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Sample records for a1 a2a a2b

  1. Ligand-, structure- and pharmacophore-based molecular fingerprints: a case study on adenosine A1, A2A, A2B, and A3 receptor antagonists

    NASA Astrophysics Data System (ADS)

    Sirci, Francesco; Goracci, Laura; Rodríguez, David; van Muijlwijk-Koezen, Jacqueline; Gutiérrez-de-Terán, Hugo; Mannhold, Raimund

    2012-11-01

    FLAP fingerprints are applied in the ligand-, structure- and pharmacophore-based mode in a case study on antagonists of all four adenosine receptor (AR) subtypes. Structurally diverse antagonist collections with respect to the different ARs were constructed by including binding data to human species only. FLAP models well discriminate "active" (=highly potent) from "inactive" (=weakly potent) AR antagonists, as indicated by enrichment curves, numbers of false positives, and AUC values. For all FLAP modes, model predictivity slightly decreases as follows: A2BR > A2AR > A3R > A1R antagonists. General performance of FLAP modes in this study is: ligand- > structure- > pharmacophore- based mode. We also compared the FLAP performance with other common ligand- and structure-based fingerprints. Concerning the ligand-based mode, FLAP model performance is superior to ECFP4 and ROCS for all AR subtypes. Although focusing on the early first part of the A2A, A2B and A3 enrichment curves, ECFP4 and ROCS still retain a satisfactory retrieval of actives. FLAP is also superior when comparing the structure-based mode with PLANTS and GOLD. In this study we applied for the first time the novel FLAPPharm tool for pharmacophore generation. Pharmacophore hypotheses, generated with this tool, convincingly match with formerly published data. Finally, we could demonstrate the capability of FLAP models to uncover selectivity aspects although single AR subtype models were not trained for this purpose.

  2. Adenosine A2A and A2B Receptors Differentially Modulate Keratinocyte Proliferation: Possible Deregulation in Psoriatic Epidermis.

    PubMed

    Andrés, Rosa M; Terencio, María Carmen; Arasa, Jorge; Payá, Miguel; Valcuende-Cavero, Francisca; Navalón, Pedro; Montesinos, María Carmen

    2017-01-01

    Adenosine is a potent regulator of inflammation and immunity, but the role of adenosine receptors in keratinocytes remains controversial. We determined that in addition to A2B receptors, human epidermal keratinocytes also express A2A receptors, although to a lower extent. Through the use of selective adenosine receptor agonists and antagonists, we showed that physiological concentrations of adenosine activate A2B receptors in normal human keratinocytes, inducing cell cycle arrest through the increase of intracellular calcium but not through cAMP signaling. In contrast, the selective activation of A2A receptors by CGS-21680 induces keratinocyte proliferation via p38-mitogen-activated protein kinase activation. Adenosine and selective A2A and A2B agonists presented anti-inflammatory profiles independent of adenosine receptors but mediated by membrane phosphatase activation. Finally, keratinocyte exposure to diverse inflammatory cytokines altered adenosine receptor expression by reducing A2B and increasing A2A, a pattern also observed in psoriatic epidermis. Because increased epidermal turnover and inflammatory response are characteristics of psoriatic disease, further studies are needed to assess the role and consequences of the altered adenosine receptor expression in lesional and nonlesional psoriatic keratinocytes. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

  3. Potential therapeutic relevance of adenosine A2B and A2A receptors in the central nervous system.

    PubMed

    Popoli, Patrizia; Pepponi, Rita

    2012-09-01

    Adenosine A2B and, much more importantly, adenosine A2A receptors modulate many physiological and pathological processes in the brain. In this review, the most recent evidence concerning the role of such receptors and their potential therapeutic relevance is discussed. The low affinity of A2B receptors for adenosine implies that they might represent a good therapeutic target, since they are activated only under pathological conditions (when adenosine levels raise up to micromolar concentrations). The availability of selective ligands for A2B receptors would allow exploration of such an hypothesis. Since adenosine A2A receptors mediate both potentially neuroprotective and potentially neurotoxic effects, their role in neurodegenerative diseases is highly controversial. Nevertheless, A2A receptor antagonists have shown clear antiparkinsonian effects, and a great interest exists on the role of A2A receptors in Alzheimer's disease, brain ischaemia, spinal cord injury, drug addiction and other conditions. In order to establish whether such receptors represent a target for CNS diseases, at least two conditions are needed: the full comprehension of A2A-dependent mechanisms and the availability of ligands capable of discriminating among the different receptor populations.

  4. A2a and a2b adenosine receptors affect HIF-1α signaling in activated primary microglial cells.

    PubMed

    Merighi, Stefania; Borea, Pier Andrea; Stefanelli, Angela; Bencivenni, Serena; Castillo, Carlos Alberto; Varani, Katia; Gessi, Stefania

    2015-05-15

    Microglia are central nervous system (CNS)-resident immune cells, that play a crucial role in neuroinflammation. Hypoxia-inducible factor-1 (HIF-1), the main transcription factor of hypoxia-inducible genes, is also involved in the immune response, being regulated in normoxia by inflammatory mediators. Adenosine is an ubiquitous nucleoside that has an influence on many immune properties of microglia through interaction with four receptor subtypes. The aim of this study was to investigate whether adenosine may affect microglia functions by acting on HIF-1α modulation. Primary murine microglia were activated with lipopolysaccharide (LPS) with or without adenosine, adenosine receptor agonists and antagonists and HIF-1α accumulation and downstream genes regulation were determined. Adenosine increased LPS-induced HIF-1α accumulation leading to an increase in HIF-1α target genes involved in cell metabolism [glucose transporter-1 (GLUT-1)] and pathogens killing [inducible nitric-oxide synthase (iNOS)] but did not induce HIF-1α dependent genes related to angiogenesis [vascular endothelial growth factor (VEGF)] and inflammation [tumor necrosis factor-α (TNF-α)]. The stimulatory effect of adenosine on HIF-1α and its target genes was essentially exerted by activation of A2A through p44/42 and A2B subtypes via p38 mitogen-activated protein kinases (MAPKs) and Akt phosphorylation. Furthermore the nucleoside raised VEGF and decreased TNF-α levels, by activating A2B subtypes. In conclusion adenosine increases GLUT-1 and iNOS gene expression in a HIF-1α-dependent way, through A2A and A2B receptors, suggesting their role in the regulation of microglial cells function following injury. However, inhibition of TNF-α adds an important anti-inflammatory effect only for the A2B subtype. GLIA 2015.

  5. Human monocytes respond to extracellular cAMP through A2A and A2B adenosine receptors

    PubMed Central

    Sciaraffia, Ester; Riccomi, Antonella; Lindstedt, Ragnar; Gesa, Valentina; Cirelli, Elisa; Patrizio, Mario; De Magistris, Maria Teresa; Vendetti, Silvia

    2014-01-01

    In this study, we test the hypothesis that cAMP, acting as an extracellular mediator, affects the physiology and function of human myeloid cells. The cAMP is a second messenger recognized as a universal regulator of several cellular functions in different organisms. Many studies have shown that extracellular cAMP exerts regulatory functions, acting as first mediator in multiple tissues. However, the impact of extracellular cAMP on cells of the immune system has not been fully investigated. We found that human monocytes exposed to extracellular cAMP exhibit higher expression of CD14 and lower amount of MHC class I and class II molecules. When cAMP-treated monocytes are exposed to proinflammatory stimuli, they exhibit an increased production of IL-6 and IL-10 and a lower amount of TNF-α and IL-12 compared with control cells, resembling the features of the alternative-activated macrophages or M2 macrophages. In addition, we show that extracellular cAMP affects monocyte differentiation into DCs, promoting the induction of cells displaying an activated, macrophage-like phenotype with reduced capacity of polarized, naive CD4+ T cells into IFN-γ-producing lymphocytes compared with control cells. The effects of extracellular cAMP on monocytes are mediated by CD73 ecto-5′-nucleotidase and A2A and A2B adenosine receptors, as selective antagonists could reverse its effects. Of note, the expression of CD73 molecules has been found on the membrane of a small population of CD14+CD16+ monocytes. These findings suggest that an extracellular cAMP-adenosine pathway is active in cells of the immune systems. PMID:24652540

  6. The Effect of Adenosine A2A and A2B Antagonists on Tracheal Responsiveness, Serum Levels of Cytokines and Lung Inflammation in Guinea Pig Model of Asthma

    PubMed Central

    Pejman, Laleh; Omrani, Hasan; Mirzamohammadi, Zahra; Shahbazfar, Amir Ali; Khalili, Majid; Keyhanmanesh, Rana

    2014-01-01

    Purpose: Nowadays adenosine is specified as an important factor in the pathophysiology of asthma. For determining the effect of different A2 receptors, in this investigation the effect of single dose of selective adenosine A2A and A2B antagonists (ZM241385 and MRS1706) on different inflammatory parameters; tracheal responsiveness to methacholine and ovalbumin, total and differential cell count in bronchoalveolar lavage (BAL), blood levels of IL-4 and IFN-γ and lung pathology of guinea pig model of asthma were assessed. Methods: All mentioned parameters were evaluated in two sensitized groups of guinea pigs pretreated with A2A and A2B antagonists (S+Anta A2A, S+Anta A2B) compared with sensitized (S) and control (C) groups. Results: The tracheal responsiveness to methacholine and OA, total cell and eosinophil and basophil count in BAL, blood IL-4 level and pathological changes in pre-treated group with MRS1706 (S+Anta A2B) was significantly lower than those of sensitized group (p<0.01 to p<0.05). In pretreated group with Anta A2A(S+Anta A2A), all the above changes were reversed. Conclusion: These results showed a preventive effect of A2B antagonist (MRS1706) on tracheal responsiveness to methacholine and OA, total and differential cell count in bronchoalveolar lavage, blood cytokines and pathological changes. Administration of ZM241385, selective A2A antagonist, deteriorated the induction effect of ovalbumin. PMID:24511476

  7. StyA1 and StyA2B from Rhodococcus opacus 1CP: a Multifunctional Styrene Monooxygenase System▿

    PubMed Central

    Tischler, Dirk; Kermer, René; Gröning, Janosch A. D.; Kaschabek, Stefan R.; van Berkel, Willem J. H.; Schlömann, Michael

    2010-01-01

    Two-component flavoprotein monooxygenases are emerging biocatalysts that generally consist of a monooxygenase and a reductase component. Here we show that Rhodococcus opacus 1CP encodes a multifunctional enantioselective flavoprotein monooxygenase system composed of a single styrene monooxygenase (SMO) (StyA1) and another styrene monooxygenase fused to an NADH-flavin oxidoreductase (StyA2B). StyA1 and StyA2B convert styrene and chemical analogues to the corresponding epoxides at the expense of FADH2 provided from StyA2B. The StyA1/StyA2B system presents the highest monooxygenase activity in an equimolar ratio of StyA1 and StyA2B, indicating (transient) protein complex formation. StyA1 is also active when FADH2 is supplied by StyB from Pseudomonas sp. VLB120 or PheA2 from Rhodococcus opacus 1CP. However, in both cases the reductase produces an excess of FADH2, resulting in a high waste of NADH. The epoxidation rate of StyA1 heavily depends on the type of reductase. This supports that the FADH2-induced activation of StyA1 requires interprotein communication. We conclude that the StyA1/StyA2B system represents a novel type of multifunctional flavoprotein monooxygenase. Its unique mechanism of cofactor utilization provides new opportunities for biotechnological applications and is highly relevant from a structural and evolutionary point of view. PMID:20675468

  8. Downregulation of A(1) and A(2B) adenosine receptors in human trisomy 21 mesenchymal cells from first-trimester chorionic villi.

    PubMed

    Gessi, Stefania; Merighi, Stefania; Stefanelli, Angela; Mirandola, Prisco; Bonfatti, Alessandra; Fini, Sergio; Sensi, Alberto; Marci, Roberto; Varani, Katia; Borea, Pier Andrea; Vesce, Fortunato

    2012-11-01

    Human reproduction is complex and prone to failure. Though causes of miscarriage remain unclear, adenosine, a proangiogenic nucleoside, may help determine pregnancy outcome. Although adenosine receptor (AR) expression has been characterized in euploid pregnancies, no information is available for aneuploidies, which, as prone to spontaneous abortion (SA), are a potential model for shedding light on the mechanism regulating this event. AR expression was investigated in 71 first-trimester chorionic villi (CV) samples and cultured mesenchymal cells (MC) from euploid and TR21 pregnancies, one of the most frequent autosomal aneuploidy, with a view to elucidating their potential role in the modulation of vascular endothelial growth factor (VEGF) and nitric oxide (NO). Compared to euploid cells, reduced A(1) and A(2B) expression was revealed in TR21 CV and MCs. The non-selective adenosine agonist 5'-N-ethylcarboxamidoadenosine (NECA) increased NO, by activating, predominantly, A(1)AR and A(2A)AR through a molecular pathway involving hypoxia-inducible-factor-1 (HIF-1α), and increased VEGF, mainly through A(2B). In conclusion the adenosine transduction cascade appears to be disturbed in TR21 through reduced expression of A(2B) and A(1)ARs. These anomalies may be implicated in complications such as fetal growth restriction, malformation and/or SA, well known features of aneuploid pregnancies. Therefore A(1) and A(2B)ARs could be potential biomarkers able to provide an early indication of SA risk and their stimulation may turn out to improve fetoplacental perfusion by increasing NO and VEGF.

  9. Successful unintentional ABO-incompatible renal transplantation: Blood group A1B donor into an A2B recipient.

    PubMed

    Fadeyi, Emmanuel A; Stratta, Robert J; Farney, Alan C; Pomper, Gregory J

    2014-05-01

    To report a successful unintentional transplantation of a deceased donor kidney from an "incompatible" A1B donor into a recipient who was blood group A2B with unsuspected preformed anti-A1 antibodies. The donor and recipient were both typed for ABO antigens. The recipient was tested for ABO and non-ABO antibodies. The recipient was typed for HLA class I and class II antigens, including HLA antibody screen. The T-and B-flow cytometry crossmatch test was performed using standard protocol. The donor-recipient pair was a complete six-antigen human leukocyte antigen mismatch, but final T- and B-flow cytometry cross-match tests were compatible. The recipient was a 65-year-old woman with a medical history of end-stage renal disease secondary to diabetic nephropathy who underwent kidney transplantation from a 46-year-old brain-dead standard criteria donor. The recipient's RBCs were negative with A1 lectin, and the recipient was thus typed as an A2 subgroup. Anti-A1 could be demonstrated in the recipient's plasma. The donor's RBCs were positive with A1 lectin, thereby conferring an A1 blood type. It is safe to transplant across the A1/A2 blood group barrier provided that the preformed antibodies are not reactive at 37°C and with anti-human globulin.

  10. Homology modelling of the human adenosine A2B receptor based on X-ray structures of bovine rhodopsin, the β2-adrenergic receptor and the human adenosine A2A receptor

    NASA Astrophysics Data System (ADS)

    Sherbiny, Farag F.; Schiedel, Anke C.; Maaß, Astrid; Müller, Christa E.

    2009-11-01

    A three-dimensional model of the human adenosine A2B receptor was generated by means of homology modelling, using the crystal structures of bovine rhodopsin, the β2-adrenergic receptor, and the human adenosine A2A receptor as templates. In order to compare the three resulting models, the binding modes of the adenosine A2B receptor antagonists theophylline, ZM241385, MRS1706, and PSB601 were investigated. The A2A-based model was much better able to stabilize the ligands in the binding site than the other models reflecting the high degree of similarity between A2A and A2B receptors: while the A2B receptor shares about 21% of the residues with rhodopsin, and 31% with the β2-adrenergic receptor, it is 56% identical to the adenosine A2A receptor. The A2A-based model was used for further studies. The model included the transmembrane domains, the extracellular and the intracellular hydrophilic loops as well as the terminal domains. In order to validate the usefulness of this model, a docking analysis of several selective and nonselective agonists and antagonists was carried out including a study of binding affinities and selectivities of these ligands with respect to the adenosine A2A and A2B receptors. A common binding site is proposed for antagonists and agonists based on homology modelling combined with site-directed mutagenesis and a comparison between experimental and calculated affinity data. The new, validated A2B receptor model may serve as a basis for developing more potent and selective drugs.

  11. Successful ABO-Incompatible Renal Transplantation:  Blood Group A1B Donor Into A2B Recipient With Anti-A1 Isoagglutinins.

    PubMed

    Fadeyi, Emmanuel A; Stratta, Robert J; Farney, Alan C; Pomper, Gregory J

    2016-08-01

    Transplantation of the blood group A2B in a recipient was successfully performed in the setting of receiving a deceased donor kidney from an "incompatible" A1B donor. The donor and recipient were both typed for ABO blood group, including ABO genotyping. The donor and recipient were tested for ABO, non-ABO, and human leukocyte antigen (HLA) antibodies. The donor and recipient were typed for HLA antigens, including T- and B-flow cytometry crossmatch tests. The recipient's RBCs were negative with A1 lectin, and immunoglobulin G anti-A1 was demonstrated in the recipient's plasma. The donor-recipient pair was a four-antigen HLA mismatch, but final T- and B-flow cytometry crossmatch tests were compatible. The transplant procedure was uneventful; the patient experienced immediate graft function with no episodes of rejection or readmissions more than 2 years later. It may be safe to transplant across the A1/A2 blood group AB mismatch barrier in the setting of low titer anti-A1 isoagglutinins without the need for pretransplant desensitization even if the antibody produced reacts with anti-human globulin. © American Society for Clinical Pathology, 2016. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  12. Dual A1/A2B Receptor Blockade Improves Cardiac and Renal Outcomes in a Rat Model of Heart Failure with Preserved Ejection Fraction

    PubMed Central

    Tofovic, Stevan P.; Salah, Eman M.; Smits, Glenn J.; Whalley, Eric T.; Ticho, Barry; Deykin, Aaron

    2016-01-01

    Heart failure with preserved ejection fraction (HFpEF) is prevalent and often accompanied by metabolic syndrome. Current treatment options are limited. Here, we test the hypothesis that combined A1/A2B adenosine receptor blockade is beneficial in obese ZSF1 rats, an animal model of HFpEF with metabolic syndrome. The combined A1/A2B receptor antagonist 3-[4-(2,6-dioxo-1,3-dipropyl-7H-purin-8-yl)-1-bicyclo[2.2.2]octanyl]propanoic acid (BG9928) was administered orally (10 mg/kg/day) to obese ZSF1 rats (n = 10) for 24 weeks (from 20 to 44 weeks of age). Untreated ZSF1 rats (n = 9) served as controls. After 24 weeks of administration, BG9928 significantly lowered plasma triglycerides (in mg/dl: control group, 4351 ± 550; BG9928 group, 2900 ± 551) without adversely affecting plasma cholesterol or activating renin release. BG9928 significantly decreased 24-hour urinary glucose excretion (in mg/kg/day: control group, 823 ± 179; BG9928 group, 196 ± 80) and improved oral glucose tolerance, polydipsia, and polyuria. BG9928 significantly augmented left ventricular diastolic function in association with a reduction in cardiac vasculitis and cardiac necrosis. BG9928 significantly reduced 24-hour urinary protein excretion (in mg/kg/day: control group, 1702 ± 263; BG9928 group, 1076 ± 238), and this was associated with a reduction in focal segmental glomerulosclerosis, tubular atrophy, tubular dilation, and deposition of proteinaceous material in the tubules. These findings show that, in a model of HFpEF with metabolic syndrome, A1/A2B receptor inhibition improves hyperlipidemia, exerts antidiabetic actions, reduces HFpEF, improves cardiac histopathology, and affords renal protection. We conclude that chronic administration of combined A1/A2B receptor antagonists could be beneficial in patients with HFpEF, in particular those with comorbidities such as obesity, diabetes, and dyslipidemias. PMID:26585572

  13. Remifentanil-induced preconditioning has cross-talk with A1 and A2B adenosine receptors in ischemic-reperfused rat heart.

