuclear factor ΚB and adenosine monophosphate-activated protein kinase signal pathways in human synoviocyte MH7A cells

    Li J

    2014-03-01

    inhibit IL-1β, IL-6, and IL-8 production induced by TNF-α. In addition, we also found that pretreatment with the adenosine monophosphate-activated protein kinase (AMPK agonist 5-aminoimidazole-4-carboxamide-1-β-D-ribofuranoside obviously inhibited TNF-α-induced proinflammatory cytokine production. These observations suggest that the inhibitory effect of genistein on TNF-α-induced proinflammatory cytokine production is dependent on AMPK activation. Conclusion: These findings indicate that genistein suppressed TNF-α-induced inflammation by inhibiting the ROS/Akt/NF-ΚB pathway and promoting AMPK activation in MH7A cells. Keywords: genistein, rheumatoid arthritis, cytokine, signal transduction, inflammation

  5. Post-Meal Responses of Elongation Factor 2 (eEF2 and Adenosine Monophosphate-Activated Protein Kinase (AMPK to Leucine and Carbohydrate Supplements for Regulating Protein Synthesis Duration and Energy Homeostasis in Rat Skeletal Muscle

    Donald K. Layman

    2012-11-01

    Full Text Available Previous research demonstrates that the anabolic response of muscle protein synthesis (MPS to a meal is regulated at the level of translation initiation with signals derived from leucine (Leu and insulin to activate mTORC1 signaling. Recent evidence suggests that the duration of the meal response is limited by energy status of the cell and inhibition of translation elongation factor 2 (eEF2. This study evaluates the potential to extend the anabolic meal response with post-meal supplements of Leu or carbohydrates. Adult (~256 g male Sprague-Dawley rats were food deprived for 12 h, then either euthanized before a standard meal (time 0 or at 90 or 180 min post-meal. At 135 min post-meal, rats received one of five oral supplements: 270 mg leucine (Leu270, 80:40:40 mg leucine, isoleucine, and valine (Leu80, 2.63 g carbohydrates (CHO2.6, 1 g carbohydrates (CHO1.0, or water (Sham control. Following the standard meal, MPS increased at 90 min then declined to pre-meal baseline at 180 min. Rats administered Leu270, Leu80, CHO2.6, or CHO1.0 maintained elevated rates of MPS at 180 min, while Sham controls declined from peak values. Leu80 and CHO1.0 treatments maintained MPS, but with values intermediate between Sham controls and Leu270 and CHO2.6 supplements. Consistent with MPS findings, the supplements maintained elongation activity and cellular energy status by preventing increases in AMP/ATP and phosphorylation of adenosine monophosphate-activated protein kinase (AMPK, acetyl-CoA carboxylase ACC and eEF2. The impact of the supplements on MPS and cellular energy status was in proportion to the energy content within the individual treatments (i.e., Leu270 > Leu80; CHO2.6 > CHO1.0, but the Leu supplements produced a disproportionate anabolic stimulation of MPS, eEF2 and energy status with significantly lower energy content. In summary, the incongruity between MPS and translation initiation at 180 min reflects a block in translation elongation due to

  6. Structural basis of the substrate specificity of Bacillus cereus adenosine phosphorylase

    Dessanti, Paola [Cornell University, Ithaca, NY 14853-1301 (United States); Università di Sassari, (Italy); Zhang, Yang [Cornell University, Ithaca, NY 14853-1301 (United States); Allegrini, Simone [Università di Sassari, (Italy); Tozzi, Maria Grazia [Università di Pisa, (Italy); Sgarrella, Francesco [Università di Sassari, (Italy); Ealick, Steven E., E-mail: see3@cornell.edu [Cornell University, Ithaca, NY 14853-1301 (United States)

    2012-03-01

    Adenosine phosphorylase from B. cereus shows a strong preference for adenosine over other 6-oxopurine nucleosides. Mutation of Asp204 to asparagine reduces the efficiency of adenosine cleavage but does not affect inosine cleavage, effectively reversing the substrate specificity. The structures of D204N complexes explain these observations. Purine nucleoside phosphorylases catalyze the phosphorolytic cleavage of the glycosidic bond of purine (2′-deoxy)nucleosides, generating the corresponding free base and (2′-deoxy)ribose 1-phosphate. Two classes of PNPs have been identified: homotrimers specific for 6-oxopurines and homohexamers that accept both 6-oxopurines and 6-aminopurines. Bacillus cereus adenosine phosphorylase (AdoP) is a hexameric PNP; however, it is highly specific for 6-aminopurines. To investigate the structural basis for the unique substrate specificity of AdoP, the active-site mutant D204N was prepared and kinetically characterized and the structures of the wild-type protein and the D204N mutant complexed with adenosine and sulfate or with inosine and sulfate were determined at high resolution (1.2–1.4 Å). AdoP interacts directly with the preferred substrate through a hydrogen-bond donation from the catalytically important residue Asp204 to N7 of the purine base. Comparison with Escherichia coli PNP revealed a more optimal orientation of Asp204 towards N7 of adenosine and a more closed active site. When inosine is bound, two water molecules are interposed between Asp204 and the N7 and O6 atoms of the nucleoside, thus allowing the enzyme to find alternative but less efficient ways to stabilize the transition state. The mutation of Asp204 to asparagine led to a significant decrease in catalytic efficiency for adenosine without affecting the efficiency of inosine cleavage.

  7. Structural basis of the substrate specificity of Bacillus cereus adenosine phosphorylase

    Adenosine phosphorylase from B. cereus shows a strong preference for adenosine over other 6-oxopurine nucleosides. Mutation of Asp204 to asparagine reduces the efficiency of adenosine cleavage but does not affect inosine cleavage, effectively reversing the substrate specificity. The structures of D204N complexes explain these observations. Purine nucleoside phosphorylases catalyze the phosphorolytic cleavage of the glycosidic bond of purine (2′-deoxy)nucleosides, generating the corresponding free base and (2′-deoxy)ribose 1-phosphate. Two classes of PNPs have been identified: homotrimers specific for 6-oxopurines and homohexamers that accept both 6-oxopurines and 6-aminopurines. Bacillus cereus adenosine phosphorylase (AdoP) is a hexameric PNP; however, it is highly specific for 6-aminopurines. To investigate the structural basis for the unique substrate specificity of AdoP, the active-site mutant D204N was prepared and kinetically characterized and the structures of the wild-type protein and the D204N mutant complexed with adenosine and sulfate or with inosine and sulfate were determined at high resolution (1.2–1.4 Å). AdoP interacts directly with the preferred substrate through a hydrogen-bond donation from the catalytically important residue Asp204 to N7 of the purine base. Comparison with Escherichia coli PNP revealed a more optimal orientation of Asp204 towards N7 of adenosine and a more closed active site. When inosine is bound, two water molecules are interposed between Asp204 and the N7 and O6 atoms of the nucleoside, thus allowing the enzyme to find alternative but less efficient ways to stabilize the transition state. The mutation of Asp204 to asparagine led to a significant decrease in catalytic efficiency for adenosine without affecting the efficiency of inosine cleavage

  8. Adenosine induces cell cycle arrest and apoptosis via cyclinD1/Cdk4 and Bcl-2/Bax pathways in human ovarian cancer cell line OVCAR-3.

    Shirali, Saeid; Aghaei, Mahmoud; Shabani, Mahdi; Fathi, Mojtaba; Sohrabi, Majid; Moeinifard, Marzieh

    2013-04-01

    Adenosine is a regulatory molecule with widespread physiological effects in almost every cells and acts as a potent regulator of cell growth. Adenosine has been shown to inhibit cell growth and induce apoptosis in the several cancer cells via caspase activation and Bcl-2/Bax pathway. The present study was designed to understand the mechanism underlying adenosine-induced apoptosis in the OVCAR-3 human ovarian cancer cells. MTT viability, BrdU and cell counting assays were used to study the cell proliferation effect of adenosine in presence of adenosine deaminase inhibitor and the nucleoside transporter inhibitor. Cell cycle analysis, propidium iodide and annexin V staining, caspase-3 activity assay, cyclinD1, Cdk4, Bcl-2 and Bax protein expressions were assessed to detect apoptosis. Adenosine significantly inhibited cell proliferation in a concentration-dependent manner in OVCAR-3 cell line. Adenosine induced cell cycle arrest in G0/G1 phase via Cdk4/cyclinD1-mediated pathway. Adenosine induced apoptosis, which was determined by Annexin V-FITC staining and increased sub-G1 population. Moreover, down-regulation of Bcl-2 protein expression, up-regulation of Bax protein expression and activation of caspase-3 were observed in response to adenosine treatment. The results of this study suggest that extracellular adenosine induced G1 cell cycle arrest and apoptosis in ovarian cancer cells via cyclinD1/ Cdk4 and Bcl-2/Bax pathways and caspase-3 activation. These data might suggest that adenosine could be used as an agent for the treatment of ovarian cancer. PMID:23345014

  9. 2'-O methylation of internal adenosine by flavivirus NS5 methyltransferase.

    Hongping Dong

    Full Text Available RNA modification plays an important role in modulating host-pathogen interaction. Flavivirus NS5 protein encodes N-7 and 2'-O methyltransferase activities that are required for the formation of 5' type I cap (m(7GpppAm of viral RNA genome. Here we reported, for the first time, that flavivirus NS5 has a novel internal RNA methylation activity. Recombinant NS5 proteins of West Nile virus and Dengue virus (serotype 4; DENV-4 specifically methylates polyA, but not polyG, polyC, or polyU, indicating that the methylation occurs at adenosine residue. RNAs with internal adenosines substituted with 2'-O-methyladenosines are not active substrates for internal methylation, whereas RNAs with adenosines substituted with N⁶-methyladenosines can be efficiently methylated, suggesting that the internal methylation occurs at the 2'-OH position of adenosine. Mass spectroscopic analysis further demonstrated that the internal methylation product is 2'-O-methyladenosine. Importantly, genomic RNA purified from DENV virion contains 2'-O-methyladenosine. The 2'-O methylation of internal adenosine does not require specific RNA sequence since recombinant methyltransferase of DENV-4 can efficiently methylate RNAs spanning different regions of viral genome, host ribosomal RNAs, and polyA. Structure-based mutagenesis results indicate that K61-D146-K181-E217 tetrad of DENV-4 methyltransferase forms the active site of internal methylation activity; in addition, distinct residues within the methyl donor (S-adenosyl-L-methionine pocket, GTP pocket, and RNA-binding site are critical for the internal methylation activity. Functional analysis using flavivirus replicon and genome-length RNAs showed that internal methylation attenuated viral RNA translation and replication. Polymerase assay revealed that internal 2'-O-methyladenosine reduces the efficiency of RNA elongation. Collectively, our results demonstrate that flavivirus NS5 performs 2'-O methylation of internal adenosine of

  10. Lack of effect of adenosine on the function of rodent osteoblasts and osteoclasts in vitro.

    Hajjawi, Mark O R; Patel, Jessal J; Corcelli, Michelangelo; Arnett, Timothy R; Orriss, Isabel R

    2016-06-01

    Extracellular ATP, signalling through P2 receptors, exerts well-documented effects on bone cells, inhibiting mineral deposition by osteoblasts and stimulating the formation and resorptive activity of osteoclasts. The aims of this study were to determine the potential osteotropic effects of adenosine, the hydrolysis product of ATP, on primary bone cells in vitro. We determined the effect of exogenous adenosine on (1) the growth, alkaline phosphatase (TNAP) activity and bone-forming ability of osteoblasts derived from the calvariae of neonatal rats and mice and the marrow of juvenile rats and (2) the formation and resorptive activity of osteoclasts from juvenile mouse marrow. Reverse transcription polymerase chain reaction (RT-PCR) analysis showed marked differences in the expression of P1 receptors in osteoblasts from different sources. Whilst mRNA for the A1 and A2B receptors was expressed by all primary osteoblasts, A2A receptor expression was limited to rat bone marrow and mouse calvarial osteoblasts and the A3 receptor to rat bone marrow osteoblasts. We found that adenosine had no detectable effects on cell growth, TNAP activity or bone formation by rodent osteoblasts in vitro. The analogue 2-chloroadenosine, which is hydrolysed more slowly than adenosine, had no effects on rat or mouse calvarial osteoblasts but increased TNAP activity and bone formation by rat bone marrow osteoblasts by 30-50 % at a concentration of 1 μM. Osteoclasts were found to express the A2A, A2B and A3 receptors; however, neither adenosine (≤100 μM) nor 2-chloroadenosine (≤10 μM) had any effect on the formation or resorptive activity of mouse osteoclasts in vitro. These results suggest that adenosine, unlike ATP, is not a major signalling molecule in the bone. PMID:26861849

  11. Effects of synthetic and naturally occurring flavonoids on Na+, K+-ATPase: Aspects of the structure-activity relationship and action mechanism

    A comparative study was made of the effects of 15 synthetic and naturally occurring flavonoids on the hydrolytic activity of Na+, K+ -adenosine triphosphatase (ATPase). Twelve of the flavonoids examined were mono-hydroxy or mono-methoxy derivatives. All inhibited Na+, K+ -ATPase from dog kidney cortex when present at concentrations from 40-1000 μM. Flavones possessing cyclohexyl instead of the phenyl group were the most potent with IC50 at 257-320 μM. Structure-activity relationships were observed among the following mono-substituted flavones as: (i) 2-cyclohexyl-benzopyran-4-one much-gt 2-phenyl-benzopyran-4-one; (ii) 2-cyclohexyl-7-hydroxybenzopyran-4-one > 2-cyclohexyl-6-hydroxy-benzopyran-4-one > 2-cyclohexyl-5-hydroxybenzopyran-4-one. Some flavonoids showing potent inhibitory activity were also examined for ouabain-displacement activity on human erythrocytes. Hardly and of the flavonoids were able to block [3H] ouabain binding to erythrocytes. These results suggest that the mechanism by which flavonoid block Na+, K+ -ATPase is not related to the cardiac glycoside-specific binding site(s) of this enzyme

  12. Internalization and desensitization of adenosine receptors

    Klaasse, Elisabeth C.; IJzerman, Adriaan P.; de Grip, Willem J.; Beukers, Margot W.

    2007-01-01

    Until now, more than 800 distinct G protein-coupled receptors (GPCRs) have been identified in the human genome. The four subtypes of the adenosine receptor (A1, A2A, A2B and A3 receptor) belong to this large family of GPCRs that represent the most widely targeted pharmacological protein class. Since adenosine receptors are widespread throughout the body and involved in a variety of physiological processes and diseases, there is great interest in understanding how the different subtypes are re...

  13. Bradykinin and adenosine receptors mediate desflurane induced postconditioning in human myocardium: role of reactive oxygen species

    Gérard Jean-Louis

    2010-07-01

    Full Text Available Abstract Background Desflurane during early reperfusion has been shown to postcondition human myocardium, in vitro. We investigated the role of adenosine and bradykinin receptors, and generation of radical oxygen species in desflurane-induced postconditioning in human myocardium. Methods We recorded isometric contraction of human right atrial trabeculae hanged in an oxygenated Tyrode's solution (34 degrees Celsius, stimulation frequency 1 Hz. After a 30-min hypoxic period, desflurane 6% was administered during the first 5 min of reoxygenation. Desflurane was administered alone or with pretreatment of N-mercaptopropionylglycine, a reactive oxygen species scavenger, 8-(p-Sulfophenyltheophylline, an adenosine receptor antagonist, HOE140, a selective B2 bradykinin receptor antagonist. In separate groups, adenosine and bradykinin were administered during the first minutes of reoxygenation alone or in presence of N-mercaptopropionylglycine. The force of contraction of trabeculae was recorded continuously. Developed force at the end of a 60-min reoxygenation period was compared (mean ± standard deviation between the groups by a variance analysis and post hoc test. Results Desflurane 6% (84 ± 6% of baseline enhanced the recovery of force after 60-min of reoxygenation as compared to control group (51 ± 8% of baseline, P N-mercaptopropionylglycine (54 ± 3% of baseline, 8-(p-Sulfophenyltheophylline (62 ± 9% of baseline, HOE140 (58 ± 6% of baseline abolished desflurane-induced postconditioning. Adenosine (80 ± 9% of baseline and bradykinin (83 ± 4% of baseline induced postconditioning (P vs control, N-mercaptopropionylglycine abolished the beneficial effects of adenosine and bradykinin (54 ± 8 and 58 ± 5% of baseline, respectively. Conclusions In vitro, desflurane-induced postconditioning depends on reactive oxygen species production, activation of adenosine and bradykinin B2 receptors. And, the cardioprotective effect of adenosine and bradykinin