    PubMed

    Lee, Yong-Cheol; Jung, Jiyoon; Park, Sang-Jin

    2016-01-01

    The purpose of this study was to determine whether there is a cross-talk between opioid receptors (OPRs) and adenosine receptors (ADRs) in remifentanil preconditioning (R-Pre) and, if so, to investigate the types of ADRs involved in the cross-talk. Isolated rat hearts received 30 min of regional ischemia followed by 2 hr of reperfusion. OPR and ADR antagonists were perfused from 10 min before R-Pre until the end of R-Pre. The heart rate, left ventricular developed pressure (LVDP),velocity of contraction (+dP/dtmax), and coronary flow (CF) were recorded. The area at risk and area of necrosis were measured. After reperfusion, the LVDP, +dP/dtmax,and CF showed a significant increase in the R-Pre group compared with the control group (no intervention before or after regional ischemia). These increases in the R-Pre group were blocked by naloxone, a nonspecific ADR antagonist, an A1 ADR antagonist, and an A2B ADR antagonist. The infarct size was reduced significantly in the R-Pre group compared with the control group. The infarct-reducing effect in the R-Pre group was blocked by naloxone, the nonspecific ADR antagonist, the A1 ADR antagonist, and the A2B ADR antagonist. The results of this study demonstrate that there is cross-talk between ADRs and OPRs in R-Pre and that A1 ADR and A2B ADR appear to be involved in the cross-talk.

  14. A widespread sequence-specific mRNA decay pathway mediated by hnRNPs A1 and A2/B1

    PubMed Central

    Geissler, Rene; Simkin, Alfred; Floss, Doreen; Patel, Ravi; Fogarty, Elizabeth A.; Scheller, Jürgen; Grimson, Andrew

    2016-01-01

    3′-untranslated regions (UTRs) specify post-transcriptional fates of mammalian messenger RNAs (mRNAs), yet knowledge of the underlying sequences and mechanisms is largely incomplete. Here, we identify two related novel 3′ UTR motifs in mammals that specify transcript degradation. These motifs are interchangeable and active only within 3′ UTRs, where they are often preferentially conserved; furthermore, they are found in hundreds of transcripts, many encoding regulatory proteins. We found that degradation occurs via mRNA deadenylation, mediated by the CCR4–NOT complex. We purified trans factors that recognize the motifs and identified heterogeneous nuclear ribonucleoproteins (hnRNPs) A1 and A2/B1, which are required for transcript degradation, acting in a previously unknown manner. We used RNA sequencing (RNA-seq) to confirm hnRNP A1 and A2/B1 motif-dependent roles genome-wide, profiling cells depleted of these factors singly and in combination. Interestingly, the motifs are most active within the distal portion of 3′ UTRs, suggesting that their role in gene regulation can be modulated by alternative processing, resulting in shorter 3′ UTRs. PMID:27151978

  15. In vitro digestion of purified β-casein variants A(1), A(2), B, and I: effects on antioxidant and angiotensin-converting enzyme inhibitory capacity.

    PubMed

    Petrat-Melin, B; Andersen, P; Rasmussen, J T; Poulsen, N A; Larsen, L B; Young, J F

    2015-01-01

    Genetic polymorphisms of bovine milk proteins affect the protein profile of the milk and, hence, certain technological properties, such as casein (CN) number and cheese yield. However, reports show that such polymorphisms may also affect the health-related properties of milk. Therefore, to gain insight into their digestion pattern and bioactive potential, β-CN was purified from bovine milk originating from cows homozygous for the variants A(1), A(2), B, and I by a combination of cold storage, ultracentrifugation, and acid precipitation. The purity of the isolated β-CN was determined by HPLC, variants were verified by mass spectrometry, and molar extinction coefficients at λ=280nm were determined. β-Casein from each of the variants was subjected to in vitro digestion using pepsin and pancreatic enzymes. Antioxidant and angiotensin-converting enzyme (ACE) inhibitory capacities of the hydrolysates were assessed at 3 stages of digestion and related to that of the undigested samples. Neither molar extinction coefficients nor overall digestibility varied significantly between these 4 variants; however, clear differences in digestion pattern were indicated by gel electrophoresis. In particular, after 60min of pepsin followed by 5min of pancreatic enzyme digestion, one ≈4kDa peptide with the N-terminal sequence (106)H-K-E-M-P-F-P-K- was absent from β-CN variant B. This is likely a result of the (122)Ser to (122)Arg substitution in variant B introducing a novel trypsin cleavage site, leading to the changed digestion pattern. All investigated β-CN variants exhibited a significant increase in antioxidant capacity upon digestion, as measured by the Trolox-equivalent antioxidant capacity assay. After 60min of pepsin + 120min of pancreatic enzyme digestion, the accumulated increase in antioxidant capacity was ≈1.7-fold for the 4 β-CN variants. The ACE inhibitory capacity was also significantly increased by digestion, with the B variant reaching the highest inhibitory

  16. Breadth of neutralization and synergy of clinically relevant human monoclonal antibodies against HCV genotypes 1a, 1b, 2a, 2b, 2c, and 3a.

    PubMed

    Carlsen, Thomas H R; Pedersen, Jannie; Prentoe, Jannick C; Giang, Erick; Keck, Zhen-Yong; Mikkelsen, Lotte S; Law, Mansun; Foung, Steven K H; Bukh, Jens

    2014-11-01

    Human monoclonal antibodies (HMAbs) with neutralizing capabilities constitute potential immune-based treatments or prophylaxis against hepatitis C virus (HCV). However, lack of cell culture-derived HCV (HCVcc) harboring authentic envelope proteins (E1/E2) has hindered neutralization investigations across genotypes, subtypes, and isolates. We investigated the breadth of neutralization of 10 HMAbs with therapeutic potential against a panel of 16 JFH1-based HCVcc-expressing patient-derived Core-NS2 from genotypes 1a (strains H77, TN, and DH6), 1b (J4, DH1, and DH5), 2a (J6, JFH1, and T9), 2b (J8, DH8, and DH10), 2c (S83), and 3a (S52, DBN, and DH11). Virus stocks used for in vitro neutralization analysis contained authentic E1/E2, with the exception of full-length JFH1 that acquired the N417S substitution in E2. The 50% inhibition concentration (IC50) for each HMAb against the HCVcc panel was determined by dose-response neutralization assays in Huh7.5 cells with antibody concentrations ranging from 0.0012 to 100 μg/mL. Interestingly, IC50 values against the different HCVcc's exhibited large variations among the HMAbs, and only three HMAbs (HC-1AM, HC84.24, and AR4A) neutralized all 16 HCVcc recombinants. Furthermore, the IC50 values for a given HMAb varied greatly with the HCVcc strain, which supports the use of a diverse virus panel. In cooperation analyses, HMAbs HC84.24, AR3A, and, especially HC84.26, demonstrated synergistic effects towards the majority of the HCVcc's when combined individually with AR4A. Through a neutralization analysis of 10 clinically relevant HMAbs against 16 JFH1-based Core-NS2 recombinants from genotypes 1a, 1b, 2a, 2b, 2c, and 3a, we identified at least three HMAbs with potent and broad neutralization potential. The neutralization synergism obtained when pooling the most potent HMAbs could have significant implications for developing novel strategies to treat and control HCV. © 2014 by the American Association for the Study of Liver

  17. Human natural killer cell microRNA: differential expression of MIR181A1B1 and MIR181A2B2 genes encoding identical mature microRNAs.

    PubMed

    Presnell, S R; Al-Attar, A; Cichocki, F; Miller, J S; Lutz, C T

    2015-01-01

    Natural killer (NK) and T lymphocytes share many properties, yet only NK cells respond rapidly to infection and cancer without pre-activation. We found that few microRNAs (miRNAs) differed significantly between human NK and T cells. Among those miRNAs, miR-181a and miR-181b levels rose during NK cell differentiation. Prior studies indicate that miR-181a and miR-181b are critical for human NK cell development and are co-transcribed from genes on chromosome 1 (MIR181A1B1) and on chromosome 9 (MIR181A2B2). We mapped human MIR181A1B1 and MIR181A2B2 transcription start sites to 78.3 kb and 34.0 kb upstream of the mature miRNAs, generating predominantly unspliced transcripts of 80-127 kb and ~60 kb, respectively. Unlike mouse thymocytes, human T cells expressed both MIR181A1B1 and MIR181A2B2. We tested the hypothesis that NK cells differentially transcribe the two genes during development and in response to immune regulatory cytokines. During NK-cell differentiation, MIR181A2B2 expression rose markedly and exceeded that of MIR181A1B1. TGF-β treatment increased NK-cell MIR181A2B2 transcription, whereas IL-2, IL-15 and IL-12/IL-18 treatments upregulated MIR181A1B1. The MIR181A2B2 promoter was strongly transactivated by SMAD3 and SMAD4 transcription factors, suggesting that TGF-β signaling upregulates MIR181A2B2 expression, at least in part, through SMAD-dependent promoter activation.

  18. Optimization of arylindenopyrimidines as potent adenosine A(2A)/A(1) antagonists.

    PubMed

    Shook, Brian C; Rassnick, Stefanie; Chakravarty, Devraj; Wallace, Nathaniel; Ault, Mark; Crooke, Jeffrey; Barbay, J Kent; Wang, Aihua; Leonard, Kristi; Powell, Mark T; Alford, Vernon; Hall, Daniel; Rupert, Kenneth C; Heintzelman, Geoffrey R; Hansen, Kristen; Bullington, James L; Scannevin, Robert H; Carroll, Karen; Lampron, Lisa; Westover, Lori; Russell, Ronald; Branum, Shawn; Wells, Kenneth; Damon, Sandra; Youells, Scott; Beauchamp, Derek; Li, Xun; Rhodes, Kenneth; Jackson, Paul F

    2010-05-01

    Two reactive metabolites were identified in vivo for the dual A(2A)/A(1) receptor antagonist 1. Two strategies were implemented to successfully mitigate the metabolic liabilities associated with 1. Optimization of the arylindenopyrimidines led to a number of amide, ether, and amino analogs having comparable in vitro and in vivo activity. 2010 Elsevier Ltd. All rights reserved.

  19. Physiological roles of A1 and A2A adenosine receptors in regulating heart rate, body temperature, and locomotion as revealed using knockout mice and caffeine.

    PubMed

    Yang, Jiang-Ning; Chen, Jiang-Fan; Fredholm, Bertil B

    2009-04-01

    Heart rate (HR), body temperature (Temp), locomotor activity (LA), and oxygen consumption (O(2)C) were studied in awake mice lacking one or both of the adenosine A(1) or A(2A) receptors (A(1)R or A(2A)R, respectively) using telemetry and respirometry, before and after caffeine administration. All parameters were lower during day than night and higher in females than males. When compared with wild-type (WT) littermates, HR was higher in male A(1)R knockout (A(1)RKO) mice but lower in A(2A)RKO mice and intermediate in A(1)-A(2A)R double KO mice. A single dose of an unselective beta-blocker (timolol; 1 mg/kg) abolished the HR differences between these genotypes. Deletion of A(1)Rs had little effect on Temp, whereas deletion of A(2A)Rs increased it in females and decreased it in males. A(1)-A(2A)RKO mice had lower Temp than WT mice. LA was unaltered in A(1)RKO mice and lower in A(2A)RKO and A(1)-A(2A)RKO mice than in WT mice. Caffeine injection increased LA but only in mice expressing A(2A)R. Caffeine ingestion also increased LA in an A(2A)R-dependent manner in male mice. Caffeine ingestion significantly increased O(2)C in WT mice, but less in the different KO mice. Injection of 30 mg/kg caffeine decreased Temp, especially in KO mice, and hence in a manner unrelated to A(1)R or A(2A)R blockade. Selective A(2B) antagonism had little or no effect. Thus A(1)R and A(2A)R influence HR, Temp, LA, and O(2)C in mice in a sex-dependent manner, indicating effects of endogenous adenosine. The A(2A)R plays an important role in the modulation of O(2)C and LA by acute and chronic caffeine administration. There is also evidence for effects of higher doses of caffeine being independent of both A(1)R and A(2A)R.

  20. Mass spectrometry-based ligand binding assays on adenosine A1 and A2A receptors.

    PubMed

    Massink, A; Holzheimer, M; Hölscher, A; Louvel, J; Guo, D; Spijksma, G; Hankemeier, T; IJzerman, A P

    2015-12-01

    Conventional methods to measure ligand-receptor binding parameters typically require radiolabeled ligands as probes. Despite the robustness of radioligand binding assays, they carry inherent disadvantages in terms of safety precautions, expensive synthesis, special lab requirements, and waste disposal. Mass spectrometry (MS) is a method that can selectively detect ligands without the need of a label. The sensitivity of MS equipment increases progressively, and currently, it is possible to detect low ligand quantities that are usually found in ligand binding assays. We developed a label-free MS ligand binding (MS binding) assay on the adenosine A(1) and A(2A) receptors (A(1)AR and A(2A)AR), which are well-characterized members of the class A G protein-coupled receptor (GPCR) family. Radioligand binding assays for both receptors are well established, and ample data is available to compare and evaluate the performance of an MS binding assay. 1,3-Dipropyl-8-cyclopentyl-xanthine (DPCPX) and 4-(2-((7-amino-2-(furan-2-yl)-[1,2,4]triazolo[1,5-a]-[1,3,5]triazin-5-yl)amino)ethyl)phenol (ZM-241,385) are high-affinity ligands selective for the A(1)AR and A(2A)AR, respectively. To proof the feasibility of MS binding on the A(1)AR and A(2A)AR, we first developed an MS detection method for unlabeled DPCPX and ZM-241,385. To serve as internal standards, both compounds were also deuterium-labeled. Subsequently, we investigated whether the two unlabeled compounds could substitute for their radiolabeled counterparts as marker ligands in binding experiments, including saturation, displacement, dissociation, and competition association assays. Furthermore, we investigated the accuracy of these assays if the use of internal standards was excluded. The results demonstrate the feasibility of the MS binding assay, even in the absence of a deuterium-labeled internal standard, and provide great promise for the further development of label-free assays based on MS for other GPCRs.

  1. 47 CFR 80.1093 - Ship radio equipment-Sea areas A1, A2, A3, and A4.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... 47 Telecommunication 5 2013-10-01 2013-10-01 false Ship radio equipment-Sea areas A1, A2, A3, and... AND SPECIAL RADIO SERVICES STATIONS IN THE MARITIME SERVICES Global Maritime Distress and Safety System (GMDSS) Equipment Requirements for Ship Stations § 80.1093 Ship radio equipment—Sea areas A1, A2...

  2. 47 CFR 80.1093 - Ship radio equipment-Sea areas A1, A2, A3, and A4.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... 47 Telecommunication 5 2014-10-01 2014-10-01 false Ship radio equipment-Sea areas A1, A2, A3, and... AND SPECIAL RADIO SERVICES STATIONS IN THE MARITIME SERVICES Global Maritime Distress and Safety System (GMDSS) Equipment Requirements for Ship Stations § 80.1093 Ship radio equipment—Sea areas A1, A2...

  3. 47 CFR 80.1093 - Ship radio equipment-Sea areas A1, A2, A3, and A4.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... 47 Telecommunication 5 2012-10-01 2012-10-01 false Ship radio equipment-Sea areas A1, A2, A3, and... AND SPECIAL RADIO SERVICES STATIONS IN THE MARITIME SERVICES Global Maritime Distress and Safety System (GMDSS) Equipment Requirements for Ship Stations § 80.1093 Ship radio equipment—Sea areas A1, A2...

  4. 47 CFR 80.1093 - Ship radio equipment-Sea areas A1, A2, A3, and A4.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... 47 Telecommunication 5 2010-10-01 2010-10-01 false Ship radio equipment-Sea areas A1, A2, A3, and... AND SPECIAL RADIO SERVICES STATIONS IN THE MARITIME SERVICES Global Maritime Distress and Safety System (GMDSS) Equipment Requirements for Ship Stations § 80.1093 Ship radio equipment—Sea areas A1, A2...

  5. 47 CFR 80.1093 - Ship radio equipment-Sea areas A1, A2, A3, and A4.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... 47 Telecommunication 5 2011-10-01 2011-10-01 false Ship radio equipment-Sea areas A1, A2, A3, and... AND SPECIAL RADIO SERVICES STATIONS IN THE MARITIME SERVICES Global Maritime Distress and Safety System (GMDSS) Equipment Requirements for Ship Stations § 80.1093 Ship radio equipment—Sea areas A1, A2...

  6. Activation of A1, A2A, or A3 adenosine receptors attenuates lung ischemia-reperfusion injury

    PubMed Central

    Gazoni, Leo M.; Walters, Dustin M.; Unger, Eric B.; Linden, Joel; Kron, Irving L.; Laubach, Victor E.

    2010-01-01

    Objective Adenosine and the activation of specific adenosine receptors are implicated in the attenuation of inflammation and organ ischemia-reperfusion (IR) injury. We hypothesized that activation of A1, A2A, or A3 adenosine receptors would provide protection against lung IR injury. Methods Using an isolated, ventilated, blood-perfused rabbit lung model, lungs underwent 18 hours cold ischemia followed by 2 hours reperfusion. Lungs were administered either vehicle, adenosine, or selective A1, A2A, or A3 receptor agonists (CCPA, ATL-313, or IB-MECA, respectively) alone or with their respective antagonists (DPCPX, ZM241385, or MRS1191) during reperfusion. Results Compared to the vehicle-treated control group, treatment with A1, A2A, or A3 agonists significantly improved function (increased lung compliance and oxygenation and decreased pulmonary artery pressure), decreased neutrophil infiltration by myeloperoxidase activity, decreased edema, and reduced TNF-α production. Adenosine treatment was also protective but not to the level of the agonists. When each agonist was paired with its respective antagonist, all protective effects were blocked. The A2A agonist reduced pulmonary artery pressure and myeloperoxidase activity and increased oxygenation to a greater degree than the A1 or A3 agonists. Conclusions Selective activation of A1, A2A, or A3 adenosine receptors provides significant protection against lung IR injury. The decreased elaboration of the potent proinflammatory cytokine, TNF-α, and decreased neutrophil sequestration likely contribute to the overall improvement in pulmonary function. These results provide evidence for the therapeutic potential of specific adenosine receptor agonists in lung transplant recipients. PMID:20398911

  7. Bioactive saponins and glycosides. IV. Four methyl-migrated 16,17-seco-dammarane triterpene gylcosides from Chinese natural medicine, hoveniae semen seu fructus, the seeds and fruit of Hovenia dulcis THUNB.: absolute stereostructures and inhibitory activity on histamine release of hovenidulciosides A1, A2, B1, and B2.

    PubMed

    Yoshikawa, M; Murakami, T; Ueda, T; Matsuda, H; Yamahara, J; Murakami, N

    1996-09-01

    Four bioactive methyl-migrated 16,17-seco-dammarane type triterpene glycosides called hovenidulciosides A1, A2, B1, and B2 were isolated from a Chinese natural medicine, Hoveniae Semen Seu Fructus, the seeds and fruit of Hovenia dulcis THUNB. (Rhamnaceae) together with hoduloside III and (+)-gallocatechin. The absolute stereostructures of hovenidulciosides A1, A2, B1, and B2 have been elucidated by chemical and physicochemical evidence. All were found to inhibit the histamine release from rat peritoneal exudate cells induced by compound 48/80 and calcium ionophore A-23187.

  8. The Functions of the A1A2A3 Domains in Von Willebrand Factor Include Multimerin 1 Binding

    PubMed Central

    Parker, D’Andra N.; Tasneem, Subia; Farndale, Richard W.; Bihan, Dominique; Sadler, J. Evan; Sebastian, Silvie; De Groot, Philip G.