  14. Lower frequency of the low activity adenosine deaminase allelic variant (ADA1*2 in schizophrenic patients Diminuição da frequência da variante alélica de baixa atividade da adenosina desaminase (ADA1*2 em pacientes esquizofrênicos

    Gustavo Pimentel Dutra

    2010-09-01

    Full Text Available OBJECTIVE: Adenosine may play a role in the pathophysiology of schizophrenia, since it modulates the release of several neurotransmitters such as glutamate, dopamine, serotonin and acetylcholine, decreases neuronal activity by pos-synaptic hyperpolarization and inhibits dopaminergic activity. Adenosine deaminase participates in purine metabolism by converting adenosine into inosine. The most frequent functional polymorphism of adenosine deaminase (22G→A (ADA1*2 exhibits 20-30% lower enzymatic activity in individuals with the G/A genotype than individuals with the G/G genotype. The aim of this study was to evaluate the ADA polymorphism 22G→A (ADA1*2 in schizophrenic patients and healthy controls. METHOD: The genotypes of the ADA 22G→A were identified with allele-specific PCR strategy in 152 schizophrenic patients and 111 healthy individuals. RESULTS: A significant decrease in the frequency of the G/A genotype was seen in schizophrenic patients (7/152 - 4.6% relative to controls (13/111 - 11.7%, p = 0.032, OR = 2.6. CONCLUSION: These results suggest that the G/A genotype associated with low adenosine deaminase activity and, supposingly, with higher adenosine levels is less frequent among schizophrenic patients.OBJETIVO: A adenosina pode ter um papel importante na fisiopatologia da esquizofrenia, uma vez que modula a liberação de vários neurotransmissores, tais como glutamato, dopamina, serotonina e acetilcolina, diminui a atividade neuronal por hiperpolarização pós-sináptica e inibe a atividade dopaminérgica. A adenosina desaminase participa do metabolismo das purinas pela conversão de adenosina em inosina. O mais frequente polimorfismo funcional da adenosina desaminase (22G →A (ADA1*2 exibe uma diminuição de 20-30% da atividade funcional em indivíduos com genótipo G/A quando comparados com indivíduos com o genótipo G/G. O objetivo deste estudo foi avaliar o polimorfismo 22G→A (ADA1*2 em pacientes esquizofrênicos e em

  15. 21 CFR 864.7040 - Adenosine triphosphate release assay.

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Adenosine triphosphate release assay. 864.7040 Section 864.7040 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES... Adenosine triphosphate release assay. (a) Identification. An adenosine triphosphate release assay is...

  16. Study of Ectonucleotidases and Adenosine Deaminases in Drosophila

    PREUER, Kristina

    2013-01-01

    Extracellular adenosine triphosphate and extracellular adenosine are important regulatory molecules in the human immune system. The concentrations of these molecules are in turn regulated by ectonucleotidases and adenosine deaminases. In this thesis I attempt to test the gene silencing efficiency of RNA interference for three different genes coding for such enzymes in the model organism Drosophila melanogaster.

  17. Is adenosine a modulator of peripheral vasoconstrictor responses?

    Dayan, Lior; Brill, Silviu; Hochberg, Uri; Jacob, Giris

    2016-01-01

    Background Local vasoconstrictor reflexes, the vascular myogenic response (VMR) and the veno-arterial reflex (VAR) are necessary for the maintenance of regional blood flow and systemic arterial blood pressure during orthostatic stress. Their molecular mechanism is unknown. We postulated that adenosine is involved in the activation of these local reflexes. Methods This hypothesis was tested in 10 healthy male volunteers (age 29 ± 3 years, BMI 24 ± 1 kg/m2). We used veno-occlusive plethysmograp...

  18. Internalization and desensitization of adenosine receptors.

    Klaasse, E.C.; IJzerman, A.P.; Grip, W.J. de; Beukers, M.W.

    2008-01-01

    Until now, more than 800 distinct G protein-coupled receptors (GPCRs) have been identified in the human genome. The four subtypes of the adenosine receptor (A(1), A(2A), A(2B) and A(3) receptor) belong to this large family of GPCRs that represent the most widely targeted pharmacological protein clas

  19. Differential modulation of ATP-induced calcium signalling by A1 and A2 adenosine receptors in cultured cortical astrocytes

    Alloisio, Susanna; Cugnoli, Carlo; Ferroni, Stefano; Nobile, Mario

    2004-01-01

    Despite the accumulating evidence that under various pathological conditions the extracellular elevation of adenine-based nucleotides and nucleosides plays a key role in the control of astroglial reactivity, how these signalling molecules interact in the regulation of astrocyte function is still largely elusive.The action of the nucleoside adenosine in the modulation of the intracellular calcium signalling ([Ca2+]i) elicited by adenosine 5′-triphosphate (ATP)-induced activation of P2 purinoce...

  20. Adenosine inhibitory effect on enhanced growth of aortic smooth muscle cells from streptozotocin-induced diabetic rats.

    Parés-Herbuté, N.; Hillaire-Buys, D.; Etienne, P.; Gross, R.; Loubatières-Mariani, M. M.; MONNIER, L.

    1996-01-01

    1. There is evidence to suggest that adenosine may regulate arterial smooth muscle cell (SMC) growth and proliferation, which is a key event in atherogenesis. This regulation may be mediated via adenylate cyclase. As diabetes is a known risk factor for atherosclerosis, we investigated the growth of aortic SMC from diabetic rats in primary culture and their sensitivity to adenosine and to adenylate cyclase activity. 2. Diabetes was induced with streptozotocin (STZ, 66 mg kg-1, i.p.) Aortic SMC...

  1. Radio-chromatographic determination of plasmatic adenosine deaminase (A.D.); Determination radiochromatographique de l'adenosine deaminase (A.D.)

    Chivot, J.J.; Depernet, D.; Caen, J. [Commissariat a l' Energie Atomique, Bruyeres-le-Chatel (France). Centre d' Etudes

    1970-07-01

    We were able, by using a radio-chromatographic method, to measure an adenosine deaminase activity in normal human heparinized platelet-poor plasma, which can degrade 0.016 {mu}M adenosine. This activity suppressed by heating 56 C for 30 minutes is inhibited by high concentrations of urea and is proportional to the amount of plasma, source of enzyme, in the systems. (authors) [French] Nous avons pu, en utilisant une methode radiochromatographique, mesurer une activite adenosine deaminasique dans le plasma humain pauvre en plaquettes heparine qui peut degrader 0,016 {mu}M d'adenosine. Cette activite qui est supprimee par chauffage a 56 degres pendant 30 minutes, est reduite par conservation a -20 C pendant une semaine, est inhibee par d'importantes concentrations d'uree et ne l'est pas, ni par le dipyridamol, ni par le pHMB. Cette activite est proportionnelle a la quantite de plasma, source d'enzyme, mise dans les differents systemes reactifs. (auteur)

  2. Stimulation of Glia Reveals Modulation of Mammalian Spinal Motor Networks by Adenosine.

    David Acton

    Full Text Available Despite considerable evidence that glia can release modulators to influence the excitability of neighbouring neurons, the importance of gliotransmission for the operation of neural networks and in shaping behaviour remains controversial. Here we characterise the contribution of glia to the modulation of the mammalian spinal central pattern generator for locomotion, the output of which is directly relatable to a defined behaviour. Glia were stimulated by specific activation of protease-activated receptor-1 (PAR1, an endogenous G-protein coupled receptor preferentially expressed by spinal glia during ongoing activity of the spinal central pattern generator for locomotion. Selective activation of PAR1 by the agonist TFLLR resulted in a reversible reduction in the frequency of locomotor-related bursting recorded from ventral roots of spinal cord preparations isolated from neonatal mice. In the presence of the gliotoxins methionine sulfoximine or fluoroacetate, TFLLR had no effect, confirming the specificity of PAR1 activation to glia. The modulation of burst frequency upon PAR1 activation was blocked by the non-selective adenosine-receptor antagonist theophylline and by the A1-receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine, but not by the A2A-receptor antagonist SCH5826, indicating production of extracellular adenosine upon glial stimulation, followed by A1-receptor mediated inhibition of neuronal activity. Modulation of network output following glial stimulation was also blocked by the ectonucleotidase inhibitor ARL67156, indicating glial release of ATP and its subsequent degradation to adenosine rather than direct release of adenosine. Glial stimulation had no effect on rhythmic activity recorded following blockade of inhibitory transmission, suggesting that glial cell-derived adenosine acts via inhibitory circuit components to modulate locomotor-related output. Finally, the modulation of network output by endogenous adenosine was found to

  3. Effects of adenosine infusion into renal interstitium on renal hemodynamics

    This study was designed to investigate the hemodynamic effects of exogenous adenosine in the interstitium of the rat kidney. Adenosine or its analogues were infused into the renal interstitium by means of chronically implanted capsules. In fusion of adenosine decreased glomerular filtration rate (GFR) from 0.81 +/- 0.06 to 0.37 +/- 0.06 ml/min while having no effect on renal blood flow (RBF). The metabolically stable analogue, 2-chloradenosine (2-ClAdo), decreased GFR from 0.73 +/- 0.07 to 021 +/- 0.06 ml/min. Interstitial infusion of theophylline, an adenosine receptor antagonist, completely abolished the effects of adenosine and 2-ClAdo on GFR. The distribution of adenosine, when infused into the renal interstitium, was determined using radiolabeled 5'-(N-ethyl)-carboxamidoadenosine (NECA), a metabolically stable adenosine agonist. After continuous infusion, [3H]NECA was distributed throughout the kidney. The effects of NECA to reduce GFR were similar to those of adenosine and 2-ClAdo. They conclude that increased levels of adenosine in the renal interstitium markedly decrease GFR without affecting RBF in steady-state conditions. The marked effects of adenosine agonists during their infusion into the renal interstitium and the complete blockade of these effects by theophylline suggest an extracellular action of adenosine

  4. Adenosine A1 Receptor Mediates Delayed Cardioprotective Effect of Sildenafil in Mouse

    Salloum, Fadi N.; Das, Anindita; Thomas, Christopher S; Yin, Chang; Kukreja, Rakesh C.

    2007-01-01

    Sildenafil induces powerful cardioprotection against ischemia/reperfusion (I/R) injury. Since adenosine is known to be major trigger of ischemic preconditioning, we hypothesized that A1 adenosine receptor (A1AR) activation plays a role in sildenafil-induced cardioprotective signaling. Adult male C57BL-wild type (WT) mice or their corresponding A1AR knockout (A1AR-KO) mice were treated intraperitoneally (i.p.) with either sildenafil (0.71 mg/kg, equivalent to 50 mg dose for a 70 kg patient) or...

  5. Regulation of photoreceptor gap junction phosphorylation by adenosine in zebrafish retina.

    Li, Hongyan; Chuang, Alice Z; O'Brien, John

    2014-05-01

    Electrical coupling of photoreceptors through gap junctions suppresses voltage noise, routes rod signals into cone pathways, expands the dynamic range of rod photoreceptors in high scotopic and mesopic illumination, and improves detection of contrast and small stimuli. In essentially all vertebrates, connexin 35/36 (gene homologs Cx36 in mammals, Cx35 in other vertebrates) is the major gap junction protein observed in photoreceptors, mediating rod-cone, cone-cone, and possibly rod-rod communication. Photoreceptor coupling is dynamically controlled by the day/night cycle and light/dark adaptation, and is directly correlated with phosphorylation of Cx35/36 at two sites, serine110 and serine 276/293 (homologous sites in teleost fish and mammals, respectively). Activity of protein kinase A (PKA) plays a key role during this process. Previous studies have shown that activation of dopamine D4 receptors on photoreceptors inhibits adenylyl cyclase, down-regulates cAMP and PKA activity, and leads to photoreceptor uncoupling, imposing the daytime/light condition. In this study, we explored the role of adenosine, a nighttime signal with a high extracellular concentration at night and a low concentration in the day, in regulating photoreceptor coupling by examining photoreceptor Cx35 phosphorylation in zebrafish retina. Adenosine enhanced photoreceptor Cx35 phosphorylation in daytime, but with a complex dose-response curve. Selective pharmacological manipulations revealed that adenosine A2a receptors provide a potent positive drive to phosphorylate photoreceptor Cx35 under the influence of endogenous adenosine at night. A2a receptors can be activated in the daytime as well by micromolar exogenous adenosine. However, the higher affinity adenosine A1 receptors are also present and have an antagonistic though less potent effect. Thus, the nighttime/darkness signal adenosine provides a net positive drive on Cx35 phosphorylation at night, working in opposition to dopamine to

  6. Regulation of adenosine triphosphate-sensitive potassium channels suppresses the toxic effects of amyloid-beta peptide (25-35)

    Min Kong; Maowen Ba; Hui Liang; Peng Shao; Tianxia Yu; Ying Wang

    2013-01-01

    In this study, we treated PC12 cells with 0-20 μM amyloid-β peptide (25-35) for 24 hours to induce cytotoxicity, and found that 5-20 μM amyloid-β peptide (25-35) decreased PC12 cell viability, but adenosine triphosphate-sensitive potassium channel activator diazoxide suppressed the decrease reactive oxygen species levels. These protective effects were reversed by the selective mitochondrial adenosine triphosphate-sensitive potassium channel blocker 5-hydroxydecanoate. An inducible nitric oxide synthase inhibitor, Nω-nitro-L-arginine, also protected PC12 cells from intracellular reactive oxygen species levels. However, the H2O2-degrading enzyme catalase could that the increases in both mitochondrial membrane potential and reactive oxygen species levels adenosine triphosphate-sensitive potassium channels and nitric oxide. Regulation of adenosine triphosphate-sensitive potassium channels suppresses PC12 cell cytotoxicity induced by amyloid-β

  7. Adenosine receptor antagonists alter the stability of human epileptic GABAA receptors

    Roseti, Cristina; Martinello, Katiuscia; Fucile, Sergio; Piccari, Vanessa; Mascia, Addolorata; Di Gennaro, Giancarlo; Quarato, Pier Paolo; Manfredi, Mario; Esposito, Vincenzo; Cantore, Gianpaolo; Arcella, Antonella; Simonato, Michele; Fredholm, Bertil B.; Limatola, Cristina; Miledi, Ricardo; Eusebi, Fabrizio