    2016-01-01

    Summary Multimerin 1 (MMRN1) is a massive, homopolymeric protein that is stored in platelets and endothelial cells for activation-induced release. In vitro, MMRN1 binds to the outer surfaces of activated platelets and endothelial cells, the extracellular matrix (including collagen) and von Willebrand factor (VWF) to support platelet adhesive functions. VWF associates with MMRN1 at high shear, not static conditions, suggesting that shear exposes cryptic sites within VWF that support MMRN1 binding. Modified ELISA and surface plasmon resonance were used to study the structural features of VWF that support MMRN1 binding, and determine the affinities for VWF-MMRN1 binding. High shear microfluidic platelet adhesion assays determined the functional consequences for VWF-MMRN1 binding. VWF binding to MMRN1 was enhanced by shear exposure and ristocetin, and required VWF A1A2A3 region, specifically the A1 and A3 domains. VWF A1A2A3 bound to MMRN1 with a physiologically relevant binding affinity (KD: 2.0 ± 0.4 nM), whereas the individual VWF A1 (KD: 39.3 ± 7.7 nM) and A3 domains (KD: 229 ± 114 nM) bound to MMRN1 with lower affinities. VWF A1A2A3 was also sufficient to support the adhesion of resting platelets to MMRN1 at high shear, by a mechanism dependent on VWF-GPIbα binding. Our study provides new information on the molecular basis of MMRN1 binding to VWF, and its role in supporting platelet adhesion at high shear. We propose that at sites of vessel injury, MMRN1 that is released following activation of platelets and endothelial cells, binds to VWF A1A2A3 region to support platelet adhesion at arterial shear rates. PMID:27052467

  9. A1 and A2a receptors mediate inhibitory effects of adenosine on the motor activity of human colon.

    PubMed

    Fornai, M; Antonioli, L; Colucci, R; Ghisu, N; Buccianti, P; Marioni, A; Chiarugi, M; Tuccori, M; Blandizzi, C; Del Tacca, M

    2009-04-01

    Experimental evidence in animal models suggests that adenosine is involved in the regulation of digestive functions. This study examines the influence of adenosine on the contractile activity of human colon. Reverse transcription-polymerase chain reaction revealed A(1) and A(2a) receptor expression in colonic neuromuscular layers. Circular muscle preparations were connected to isotonic transducers to determine the effects of 8-cyclopentyl-1,3-dipropylxanthine (DPCPX; A(1) receptor antagonist), ZM 241385 (A(2a) receptor antagonist), CCPA (A(1) receptor agonist) and 2-[(p-2-carboxyethyl)-phenethylamino]-5'-N-ethyl-carboxamide-adenosine (CGS 21680; A(2a) receptor agonist) on motor responses evoked by electrical stimulation or carbachol. Electrically evoked contractions were enhanced by DPCPX and ZM 241385, and reduced by CCPA and CGS 21680. Similar effects were observed when colonic preparations were incubated with guanethidine (noradrenergic blocker), L-732,138, GR-159897 and SB-218795 (NK receptor antagonists). However, in the presence of guanethidine, NK receptor antagonists and N(omega)-propyl-L-arginine (NPA; neuronal nitric oxide synthase inhibitor), the effects of DPCPX and CCPA were still evident, while those of ZM 241385 and CGS 21680 no longer occurred. Carbachol-induced contractions were unaffected by A(2a) receptor ligands, but they were enhanced or reduced by DPCPX and CCPA, respectively. When colonic preparations were incubated with guanethidine, NK antagonists and atropine, electrically induced relaxations were partly reduced by ZM 241385 or NPA, but unaffected by DPCPX. Dipyridamole or application of exogenous adenosine reduced electrically and carbachol-evoked contractions, whereas adenosine deaminase enhanced such motor responses. In conclusion, adenosine exerts an inhibitory control on human colonic motility. A(1) receptors mediate direct modulating actions on smooth muscle, whereas A(2a) receptors operate through inhibitory nitrergic nerve pathways.

  10. Physiological roles of A1 and A2A adenosine receptors in regulating heart rate, body temperature, and locomotion as revealed using knockout mice and caffeine

    PubMed Central

    Yang, Jiang-Ning; Chen, Jiang-Fan; Fredholm, Bertil B.

    2009-01-01

    Heart rate (HR), body temperature (Temp), locomotor activity (LA), and oxygen consumption (O2C) were studied in awake mice lacking one or both of the adenosine A1 or A2A receptors (A1R or A2AR, respectively) using telemetry and respirometry, before and after caffeine administration. All parameters were lower during day than night and higher in females than males. When compared with wild-type (WT) littermates, HR was higher in male A1R knockout (A1RKO) mice but lower in A2ARKO mice and intermediate in A1-A2AR double KO mice. A single dose of an unselective β-blocker (timolol; 1 mg/kg) abolished the HR differences between these genotypes. Deletion of A1Rs had little effect on Temp, whereas deletion of A2ARs increased it in females and decreased it in males. A1-A2ARKO mice had lower Temp than WT mice. LA was unaltered in A1RKO mice and lower in A2ARKO and A1-A2ARKO mice than in WT mice. Caffeine injection increased LA but only in mice expressing A2AR. Caffeine ingestion also increased LA in an A2AR-dependent manner in male mice. Caffeine ingestion significantly increased O2C in WT mice, but less in the different KO mice. Injection of 30 mg/kg caffeine decreased Temp, especially in KO mice, and hence in a manner unrelated to A1R or A2AR blockade. Selective A2B antagonism had little or no effect. Thus A1R and A2AR influence HR, Temp, LA, and O2C in mice in a sex-dependent manner, indicating effects of endogenous adenosine. The A2AR plays an important role in the modulation of O2C and LA by acute and chronic caffeine administration. There is also evidence for effects of higher doses of caffeine being independent of both A1R and A2AR. PMID:19218506

  11. Recent improvements in the development of A2B adenosine receptor agonists

    PubMed Central

    Tabrizi, Mojgan Aghazadeh; Fruttarolo, Francesca; Romagnoli, Romeo; Preti, Delia

    2008-01-01

    Adenosine is known to exert most of its physiological functions by acting as local modulator at four receptor subtypes named A1, A2A, A2B and A3 (ARs). Principally as a result of the difficulty in identifying potent and selective agonists, the A2B AR is the least extensively characterised of the adenosine receptors family. Despite these limitations, growing understanding of the physiological meaning of this target indicates promising therapeutic perspectives for specific ligands. As A2B AR signalling seems to be associated with pre/postconditioning cardioprotective and anti-inflammatory mechanisms, selective agonists may represent a new therapeutic group for patients suffering from coronary artery disease. Herein we present an overview of the recent advancements in identifying potent and selective A2B AR agonists reported in scientific and patent literature. These compounds can be classified into adenosine-like and nonadenosine ligands. Nucleoside-based agonists are the result of modifying adenosine by substitution at the N6-, C2-positions of the purine heterocycle and/or at the 5′-position of the ribose moiety or combinations of these substitutions. Compounds 1-deoxy-1-{6-[N′-(furan-2-carbonyl)-hydrazino]-9H-purin-9-yl}-N-ethyl-β-D-ribofuranuronamide (19, hA1Ki = 1050 nM, hA2AKi = 1550 nM, hA2B EC50 = 82 nM, hA3Ki > 5 μM) and its 2-chloro analogue 23 (hA1Ki = 3500 nM, hA2AKi = 4950 nM, hA2B EC50 = 210 nM, hA3Ki > 5 μM) were confirmed to be potent and selective full agonists in a cyclic adenosine monophosphate (cAMP) functional assay in Chinese hamster ovary (CHO) cells expressing hA2B AR. Nonribose ligands are represented by conveniently substituted dicarbonitrilepyridines, among which 2-[6-amino-3,5-dicyano-4-[4-(cyclopropylmethoxy)phenyl]pyridin-2-ylsulfanyl]acetamide (BAY-60–6583, hA1, hA2A, hA3 EC50 > 10 μM; hA2B EC50 = 3 nM) is currently under preclinical-phase investigation for treating coronary

  12. Recent improvements in the development of A2B adenosine receptor agonists

    PubMed Central

    Tabrizi, Mojgan Aghazadeh; Fruttarolo, Francesca; Romagnoli, Romeo; Preti, Delia

    2009-01-01

    Adenosine is known to exert most of its physiological functions by acting as local modulator at four receptor subtypes named A1, A2A, A2B and A3 (ARs). Principally as a result of the difficulty in identifying potent and selective agonists, the A2B AR is the least extensively characterised of the adenosine receptors family. Despite these limitations, growing understanding of the physiological meaning of this target indicates promising therapeutic perspectives for specific ligands. As A2B AR signalling seems to be associated with pre/postconditioning cardioprotective and anti-inflammatory mechanisms, selective agonists may represent a new therapeutic group for patients suffering from coronary artery disease. Herein we present an overview of the recent advancements in identifying potent and selective A2B AR agonists reported in scientific and patent literature. These compounds can be classified into adenosine-like and nonadenosine ligands. Nucleoside-based agonists are the result of modifying adenosine by substitution at the N6-, C2-positions of the purine heterocycle and/or at the 5′-position of the ribose moiety or combinations of these substitutions. Compounds 1-deoxy-1-{6-[N′-(furan-2-carbonyl)-hydrazino]-9H-purin-9-yl}-N-ethyl-β-D-ribofuranuronamide (19, hA1Ki = 1050 nM, hA2AKi = 1550 nM, hA2B EC50 = 82 nM, hA3Ki > 5 μM) and its 2-chloro analogue 23 (hA1Ki = 3500 nM, hA2AKi = 4950 nM, hA2B EC50 = 210 nM, hA3Ki > 5 μM) were confirmed to be potent and selective full agonists in a cyclic adenosine monophosphate (cAMP) functional assay in Chinese hamster ovary (CHO) cells expressing hA2B AR. Nonribose ligands are represented by conveniently substituted dicarbonitrilepyridines, among which 2-[6-amino-3,5-dicyano-4-[4-(cyclopropylmethoxy)phenyl]pyridin-2-ylsulfanyl]acetamide (BAY-60–6583, hA1, hA2A, hA3 EC50 > 10 μM; hA2B EC50 = 3 nM) is currently under preclinical-phase investigation for treating coronary

  13. Time and Sex-Dependent Effects of an Adenosine A2A/A1 Receptor Antagonist on Motivation to Self-Administer Cocaine in Rats

    PubMed Central

    Doyle, Susan E.; Breslin, Florence J.; Rieger, Jayson M.; Beauglehole, Anthony; Lynch, Wendy J.

    2012-01-01

    Adenosine is an important neuromodulator, known to interact with both dopaminergic and glutamatergic systems to influence psychostimulant action. In the present study, we examined the effects of ATL444, a novel adenosine receptor antagonist, on motivation for cocaine in male and female rats. Adult male and female Sprague-Dawley rats were trained to self-administer cocaine (1.5 mg/kg/infusion) on a fixed-ratio 1 schedule with a daily maximum of 20 infusions. Following 5 consecutive sessions during which all 20 available infusions were obtained, motivation for cocaine (0.5 mg/kg/infusion) was assessed under a progressive ratio (PR) schedule, and once responding stabilized, the effect of treatment with ATL444 (0, 15, and 30 mg/kg, i.p.) was examined. As a control, we also assessed its effects on PR responding for sucrose. Binding studies revealed that ATL 444 was 3-fold, 25-fold, and 400-fold more selective for the A2A receptor as compared to A1, A2B, and A3 receptors, respectively. ATL444 produced a significant increase in motivation for cocaine on the day of treatment in females with a trend for an increase in males. In addition, over the two PR sessions following ATL444 treatment a significant decrease in responding was observed in males but not females. Responding for sucrose was unaffected by ATL444 treatment. Our results reveal that adenosine receptor blockade may mediate both acute increases in the reinforcing effects of cocaine, and longer term inhibitory effects on cocaine reinforcement that differ according to sex. PMID:22579716

  14. Dipyridamole attenuates ischemia reperfusion induced acute kidney injury through adenosinergic A1 and A2A receptor agonism in rats.

    PubMed

    Puri, Nikkita; Mohey, Vinita; Singh, Manjinder; Kaur, Tajpreet; Pathak, Devendra; Buttar, Harpal Singh; Singh, Amrit Pal

    2016-04-01

    Dipyridamole (DYP) is an anti-platelet agent with marked vasodilator, anti-oxidant, and anti-inflammatory activity. The present study investigated the role of adenosine receptors in DYP-mediated protection against ischemia reperfusion-induced acute kidney injury (AKI) in rats. The rats were subjected to bilateral renal ischemia for 40 min followed by reperfusion for 24 h. The renal damage induced by ischemia reperfusion injury (IRI) was assessed by measuring creatinine clearance, blood urea nitrogen, uric acid, plasma potassium, fractional excretion of sodium, and microproteinuria in rats. The oxidative stress in renal tissues was assessed by quantification of thiobarbituric acid-reactive substances, superoxide anion generation, and reduced glutathione level. The hematoxylin-eosin staining was carried out to observe histopathological changes in renal tissues. DYP (10 and 30 mg/kg, intraperitoneal, i.p.) was administered 30 min before subjecting the rats to renal IRI. In separate groups, caffeine (50 mg/kg, i.p.), an adenosinergic A1 and A2A receptor antagonist was administered with and without DYP treatment before subjecting the rats to renal IRI. The ischemia reperfusion-induced AKI was demonstrated by significant changes in serum as well as urinary parameters, enhanced oxidative stress, and histopathological changes in renal tissues. The administration of DYP demonstrated protection against AKI. The prior treatment with caffeine abolished DYP-mediated reno-protection suggesting role of A1 and A2A adenosine receptors in DYP-mediated reno-protection in rats. It is concluded that adenosine receptors find their definite involvement in DYP-mediated anti-oxidative and reno-protective effect against ischemia reperfusion-induced AKI.

  15. The role of 5-arylalkylamino- and 5-piperazino- moieties on the 7-aminopyrazolo[4,3-d]pyrimidine core in affecting adenosine A1 and A2A receptor affinity and selectivity profiles.

    PubMed

    Squarcialupi, Lucia; Betti, Marco; Catarzi, Daniela; Varano, Flavia; Falsini, Matteo; Ravani, Annalisa; Pasquini, Silvia; Vincenzi, Fabrizio; Salmaso, Veronica; Sturlese, Mattia; Varani, Katia; Moro, Stefano; Colotta, Vittoria

    2017-12-01

    New 7-amino-2-phenylpyrazolo[4,3-d]pyrimidine derivatives, substituted at the 5-position with aryl(alkyl)amino- and 4-substituted-piperazin-1-yl- moieties, were synthesized with the aim of targeting human (h) adenosine A1 and/or A2A receptor subtypes. On the whole, the novel derivatives 1-24 shared scarce or no affinities for the off-target hA2B and hA3 ARs. The 5-(4-hydroxyphenethylamino)- derivative 12 showed both good affinity (Ki = 150 nM) and the best selectivity for the hA2A AR while the 5-benzylamino-substituted 5 displayed the best combined hA2A (Ki = 123 nM) and A1 AR affinity (Ki = 25 nM). The 5-phenethylamino moiety (compound 6) achieved nanomolar affinity (Ki = 11 nM) and good selectivity for the hA1 AR. The 5-(N(4)-substituted-piperazin-1-yl) derivatives 15-24 bind the hA1 AR subtype with affinities falling in the high nanomolar range. A structure-based molecular modeling study was conducted to rationalize the experimental binding data from a molecular point of view using both molecular docking studies and Interaction Energy Fingerprints (IEFs) analysis.[Formula: see text].

  16. Deletion of adenosine A1 or A2A receptors reduces L-3,4-dihydroxyphenylalanine-induced dyskinesia in a model of Parkinson’s disease

    PubMed Central

    Xiao, Danqing; Cassin, Jared J.; Healy, Brian; Burdett, Thomas C.; Chen, Jiang-Fan; Fredholm, Bertil B.; Schwarzschild, Michael A.

    2010-01-01

    Adenosine A2A receptor antagonism provides a promising approach to developing nondopaminergic therapy for Parkinson’s disease (PD). Clinical trials of A2A antagonists have targeted PD patients with L-3,4-dihydroxyphenylalanine (L-DOPA)-induced dyskinesia (LID) in an effort to improve parkinsonian symptoms. The role of adenosine in the development of LID is little known, especially regarding its actions via A1 receptors. We aimed to examine the effects of genetic deletion and pharmacological blockade of A1 and/or A2A receptors on the development of LID, on the induction of molecular markers of LID including striatal preprodynorphin and preproenkephalin (PPE), and on the integrity of dopaminergic nigrostriatal neurons in hemiparkinsonian mice. Following a unilateral 6-hydroxydopamine lesion A1, A2A and double A1-A2A knockout (KO) and wild-type littermate mice, and mice pretreated with caffeine (an antagonist of both A1 and A2A receptors) or saline were treated daily for 18–21 days with a low dose of L-DOPA. Total abnormal involuntary movements (AIMs, a measure of LID) were significantly attenuated (p<0.05) in A1 and A2A KOs, but not in A1-A2A KOs and caffeine-pretreated mice. An elevation of PPE mRNA ipsilateral to the lesion in WT mice was reduced in all KO mice. In addition, neuronal integrity assessed by striatal dopamine content was similar in all KOs and caffeine-pretreated mice following 6-hydroxydopamine lesioning. Our findings raise the possibility that A1 or A2A receptors blockade might also confer a disease-modifying benefit of reduced risk of disabling LID, whereas the effect of their combined inactivation is less clear. PMID:20828543

  17. Inosine attenuates spontaneous activity in the rat neurogenic bladder through an A2B pathway

    PubMed Central

    Doyle, Claire; Cristofaro, Vivian; Sack, Bryan S.; Lukianov, Stefan N.; Schäfer, Mattias; Chung, Yeun Goo; Sullivan, Maryrose P.; Adam, Rosalyn M.

    2017-01-01

    Neurogenic detrusor overactivity (NDO) is among the most challenging complications of spinal cord injury (SCI). A recent report by us demonstrated an improvement in NDO in SCI rats following chronic systemic treatment with the purine nucleoside inosine. The objective of this study was to investigate the mechanism of action of inosine underlying improvement of NDO. Male Sprague-Dawley rats underwent complete spinal cord transection at T8. Inosine (1 mM) delivered intravesically to SCI rats during conscious cystometry significantly decreased the frequency of spontaneous non-voiding contractions. In isolated tissue assays, inosine (1 mM) significantly decreased the amplitude of spontaneous activity (SA) in SCI bladder muscle strips. This effect was prevented by a pan-adenosine receptor antagonist CGS15943, but not by A1 or A3 receptor antagonists. The A2A antagonist ZM241385 and A2B antagonist PSB603 prevented the effect of inosine. The effect of inosine was mimicked by the adenosine receptor agonist NECA and the A2B receptor agonist BAY60-6583. The inhibition of SA by inosine was not observed in the presence of the BK antagonist, iberiotoxin, but persisted in the presence of KATP and SK antagonists. These findings demonstrate that inosine acts via an A2B receptor-mediated pathway that impinges on specific potassium channel effectors. PMID:28294142

  18. Hyperthermia-induced seizures alter adenosine A1 and A2A receptors and 5'-nucleotidase activity in rat cerebral cortex.