    2008-01-01

    We examined how the endogenous anticonvulsant adenosine might influence γ-aminobutyric acid type A (GABAA) receptor stability and which adenosine receptors (ARs) were involved. Upon repetitive activation (GABA 500 μM), GABAA receptors, microtransplanted into Xenopus oocytes from neurosurgically resected epileptic human nervous tissues, exhibited an obvious GABAA-current (IGABA) run-down, which was consistently and significantly reduced by treatment with the nonselective adenosine receptor antagonist CGS15943 (100 nM) or with adenosine deaminase (ADA) (1 units/ml), that inactivates adenosine. It was also found that selective antagonists of A2B (MRS1706, 10 nM) or A3 (MRS1334, 30 nM) receptors reduced IGABA run-down, whereas treatment with the specific A1 receptor antagonist DPCPX (10 nM) was ineffective. The selective A2A receptor antagonist SCH58261 (10 nM) reduced or potentiated IGABA run-down in ≈40% and ≈20% of tested oocytes, respectively. The ADA-resistant, AR agonist 2-chloroadenosine (2-CA) (10 μM) potentiated IGABA run-down but only in ≈20% of tested oocytes. CGS15943 administration again decreased IGABA run-down in patch-clamped neurons from either human or rat neocortex slices. IGABA run-down in pyramidal neurons was equivalent in A1 receptor-deficient and wt neurons but much larger in neurons from A2A receptor-deficient mice, indicating that, in mouse cortex, GABAA-receptor stability is tonically influenced by A2A but not by A1 receptors. IGABA run-down from wt mice was not affected by 2-CA, suggesting maximal ARs activity by endogenous adenosine. Our findings strongly suggest that cortical A2–A3 receptors alter the stability of GABAA receptors, which could offer therapeutic opportunities. PMID:18809912

  8. Adenosine elicits an eNOS-independent reduction in arterial blood pressure in conscious mice that involves adenosine A(2A) receptors

    Andersen, Henrik; Jaff, Mohammad G; Høgh, Ditte;

    2011-01-01

    Aims:  Adenosine plays an important role in the regulation of heart rate and vascular reactivity. However, the mechanisms underlying the acute effect of adenosine on arterial blood pressure in conscious mice are unclear. Therefore, the present study investigated the effect of the nucleoside on mean...... arterial blood pressure (MAP) and heart rate (HR) in conscious mice. Methods:  Chronic indwelling catheters were placed in C57Bl/6J (WT) and endothelial nitric oxide synthase knock-out (eNOS(-/-) ) mice for continuous measurements of MAP and HR. Using PCR and myograph analysis involment of adenosine...... receptors was investigated in human and mouse renal blood vessels Results:  Bolus infusion of 0.5 mg/kg adenosine elicited significant transient decreases in MAP (99.3±2.3 to 70.4±4.5 mmHg) and HR (603.2±18.3 to 364.3±49.2 min(-1) ) which were inhibited by the A(2A) receptor antagonist ZM 241385. Activation...

  9. Adenosine as a signaling molecule in the retina: biochemical and developmental aspects

    ROBERTO PAES-DE-CARVALHO

    2002-09-01

    Full Text Available The nucleoside adenosine plays an important role as a neurotransmitter or neuromodulator in the central nervous system, including the retina. In the present paper we review compelling evidence showing that adenosine is a signaling molecule in the developing retina. In the chick retina, adenosine transporters are present since early stages of development before the appearance of adenosine A1 receptors modulating dopamine-dependent adenylate cyclase activity or A2 receptors that directly activate the enzyme. Experiments using retinal cell cultures revealed that adenosine is taken up by specific cell populations that when stimulated by depolarization or neurotransmitters such as dopamine or glutamate, release the nucleoside through calcium-dependent transporter-mediated mechanisms. The presence of adenosine in the extracellular medium and the long-term activation of adenosine receptors is able to regulate the survival of retinal neurons and blocks glutamate excitoxicity. Thus, adenosine besides working as a neurotransmitter or neuromodulator in the mature retina, is considered as an important signaling molecule during retinal development having important functions such as regulation of neuronal survival and differentiation.O nucleosídeo adenosina apresenta um importante papel como neurotransmissor ou neuromodulador no sistema nervoso central, inclusive na retina. Neste artigo apresentamos uma revisão das evidências que mostram que a adenosina é uma molécula sinalizadora na retina em desenvolvimento. Na retina de pinto, transportadores de adenosina estão presentes desde estágios precoces do desenvolvimento, antes do aparecimento dos receptores A1 que modulam a atividade adenilato ciclase dependente de dopamina ou dos receptores A2 que ativam diretamente a enzima. Experimentos usando culturas de células de retina revelaram que a adenosina é captada por populações celulares específicas que, quando estimuladas por despolarização ou por

  10. Characterization of the binding of a novel nonxanthine adenosine antagonist radioligand, ( sup 3 H)CGS 15943, to multiple affinity states of the adenosine A1 receptor in the rat cortex

    Jarvis, M.F.; Williams, M.; Do, U.H.; Sills, M.A. (CIBA-GEIGY Corp., Summit, NJ (USA))

    1991-01-01

    The triazoloquinazoline CGS 15943 is the first reported nonxanthine adenosine antagonist that has high affinity for brain adenosine receptors. In the present study, the binding of (3H) CGS 15943 to recognition sites in rat cortical membranes was characterized. Saturation experiments revealed that (3H)CGS 15943 labeled a single class of recognition sites with high affinity and limited capacity. Competition studies revealed that the binding of (3H)CGS 15943 was consistent with the labeling of brain adenosine A1 receptors. Adenosine agonists inhibited 1 nM (3H)CGS 15943 binding with the following order of activity N6-cyclopentyladenosine (IC50 = 15 nM) greater than 2-chloroadenosine greater than (R)-N6-phenylisopropyladenosine greater than 5'-N6-ethylcarboxamidoadenosine greater than (S)N6-phenylisopropyladenosine greater than CGS 21680 greater than CV 1808 (IC50 greater than 10,000 nM). The potency order for adenosine antagonists was CGS 15943 (IC50 = 5 nM) greater than 8-phenyltheophylline greater than 1,3-dipropyl-8-(4-amino-2-chloro)phenylxanthine greater than 1,3-diethyl-8-phenylxanthine greater than theophylline = caffeine (IC50 greater than 10,000 nM). Antagonist inhibition curves were steep and best described by a one-site binding model. In contrast, adenosine A1 agonist competition curves were shallow, as indicated by Hill coefficients less than unity. Computer analysis revealed that these inhibition curves were best described by a two-site binding model. Agonist competition curves generated in the presence of 1 mM GTP resulted in a rightward shift and steepening of the inhibition-concentration curves, whereas antagonist binding was not altered in the presence of GTP. The complex binding interactions found with adenosine agonists indicate that (3H)CGS 15943 labels both high and low affinity components of the adenosine A1 receptor in the rat cortex.

  11. Magnetically assisted fluorescence ratiometric assays for adenosine deaminase using water-soluble conjusated polymers

    HE Fang; YU MingHui; WANG Shu

    2009-01-01

    A magnetically assisted fluorescence ratiometric technique has been developed for adenosine deami-nase assays with high sensitivity using water-soluble cationic conjugated polymers (CCPs).The assay contains three elements:a biotin-labeled aptamer of adenosine (biotin-aptamer),a signaling probe single-stranded DNA-tagged fiuorescein at terminus (ssDNA-FI) and a CCP.The specific binding of adenosine to biotin-aptamer makes biotin-aptamer and ssDNA-FI unhybridized,and the ssDNA-FI is washed out after streptavidin-coated magnetic beads are added and separated from the assay solution under magnetic field.In this case,after the addition of CCP to the magnetic beads solution,the fluo-rescence resonance energy transfer (FRET) from CCP to fluorescein is inefficient.Upon adding adenosine deaminase,the adenosine is converted into inosine,and the biotin-aptamer is hybridized with ssDNA-FI to form doubled stranded DNA (biotin-dsDNA-FI).The ssONA-FI is attached to the mag-netic beads at the separation step,and the addition of CCP to the magnetic beads solution leads to efficient FRET from CCP to fluorescein.Thus the adenosine deaminase activity can be monitored by fluorescence spectra in view of the intensity decrease of CCP emission or the increase of fluorescein emission in aqueous solutions.The assay integrates surface-functionalized magnetic particles with significant amplification of detection signal of water-soluble cationic conjugated polymers.

  12. Increase of adenosine plasma levels after oral trimetazidine: a pharmacological preconditioning?

    Blardi, Patrizia; de Lalla, Arianna; Volpi, Luciana; Auteri, Alberto; Di Perri, Tullio

    2002-01-01

    Trimetazidine (1-[2,3,4-trimethoxybenzyl] piperazine) (TMZ) is a cellular anti-ischemic agent able to prevent intracellular ATP decrease, limit intracellular acidosis, protect against oxygen-free radical-induced toxicity and inhibit neutrophil infiltration. However, its definitive mechanism of action had not been identified. Recent studies showed the existence of an endogenous mechanism of cellular protection against ischemia, defined as 'ischemic preconditioning'. This mechanism was related mainly to cellular liberation of adenosine, a nucleoside with protective effects in myocardial ischemia. Since TMZ acts by increasing cell tolerance to ischemia and adenosine is the mediator of ischemic preconditioning, in this study we investigated a possible interaction between TMZ and adenosine. Two groups of patients affected by angina pectoris, were admitted to the study. They received a single oral dose of TMZ. One group was treated, during different sessions, with TMZ 10 and 20 mg, the other group with TMZ 40 and 80 mg. After a 3 day wash-out from drug administration, each group received a placebo. Blood samples were collected at baseline (time 0) and 1, 2, 3, 4, 6, 8 h after drug administration, in order to detect plasma levels of adenosine by a high-performance liquid chromatography method. We observed that the administration of TMZ at doses of 10, 20, 40 and 80 mg induced an increase of adenosine plasma levels of 19, 50, 62 and 62%, respectively. We hypothesized that the activity of TMZ could depend, at least in part, on adenosine mediation and this interaction opens a new interpretation of the drug antischemic effect. PMID:11820865

  13. Adenosine receptors and asthma in humans

    Wilson, C N

    2008-01-01

    According to an executive summary of the GINA dissemination committee report, it is now estimated that approximately 300 million people (5% of the global population or 1 in 20 persons) have asthma. Despite the scientific progress made over the past several decades toward improving our understanding of the pathophysiology of asthma, there is still a great need for improved therapies, particularly oral therapies that enhance patient compliance and that target new mechanisms of action. Adenosine...

  14. Aminopyrimidine derivatives as adenosine antagonists / Janke Kleynhans

    Kleynhans, Janke

    2013-01-01

    Aims of this project - The aim of this study was to design and synthesise novel 2-aminopyrimidine derivatives as potential adenosine A1 and A2A receptor antagonists. Background and rationale - Parkinson’s disease is the second most common neurodegenerative disorder (after Alzheimer’s disease) and is characterised by the selective death of the dopaminergic neurons of the nigro-striatal pathway. Distinctive motor symptoms include bradykinesia, muscle rigidity and tremor, while non-m...

  15. Hyperalgesia, anxiety, and decreased hypoxic neuroprotection in mice lacking the adenosine A1 receptor.

    Johansson, B; Halldner, L; Dunwiddie, T V; Masino, S A; Poelchen, W; Giménez-Llort, L; Escorihuela, R M; Fernández-Teruel, A; Wiesenfeld-Hallin, Z; Xu, X J; Hårdemark, A; Betsholtz, C; Herlenius, E; Fredholm, B B

    2001-07-31

    Caffeine is believed to act by blocking adenosine A(1) and A(2A) receptors (A(1)R, A(2A)R), indicating that some A(1) receptors are tonically activated. We generated mice with a targeted disruption of the second coding exon of the A(1)R (A(1)R(-/-)). These animals bred and gained weight normally and had a normal heart rate, blood pressure, and body temperature. In most behavioral tests they were similar to A(1)R(+/+) mice, but A(1)R(-/-) mice showed signs of increased anxiety. Electrophysiological recordings from hippocampal slices revealed that both adenosine-mediated inhibition and theophylline-mediated augmentation of excitatory glutamatergic neurotransmission were abolished in A(1)R(-/-) mice. In A(1)R(+/-) mice the potency of adenosine was halved, as was the number of A(1)R. In A(1)R(-/-) mice, the analgesic effect of intrathecal adenosine was lost, and thermal hyperalgesia was observed, but the analgesic effect of morphine was intact. The decrease in neuronal activity upon hypoxia was reduced both in hippocampal slices and in brainstem, and functional recovery after hypoxia was attenuated. Thus A(1)Rs do not play an essential role during development, and although they significantly influence synaptic activity, they play a nonessential role in normal physiology. However, under pathophysiological conditions, including noxious stimulation and oxygen deficiency, they are important. PMID:11470917

  16. Role of adenosine in oligodendrocyte precursor maturation

    Elisabetta Coppi

    2015-04-01

    Full Text Available Differentiation and maturation of oligodendroglial cells are postnatal processes involving specific morphological changes correlated with the expression of stage-specific surface antigens and functional voltage-gated ion channels. A small fraction of oligodendrocyte progenitor cells (OPCs generated during development are maintained in an immature and slowly proliferative or quiescent state in the adult central nervous system (CNS representing an endogenous reservoir of immature cells. Adenosine receptors are expressed by OPCs and a key role of adenosine in oligodendrocyte maturation has been recently recognised. As evaluated on OPC cultures, adenosine by stimulating A1 receptors, promotes oligodendrocyte maturation and inhibits their proliferation; on the contrary by stimulating A2A receptors, it inhibits oligodendrocyte maturation. A1 and A2A receptor-mediated effects are related to opposite modifications of outward delayed rectifying membrane K+ currents (IK that are involved in regulation of oligodendrocyte differentiation. Brain A1 and A2A receptors might represent new molecular targets for drugs useful in demyelinating pathologies, such as multiple sclerosis (MS, stroke and brain trauma.

  17. Nucleus tractus solitarii A(2a) adenosine receptors inhibit cardiopulmonary chemoreflex control of sympathetic outputs.

    Minic, Zeljka; O'Leary, Donal S; Scislo, Tadeusz J

    2014-02-01

    Previously we have shown that stimulation of inhibitory A1 adenosine receptors located in the nucleus tractus solitarii (NTS) attenuates cardiopulmonary chemoreflex (CCR) evoked inhibition of renal, adrenal and lumbar sympathetic nerve activity and reflex decreases in arterial pressure and heart rate. Activation of facilitatory A2a adenosine receptors, which dominate over A1 receptors in the NTS, contrastingly alters baseline activity of regional sympathetic outputs: it decreases renal, increases adrenal and does not change lumbar nerve activity. Considering that NTS A2a receptors may facilitate release of inhibitory transmitters we hypothesized that A2a receptors will act in concert with A1 receptors differentially inhibiting regional sympathetic CCR responses (adrenal>lumbar>renal). In urethane/chloralose anesthetized rats (n=38) we compared regional sympathetic responses evoked by stimulation of the CCR with right atrial injections of serotonin 5HT3 receptor agonist, phenylbiguanide, (1-8μg/kg) before and after selective stimulation, blockade or combined blockade and stimulation of NTS A2a adenosine receptors (microinjections into the NTS of CGS-21680 0.2-20pmol/50nl, ZM-241385 40pmol/100nl or ZM-241385+CGS-21680, respectively). We found that stimulation of A2a adenosine receptors uniformly inhibited the regional sympathetic and hemodynamic reflex responses and this effect was abolished by the selective blockade of NTS A2a receptors. This indicates that A2a receptor triggered inhibition of CCR responses and the contrasting shifts in baseline sympathetic activity are mediated via different mechanisms. These data implicate that stimulation of NTS A2a receptors triggers unknown inhibitory mechanism(s) which in turn inhibit transmission in the CCR pathway when adenosine is released into the NTS during severe hypotension. PMID:24216055

  18. Different cellular sources and different roles of adenosine: A1 receptor-mediated inhibition through astrocytic-driven volume transmission and synapse-restricted A2A receptor-mediated facilitation of plasticity

    Cunha, Rodrigo A.

    2008-01-01

    Adenosine is a prototypical neuromodulator, which mainly controls excitatory transmission through the activation of widespread inhibitory A1 receptors and synaptically located A2A receptors. It was long thought that the predominant A1 receptor-meditated modulation by endogenous adenosine was a homeostatic process intrinsic to the synapse. New studies indicate that endogenous extracellular adenosine is originated as a consequence of the release of gliotransmitters, namely ATP, which sets a glo...