    PubMed

    León-Navarro, David Agustín; Albasanz, José L; Martín, Mairena

    2015-08-01

    Febrile seizure is one of the most common convulsive disorders in children. The neuromodulator adenosine exerts anticonvulsant actions through binding adenosine receptors. Here, the impact of hyperthermia-induced seizures on adenosine A1 and A2A receptors and 5'-nucleotidase activity has been studied at different periods in the cerebral cortical area by using radioligand binding, real-time PCR, and 5'-nucleotidase activity assays. Hyperthermic seizures were induced in 13-day-old rats using a warmed air stream from a hair dryer. Neonates exhibited rearing and falling over associated with hindlimb clonus seizures (stage 5 on Racine scale criteria) after hyperthermic induction. A significant increase in A1 receptor density was observed using [(3) H]DPCPX as radioligand, and mRNA coding A1 was observed 48 h after hyperthermia-induced seizures. In contrast, a significant decrease in A2A receptor density was detected, using [(3) H]ZM241385 as radioligand, 48 h after hyperthermia-evoked convulsions. These short-term changes in A1 and A2A receptors were also accompanied by a loss of 5'-nucleotidase activity. No significant variations either in A1 or A2A receptor density or 5'-nucleotidase were observed 5 and 20 days after hyperthermic seizures. Taken together, both regulation of A1 and A2A receptors and loss of 5'-nucleotidase in the cerebral cortex suggest the existence of a neuroprotective mechanism against seizures. Febrile seizure is one of the most common convulsive disorders in children. The consequences of hyperthermia-induced seizures (animal model of febrile seizures) on adenosine A1 and A2A receptors and 5'-nucleotidase activity have been studied at different periods in cerebral cortical area. A significant increase in A1 receptor density and mRNA coding A1 was observed 48 h after hyperthermia-induced seizures. In contrast, a significant decrease in A2A receptor density and 5'-nucleotidase activity was detected 48 h after convulsions evoked by hyperthermia

  19. A dispersed fluorescence and ab initio investigation of the X2B1 and A2A1 electronic states of the PH2 molecule.

    PubMed

    Jakubek, Z J; Bunker, P R; Zachwieja, M; Nakhate, S G; Simard, B; Yurchenko, S N; Thiel, W; Jensen, Per

    2006-03-07

    In this work, the X2B1 and A2A1 electronic states of the phosphino (PH2) free radical have been studied by dispersed fluorescence and ab initio methods. PH2 molecules were produced in a molecular free-jet apparatus by laser vaporizing a silicon rod in the presence of phosphine (PH3) gas diluted in helium. The laser-induced fluorescence, from the excited A2A1 electronic state down to the ground electronic state, was dispersed and analyzed. Ten (upsilon1upsilon2upsilon3) vibrationally excited levels of the ground electronic state, with upsilon1 < or = 2, upsilon2 < or = 6, and upsilon3 = 0, have been observed. Ab initio potential-energy surfaces for the X2B1 and A2A1 electronic states have been calculated at 210 points. These two states correlate with a 2Pi(u) state at linearity and they interact by the Renner-Teller coupling and spin-orbit coupling. Using the ab initio potential-energy surfaces with our RENNER computer program system, the vibronic structure and relative intensities of the A2A1 --> X2B1 emission band system have been calculated in order to corroborate the experimental assignments.

  20. Differential Expression of Adenosine A1 and A2A Receptors After Upper Cervical (C2) Spinal Cord Hemisection in Adult Rats

    PubMed Central

    Petrov, Theodor; Kreipke, Christian; Alilain, Warren; Nantwi, Kwaku D

    2007-01-01

    Background: In an animal model of spinal cord injury, a latent respiratory motor pathway can be pharmacologically activated via adenosine receptors to restore respiratory function after cervical (C2) spinal cord hemisection that paralyzes the hemidiaphragm ipsilateral to injury. Although spinal phrenic motoneurons immunopositive for adenosine receptors have been demonstrated (C3–C5), it is unclear if adenosine receptor protein levels are altered after C2 hemisection and theophylline administration. Objective: To assess the effects of C2 spinal cord hemisection and theophylline administration on the expression of adenosine receptor proteins. Methods: Adenosine A1 and A2A receptor protein levels were assessed in adult rats classified as (a) noninjured and theophylline treated, (b) C2 hemisected, (c) C2 hemisected and administered theophylline orally (3× daily) for 3 days only, and (d) C2 hemisected and administered theophylline (3× daily for 3 days) and assessed 12 days after drug administration. Assessment of A1 protein levels was carried out via immunohistochemistry and A2A protein levels by densitometry. Results: Adenosine A1 protein levels decreased significantly (both ipsilateral and contralateral to injury) after C2 hemisection; however, the decrease was attenuated in hemisected and theophylline-treated animals. Attenuation in adenosine A1 receptor protein levels persisted when theophylline administration was stopped for 12 days prior to assessment. Adenosine A2A protein levels were unchanged by C2 hemisection; however, theophylline reduced the levels within the phrenic motoneurons. Furthermore, the decrease in A2A levels persisted 12 days after theophylline was withdrawn. Conclusion: Our findings suggest that theophylline mitigates the effects of C2 hemisection by attenuating the C2 hemisection–induced decrease in A1 protein levels. Furthermore, A2A protein levels are unaltered by C2 hemisection but decrease after continuous or interrupted theophylline

  1. Adenosine A2B receptor blockade slows growth of bladder and breast tumors.

    PubMed

    Cekic, Caglar; Sag, Duygu; Li, Yuesheng; Theodorescu, Dan; Strieter, Robert M; Linden, Joel

    2012-01-01

    The accumulation of high levels of adenosine in tumors activates A(2A) and A(2B) receptors on immune cells and inhibits their ability to suppress tumor growth. Deletion of adenosine A(2A) receptors (A(2A)ARs) has been reported to activate antitumor T cells, stimulate dendritic cell (DC) function, and inhibit angiogenesis. In this study, we evaluated the effects of intermittent intratumor injection of a nonselective adenosine receptor antagonist, aminophylline (AMO; theophylline ethylenediamine) and, for the first time to our knowledge, a selective A(2B)AR antagonist, ATL801. AMO and ATL801 slowed the growth of MB49 bladder and 4T1 breast tumors in syngeneic mice and reduced by 85% metastasizes of breast cancer cells from mammary fat to lung. Based on experiments with A(2A)AR(-/-) or adenosine A(2B) receptor(-/-) mice, the effect of AMO injection was unexpectedly attributed to A(2B)AR and not to A(2A)AR blockade. AMO and ATL801 significantly increased tumor levels of IFN-γ and the IFN-inducible chemokine CXCL10, which is a ligand for CXCR3. This was associated with an increase in activated tumor-infiltrating CXCR3(+) T cells and a decrease in endothelial cell precursors within tumors. Tumor growth inhibition by AMO or ATL801 was eliminated in CXCR3(-/-) mice and RAG1(-/-) mice that lack mature T cells. In RAG1(-/-) mice, A(2B)AR deletion enhanced CD86 expression on CD11b(-) DCs. Bone marrow chimera experiments demonstrated that CXCR3 and A(2B)AR expression on bone marrow cells is required for the antitumor effects of AMO. The data suggest that blockade of A(2B)ARs enhances DC activation and CXCR3-dependent antitumor responses.

  2. Selected C8 two-chain linkers enhance the adenosine A1/A2A receptor affinity and selectivity of caffeine.

    PubMed

    van der Walt, M M; Terre'Blanche, G

    2017-01-05

    Recent research exploring C8 substitution on the caffeine core identified 8-(2-phenylethyl)-1,3,7-trimethylxanthine as a non-selective adenosine receptor antagonist. To elaborate further, we included various C8 two-chain-length linkers to enhance adenosine receptor affinity. The results indicated that the unsubstituted benzyloxy linker (1e A1Ki = 1.52 μM) displayed the highest affinity for the A1 adenosine receptor and the para-chloro-substituted phenoxymethyl (1d A2AKi = 1.33 μM) linker the best A2A adenosine receptor affinity. The position of the oxygen revealed that the phenoxymethyl linker favoured A1 adenosine receptor selectivity over the benzyloxy linker and, by introducing a para-chloro substituent, A2A adenosine receptor selectivity was obtained. Selected compounds (1c, 1e) behaved as A1 adenosine receptor antagonists in GTP shift assays and therefore represent selective and non-selective A1 and A2A adenosine receptor antagonists that may have potential for treating neurological disorders.

  3. Modulation of Ca2+-currents by sequential and simultaneous activation of adenosine A1 and A 2A receptors in striatal projection neurons.

    PubMed

    Hernández-González, O; Hernández-Flores, T; Prieto, G A; Pérez-Burgos, A; Arias-García, M A; Galarraga, E; Bargas, J

    2014-01-01

    D(1)- and D(2)-types of dopamine receptors are located separately in direct and indirect pathway striatal projection neurons (dSPNs and iSPNs). In comparison, adenosine A(1)-type receptors are located in both neuron classes, and adenosine A(2A)-type receptors show a preferential expression in iSPNs. Due to their importance for neuronal excitability, Ca(2+)-currents have been used as final effectors to see the function of signaling cascades associated with different G protein-coupled receptors. For example, among many other actions, D(1)-type receptors increase, while D(2)-type receptors decrease neuronal excitability by either enhancing or reducing, respectively, CaV1 Ca(2+)-currents. These actions occur separately in dSPNs and iSPNs. In the case of purinergic signaling, the actions of A(1)- and A(2A)-receptors have not been compared observing their actions on Ca(2+)-channels of SPNs as final effectors. Our hypotheses are that modulation of Ca(2+)-currents by A(1)-receptors occurs in both dSPNs and iSPNs. In contrast, iSPNs would exhibit modulation by both A(1)- and A2A-receptors. We demonstrate that A(1)-type receptors reduced Ca(2+)-currents in all SPNs tested. However, A(2A)-type receptors enhanced Ca(2+)-currents only in half tested neurons. Intriguingly, to observe the actions of A(2A)-type receptors, occupation of A(1)-type receptors had to occur first. However, A(1)-receptors decreased Ca(V)2 Ca(2+)-currents, while A(2A)-type receptors enhanced current through Ca(V)1 channels. Because these channels have opposing actions on cell discharge, these differences explain in part why iSPNs may be more excitable than dSPNs. It is demonstrated that intrinsic voltage-gated currents expressed in SPNs are effectors of purinergic signaling that therefore play a role in excitability.

  4. Adenosine receptors and diabetes: Focus on the A(2B) adenosine receptor subtype.

    PubMed

    Merighi, Stefania; Borea, Pier Andrea; Gessi, Stefania

    2015-09-01

    Over the last two decades, diabetes mellitus has become one of the most challenging health problems worldwide. Diabetes mellitus, classified as type I and II, is a pathology concerning blood glucose level in the body. The nucleoside adenosine has long been known to affect insulin secretion, glucose homeostasis and lipid metabolism, through activation of four G protein coupled adenosine receptors (ARs), named A1, A2A, A2B and A3. Currently, the novel promising subtype to develop new drugs for diabetes treatment is the A2BAR subtype. The use of selective agonists and antagonists for A2BAR subtype in various diabetic animal models allowed us to identify several effects of A2BAR signaling in cell metabolism. In particular, the focus of this review is to summarize the studies on purinergic signaling associated with diabetes through A2BARs modulation.

  5. The impact of adenosine and A(2B) receptors on glucose homoeostasis.

    PubMed

    Rüsing, D; Müller, C E; Verspohl, E J

    2006-12-01

    Adenosine and adenosine receptor antagonists are involved in glucose homoeostasis. The participating receptors are not known, mainly due to a lack of specific agonists and antagonists, but are reasonable targets for anti-diabetic therapy. The stable, albeit nonselective, adenosine analogue NECA (5'-N-ethylcarboxamidoadenosine) (10 microM) reduced glucose-stimulated insulin release from INS-1 cells. This was mimicked by A(1)-(CHA), A(2A)-(CGS-21680) and A(3)-receptor agonists (Cl-IB-MECA). Two newly synthesized A(2B)-receptor antagonists, PSB-53 and PSB-1115, counteracted the inhibitory effect of NECA. These in-vitro effects were mirrored by in-vivo data with respect to CHA, CGS and Cl-IB-MECA. Distinct concentrations of either PSB-53 or PSB-1115 reversed the decrease in plasma insulin induced by NECA. This was not mimicked by a corresponding change in blood glucose. The effect of PSB-1115 was also obvious in diabetic GotoKakizaki rats: plasma insulin was increased whereas blood glucose was unchanged. During most experiments the effects on blood glucose were not impressive probably because of the physiologically necessary homoeostasis. The adenosine levels were not different in normal Wistar rats and in diabetic GotoKakzaki rats. Altogether the A(2B)-receptor antagonists showed an anti-diabetic potential mainly by increasing plasma insulin levels under conditions when the adenosine tonus was elevated in-vivo and increased insulin release in-vitro.

  6. Involvement of adenosine A1 and A2A receptors in the adenosinergic modulation of the discriminative-stimulus effects of cocaine and methamphetamine in rats.

    PubMed

    Justinova, Zuzana; Ferre, Sergi; Segal, Pavan N; Antoniou, Katerina; Solinas, Marcello; Pappas, Lara A; Highkin, Jena L; Hockemeyer, Jorg; Munzar, Patrik; Goldberg, Steven R

    2003-12-01

    Adenosine, by acting on adenosine A1 and A2A receptors, is known to antagonistically modulate dopaminergic neurotransmission. We have recently reported that nonselective adenosine receptor antagonists (caffeine and 3,7-dimethyl-1-propargylxanthine) can partially substitute for the discriminative-stimulus effects of methamphetamine. In the present study, by using more selective compounds, we investigated the involvement of A1 and A2A receptors in the adenosinergic modulation of the discriminative-stimulus effects of both cocaine and methamphetamine. The effects of the A1 receptor agonist N6-cyclopentyladenosine (CPA; 0.01-0.1 mg/kg) and antagonist 8-cyclopentyl-1,3-dimethylxanthine (CPT; 1.3-23.7 mg/kg) and the A2A receptor agonist 2-p-(2-carboxyethyl)phenethylamino-5'-N-ethylcarboxamidoadenosine hydrochloride (CGS 21680; 0.03-0.18 mg/kg) and antagonist 3-(3-hydroxypropyl)-8-(3-methoxystyryl)-7-methyl-1-propargylxanthin phosphate disodium salt (MSX-3; 1-56 mg/kg) were evaluated in rats trained to discriminate either 1 mg/kg methamphetamine or 10 mg/kg cocaine from saline under a fixed-ratio 10 schedule of food presentation. The A1 and A2A receptor antagonists (CPT and MSX-3) both produced high levels of drug-lever selection when substituted for either methamphetamine or cocaine and significantly shifted dose-response curves of both psychostimulants to the left. Unexpectedly, the A2A receptor agonist CGS 21680 also produced drug-appropriate responding (although at lower levels) when substituted for the cocaine-training stimulus, and both CGS 21680 and the A1 receptor agonist CPA significantly shifted the cocaine dose-response curve to the left. In contrast, both agonists did not produce significant levels of drug-lever selection when substituted for the methamphetamine-training stimulus and failed to shift the methamphetamine dose-response curve. Therefore, adenosine A1 and A2A receptors appear to play important but differential roles in the modulation of the

  7. Bioactive constituents from chinese natural medicines. XXIV. Hypoglycemic effects of Sinocrassula indica in sugar-loaded rats and genetically diabetic KK-A(y) mice and structures of new acylated flavonol glycosides, sinocrassosides A(1), A(2), B(1), and B(2).

    PubMed

    Yoshikawa, Masayuki; Wang, Tao; Morikawa, Toshio; Xie, Haihui; Matsuda, Hisashi

    2007-09-01

    The methanolic extract from the whole plant of Sinocrassula indica (Crassulaceae) was found to inhibit the increase in serum glucose levels in oral administration of sucrose and glucose in rats at a dose of 250 mg/kg (p.o.). However, the extract did not inhibit the increase in serum glucose levels after intraperitoneal administration of glucose in these animals but did partly inhibit the gastric emptying. On the other hand, this extract significantly inhibited the increase in serum glucose levels after administration for 2 weeks in KK-A(y) mice, a genetically type II diabetic mice, at a dose of 250 mg/kg/d (p.o.) without significant changes of the weights of body, liver, and visceral fat. From the extract, four new acylated flavonol glycosides, sinocrassosides A(1), A(2), B(1), and B(2), were isolated together with 11 flavonoids and 2 megastigmanes. The absolute stereostructures of the four new compounds were elucidated on the basis of chemical and physicochemical evidence.

  8. Persistent reduction of cocaine seeking by pharmacological manipulation of adenosine A1 and A2A receptors during extinction training in rats

    PubMed Central

    O’Neill, Casey E.; Hobson, Benjamin D.; Levis, Sophia C.; Bachtell, Ryan K.

    2014-01-01

    Rationale Adenosine receptor stimulation and blockade has been shown to modulate a variety of cocaine related behaviors. Objectives These studies identify the direct effects of adenosine receptor stimulation on cocaine seeking during extinction training and the persistent effects on subsequent reinstatement to cocaine seeking. Methods Rats self-administered cocaine on a fixed-ratio 1 schedule in daily sessions over 3 weeks. Following 1 week withdrawal, the direct effects of adenosine receptor modulation were tested by administering the adenosine A1 receptor agonist, CPA (0.03 mg/kg and 0.1 mg/kg), the adenosine A2A agonist, CGS 21680 (0.03 mg/kg and 0.1 mg/kg), the presynaptic adenosine A2A receptor antagonist, SCH 442416 (0.3 mg/kg, 1 mg/kg, and 3 mg/kg), or vehicle prior to each of 6 daily extinction sessions. The persistent effects of adenosine receptor modulation during extinction training were subsequently tested on reinstatement to cocaine seeking induced by cues, cocaine, and the dopamine D2 receptor agonist, quinpirole. Results All doses of CPA and CGS 21680 impaired initial extinction responding, however only CPA treatment during extinction produced persistent impairment in subsequent cocaine- and quinpirole-induced seeking. Dissociating CPA treatment from extinction did not alter extinction responding or subsequent reinstatement. Administration of SCH 442416 had no direct effects on extinction responding, but produced dose-dependent persistent impairment of cocaine- and quinpirole-induced seeking. Conclusions These findings demonstrate that adenosine A1 or A2A receptor stimulation directly impair extinction responding. Interestingly, adenosine A1 receptor stimulation or presynaptic adenosine A2A receptor blockade during extinction produces lasting changes in relapse susceptibility. PMID:24562064

  9. Ethanol and Caffeine Effects on Social Interaction and Recognition in Mice: Involvement of Adenosine A2A and A1 Receptors.

    PubMed

    López-Cruz, Laura; San-Miguel, Noemí; Bayarri, Pilar; Baqi, Younis; Müller, Christa E; Salamone, John D; Correa, Mercé

    2016-01-01

    Ethanol and caffeine are frequently consumed in combination and have opposite effects on the adenosine system: ethanol metabolism leads to an increase in adenosine levels, while caffeine is a non-selective adenosine A1/A2A receptor antagonist. These receptors are highly expressed in striatum and olfactory tubercle, brain areas involved in exploration and social interaction in rodents. Ethanol modulates social interaction processes, but the role of adenosine in social behavior is still poorly understood. The present work was undertaken to study the impact of ethanol, caffeine and their combination on social behavior, and to explore the involvement of A1 and A2A receptors on those actions. Male CD1 mice were evaluated in a social interaction three-chamber paradigm, for preference of conspecific vs. object, and also for long-term recognition memory of familiar vs. novel conspecific. Ethanol showed a biphasic effect, with low doses (0.25 g/kg) increasing social contact and higher doses (1.0-1.5 g/kg) reducing social interaction. However, no dose changed social preference; mice always spent more time sniffing the conspecific than the object, independently of the ethanol dose. Ethanol, even at doses that did not change social exploration, produced amnestic effects on social recognition the following day. Caffeine reduced social contact (15.0-60.0 mg/kg), and even blocked social preference at higher doses (30.0-60.0 mg/kg). The A1 antagonist Cyclopentyltheophylline (CPT; 3-9 mg/kg) did not modify social contact or preference on its own, and the A2A antagonist MSX-3 (1.5-6 mg/kg) increased social interaction at all doses. Ethanol at intermediate doses (0.5-1.0 g/kg) was able to reverse the reduction in social exploration induced by caffeine (15.0-30.0 mg/kg). Although there was no interaction between ethanol and CPT or MSX-3 on social exploration in the first day, MSX-3 blocked the amnestic effects of ethanol observed on the following day. Thus, ethanol impairs the

  10. Ethanol and Caffeine Effects on Social Interaction and Recognition in Mice: Involvement of Adenosine A2A and A1 Receptors

    PubMed Central

    López-Cruz, Laura; San-Miguel, Noemí; Bayarri, Pilar; Baqi, Younis; Müller, Christa E.; Salamone, John D.; Correa, Mercé

    2016-01-01

    Ethanol and caffeine are frequently consumed in combination and have opposite effects on the adenosine system: ethanol metabolism leads to an increase in adenosine levels, while caffeine is a non-selective adenosine A1/A2A receptor antagonist. These receptors are highly expressed in striatum and olfactory tubercle, brain areas involved in exploration and social interaction in rodents. Ethanol modulates social interaction processes, but the role of adenosine in social behavior is still poorly understood. The present work was undertaken to study the impact of ethanol, caffeine and their combination on social behavior, and to explore the involvement of A1 and A2A receptors on those actions. Male CD1 mice were evaluated in a social interaction three-chamber paradigm, for preference of conspecific vs. object, and also for long-term recognition memory of familiar vs. novel conspecific. Ethanol showed a biphasic effect, with low doses (0.25 g/kg) increasing social contact and higher doses (1.0–1.5 g/kg) reducing social interaction. However, no dose changed social preference; mice always spent more time sniffing the conspecific than the object, independently of the ethanol dose. Ethanol, even at doses that did not change social exploration, produced amnestic effects on social recognition the following day. Caffeine reduced social contact (15.0–60.0 mg/kg), and even blocked social preference at higher doses (30.0–60.0 mg/kg). The A1 antagonist Cyclopentyltheophylline (CPT; 3–9 mg/kg) did not modify social contact or preference on its own, and the A2A antagonist MSX-3 (1.5–6 mg/kg) increased social interaction at all doses. Ethanol at intermediate doses (0.5–1.0 g/kg) was able to reverse the reduction in social exploration induced by caffeine (15.0–30.0 mg/kg). Although there was no interaction between ethanol and CPT or MSX-3 on social exploration in the first day, MSX-3 blocked the amnestic effects of ethanol observed on the following day. Thus, ethanol

  11. Pharmacological postconditioning of the rabbit heart with non-selective, A1 , A2A and A3 adenosine receptor agonists.