  19. Adenosine A2A Receptors Modulate Acute Injury and Neuroinflammation in Brain Ischemia

    Felicita Pedata

    2014-01-01

    Full Text Available The extracellular concentration of adenosine in the brain increases dramatically during ischemia. Adenosine A2A receptor is expressed in neurons and glial cells and in inflammatory cells (lymphocytes and granulocytes. Recently, adenosine A2A receptor emerged as a potential therapeutic attractive target in ischemia. Ischemia is a multifactorial pathology characterized by different events evolving in the time. After ischemia the early massive increase of extracellular glutamate is followed by activation of resident immune cells, that is, microglia, and production or activation of inflammation mediators. Proinflammatory cytokines, which upregulate cell adhesion molecules, exert an important role in promoting recruitment of leukocytes that in turn promote expansion of the inflammatory response in ischemic tissue. Protracted neuroinflammation is now recognized as the predominant mechanism of secondary brain injury progression. A2A receptors present on central cells and on blood cells account for important effects depending on the time-related evolution of the pathological condition. Evidence suggests that A2A receptor antagonists provide early protection via centrally mediated control of excessive excitotoxicity, while A2A receptor agonists provide protracted protection by controlling massive blood cell infiltration in the hours and days after ischemia. Focus on inflammatory responses provides for adenosine A2A receptor agonists a wide therapeutic time-window of hours and even days after stroke.

  20. A2BR adenosine receptor modulates sweet taste in circumvallate taste buds.

    Shinji Kataoka

    Full Text Available In response to taste stimulation, taste buds release ATP, which activates ionotropic ATP receptors (P2X2/P2X3 on taste nerves as well as metabotropic (P2Y purinergic receptors on taste bud cells. The action of the extracellular ATP is terminated by ectonucleotidases, ultimately generating adenosine, which itself can activate one or more G-protein coupled adenosine receptors: A1, A2A, A2B, and A3. Here we investigated the expression of adenosine receptors in mouse taste buds at both the nucleotide and protein expression levels. Of the adenosine receptors, only A2B receptor (A2BR is expressed specifically in taste epithelia. Further, A2BR is expressed abundantly only in a subset of taste bud cells of posterior (circumvallate, foliate, but not anterior (fungiform, palate taste fields in mice. Analysis of double-labeled tissue indicates that A2BR occurs on Type II taste bud cells that also express Gα14, which is present only in sweet-sensitive taste cells of the foliate and circumvallate papillae. Glossopharyngeal nerve recordings from A2BR knockout mice show significantly reduced responses to both sucrose and synthetic sweeteners, but normal responses to tastants representing other qualities. Thus, our study identified a novel regulator of sweet taste, the A2BR, which functions to potentiate sweet responses in posterior lingual taste fields.

  1. Structural basis of the substrate specificity of Bacillus cereus adenosine phosphorylase

    Dessanti, Paola; Zhang, Yang; Allegrini, Simone; Tozzi, Maria Grazia; Sgarrella, Francesco; Ealick, Steven E. (Cornell); (Sassari); (Pisa)

    2012-10-08

    Purine nucleoside phosphorylases catalyze the phosphorolytic cleavage of the glycosidic bond of purine (2{prime}-deoxy)nucleosides, generating the corresponding free base and (2{prime}-deoxy)ribose 1-phosphate. Two classes of PNPs have been identified: homotrimers specific for 6-oxopurines and homohexamers that accept both 6-oxopurines and 6-aminopurines. Bacillus cereus adenosine phosphorylase (AdoP) is a hexameric PNP; however, it is highly specific for 6-aminopurines. To investigate the structural basis for the unique substrate specificity of AdoP, the active-site mutant D204N was prepared and kinetically characterized and the structures of the wild-type protein and the D204N mutant complexed with adenosine and sulfate or with inosine and sulfate were determined at high resolution (1.2-1.4 {angstrom}). AdoP interacts directly with the preferred substrate through a hydrogen-bond donation from the catalytically important residue Asp204 to N7 of the purine base. Comparison with Escherichia coli PNP revealed a more optimal orientation of Asp204 towards N7 of adenosine and a more closed active site. When inosine is bound, two water molecules are interposed between Asp204 and the N7 and O6 atoms of the nucleoside, thus allowing the enzyme to find alternative but less efficient ways to stabilize the transition state. The mutation of Asp204 to asparagine led to a significant decrease in catalytic efficiency for adenosine without affecting the efficiency of inosine cleavage.

  2. Silk Fibroin Encapsulated Powder Reservoirs for Sustained Release of Adenosine

    Pritchard, Eleanor M.; Szybala, Cory; Boison, Detlev; Kaplan, David L.

    2010-01-01

    Due to its unique properties, silk fibroin was studied as a biodegradable polymer vehicle for sustained, local delivery of the anticonvulsant adenosine from encapsulated reservoirs. Silk is a biologically derived protein polymer that is biocompatible, mechanically strong and degrades to non-toxic products in vivo. To achieve local, sustained, controlled adenosine release from fully degradable implants, solid adenosine powder reservoirs were coated with silk fibroin. Material properties of the...

  3. Adenosine A1 receptor agonists inhibit trigeminovascular nociceptive transmission

    Goadsby, P J; Hoskin, K L; Storer, R J;

    2002-01-01

    There is a considerable literature to suggest that adenosine A1 receptor agonists may have anti-nociceptive effects, and we sought to explore the role of adenosine A1 receptors in a model of trigeminovascular nociceptive transmission. Cats were anaesthetized (alpha-chloralose 60 mg/kg, intraperit......There is a considerable literature to suggest that adenosine A1 receptor agonists may have anti-nociceptive effects, and we sought to explore the role of adenosine A1 receptors in a model of trigeminovascular nociceptive transmission. Cats were anaesthetized (alpha-chloralose 60 mg...

  4. Mechanisms of the adenosine A2A receptor-induced sensitization of esophageal C fibers.

    Brozmanova, M; Mazurova, L; Ru, F; Tatar, M; Hu, Y; Yu, S; Kollarik, M

    2016-02-01

    Clinical studies indicate that adenosine contributes to esophageal mechanical hypersensitivity in some patients with pain originating in the esophagus. We have previously reported that the esophageal vagal nodose C fibers express the adenosine A2A receptor. Here we addressed the hypothesis that stimulation of the adenosine A2A receptor induces mechanical sensitization of esophageal C fibers by a mechanism involving transient receptor potential A1 (TRPA1). Extracellular single fiber recordings of activity originating in C-fiber terminals were made in the ex vivo vagally innervated guinea pig esophagus. The adenosine A2A receptor-selective agonist CGS21680 induced robust, reversible sensitization of the response to esophageal distention (10-60 mmHg) in a concentration-dependent fashion (1-100 nM). At the half-maximally effective concentration (EC50: ≈3 nM), CGS21680 induced an approximately twofold increase in the mechanical response without causing an overt activation. This sensitization was abolished by the selective A2A antagonist SCH58261. The adenylyl cyclase activator forskolin mimicked while the nonselective protein kinase inhibitor H89 inhibited mechanical sensitization by CGS21680. CGS21680 did not enhance the response to the purinergic P2X receptor agonist α,β-methylene-ATP, indicating that CGS21680 does not nonspecifically sensitize to all stimuli. Mechanical sensitization by CGS21680 was abolished by pretreatment with two structurally different TRPA1 antagonists AP18 and HC030031. Single cell RT-PCR and whole cell patch-clamp studies in isolated esophagus-specific nodose neurons revealed the expression of TRPA1 in A2A-positive C-fiber neurons and demonstrated that CGS21682 potentiated TRPA1 currents evoked by allylisothiocyanate. We conclude that stimulation of the adenosine A2A receptor induces mechanical sensitization of nodose C fibers by a mechanism sensitive to TRPA1 antagonists indicating the involvement of TRPA1. PMID:26564719

  5. Engineering a prostate-specific membrane antigen-activated tumor endothelial cell prodrug for cancer therapy

    Denmeade, Samuel R; Mhaka, Annastasiah M; Rosen, D Marc;

    2012-01-01

    Heterogeneous expression of drug target proteins within tumor sites is a major mechanism of resistance to anticancer therapies. We describe a strategy to selectively inhibit, within tumor sites, the function of a critical intracellular protein, the sarcoplasmic/endoplasmic reticulum calcium...... adenosine triphosphatase (SERCA) pump, whose proper function is required by all cell types for viability. To achieve targeted inhibition, we took advantage of the unique expression of the carboxypeptidase prostate-specific membrane antigen (PSMA) by tumor endothelial cells within the microenvironment of...

  6. Intravenous adenosine and radiopharmaceutical injection in the same line was feasible in adenosine stress myocardial perfusion imaging

    Adenosine stress myocardial perfusion imaging was performed with an intravenous adenosine and radiopharmaceutical injection in the same line. A syringe containing 720 μ/kg of adenosine in 40 ml of saline was prepared and injected at the constant infusion rate of 400 ml/h. Adenosine was temporarily stopped by the stopcock when 1.5 ml of thallium was injected for 0.5 second from the three-way stopcock with two ways opened. Thereafter, the stopcock was returned to the original position in 0.5 second, and adenosine flow returned to the constant flow rate again. In this method, 0.75% of adenosine total dose was injected at a rate of 3.0 ml/s and adenosine was stopped for 3.6 second. There were no significant differences in either effects and adverse events of adenosine between this method and two intravenous injection line methods. Adenosine stress in one venous line method would be an easy method maintaining the dose effect and safety. (author)

  7. Effects of AMP579 and adenosine on L-type Ca2+ current in isolated rat ventricular myocytes

    Xiong WANG; Bo-wei WU; Dong-mei WU

    2005-01-01

    Aim: To compare the effects of AMP579 and adenosine on L-type Ca2+ current (ICa- L) in rat ventricular myocytes and explore the mechanism by which AMP579 acts on ICa-L. Methods: ICa-L was recorded by patch-clamp technique in whole-cell configuration. Results: Adenosine (10 nmol/L to 50 μmol/L) showed no effect on basal ICa- L, but it inhibited the ICa-L induced by isoproterenol 10 nmol/L in a concen tration-dependent manner with the IC50 of 13.06 μmol/L. Similar to adenosine,AMP579 also showed an inhibitory effect on the ICa-L induced by isoproterenol.AMP579 and adenosine (both in 10 μmol/L) suppressed isoproterenol-induced ICa-L by 11.1% and 5.2%, respectively. In addition, AMP579 had a direct inhibitory effect on basal ICa-L in a concentration-dependent manner with IC50 (1.17 μmol/L).PD116948 (30 μmol/L), an adenosine A1 receptor blocker, showed no action on the inhibitory effect of AMP579 on basal ICa-L. However, GF109203X (0.4 μmol/L), a special protein kinase C (PKC) blocker, could abolish the inhibitory effect of AMP579 on basal ICa-L. So the inhibitory effect of AMP579 on basal ICa-L was induced through activating PKC, but not linked to adenosine A1 receptor. Conclusion:AMP579 shows a stronger inhibitory effect than adenosine on the ICa-L induced by isoproterenol. AMP579 also has a strong inhibitory effect on basal ICa-L in rat ventricular myocytes. Activation of PKC is involved in the inhibitory effect of AMP579 on basal ICa-L at downstream-mechanism.

  8. Characterization of adenosine binding proteins in human placental membranes

    We have characterized two adenosine binding proteins in human placenta. In membranes, one site is detected with [3H] -N-ethylcarboxamidoadenosine ([3H]NECA). This site is similar to the adenosine A2 receptor. We call this site the adenosine A2-like binding site. In detergent extracts, the second site is detected and has the characteristics of an adenosine A1 receptor. The soluble adenosine A2-like binding site cannot be detected without a rapid assay. Binding to the adenosine A1 receptor with [3H]-2-chloroadenosine and [3H]NECA is time dependent, saturable, and reversible. Equilibrium displacement analysis with adenosine agonists reveals an A1 specificity: 2-chloroadenosine > R-phenylisopropyladenosine > 5'-N-ethylcarboxamidoadenosine. The antagonist potency order is 1,3-diethyl-8-phenylxanthine > isobutylmethylxanthine > theophylline. Competition analysis of membranes with the A,-selective ligands [3H]-cyclohexyladenosine [3H] cylopentylxanthine revealed adenosine A1 agonist and antagonist potency orders. We have purified the adenosine A2-like binding site. The adenosine A2-like binding site is an ubiquitous major cellular protein. It is glycosylated, highly asymmetric, and acidic. The native protein is an homodimer with a subunit molecular mass of 98 kDa. The sedimentation coefficient and partial specific volume of the binding complex are 6.9 s and 0.698 ml/g, respectively. The Stokes' radius is 70 Angstrom. The native molecular mass of the detergent-protein complex is 230 kDa. The adenosine A2-like binding site has an agonist potency order of 5'-N-ethylcarboxamidoadenosine > 2-chloroadenosine >> R-phenylisopropyladenosine and an antagonist potency order of isobutylmethylxanthine > theophylline >> 1,3-diethyl-8-phenylxanthine

  9. Characterization of adenosine binding proteins in human placental membranes

    Hutchison, K.A.

    1989-01-01

    We have characterized two adenosine binding proteins in human placenta. In membranes, one site is detected with ({sup 3}H) -N-ethylcarboxamidoadenosine (({sup 3}H)NECA). This site is similar to the adenosine A{sub 2} receptor. We call this site the adenosine A{sub 2}-like binding site. In detergent extracts, the second site is detected and has the characteristics of an adenosine A{sub 1} receptor. The soluble adenosine A{sub 2}-like binding site cannot be detected without a rapid assay. Binding to the adenosine A{sub 1} receptor with ({sup 3}H)-2-chloroadenosine and ({sup 3}H)NECA is time dependent, saturable, and reversible. Equilibrium displacement analysis with adenosine agonists reveals an A{sub 1} specificity: 2-chloroadenosine > R-phenylisopropyladenosine > 5{prime}-N-ethylcarboxamidoadenosine. The antagonist potency order is 1,3-diethyl-8-phenylxanthine > isobutylmethylxanthine > theophylline. Competition analysis of membranes with the A,-selective ligands ({sup 3}H)-cyclohexyladenosine ({sup 3}H) cylopentylxanthine revealed adenosine A{sub 1} agonist and antagonist potency orders. We have purified the adenosine A{sub 2}-like binding site. The adenosine A{sub 2}-like binding site is an ubiquitous major cellular protein. It is glycosylated, highly asymmetric, and acidic. The native protein is an homodimer with a subunit molecular mass of 98 kDa. The sedimentation coefficient and partial specific volume of the binding complex are 6.9 s and 0.698 ml/g, respectively. The Stokes' radius is 70 {Angstrom}. The native molecular mass of the detergent-protein complex is 230 kDa. The adenosine A{sub 2}-like binding site has an agonist potency order of 5'-N-ethylcarboxamidoadenosine > 2-chloroadenosine >> R-phenylisopropyladenosine and an antagonist potency order of isobutylmethylxanthine > theophylline >> 1,3-diethyl-8-phenylxanthine.

  10. High-dose adenosine overcomes the attenuation of myocardial perfusion reserve caused by caffeine.

    Reyes, E.; Loong, C Y; Harbinson, Mark; Donovan, J; Anagnostopoulos, C.; Underwood, S. R.

    2008-01-01

    Objectives:We studied whether an increase in adenosine dose overcomes caffeine antagonism on adenosine-mediated coronary vasodilation.Background:Caffeine is a competitive antagonist at the adenosine receptors, but it is unclear whether caffeine in coffee alters the actions of exogenous adenosine, and whether the antagonism can be surmounted by increasing the adenosine dose.Methods:Myocardial perfusion scintigraphy (MPS) was used to assess adenosine-induced hyperemia in 30 patients before (bas...