    PubMed

    Bibli, Sophia-Iris; Iliodromitis, Efstathios K; Lambertucci, Catia; Zoga, Anastasia; Lougiakis, Nikolaos; Dagres, Nikolaos; Volpini, Rosaria; Dal Ben, Diego; Kremastinos, Dimitrios Th; Tsantili Kakoulidou, Anna; Cristalli, Gloria; Andreadou, Ioanna

    2014-08-01

    We investigated the effects of novel selective and non-selective adenosine receptor agonists (ARs) on cardioprotection. Male rabbits divided into six groups were subjected to 30-min heart ischaemia and 3-h reperfusion: (1) control group, (2) postconditioning (PostC) group, (3) group A: treated with the non-selective agonist (S)-PHPNECA, (4) group B: treated with the A1 agonist CCPA, (5) group C: treated with the A2A agonist VT 7 and (6) group D: treated with the A3 agonist AR 170. The infarcted (I) and the areas at risk (R) were estimated as %I/R. In additional rabbits of all groups, heart samples were taken for determination of Akt, eNOS and STAT 3 at the 10th reperfusion minute. (S)-PHPNECA and CCPA reduced the infarct size (17.2 ± 2.9% and 17.9 ± 2.0% vs 46.8 ± 1.9% in control, P < 0.05), conferring a benefit similar to PostC (26.4 ± 0.3%). Selective A2A and A3 receptor agonists did not reduce the infarct size (39.5 ± 0.8% and 38.7 ± 3.5%, P = NS vs control). Akt, eNOS and STAT 3 were significantly activated after non-selective A1 ARs and PostC. Non-selective and A1 but not A2A and A3 ARs agonists are essential for triggering cardioprotection. The molecular mechanism involves both RISK and the JAK/STAT pathways. © 2014 Royal Pharmaceutical Society.

  12. The role of adenosine A1 and A2A receptors in the caffeine effect on MDMA-induced DA and 5-HT release in the mouse striatum.

    PubMed

    Górska, A M; Gołembiowska, K

    2015-04-01

    3,4-Methylenedioxymethamphetamine (MDMA, "ecstasy") popular as a designer drug is often used with caffeine to gain a stronger stimulant effect. MDMA induces 5-HT and DA release by interaction with monoamine transporters. Co-administration of caffeine and MDMA may aggravate MDMA-induced toxic effects on DA and 5-HT terminals. In the present study, we determined whether caffeine influences DA and 5-HT release induced by MDMA. We also tried to find out if adenosine A1 and A2A receptors play a role in the effect of caffeine by investigating the effect of the selective adenosine A1 and A2A receptor antagonists, DPCPX and KW 6002 on DA and 5-HT release induced by MDMA. Mice were treated with caffeine (10 mg/kg) and MDMA (20 or 40 mg/kg) alone or in combination. DA and 5-HT release in the mouse striatum was measured using in vivo microdialysis. Caffeine exacerbated the effect of MDMA on DA and 5-HT release. DPCPX or KW 6002 co-administered with MDMA had similar influence as caffeine, but KW 6002 was more potent than caffeine or DPCPX. To exclude the contribution of MAO inhibition by caffeine in the caffeine effect on MDMA-induced increase in DA and 5-HT, we also tested the effect of the nonxanthine adenosine receptor antagonist CGS 15943A lacking properties of MAO activity modification. Our findings indicate that adenosine A1 and A2A receptor blockade may account for the caffeine-induced exacerbation of the MDMA effect on DA and 5-HT release and may aggravate MDMA toxicity.

  13. Modulation of dopamine-mediated facilitation at the neuromuscular junction of Wistar rats: A role for adenosine A1/A2A receptors and P2 purinoceptors.

    PubMed

    Elnozahi, Neveen A; AlQot, Hadir E; Mohy El-Din, Mahmoud M; Bistawroos, Azza E; Abou Zeit-Har, Mohamed S

    2016-06-21

    This study aims to understand how dopamine and the neuromodulators, adenosine and adenosine triphosphate (ATP) modulate neuromuscular transmission. Adenosine and ATP are well-recognized for their regulatory effects on dopamine in the central nervous system. However, if similar interactions occur at the neuromuscular junction is unknown. We hypothesize that the activation of adenosine A1/A2A and/or P2 purinoceptors may influence the action of dopamine on neuromuscular transmission. Using the rat phrenic nerve hemi-diaphragm, we assessed the influence of dopamine, adenosine and ATP on the height of nerve-evoked muscle twitches. We investigated how the selective blockade of adenosine A1 receptors (2.5nM DPCPX), adenosine A2A receptors (50nM CSC) and P2 purinoceptors (100μM suramin) modified the effects of dopamine. Dopamine alone increased indirect muscle contractions while adenosine and ATP either enhanced or depressed nerve-evoked muscle twitches in a concentration-dependent manner. The facilitatory effects of 256μM dopamine were significantly reduced to 29.62±2.79% or 53.69±5.45% in the presence of DPCPX or CSC, respectively, relative to 70.03±1.57% with dopamine alone. Alternatively, the action of 256μM dopamine was potentiated from 70.03±1.57, in the absence of suramin, to 86.83±4.36%, in the presence of suramin. It can be concluded that the activation of adenosine A1 and A2A receptors and P2 purinoceptors potentially play a central role in the regulation of dopamine effects at the neuromuscular junction. Clinically this study offers new insights for the indirect manipulation of neuromuscular transmission for the treatment of disorders characterized by motor dysfunction. Copyright © 2016 IBRO. Published by Elsevier Ltd. All rights reserved.

  14. Effect of Caffeine Chronically Consumed During Pregnancy on Adenosine A1 and A2A Receptors Signaling in Both Maternal and Fetal Heart from Wistar Rats.

    PubMed

    Iglesias, Inmaculada; Albasanz, Jose Luis; Martín, Mairena

    2014-12-01

    Background: Caffeine is the most widely consumed psychoactive substance in the world, even during pregnancy. Its stimulatory effects are mainly due to antagonism of adenosine actions by blocking adenosine A1 and A2A receptors. Previous studies have shown that caffeine can cross the placenta and therefore modulate these receptors not only in the fetal brain but also in the heart. Methods: In the present work, the effect of caffeine chronically consumed during pregnancy on A1 and A2A receptors in Wistar rat heart, from both mothers and their fetuses, were studied using radioligand binding, Western-blotting, and adenylyl cyclase activity assays, as well as reverse transcription polymerase chain reaction. Results: Caffeine did not significantly alter A1R neither at protein nor at gene expression level in both the maternal and fetal heart. On the contrary, A2AR significantly decreased in the maternal heart, although mRNA was not affected. Gi and Gs proteins were also preserved. Finally, A1R-mediated inhibition of adenylyl cyclase activity did not change in the maternal heart, but A2AR mediated stimulation of this enzymatic activity significantly decreased according to the detected loss of this receptor. Conclusions: Opposite to the downregulation and desensitization of the A1R/AC pathway previously reported in the brain, these results show that this pathway is not affected in rat heart after caffeine exposure during pregnancy. In addition, A2AR is downregulated and desensitized in the maternal heart, suggesting a differential modulation of these receptor-mediated pathways by caffeine.

  15. Effect of Caffeine Chronically Consumed During Pregnancy on Adenosine A1 and A2A Receptors Signaling in Both Maternal and Fetal Heart from Wistar Rats

    PubMed Central

    Iglesias, Inmaculada; Albasanz, Jose Luis

    2014-01-01

    Background: Caffeine is the most widely consumed psychoactive substance in the world, even during pregnancy. Its stimulatory effects are mainly due to antagonism of adenosine actions by blocking adenosine A1 and A2A receptors. Previous studies have shown that caffeine can cross the placenta and therefore modulate these receptors not only in the fetal brain but also in the heart. Methods: In the present work, the effect of caffeine chronically consumed during pregnancy on A1 and A2A receptors in Wistar rat heart, from both mothers and their fetuses, were studied using radioligand binding, Western-blotting, and adenylyl cyclase activity assays, as well as reverse transcription polymerase chain reaction. Results: Caffeine did not significantly alter A1R neither at protein nor at gene expression level in both the maternal and fetal heart. On the contrary, A2AR significantly decreased in the maternal heart, although mRNA was not affected. Gi and Gs proteins were also preserved. Finally, A1R-mediated inhibition of adenylyl cyclase activity did not change in the maternal heart, but A2AR mediated stimulation of this enzymatic activity significantly decreased according to the detected loss of this receptor. Conclusions: Opposite to the downregulation and desensitization of the A1R/AC pathway previously reported in the brain, these results show that this pathway is not affected in rat heart after caffeine exposure during pregnancy. In addition, A2AR is downregulated and desensitized in the maternal heart, suggesting a differential modulation of these receptor-mediated pathways by caffeine. PMID:25538864

  16. Adenosine A1 and A2A receptors are not upstream of caffeine's dopamine D2 receptor-dependent aversive effects and dopamine-independent rewarding effects

    PubMed Central

    Sturgess, Jessica E; Ting-A-Kee, Ryan A; Podbielski, Dominik; Sellings, Laurie HL; Chen, Jiang-Fan; van der Kooy, Derek

    2010-01-01

    Caffeine is widely consumed throughout the world, yet little is known about the mechanisms underlying its rewarding and aversive properties. We show that pharmacological antagonism of dopamine not only blocks conditioned place aversions to caffeine, but reveals dopamine blockade-induced conditioned place preferences. These aversions are mediated by the dopamine D2 receptor since knockout mice showed conditioned place preferences to doses of caffeine that C57Bl/6 mice found aversive. Further, these aversions appear to be centrally-mediated since a quaternary analogue to caffeine failed to produce conditioned place aversions. While the adenosine A2A receptor is important for caffeine's physiological effects, this receptor seems only to modulate the appetitive and aversive effects of caffeine. A2A receptor knockout mice showed stronger dopamine-dependent aversions to caffeine than C57Bl/6 animals, which partially obscured the dopamine- and A2A receptor-independent preferences. Additionally, the A1 receptor, alone or in combination with the A2A receptor, does not seem to be important for caffeine's rewarding or aversive effects. Finally, excitotoxic lesions of the tegmental pedunculopontine nucleus revealed that this brain region is not involved in dopamine blockade-induced caffeine reward. This data provides surprising new information on the mechanism of action of caffeine, indicating that adenosine receptors do not mediate caffeine's appetitive and aversive effects. We show that caffeine has an atypical reward mechanism, independent of the dopaminergic system and the tegmental pedunculopontine nucleus and provide additional evidence in support of a role for the dopaminergic system in aversive learning. PMID:20576036

  17. Inosine reduces pain-related behavior in mice: involvement of adenosine A1 and A2A receptor subtypes and protein kinase C pathways.

    PubMed

    Nascimento, Francisney P; Figueredo, Sonia M; Marcon, Rodrigo; Martins, Daniel F; Macedo, Sérgio J; Lima, Denise A N; Almeida, Rúbia C; Ostroski, Rosana M; Rodrigues, Ana Lúcia S; Santos, Adair Roberto Soares

    2010-08-01

    Inosine, an endogenous purine, is the first metabolite of adenosine in a reaction catalyzed by adenosine deaminase. This study aimed to investigate the antinociceptive effects of inosine against several models of pain in mice and rats. In mice, inosine given by systemic or central routes inhibited acetic acid-induced nociception. Furthermore, inosine also decreased the late phase of formalin-induced licking and the nociception induced by glutamate. Inosine produced inhibition (for up to 4 h) of mechanical allodynia induced by complete Freund's adjuvant (CFA) injected into the mouse's paw. Given chronically for 21 days, inosine reversed the mechanical allodynia caused by CFA. Moreover, inosine also reduced the thermal (cold stimuli) and mechanical allodynia caused by partial sciatic nerve ligation (PSNL) for 4 h; when inosine was chronically administered, it decreased the mechanical allodynia induced by PSNL for 22 days. Antinociception caused by inosine in the acetic acid test was attenuated by treatment of mice with 1,3-dipropyl-8-cyclopentylxanthine (DPCPX; a selective adenosine A(1) receptor antagonist), 8-phenyltheophylline (8-PT; a nonselective adenosine A(1) receptor antagonist), and 4-{2- [7-amino-2-(2-furyl)[1,2,4]triazolo-[2,3-a][1,3,5]triazin-5-yl- amino]ethyl}phenol (ZM241385; a selective adenosine A(2A) receptor antagonist). In rats, inosine inhibited the mechanical and heat hyperalgesia induced by bradykinin and phorbol 12-myristate 13-acetate, without affecting similar responses caused by prostaglandin E(2) or forskolin. These results indicate that inosine induces antinociceptive, antiallodynic, and antihyperalgesic effects in rodents. The precise mechanisms through which inosine produces antinociception are currently under investigation, but involvement of adenosine A(1) and A(2A) receptors and blockade of the protein kinase C pathway seem to largely account for inosine's antinociceptive effect.

  18. The antidepressant-like effect of inosine in the FST is associated with both adenosine A1 and A 2A receptors.

    PubMed

    Kaster, Manuella P; Budni, Josiane; Gazal, Marta; Cunha, Mauricio P; Santos, Adair R S; Rodrigues, Ana Lúcia S

    2013-09-01

    Inosine is an endogenous purine nucleoside, which is formed during the breakdown of adenosine. The adenosinergic system was already described as capable of modulating mood in preclinical models; we now explored the effects of inosine in two predictive models of depression: the forced swim test (FST) and tail suspension test (TST). Mice treated with inosine displayed higher anti-immobility in the FST (5 and 50 mg/kg, intraperitoneal route (i.p.)) and in the TST (1 and 10 mg/kg, i.p.) when compared to vehicle-treated groups. These antidepressant-like effects started 30 min and lasted for 2 h after intraperitoneal administration of inosine and were not accompanied by any changes in the ambulatory activity in the open-field test. Both adenosine A1 and A2A receptor antagonists prevented the antidepressant-like effect of inosine in the FST. In addition, the administration of an adenosine deaminase inhibitor (1 and 10 mg/kg, i.p.) also caused an antidepressant-like effect in the FST. These results indicate that inosine possesses an antidepressant-like effect in the FST and TST probably through the activation of adenosine A1 and A2A receptors, further reinforcing the potential of targeting the purinergic system to the management of mood disorders.

  19. Two families with Leber's hereditary optic neuropathy carrying G11778A and T14502C mutations with haplogroup H2a2a1 in mitochondrial DNA.

    PubMed

    Qiao, Chen; Wei, Tanwei; Hu, Bo; Peng, Chunyan; Qiu, Xueping; Wei, Li; Yan, Ming

    2015-08-01

    The mitochondrial haplogroup has been reported to affect the clinical expression of Leber's hereditary optic neuropathy (LHON). The present study aimed to investigate the interaction between mutations and the haplogroup of mitochondrial DNA (mtDNA) in families. Two unrelated families with LHON were enrolled in the study, and clinical, genetic and molecular characterizations were determined in the affected and unaffected family members. Polymerase chain reaction direct sequencing was performed using 24 pairs of overlapping primers for whole mtDNA to screen for mutations and haplogroup. Bioinformatics analysis was performed to evaluate the pathogenic effect of these mtDNA mutations and the haplogroup. The G11778A mutation was identified in the two families. In addition, the members of family 2 exhibited the T14502C mutation and those in family 1 exhibited the T3394C and T14502C mutations, which were regarded as secondary mutations. The penetrance of visual loss in families 1 and 2 were 30.8 and 33.3%, respectively. In addition, the two families were found to be in the H2a2a1 haplogroup. In this limited sample size, it was demonstrated that the H2a2a1 haplogroup had a possible protective effect against LHON. Additional modifying factors, including environmental factors, lifestyle, estrogen levels and nuclear genes may also be important in LHON.

  20. The A2B adenosine receptor impairs the maturation and immunogenicity of dendritic cells.

    PubMed

    Wilson, Jeffrey M; Ross, William G; Agbai, Oma N; Frazier, Renea; Figler, Robert A; Rieger, Jayson; Linden, Joel; Ernst, Peter B

    2009-04-15

    The endogenous purine nucleoside adenosine is an important antiinflammatory mediator that contributes to the control of CD4(+) T cell responses. While adenosine clearly has direct effects on CD4(+) T cells, it remains to be determined whether actions on APC such as dendritic cells (DC) are also important. In this report we characterize DC maturation and function in BMDC stimulated with LPS in the presence or absence of the nonselective adenosine receptor agonist NECA (5'-N-ethylcarboxamidoadenosine). We found that NECA inhibited TNF-alpha and IL-12 in a concentration-dependent manner, whereas IL-10 production was increased. NECA-treated BMDC also expressed reduced levels of MHC class II and CD86 and were less effective at stimulating CD4(+) T cell proliferation and IL-2 production compared with BMDC exposed to vehicle control. Based on real-time RT-PCR, the A(2A) adenosine receptor (A(2A)AR) and A(2B)AR were the predominant adenosine receptors expressed in BMDC. Using adenosine receptor subtype selective antagonists and BMDC derived from A(2A)AR(-/-) and A(2B)AR(-/-)mice, it was shown that NECA modulates TNF-alpha, IL-12, IL-10, and CD86 responses predominantly via A(2B)AR. These data indicate that engagement of A(2B)AR modifies murine BMDC maturation and suggest that adenosine regulates CD4(+) T cell responses by selecting for DC with impaired immunogencity.

  1. Contribution of Adenosine A2B Receptors in Clostridium difficile Intoxication and Infection

    PubMed Central

    Li, Yuesheng; Calabrese, Gina M.; Freire, Rosemayre S.; Zaja-Milatovic, Snjezana; van Opstal, Edward; Figler, Robert A.; Linden, Joel; Guerrant, Richard L.

    2012-01-01

    Clostridium difficile toxins A (TcdA) and B (TcdB) induce a pronounced systemic and intestinal inflammatory response. A2B adenosine receptors (A2BARs) are the predominant adenosine receptors in the intestinal epithelium. We investigated whether A2BARs are upregulated in human intestinal cells by TcdA or TcdB and whether blockade of A2BARs can ameliorate C. difficile TcdA-induced enteritis and alter the outcome of C. difficile infection (CDI). Adenosine receptor subtype (A1, A2A, A2B, and A3) mRNAs were assayed in HCT-8 cells. Ileal loops from wild-type rabbits and mice and A2BAR−/− mice were treated with TcdA, with or without the selective A2BAR antagonist ATL692 or PSB1115. A murine model of CDI was used to determine the effect of A2BAR deletion or blockade with the orally available agent ATL801, on clinical outcome, histopathology and intestinal interleukin-6 (IL-6) expression from infection. TcdA and TcdB upregulated A2BAR gene expression in HCT-8 cells. ATL692 decreased TcdA-induced secretion and epithelial injury in rabbit ileum. Deletion of A2BARs reduced secretion and histopathology in TcdA-challenged mouse ileum. Deletion or blockade of A2BARs reduced histopathology, IL-6 expression, weight loss, diarrhea, and mortality in C. difficile-infected mice. A2BARs mediate C. difficile toxin-induced enteritis and disease. Inhibition of A2BAR activation may be a potential strategy to limit morbidity and mortality from CDI. PMID:23045479

  2. Presymptomatic and symptomatic ALS SOD1(G93A) mice differ in adenosine A1 and A2A receptor-mediated tonic modulation of neuromuscular transmission.

    PubMed

    Nascimento, Filipe; Sebastião, Ana M; Ribeiro, Joaquim A

    2015-12-01

    Amyotrophic lateral sclerosis (ALS) is a disease leading to neuromuscular transmission impairment. A2A adenosine receptor (A2AR) function changes with disease stage, but the role of the A(1) receptors (A1Rs) is unknown and may have a functional cross-talk with A2AR. The role of A1R in the SOD1(G93A) mouse model of ALS in presymptomatic (4-6 weeks old) and symptomatic (12-14 weeks old) phases was investigated by recording endplate potentials (EPPs), miniature endplate potentials (MEPPs), and quantal content (q.c.) of EPPs, from Mg(2+) paralyzed hemidiaphragm preparations. In presymptomatic mice, the A1R agonist, N (6)-cyclopentyladenosine (CPA) (50 nM), decreased mean EPP amplitude, MEPP frequency, and q.c. of EPPs, an effect quantitatively similar to that in age-matched wild-type (WT) mice. However, coactivation of A2AR with CGS 21680 (5 nM) prevented the effects of CPA in WT mice but not in presymptomatic SOD1(G93A) mice, suggestive of A1R/A2AR cross-talk disruption in this phase of ALS. DPCPX (50 nM) impaired CGS 21680 facilitatory action on neuromuscular transmission in WT but not in presymptomatic mice. In symptomatic animals, CPA only inhibited transmission if added in the presence of adenosine deaminase (ADA, 1 U/mL). ADA and DPCPX enhanced more transmission in symptomatic mice than in age-matched WT mice, suggestive of increase in extracellular adenosine during the symptomatic phase of ALS. The data documents that at the neuromuscular junction of presymptomatic SOD1(G93A) mice, there is a loss of A1R-A2AR functional cross-talk, while in symptomatic mice there is increased A1R tonic activation, and that with disease progression, changes in A1R-mediated adenosine modulation may act as aggravating factors during the symptomatic phase of ALS.

  3. Dual blockade of the A1 and A2A adenosine receptor prevents amyloid beta toxicity in neuroblastoma cells exposed to aluminum chloride.

    PubMed

    Giunta, Salvatore; Andriolo, Violetta; Castorina, Alessandro

    2014-09-01

    In a previous work we have shown that exposure to aluminum (Al) chloride (AlCl3) enhanced the neurotoxicity of the amyloid beta(25-35) fragment (Abeta(25-35)) in neuroblastoma cells and affected the expression of Alzheimer's disease (AD)-related genes. Caffein, a compound endowed with beneficial effects against AD, exerts neuroprotection primarily through its antagonist activity on A2A adenosine receptors (A2AR), although it also inhibits A1Rs with similar potency. Still, studies on the specific involvement of these receptors in neuroprotection in a model of combined neurotoxicity (Abeta(25-35)+AlCl3) are missing. To address this issue, cultured SH-SY5Y cells exposed to Abeta(25-35)+AlCl3 were assessed for cell viability, morphology, intracellular ROS activity and expression of apoptosis-, stress- and AD-related proteins. To define the role of A1R and A2ARs, pretreatment with caffein, specific receptor antagonists (DPCPX or SCH58261) or siRNA-mediated gene knockdown were delivered. Results indicate that AlCl3 treatment exacerbated Abeta(25-35) toxicity, increased ROS production, lipid peroxidation, β-secretase-1 (BACE1) and amyloid precursor protein (APP). Interestingly, SCH58261 successfully prevented toxicity associated to Abeta(25-35) only, whereas pretreatment with both DPCPX and SCH58261 was required to fully avert Abeta(25-35)+AlCl3-induced damage, suggesting that A1Rs might also be critically involved in protection during combined toxicity. The effects of caffein were mimicked by both N-acetyl cysteine, an antioxidant, and desferrioxamine, likely acting through distinct mechanisms. Altogether, our data establish a novel protective function associated with A1R inhibition in the setting of combined Abeta(25-35)+AlCl3 neurotoxicity, and expand our current knowledge on the potential beneficial role of caffein to prevent AD progression in subjects environmentally exposed to aluminum. Copyright © 2014 Elsevier Ltd. All rights reserved.

  4. The A2B adenosine receptor promotes Th17 differentiation via stimulation of dendritic cell IL-6.

    PubMed

    Wilson, Jeffrey M; Kurtz, Courtney C; Black, Steven G; Ross, William G; Alam, Mohammed S; Linden, Joel; Ernst, Peter B

    2011-06-15

    Adenosine is an endogenous metabolite produced during hypoxia or inflammation. Previously implicated as an anti-inflammatory mediator in CD4(+) T cell regulation, we report that adenosine acts via dendritic cell (DC) A(2B) adenosine receptor (A(2B)AR) to promote the development of Th17 cells. Mouse naive CD4(+) T cells cocultured with DCs in the presence of adenosine or the stable adenosine mimetic 5'-(N-ethylcarboximado) adenosine resulted in the differentiation of IL-17- and IL-22-secreting cells and elevation of mRNA that encode signature Th17-associated molecules, such as IL-23R and RORγt. The observed response was similar when DCs were generated from bone marrow or isolated from small intestine lamina propria. Experiments using adenosine receptor antagonists and cells from A(2B)AR(-/-) or A(2A)AR(-/-)/A(2B)AR(-/-) mice indicated that the DC A(2B)AR promoted the effect. IL-6, stimulated in a cAMP-independent manner, is an important mediator in this pathway. Hence, in addition to previously noted direct effects of adenosine receptors on regulatory T cell development and function, these data indicated that adenosine also acts indirectly to modulate CD4(+) T cell differentiation and suggested a mechanism for putative proinflammatory effects of A(2B)AR.

  5. Involvement of cAMP-PKA pathway in adenosine A1 and A2A receptor-mediated regulation of acetaldehyde-induced activation of HSCs.

    PubMed

    Yang, Yaru; Wang, He; Lv, Xiongwen; Wang, Qi; Zhao, Han; Yang, Feng; Yang, Yan; Li, Jun

    2015-08-01

    The present study was undertaken to investigate the mechanism by which adenosine receptors (ARs)-mediated the cAMP/PKA/CREB signal pathway regulates the activation of acetaldehyde-induced hepatic stellate cells (HSCs). Primary HSCs were isolated from SD rats, cultured in vitro, and activated with different concentrations of acetaldehyde at different time points. Quantitative real-time PCR and Western blotting were used to quantify both protein and mRNA levels of the four AR (A1R, A2AR, A2BR, and A3R) in rat HSCs. Selective inhibitors of PDEs and the Gi/o protein pathway, general AR agonists, and AR subtype specific agents were used to study the AR signaling. The level of cAMP was measured by radio-immunoassay, and the expression of α-SMA, collagen type I and III, PKA and p-CREB were also detected by Western blotting. Acetaldehyde could significantly promote HSC proliferation, with a maximum stimulatory effect observed at 48 h after exposure to 200 μM acetaldehyde. All four AR subtypes could be present in rat HSCs, and the mRNA and protein expression levels for A2AR and A1R in much greater abundance than those for A2BR and A3R. The expression of A2AR and A1R was significantly increased in acetaldehyde-induced HSCs as compared with that of control group, whereas the expression of A2BR and A3R remained unaffected by the addition of acetaldehyde. Curiously, there is coupling of A2AR to the Gs-AC signaling, as well as coupling of A1R to the Gi/o-AC signaling pathway in acetaldehyde-induced HSCs. Both the A2AR and A1R antagonists could suppress the activation of HSC, although they have opposing effects on cAMP signal transduction. These results suggested that a combination of cAMP/PKA/CREB signals via A2AR and A1R likely mediate the activation of acetaldehyde-induced HSCs, and A1R coupled to the Gi/o-AC signaling pathway may be masked by the more predominant A2AR that coupled to the Gs-AC signaling pathway.

  6. Down-regulation of adenosine A1 and A2A receptors in peripheral cells from idiopathic normal-pressure hydrocephalus patients.

    PubMed

    Casati, Martina; Arosio, Beatrice; Gussago, Cristina; Ferri, Evelyn; Magni, Lorenzo; Assolari, Lara; Scortichini, Valeria; Nani, Carolina; Rossi, Paolo Dionigi; Mari, Daniela

    2016-02-15

    Idiopathic normal-pressure hydrocephalus (iNPH) is a neurological disease that usually develops in the elderly. Natural history of iNPH is still unknown. It has been hypothesized that cerebrovascular diseases could have a role in etiology of chronic hydrocephalus and studies show an increased prevalence of cardiovascular diseases in iNPH patients. Moreover, evidences show a possible alteration of immune system in iNPH patients. Adenosine (Ado) is a metabolite produced in response to metabolic stress and injury. Adenosine and its receptors play an important role in vascular protection and in the modulation of inflammatory reactions and neuroinflammation. Our aim is to evaluate gene and protein expression of A1R and A2AR in the peripheral blood mononuclear cells (PBMCs) from iNPH patients compared to control subjects. We investigate if Ado system, that plays an important role in central nervous system, in vascular system, and also in inflammation, is involved in pathophysiology of iNPH disease. Our analysis showed that A1R mRNA levels and A1R density in PBMCs from iNPH patients were significantly lower than CT subjects (0.84 ± 0.12 and 2.42 ± 0.42, p<0.001 and 0.31 ± 0.02 and 0.42 ± 0.04, p=0.043; respectively). About A2AR, the gene expression in PBMCs was significantly lower in iNPH than CT (0.65 ± 0.09 and 1.5 ± 0.14, p<0.001) as well as there was a trend in protein expression: iNPH and CT (0.51 ± 0.05 and 0.62 ± 0.03; p=0.172). This preliminary study underlines the involvement of Ado system in iNPH disease whose pathophysiology is still unclear.

  7. Dynamic hip screw versus proximal femur locking compression plate in intertrochanteric femur fractures (AO 31A1 and 31A2): A prospective randomized study

    PubMed Central

    Agrawal, Prabhat; Gaba, Sahil; Das, Saubhik; Singh, Ranjit; Kumar, Arvind; Yadav, Gajanand

    2017-01-01

    Introduction: Intertrochanteric fractures are common in elderly population and pose a significant financial burden to the society. Anatomically contoured proximal femur locking compression plate (PFLCP) is the latest addition in the surgeons’ armamentarium to deal with these fractures. It creates an angular stable construct, which will theoretically lessen the risk of failure by screw cut-out and varus collapse, the common mode of DHS failure. We compared DHS with PFLCP in AO type 31A1 and 31A2 intertrochanteric fractures. Materials and Methods: A randomized prospective study was carried out between June 2011 and June 2013. 26 cases each of DHS and PFLCP were included. Results: Functional and radiological outcome was similar in both groups. Conclusion: Both DHS and PFLCP are good choices for stable intertrochanteric fractures, and both lead to excellent functional outcomes, but non-union might be more common with PFLCP.

  8. N9-benzyl-substituted 1,3-dimethyl- and 1,3-dipropyl-pyrimido[2,1-f]purinediones: synthesis and structure-activity relationships at adenosine A1 and A2A receptors.

    PubMed

    Drabczyńska, Anna; Müller, Christa E; Karolak-Wojciechowska, Janina; Schumacher, Britta; Schiedel, Anke; Yuzlenko, Olga; Kieć-Kononowicz, Katarzyna

    2007-07-15

    Synthesis and physicochemical properties of N-benzyl pyrimido[2,1-f]purinediones are described. These derivatives were synthesized by the cyclization of 7-chloropropylo-8-bromo-1,3-dimethyl- or 1,3-dipropyl xanthine derivatives with corresponding (un)substituted benzylamines. Dipropyl derivatives were obtained under microwave irradiation conditions either. The obtained compounds (1-20) were evaluated for their affinity to adenosine A1 and A2A receptors, selected compounds were additionally investigated for affinity to the A3 receptor subtype. The results of the radioligand binding assays to A1 and A2A adenosine receptors showed that most of the 1,3-dimethyl-9-benzylpyrimidopurinediones exhibited selective affinity to A2A receptors at micromolar or submicromolar concentrations (for example, derivative 9 with o-methoxy substituent displayed a Ki value of 0.699 microM at rat A2A receptor with more than 36-fold selectivity). Contrary to previously described arylpyrimido[2,1-f]purinediones dipropyl derivatives (compounds 15-20) showed affinity to both kinds of receptors increased, however A1 affinity increased to a larger extent, with the result that A2A selectivity was abolished. The best adenosine A1 receptor ligand was m-chlorobenzyl derivative 18 (Ki=0.089 microM and 5-fold A1 selectivity). Structure-activity relationships were discussed with the analysis of lipophilic and spatial properties of the investigated compounds. Pharmacophore model of adenosine A1 receptor antagonist was adopted for this purpose.

  9. Role of adenosine A2b receptor overexpression in tumor progression.

    PubMed

    Sepúlveda, Cesar; Palomo, Iván; Fuentes, Eduardo

    2016-12-01

    The adenosine A2b receptor is a G-protein coupled receptor. Its activation occurs with high extracellular adenosine concentration, for example in inflammation or hypoxia. These conditions are generated in the tumor environment. Studies show that A2b receptor is overexpressed in various tumor lines and biopsies from patients with different cancers. This suggests that A2b receptor can be used by tumor cells to promote progression. Thus A2b participates in different events, such as angiogenesis and metastasis, besides exerting immunomodulatory effects that protect tumor cells. Therefore, adenosine A2b receptor appears as an interesting therapeutic target for cancer treatment.

  10. The Macrophage A2b Adenosine Receptor Regulates Tissue Insulin Sensitivity

    PubMed Central

    Koupenova, Milka; Carroll, Shannon; Ravid, Katya

    2014-01-01

    High fat diet (HFD)-induced type 2 diabetes continues to be an epidemic with significant risk for various pathologies. Previously, we identified the A2b adenosine receptor (A2bAR), an established regulator of inflammation, as a regulator of HFD-induced insulin resistance. In particular, HFD was associated with vast upregulation of liver A2bAR in control mice, and while mice lacking this receptor showed augmented liver inflammation and tissue insulin resistance. As the A2bAR is expressed in different tissues, here, we provide the first lead to cellular mechanism by demonstrating that the receptor's influence on tissue insulin sensitivity is mediated via its expression in macrophages. This was shown using a newly generated transgenic mouse model expressing the A2bAR gene in the macrophage lineage on an otherwise A2bAR null background. Reinstatement of macrophage A2bAR expression in A2bAR null mice fed HFD restored insulin tolerance and tissue insulin signaling to the level of control mice. The molecular mechanism for this effect involves A2bAR-mediated changes in cyclic adenosine monophosphate in macrophages, reducing the expression and release of inflammatory cytokines, which downregulate insulin receptor-2. Thus, our results illustrate that macrophage A2bAR signaling is needed and sufficient for relaying the protective effect of the A2bAR against HFD-induced tissue inflammation and insulin resistance in mice. PMID:24892847

  11. A2B and A3 Adenosine Receptors Modulate Vascular Endothelial Growth Factor and Interleukin-8 Expression in Human Melanoma Cells Treated with Etoposide and Doxorubicin

    PubMed Central

    Merighi, Stefania; Simioni, Carolina; Gessi, Stefania; Varani, Katia; Mirandola, Prisco; Tabrizi, Mojgan Aghazadeh; Baraldi, Pier Giovanni; Borea, Pier Andrea

    2009-01-01

    Cancer patients undergoing treatment with systemic cancer chemotherapy drugs often have abnormal growth factor and cytokine profiles. Thus, serum levels of interleukin-8 (IL-8) are elevated in patients with malignant melanoma. In addition to IL-8, aggressive melanoma cells secrete, through its transcriptional regulator hypoxia-inducible factor 1 (HIF-1), vascular endothelial growth factor (VEGF), which promotes angiogenesis and metastasis of human cancerous cells. Whether these responses are related to adenosine, a ubiquitous mediator expressed at high concentrations in cancer and implicated in numerous inflammatory processes, is not known and is the focus of this study. We have examined whether the DNA-damaging agents etoposide (VP-16) and doxorubicin can affect IL-8, VEGF, and HIF-1 expressions in human melanoma cancer cells. In particular, we have investigated whether these responses are related to the modulation of the adenosine receptor subtypes, namely, A1, A2A, A2B, and A3. We have demonstrated that A2B receptor blockade can impair IL-8 production, whereas blocking A3 receptors, it is possible to further decrease VEGF secretion in melanoma cells treated with VP-16 and doxorubicin. This understanding may present the possibility of using adenosine antagonists to reduce chemotherapy-induced inflammatory cytokine production and to improve the ability of chemotherapeutic drugs to block angiogenesis. Consequently, we conclude that adenosine receptor modulation may be useful for refining the use of chemotherapeutic drugs to treat human cancer more effectively. PMID:19794965

  12. Altered expression of adenosine A1 and A2A receptors in the carotid body and nucleus tractus solitarius of adult male and female rats following neonatal caffeine treatment.

    PubMed

    Bairam, Aida; Joseph, Vincent; Lajeunesse, Yves; Kinkead, Richard

    2009-09-01

    Neonatal caffeine treatment (adenosine receptor antagonist, 15 mg/kg/day, between postnatal days 3 and 12) affects respiratory patterns in adult male but not female rats as shown by an increase in the respiratory frequency in the early phase of response to hypoxia and an increase in the tidal volume in the late phase of response. Here, we tested the hypothesis that these changes are correlated with modified expression of adenosine receptors in the chemoreflex pathway. Carotid bodies, nucleus tractus solitarii, and superior cervical ganglia were collected from 3-month-old male and female rats that were either naive (not manipulated during the neonatal period) or treated with caffeine (NCT) or water (NWT) between postnatal days 3 and 12 by gavage. Western blot analysis was used to assess the expression of adenosine A(1) and A(2A) receptors and tyrosine hydroxylase, the rate-limiting enzyme for dopamine synthesis. In male rats, there was a 37% increase in the level of A(2A) receptor and a 17% decrease in tyrosine hydroxylase in the carotid body of NCT (p<0.001) as compared to NWT rats. In the nucleus tractus solitarius, we found a 13% and 19% decrease in A(1) receptor expression in NWT and NCT rats (p<0.01), respectively, compared to naive rats. In the superior cervical ganglion, there was no change in A(1) receptor, A(2A) receptor, and tyrosine hydroxylase expression. In female rats, the only changes observed were decreases of 12% and 15% in A(1) receptor levels in the nucleus tractus solitarius of NWT and NCT rats (p<0.01), respectively, compared to naive rats. We conclude that NCT induces long-term changes in the adenosine receptor system. These changes may partially explain the modifications of the respiratory pattern induced by NCT in adults. The increased expression of the adenosine A(2A) receptor (specific to male rats), combined with the decreased tyrosine hydroxylase expression in the carotid body, suggests that NCT affects adenosine-dopamine interactions

  13. Binding of β4γ5 by Adenosine A1 and A2A Receptors Determined by Stable Isotope Labeling with Amino Acids in Cell Culture and Mass Spectrometry†

    PubMed Central

    Wang, Dora Bigler; Sherman, Nicholas E.; Shannon, John D.; Leonhardt, Susan A.; Mayeenuddin, Linnia H.; Yeager, Mark; McIntire, William E.

    2011-01-01

    Characterization of G protein βγ dimer isoform expression in different cellular contexts has been impeded by low levels of protein expression, broad isoform heterogeneity, and antibodies of limited specificity, sensitivity or availability. As a new approach, we used quantitative mass spectrometry to characterize native βγ dimers associated with adenosine A1:αi1 and adenosine A2A:αS receptor fusion proteins expressed in HEK-293 cells. Cells expressing A1:αi1 were cultured in media containing [13C6] Arg and [13C6] Lys, and βγ labeled with heavy isotopes purified. Heavy βγ was combined with either recombinant βγ purified from Sf9 cells, βγ purified from the A2A:αS expressed in HEK-293 cells cultured in standard media, or an enriched βγ fraction from HEK-293 cells. Samples were separated by SDS-PAGE, and protein bands containing β and γ were excised, digested with trypsin, separated by HPLC and isotope ratios analyzed by mass spectrometry. Three β isoforms, β1, β2 and β4, and seven γ isoforms, γ2, γ4, γ5, γ7, γ10, γ11 and γ12 were identified in the analysis. β1 and γ5 were most abundant in the enriched βγ fraction, and this βγ profile was generally mirrored in the fusion proteins. However, both A2A:αS and A1:αi1 bound more β4 and γ5 compared to the enriched βγ fraction; also, more β4 was associated with A2A:αS than A1:αi1. Both fusion proteins also contained less γ2, γ10 and γ12 than the enriched βγ fraction. These results suggest that preferences for particular βγ isoforms may be driven in part by structural motifs common to adenosine receptor family members. PMID:21128647

  14. A2B adenosine receptors stimulate IL-6 production in primary murine microglia through p38 MAPK kinase pathway.

    PubMed

    Merighi, Stefania; Bencivenni, Serena; Vincenzi, Fabrizio; Varani, Katia; Borea, Pier Andrea; Gessi, Stefania

    2017-03-01

    The hallmark of neuroinflammation is the activation of microglia, the immunocompetent cells of the CNS, releasing a number of proinflammatory mediators implicated in the pathogenesis of neuronal diseases. Adenosine is an ubiquitous autacoid regulating several microglia functions through four receptor subtypes named A1, A2A, A2B and A3 (ARs), that represent good targets to suppress inflammation occurring in CNS. Here we investigated the potential role of ARs in the modulation of IL-6 secretion and cell proliferation in primary microglial cells. The A2BAR agonist 2-[[6-Amino-3,5-dicyano-4-[4-(cyclopropylmethoxy)phenyl]-2-pyridinyl]thio]-acetamide (BAY60-6583) stimulated IL-6 increase under normoxia and hypoxia, in a dose- and time-dependent way. In cells incubated with the blockers of phospholipase C (PLC), protein kinase C epsilon (PKC-ε) and PKC delta (PKC-δ) the IL-6 increase due to A2BAR activation was strongly reduced, whilst it was not affected by the inhibitor of adenylyl cyclase (AC). Investigation of cellular signalling involved in the A2BAR effect revealed that only the inhibitor of p38 mitogen activated protein kinase (MAPK) was able to block the agonist's effect on IL-6 secretion, whilst inhibitors of pERK1/2, JNK1/2 MAPKs and Akt were not. Stimulation of p38 by BAY60-6583 was A2BAR-dependent, through a pathway affecting PLC, PKC-ε and PKC-δ but not AC, in both normoxia and hypoxia. Finally, BAY60-6583 increased microglial cell proliferation involving A2BAR, PLC, PKC-ε, PKC-δ and p38 signalling. In conclusion, A2BARs activation increased IL-6 secretion and cell proliferation in murine primary microglial cells, through PLC, PKC-ε, PKC-δ and p38 pathways, thus suggesting their involvement in microglial activation and neuroinflammation.

  15. Production of novel antioxidative prenyl naphthalen-ols by combinational bioconversion with dioxygenase PhnA1A2A3A4 and prenyltransferase NphB or SCO7190.

    PubMed

    Shindo, Kazutoshi; Tachibana, Ayako; Tanaka, Ayumi; Toba, Shizuka; Yuki, Emi; Ozaki, Taro; Kumano, Takuto; Nishiyama, Makoto; Misawa, Norihiko; Kuzuyama, Tomohisa

    2011-01-01

    We performed combinational bioconversion of substituted naphthalenes with PhnA1A2A3A4 (an aromatic dihydroxylating dioxygenase from marine bacterium Cycloclasticus sp. strain A5) and prenyltransferase NphB (geranyltransferase from Streptomyces sp. strain CL190) or SCO7190 (dimethylallyltransferase from Streptomyces coelicolor A3(2)) to produce prenyl naphthalen-ols. Using 2-methylnaphthalene, 1-methoxynaphthalene, and 1-ethoxynaphthalene as the starting substrates, 10 novel prenyl naphthalen-ols were produced by combinational bioconversion. These novel prenyl naphthalen-ols each showed potent antioxidative activity against a rat brain homogenate model. 2-(2,3-Dihydroxyphenyl)-5,7-dihydroxy-chromen-4-one (2',3'-dihydroxychrysin) generated with another aromatic dihydroxylating dioxygenase and subsequent dehydrogenase was also geranylated at the C-5'-carbon by the action of NphB.

  16. Adenosine A2B-receptor-mediated cyclic AMP accumulation in primary rat astrocytes.

    PubMed Central

    Peakman, M. C.; Hill, S. J.

    1994-01-01

    1. The effects of adenosine receptor agonists and antagonists on the accumulation of cyclic AMP have been investigated in primary cultures of rat astrocytes. 2. Adenosine A2-receptor stimulation caused a concentration-dependent increase in the accumulation of [3H]-cyclic AMP in cells prelabelled with [3H]-adenine. The rank order of agonist potencies was 5'-N-ethylcarboxamidoadenosine (NECA; EC50 = 1 microM) > adenosine (EC50 = 5 microM) > 2-chloroadenosine (EC50 = 20 microM) >> CGS 21680 (EC50 > 10 microM). The presence of 0.5 microM dipyridamole, an adenosine uptake blocker, had no effect on the potency of adenosine. 3. The response to 10 microM NECA was antagonized in a concentration-dependent manner by the non-selective adenosine receptor antagonists, xanthine amine congener (apparent KD = 12 nM), PD 115,199 (apparent KD = 134 nM) and 8-phenyltheophylline (apparent KD = 126 nM). However, the A1-receptor-selective antagonist, 8-cyclopentyl-1,3-dipropylxanthine, had no significant effect on the responses to NECA or 2-chloroadenosine at concentrations up to 1 microM. 4. Stimulation of A1-receptors with the selective agonist, N6-cyclopentyladenosine, did not alter the basal accumulation of [3H]-cyclic AMP but inhibited a forskolin-mediated elevation of [3H]-cyclic AMP accumulation by a maximal value of 42%. This inhibition was fully reversed in the presence of 0.1 microM, 8-cyclopentyl-1,3-dipropylxanthine. 5. The time course for NECA-mediated [3H]-cyclic AMP accumulation was investigated. The results suggest that there is a substantial efflux of cyclic AMP from the cells in addition to the rapid and sustained elevation of intracellular cyclic AMP (5 fold over basal) which was also observed. 6. These data indicate that rat astrocytes in primary culture express an A2B-adenosine receptor coupled positively to adenylyl cyclase. Furthermore, the presence of A1-receptors negatively coupled to adenylyl cyclase appears to have no significant effect on the A2B

  17. The A2B adenosine receptor modulates pulmonary hypertension associated with interstitial lung disease.

    PubMed

    Karmouty-Quintana, Harry; Zhong, Hongyan; Acero, Luis; Weng, Tingting; Melicoff, Ernestina; West, James D; Hemnes, Anna; Grenz, Almut; Eltzschig, Holger K; Blackwell, Timothy S; Xia, Yang; Johnston, Richard A; Zeng, Dewan; Belardinelli, Luiz; Blackburn, Michael R

    2012-06-01

    Development of pulmonary hypertension is a common and deadly complication of interstitial lung disease. Little is known regarding the cellular and molecular mechanisms that lead to pulmonary hypertension in patients with interstitial lung disease, and effective treatment options are lacking. The purpose of this study was to examine the adenosine 2B receptor (A(2B)R) as a regulator of vascular remodeling and pulmonary hypertension secondary to pulmonary fibrosis. To accomplish this, cellular and molecular changes in vascular remodeling were monitored in mice exposed to bleomycin in conjunction with genetic removal of the A(2B)R or treatment with the A(2B)R antagonist GS-6201. Results demonstrated that GS-6201 treatment or genetic removal of the A(2B)R attenuated vascular remodeling and hypertension in our model. Furthermore, direct A(2B)R activation on vascular cells promoted interleukin-6 and endothelin-1 release. These studies identify a novel mechanism of disease progression to pulmonary hypertension and support the development of A(2B)R antagonists for the treatment of pulmonary hypertension secondary to interstitial lung disease.

  18. Probing biased/partial agonism at the G protein-coupled A(2B) adenosine receptor.

    PubMed

    Gao, Zhan-Guo; Balasubramanian, Ramachandran; Kiselev, Evgeny; Wei, Qiang; Jacobson, Kenneth A

    2014-08-01

    G protein-coupled A(2B) adenosine receptor (AR) regulates numerous important physiological functions, but its activation by diverse A(2B)AR agonists is poorly profiled. We probed potential partial and/or biased agonism in cell lines expressing variable levels of endogenous or recombinant A(2B)AR. In cAMP accumulation assays, both 5'-substituted NECA and C2-substituted MRS3997 are full agonists. However, only 5'-substituted adenosine analogs are full agonists in calcium mobilization, ERK1/2 phosphorylation and β-arrestin translocation. A(2B)AR overexpression in HEK293 cells markedly increased the agonist potency and maximum effect in cAMP accumulation, but less in calcium and ERK1/2. A(2B)AR siRNA silencing was more effective in reducing the maximum cAMP effect of non-nucleoside agonist BAY60-6583 than NECA's. A quantitative 'operational model' characterized C2-substituted MRS3997 as either balanced (cAMP accumulation, ERK1/2) or strongly biased agonist (against calcium, β-arrestin). N⁶-substitution biased against ERK1/2 (weakly) and calcium and β-arrestin (strongly) pathways. BAY60-6583 is ERK1/2-biased, suggesting a mechanism distinct from adenosine derivatives. BAY60-6583, as A(2B)AR antagonist in MIN-6 mouse pancreatic β cells expressing low A(2B)AR levels, induced insulin release. This is the first relatively systematic study of structure-efficacy relationships of this emerging drug target. Published by Elsevier Inc.

  19. Multiple sclerosis lymphocytes upregulate A2A adenosine receptors that are antiinflammatory when stimulated.

    PubMed

    Vincenzi, Fabrizio; Corciulo, Carmen; Targa, Martina; Merighi, Stefania; Gessi, Stefania; Casetta, Ilaria; Gentile, Mauro; Granieri, Enrico; Borea, Pier Andrea; Varani, Katia

    2013-08-01

    Multiple sclerosis (MS) is an autoimmune-mediated inflammatory disease characterized by multifocal areas of demyelination. Experimental evidence indicates that A2A adenosine receptors (ARs) play a pivotal role in the inhibition of inflammatory processes. The aim of this study was to investigate the contribution of A2A ARs in the inhibition of key pro-inflammatory mediators for the pathogenesis of MS. In lymphocytes from MS patients, A1, A2A, A2B, and A3 ARs were analyzed by using RT-PCR, Western blotting, immunofluorescence, and binding assays. Moreover the effect of A2A AR stimulation on proinflammatory cytokine release such as TNF-α, IFN-γ, IL-6, IL-1β, IL-17, and on lymphocyte proliferation was evaluated. The capability of an A2A AR agonist on the modulation of very late antigen (VLA)-4 expression and NF-κB was also explored. A2A AR upregulation was observed in lymphocytes from MS patients in comparison with healthy subjects. The stimulation of these receptors mediated a significant inhibition of TNF-α, IFN-γ, IL-6, IL-1β, IL-17, and cell proliferation as well as VLA-4 expression and NF-κB activation. This new evidence highlights that A2A AR agonists could represent a novel therapeutic tool for MS treatment as suggested by the antiinflammatory role of A2A ARs in lymphocytes from MS patients.

  20. Immunodiagnosis of tumors in vivo using radiolabeled monoclonal antibody A2B5

    SciTech Connect

    Reintgen, D.S.; Shimizu, K.; Coleman, E.; Briner, W.; Kitzmiller, J.; Eisenbarth, G.; Seigler, H.F.

    1983-07-01

    Recently a murine monoclonal antibody (A2B5) has been described that reacts with a membrane associated GQ ganglioside common to peptide secreting normal cells and tumors. In vitro binding data demonstrated the presence of this ganglioside on neurons, adrenal medulla, and pancreatic islets, along with neuroendocrine tumors such as insulinomas, pheochromocytomas, melanomas and neuroblastomas. Negative binding has previously been shown for tissue sections from liver, kidney, colon, lung, stomach, and tumors not derived from the neural crest. Because of the specificity at A2B5 in vitro, this monoclonal antibody was labeled with /sup 131/I for in vivo tumor localization studies. Daily radionuclear scans were obtained in 5 KX rats bearing the radiation induced rat insulinoma with disappearance of the label from the blood pool and concentration in the tumor so that by the fourth day, the only activity present by scan was in the insulinoma. In addition A2B5 also localized to five different human melanoma cells lines grown in nude mice with high tumor/blood levels compared to normal tissues, while no localization is seen in nudes carrying osteosarcomas, colon, bladder, and renal cell carcinomas. In addition antibody A2B5 did not concentrate in any normal tissue though the antigen is present on several. The finding that A2B5 reacts across species lines (mouse, rat, man) lends itself to obvious diagnostic and therapeutic possibilities.

  1. Immunodiagnosis of tumors in vivo using radiolabeled monoclonal antibody A2B5.

    PubMed

    Reintgen, D S; Shimizu, K; Coleman, E; Briner, W; Kitzmiller, J; Eisenbarth, G; Seigler, H F

    1983-07-01

    Recently a murine monoclonal antibody (A2B5) has been described that reacts with a membrane associated GQ ganglioside common to peptide secreting normal cells and tumors. In vitro binding data demonstrated the presence of this ganglioside on neurons, adrenal medulla, and pancreatic islets, along with neuroendocrine tumors such as insulinomas, pheochromocytomas, melanomas and neuroblastomas. Negative binding has previously been shown for tissue sections from liver, kidney, colon, lung, stomach, and tumors not derived from the neural crest. Because of the specificity at A2B5 in vitro, this monoclonal antibody was labeled with 131I for in vivo tumor localization studies. Daily radionuclear scans were obtained in 5 KX rats bearing the radiation induced rat insulinoma with disappearance of the label from the blood pool and concentration in the tumor so that by the fourth day, the only activity present by scan was in the insulinoma. Tissue-counting data showed tumor/blood ratios (av +/- SE, 1.29 +/- 0.25) of A2B5 activity two to ten times the average activity found in other organs (0.28 +/- 0.05). No tumor concentration of the control nonspecific monoclonal antibody P3X63 was evident (0.27 +/- 0.04). In addition A2B5 also localized to five different human melanoma cells lines grown in nude mice with high tumor/blood levels (1.04 +/- 0.27) compared to normal tissues (0.32 +/- 0.05) (P = .0005), while no localization is seen in nudes carrying osteosarcomas, colon, bladder, and renal cell carcinomas. In addition antibody A2B5 did not concentrate in any normal tissue though the antigen is present on several. The finding that A2B5 reacts across species lines (mouse, rat, man) lends itself to obvious diagnostic and therapeutic possibilities.

  2. Textbook Evaluation: An Analysis of Listening Comprehension Parts in Top Notch 2A & 2B

    ERIC Educational Resources Information Center

    Soori, Afshin; Haghani, Elham

    2015-01-01

    Textbooks are the instruments that assist both teachers and learners in process of second language learning. With respect to the importance of textbooks in a language course, evaluation of course books is a significant issue for most researchers. The present study investigated and analyzed Listening Comprehension parts in Top Notch 2A & 2B 2nd…

  3. New chromene scaffolds for adenosine A(2A) receptors: synthesis, pharmacology and structure-activity relationships.

    PubMed

    Areias, Filipe; Costa, Marta; Castro, Marián; Brea, José; Gregori-Puigjané, Elisabet; Proença, M Fernanda; Mestres, Jordi; Loza, María I

    2012-08-01

    In silico screening of a collection of 1584 academic compounds identified a small molecule hit for the human adenosine A(2A) receptor (pK(i) = 6.2) containing a novel chromene scaffold (3a). To explore the structure-activity relationships of this new chemical series for adenosine receptors, a focused library of 43 2H-chromene-3-carboxamide derivatives was synthesized and tested in radioligand binding assays at human adenosine A(1), A(2A), A(2B) and A(3) receptors. The series was found to be enriched with bioactive compounds for adenosine receptors, with 14 molecules showing submicromolar affinity (pK(i) ≥ 6.0) for at least one adenosine receptor subtype. These results provide evidence that the chromene scaffold, a core structure present in natural products from a wide variety of plants, vegetables, and fruits, constitutes a valuable source for novel therapeutic agents. Copyright © 2012 Elsevier Masson SAS. All rights reserved.

  4. Mysterious link between iron overload and CDKN2A/2B

    PubMed Central

    Toyokuni, Shinya

    2011-01-01

    Persistent oxidative stress has been associated with carcinogenesis. Iron overload is considered one such condition that causes oxidative stress. Epidemiological studies support a close link between iron overload and carcinogenesis. Reportedly, regular semiannual phlebotomies reduced cancer risk in an otherwise normal population. More specifically, genetic hemochromatosis, chronic viral hepatitis, ovarian endometriosis and asbestosis induce iron overload, which can lead to hepatocellular carcinoma, ovarian carcinoma or mesothelioma in humans. Through a combination of animal experiments and microarray analyses, homozygous deletion of CDKN2A/2B has been recognized as one of the major target genes involved in iron overload-induced carcinogenesis. CDKN2A/2B are the second most frequently inactivated tumor suppressing genes in human cancers. Currently, when infection is becoming sufficiently controlled worldwide, iron regulation may be the next target for human longevity. PMID:21297911

  5. Draft Genome Sequences of Candida glabrata Isolates 1A, 1B, 2A, 2B, 3A, and 3B

    PubMed Central

    Håvelsrud, Othilde Elise

    2017-01-01

    ABSTRACT Here, we report the draft genome sequences of six Candida glabrata isolates. The isolates were taken from blood samples from patients after recurrent C. glabrata infection. Two isolates were taken from each of three patients a minimum 3 months apart. PMID:28280017

  6. A2B Adenosine Receptor Agonist Improves Erectile Function in Diabetic Rats.

    PubMed

    Wen, Jiaming; Wang, Bohan; Du, Chuanjun; Xu, Gang; Zhang, Zhewei; Li, Yi; Zhang, Nan

    2015-10-01

    Diabetes is an important risk factor for erectile dysfunction (ED). Recent studies have indicated that A2B adenosine receptor (ADORA2B) signaling is essential for penile erection. Thus, we hypothesize that diabetic ED may be attributed to impaired A2B adenosine signaling. To test this hypothesis, we generated diabetic rats by injecting streptozocin as animal model. After 12 weeks, immunohistochemistry staining was used to localize the expression of ADORA2B. Western Blot and quantitative PCR were employed to determine ADORA2B expression level. Intracavernosal pressure (ICP) measurement was used to evaluate erectile function. Diabetic rats received a single intravenous injection of BAY 60-6583, an ADORA2B agonist, or vehicle solution, at 60 min before the ICP measurement. The results showed that ADORA2B expressed in the nerve bundle, smooth muscle, and endothelium in penile tissue of control mice. Western Blot and quantitative PCR results indicated that the expression levels of ADORA2B protein and mRNA were significantly reduced in penile tissues of diabetic rats. Functional studies showed that the erectile response induced by electrical stimulation was remarkably decreased in diabetic rats, compared with age-matched control rats. However, at 60 min after BAY 60-6583 treatment, the erectile function was improved in diabetic rats, suggesting that enhancement of ADORA2B signaling may improve erectile function in diabetic ED. This preclinical study has revealed a previously unrecognized therapeutic possibility of BAY 60-6583 as an effective and mechanism-based drug to treat diabetic ED. In conclusion, we propose that impaired A2B adenosine signaling is one of the pathological mechanisms of diabetic ED.

  7. A2A adenosine receptors are up-regulated in lymphocytes from amyotrophic lateral sclerosis patients.

    PubMed

    Vincenzi, Fabrizio; Corciulo, Carmen; Targa, Martina; Casetta, Ilaria; Gentile, Mauro; Granieri, Enrico; Borea, Pier Andrea; Popoli, Patrizia; Varani, Katia

    2013-09-01

    Adenosine, a purine nucleoside interacting with A1, A2A, A2B and A3 adenosine receptors (ARs), is a potent endogenous modulator of inflammatory and neuronal processes involved in the pathophysiology of several neurodegenerative diseases. In the present study, ARs were investigated in lymphocytes from patients with amyotrophic lateral sclerosis (ALS) and compared with age-matched healthy subjects. In ALS patients A2AARs were analysed by using RT-PCR, Western blotting and saturation binding experiments. The effect of A2AAR stimulation on cyclic AMP levels was evaluated in lymphocytes from ALS patients and healthy subjects. An up-regulation of A2AARs was observed in ALS patients with respect to healthy subjects while A1, A2B and A3AR affinity and density did not change. In ALS patients, the A2AAR density values correlated with the Amyotrophic Lateral Sclerosis Functional Rating Scale-Revised (ALSFRS-R) scores. Furthermore, the stimulation of A2AARs mediated a significant increase in cyclic AMP levels in lymphocytes from ALS patients, with a higher potency than in lymphocytes from healthy subjects. In conclusion, the positive correlation between A2AAR density and ALSFRS-R scores could indicate a possible protective effect of this receptor subtype, representing an interesting starting point for the study of alternative therapeutic approaches for ALS based on A2AAR modulation.

  8. Effects of hnRNP A2/B1 Knockdown on Inhibition of Glioblastoma Cell Invasion, Growth and Survival.

    PubMed

    Deng, Jinmu; Chen, Song; Wang, Feng; Zhao, Hongxin; Xie, Zongyi; Xu, Zhongye; Zhang, Qingtao; Liang, Ping; Zhai, Xuan; Cheng, Yuan

    2016-03-01

    Heterogeneous nuclear ribonucleoprotein A2/B1 (hnRNP A2/B1) plays an important role in influence of pre-messenger RNA (pre-mRNA) processing and mRNA metabolism and transportation in cells. Increasing evidence indicates that hnRNP A2/B1 played an important role in development and progression of various human cancers. Forty cases of normal and human glioma tissue samples were analyzed using immunohistochemistry to reveal the expression of hnRNP A2/B1 protein in the samples. Then, knockdown of hnRNP A2/B1 expression induced by RNA interference (RNAi) method was used to analyze the role of hnRNP A2/B1 in glioblastoma cell viability, adhesion, migration, invasion, and chemoresistance for temozolomide (TMZ). The data showed that hnRNP A2/B1 protein was overexpressed in glioma tissue specimens and associated with advanced glioma grades. Knockdown of hnRNP A2/B1 could reduce glioblastoma cell viability, adhesion, migration, invasion, and chemoresistance for TMZ capacity, but induced tumor cells to apoptosis and reactive oxygen species (ROS) generation in glioma U251 and SHG44 cells. Molecularly, hnRNP A2/B1 knockdown reduced expression of phospho-STAT3 and MMP-2. Detection of hnRNP A2/B1 expression may be useful as a biomarker for prediction of glioma progression and knockdown of hnRNP A2/B1 expression as a novel strategy in future control of glioblastoma in clinic.

  9. Elevated Adenosine Induces Placental DNA Hypomethylation Independent of A2B Receptor Signaling in Preeclampsia.

    PubMed

    Huang, Aji; Wu, Hongyu; Iriyama, Takayuki; Zhang, Yujin; Sun, Kaiqi; Song, Anren; Liu, Hong; Peng, Zhangzhe; Tang, Lili; Lee, Minjung; Huang, Yun; Ni, Xin; Kellems, Rodney E; Xia, Yang

    2017-07-01

    Preeclampsia is a prevalent pregnancy hypertensive disease with both maternal and fetal morbidity and mortality. Emerging evidence indicates that global placental DNA hypomethylation is observed in patients with preeclampsia and is linked to altered gene expression and disease development. However, the molecular basis underlying placental epigenetic changes in preeclampsia remains unclear. Using 2 independent experimental models of preeclampsia, adenosine deaminase-deficient mice and a pathogenic autoantibody-induced mouse model of preeclampsia, we demonstrate that elevated placental adenosine not only induces hallmark features of preeclampsia but also causes placental DNA hypomethylation. The use of genetic approaches to express an adenosine deaminase minigene specifically in placentas, or adenosine deaminase enzyme replacement therapy, restored placental adenosine to normal levels, attenuated preeclampsia features, and abolished placental DNA hypomethylation in adenosine deaminase-deficient mice. Genetic deletion of CD73 (an ectonucleotidase that converts AMP to adenosine) prevented the elevation of placental adenosine in the autoantibody-induced preeclampsia mouse model and ameliorated preeclampsia features and placental DNA hypomethylation. Immunohistochemical studies revealed that elevated placental adenosine-mediated DNA hypomethylation predominantly occurs in spongiotrophoblasts and labyrinthine trophoblasts and that this effect is independent of A2B adenosine receptor activation in both preeclampsia models. Extending our mouse findings to humans, we used cultured human trophoblasts to demonstrate that adenosine functions intracellularly and induces DNA hypomethylation without A2B adenosine receptor activation. Altogether, both mouse and human studies reveal novel mechanisms underlying placental DNA hypomethylation and potential therapeutic approaches for preeclampsia. © 2017 American Heart Association, Inc.

  10. Summary report on the fuel performance modeling of the AFC-2A, 2B irradiation experiments

    SciTech Connect

    Pavel G. Medvedev

    2013-09-01

    The primary objective of this work at the Idaho National Laboratory (INL) is to determine the fuel and cladding temperature history during irradiation of the AFC-2A, 2B transmutation metallic fuel alloy irradiation experiments containing transuranic and rare earth elements. Addition of the rare earth elements intends to simulate potential fission product carry-over from pyro-metallurgical reprocessing. Post irradiation examination of the AFC-2A, 2B rodlets revealed breaches in the rodlets and fuel melting which was attributed to the release of the fission gas into the helium gap between the rodlet cladding and the capsule which houses six individually encapsulated rodlets. This release is not anticipated during nominal operation of the AFC irradiation vehicle that features a double encapsulated design in which sodium bonded metallic fuel is separated from the ATR coolant by the cladding and the capsule walls. The modeling effort is focused on assessing effects of this unanticipated event on the fuel and cladding temperature with an objective to compare calculated results with the temperature limits of the fuel and the cladding.

  11. 180-Nucleotide Duplication in the G Gene of Human metapneumovirus A2b Subgroup Strains Circulating in Yokohama City, Japan, since 2014

    PubMed Central

    Saikusa, Miwako; Kawakami, Chiharu; Nao, Naganori; Takeda, Makoto; Usuku, Shuzo; Sasao, Tadayoshi; Nishimoto, Kimiko; Toyozawa, Takahiro

    2017-01-01

    Human metapneumovirus (HMPV), a member of the family Paramyxoviridae, was first isolated in 2001. Seroepidemiological studies have shown that HMPV has been a major etiological agent of acute respiratory infections in humans for more than 50 years. Molecular epidemiological, genetic, and antigenetic evolutionary studies of HMPV will strengthen our understanding of the epidemic behavior of the virus and provide valuable insight for the control of HMPV and the development of vaccines and antiviral drugs against HMPV infection. In this study, the nucleotide sequence of and genetic variations in the G gene were analyzed in HMPV strains prevalent in Yokohama City, in the Kanto area, Japan, between January 2013 and June 2016. As a part of the National Epidemiological Surveillance of Infectious Diseases, Japan, 1308 clinical specimens (throat swabs, nasal swabs, nasal secretions, and nasal aspirate fluids) collected at 24 hospitals or clinics in Yokohama City were screened for 15 major respiratory viruses with a multiplex reverse transcription–PCR assay. HMPV was detected in 91 specimens, accounting for 7.0% of the total specimens, and the nucleotide sequences of the G genes of 84 HMPV strains were determined. Among these 84 strains, 6, 43, 10, and 25 strains were classified into subgroups A2a, A2b, B1, and B2, respectively. Approximately half the HMPV A2b subgroup strains detected since 2014 had a 180-nucleotide duplication (180nt-dup) in the G gene and clustered on a phylogenic tree with four classical 180nt-dup-lacking HMPV A2b strains prevalent between 2014 and 2015. The 180nt-dup causes a 60-amino-acid duplication (60aa-dup) in the G protein, creating 23–25 additional potential acceptor sites for O-linked sugars. Our data suggest that 180nt-dup occurred between 2011 and 2013 and that HMPV A2b strains with 180nt-dup (A2b180nt-dup HMPV) became major epidemic strains within 3 years. The detailed mechanism by which the A2b180nt-dup HMPV strains gained an advantage

  12. Adenosine A2A receptors in diffuse dermal fibrosis: pathogenic role in human dermal fibroblasts and in a murine model of scleroderma.

    PubMed

    Chan, E S L; Fernandez, P; Merchant, A A; Montesinos, M C; Trzaska, S; Desai, A; Tung, C F; Khoa, D N; Pillinger, M H; Reiss, A B; Tomic-Canic, M; Chen, J F; Schwarzschild, M A; Cronstein, B N

    2006-08-01

    Adenosine regulates inflammation and tissue repair, and adenosine A2A receptors promote wound healing by stimulating collagen matrix production. We therefore examined whether adenosine A2A receptors contribute to the pathogenesis of dermal fibrosis. Collagen production by primary human dermal fibroblasts was analyzed by real-time polymerase chain reaction, 14C-proline incorporation, and Sircol assay. Intracellular signaling for dermal collagen production was investigated using inhibitors of MEK-1 and by demonstration of ERK phosphorylation. In vivo effects were studied in a bleomycin-induced dermal fibrosis model using adenosine A2A receptor-deficient wild-type littermate mice, C57BL/6 mice, and mice treated with adenosine A2A receptor antagonist. Morphometric features and levels of hydroxyproline were determined as measures of dermal fibrosis. Adenosine A2A receptor occupancy promoted collagen production by primary human dermal fibroblasts, which was blocked by adenosine A2A, but not A1 or A2B, receptor antagonism. Adenosine A2A receptor ligation stimulated ERK phosphorylation, and A2A receptor-mediated collagen production by dermal fibroblasts was blocked by MEK-1 inhibitors. Adenosine A2A receptor-deficient and A2A receptor antagonist-treated mice were protected from developing bleomycin-induced dermal fibrosis. These results demonstrate that adenosine A2A receptors play an active role in the pathogenesis of dermal fibrosis and suggest a novel therapeutic target in the treatment and prevention of dermal fibrosis in diseases such as scleroderma.

  13. Adenosine A2B receptor: from cell biology to human diseases

    NASA Astrophysics Data System (ADS)

    Sun, Ying; Huang, Pingbo

    2016-08-01

    Extracellular adenosine is a ubiquitous signaling molecule that modulates a wide array of biological processes. Recently, significant advances have been made in our understanding of A2B adenosine receptor (A2BAR). In this review, we first summarize some of the general characteristics of A2BAR, and then we describe the multiple binding partners of the receptor, such as newly identified α-actinin-1 and p105, and discuss how these associated proteins could modulate A2BAR’s functions, including certain seemingly paradoxical functions of the receptor. Growing evidence indicates a critical role of A2BAR in cancer, renal disease, and diabetes, in addition to its importance in the regulation of vascular diseases and lung disease. Here, we also discuss the role of A2BAR in cancer, renal disease, and diabetes and the potential of the receptor as a target for treating these three diseases.

  14. Adenosine A2B Receptor: From Cell Biology to Human Diseases

    PubMed Central

    Sun, Ying; Huang, Pingbo

    2016-01-01

    Extracellular adenosine is a ubiquitous signaling molecule that modulates a wide array of biological processes. Recently, significant advances have been made in our understanding of A2B adenosine receptor (A2BAR). In this review, we first summarize some of the general characteristics of A2BAR, and then we describe the multiple binding partners of the receptor, such as newly identified α-actinin-1 and p105, and discuss how these associated proteins could modulate A2BAR's functions, including certain seemingly paradoxical functions of the receptor. Growing evidence indicates a critical role of A2BAR in cancer, renal disease, and diabetes, in addition to its importance in the regulation of vascular diseases, and lung disease. Here, we also discuss the role of A2BAR in cancer, renal disease, and diabetes and the potential of the receptor as a target for treating these three diseases. PMID:27606311

  15. Adenosine A2B Receptor Deficiency Promotes Host Defenses against Gram-Negative Bacterial Pneumonia

    PubMed Central

    Barletta, Kathryn E.; Cagnina, R. Elaine; Burdick, Marie D.; Linden, Joel

    2012-01-01

    Rationale: Activation of the adenosine A2B receptor (A2BR) promotes antiinflammatory effects in diverse biological settings, but the role of this receptor in antimicrobial host defense in the lung has not been established. Gram-negative bacillary pneumonia is a common and serious illness associated with high morbidity and mortality, the treatment of which is complicated by increasing rates of antibiotic resistance. Objectives: To test the hypothesis that absence of adenosine A2B receptor signaling promotes host defense against bacterial pneumonia. Methods: We used a model of Klebsiella pneumoniae pneumonia in wild-type mice and mice with targeted deletion of the A2BR. Host responses were compared in vivo and leukocyte responses to the bacteria were examined in vitro. Measurements and Main Results: A2BR–/– mice demonstrated enhanced bacterial clearance from the lung and improved survival after infection with K. pneumoniae compared with wild-type controls, an effect that was mediated by bone marrow–derived cells. Leukocyte recruitment to the lungs and expression of inflammatory cytokines did not differ between A2BR–/– and wild-type mice, but A2BR–/– neutrophils exhibited sixfold greater bactericidal activity and enhanced production of neutrophil extracellular traps compared with wild-type neutrophils when incubated with K. pneumoniae. Consistent with this finding, bronchoalveolar lavage fluid from A2BR–/– mice with Klebsiella pneumonia contained more extracellular DNA compared with wild-type mice with pneumonia. Conclusions: These data suggest that the absence of A2BR signaling enhances antimicrobial activity in gram-negative bacterial pneumonia. PMID:22997203

  16. Heterogeneous Nuclear Ribonucleoprotein A2/B1 as a Target Antigen in Han Chinese for BD Patients.

    PubMed

    Liang, Jinghui; Yang, Weikang; Meng, Xiangyu; Chen, Peng; Du, Hongwu

    2015-01-01

    Behcet's disease (BD) is a recurrent pathema with a typical symptom of inflammation involved in many organs. Previous report indicated that the serum of Korean patients with BD stimulates membrane expression of hnRNP A2/B1 in endothelial cells. In this study, the target 35 kDa recombinant human hnRNP A2/B1 were over-expressed and purified, then sequenced with MALDI-TOF- TOF mass spectrometry. Western blotting and ELISA were applied to detect serum reactivity against hnRNP A2/B1 respectively. The results demonstrate that hnRNP A2/B1 is an autoantigen of BD in Han Chinese population.

  17. Role of A2B adenosine receptor signaling in adenosine-dependent pulmonary inflammation and injury

    PubMed Central

    Sun, Chun-Xiao; Zhong, Hongyan; Mohsenin, Amir; Morschl, Eva; Chunn, Janci L.; Molina, Jose G.; Belardinelli, Luiz; Zeng, Dewan; Blackburn, Michael R.

    2006-01-01

    Adenosine has been implicated in the pathogenesis of chronic lung diseases such as asthma and chronic obstructive pulmonary disease. In vitro studies suggest that activation of the A2B adenosine receptor (A2BAR) results in proinflammatory and profibrotic effects relevant to the progression of lung diseases; however, in vivo data supporting these observations are lacking. Adenosine deaminase–deficient (ADA-deficient) mice develop pulmonary inflammation and injury that are dependent on increased lung adenosine levels. To investigate the role of the A2BAR in vivo, ADA-deficient mice were treated with the selective A2BAR antagonist CVT-6883, and pulmonary inflammation, fibrosis, and airspace integrity were assessed. Untreated and vehicle-treated ADA-deficient mice developed pulmonary inflammation, fibrosis, and enlargement of alveolar airspaces; conversely, CVT-6883–treated ADA-deficient mice showed less pulmonary inflammation, fibrosis, and alveolar airspace enlargement. A2BAR antagonism significantly reduced elevations in proinflammatory cytokines and chemokines as well as mediators of fibrosis and airway destruction. In addition, treatment with CVT-6883 attenuated pulmonary inflammation and fibrosis in wild-type mice subjected to bleomycin-induced lung injury. These findings suggest that A2BAR signaling influences pathways critical for pulmonary inflammation and injury in vivo. Thus in chronic lung diseases associated with increased adenosine, antagonism of A2BAR-mediated responses may prove to be a beneficial therapy. PMID:16841096

  18. The A2B adenosine receptor modulates pulmonary hypertension associated with interstitial lung disease

    PubMed Central

    Karmouty-Quintana, Harry; Zhong, Hongyan; Acero, Luis; Weng, Tingting; Melicoff, Ernestina; West, James D.; Hemnes, Anna; Grenz, Almut; Eltzschig, Holger K.; Blackwell, Timothy S.; Xia, Yang; Johnston, Richard A.; Zeng, Dewan; Belardinelli, Luiz; Blackburn, Michael R.

    2012-01-01

    Development of pulmonary hypertension is a common and deadly complication of interstitial lung disease. Little is known regarding the cellular and molecular mechanisms that lead to pulmonary hypertension in patients with interstitial lung disease, and effective treatment options are lacking. The purpose of this study was to examine the adenosine 2B receptor (A2BR) as a regulator of vascular remodeling and pulmonary hypertension secondary to pulmonary fibrosis. To accomplish this, cellular and molecular changes in vascular remodeling were monitored in mice exposed to bleomycin in conjunction with genetic removal of the A2BR or treatment with the A2BR antagonist GS-6201. Results demonstrated that GS-6201 treatment or genetic removal of the A2BR attenuated vascular remodeling and hypertension in our model. Furthermore, direct A2BR activation on vascular cells promoted interleukin-6 and endothelin-1 release. These studies identify a novel mechanism of disease progression to pulmonary hypertension and support the development of A2BR antagonists for the treatment of pulmonary hypertension secondary to interstitial lung disease.—Karmouty-Quintana, H., Zhong, H., Acero, L., Weng, T., Melicoff, E., West, J. D., Hemnes, A., Grenz, A., Eltzschig, H. K., Blackwell, T. S., Xia, Y., Johnston, R. A., Zeng, D., Belardinelli, L., Blackburn, M. R. The A2B adenosine receptor modulates pulmonary hypertension associated with interstitial lung disease. PMID:22415303

  19. The A2B adenosine receptor protects against inflammation and excessive vascular adhesion

    PubMed Central

    Yang, Dan; Zhang, Ying; Nguyen, Hao G.; Koupenova, Milka; Chauhan, Anil K.; Makitalo, Maria; Jones, Matthew R.; Hilaire, Cynthia St.; Seldin, David C.; Toselli, Paul; Lamperti, Edward; Schreiber, Barbara M.; Gavras, Haralambos; Wagner, Denisa D.; Ravid, Katya

    2006-01-01

    Adenosine has been described as playing a role in the control of inflammation, but it has not been certain which of its receptors mediate this effect. Here, we generated an A2B adenosine receptor–knockout/reporter gene–knock-in (A2BAR-knockout/reporter gene–knock-in) mouse model and showed receptor gene expression in the vasculature and macrophages, the ablation of which causes low-grade inflammation compared with age-, sex-, and strain-matched control mice. Augmentation of proinflammatory cytokines, such as TNF-α, and a consequent downregulation of IκB-α are the underlying mechanisms for an observed upregulation of adhesion molecules in the vasculature of these A2BAR-null mice. Intriguingly, leukocyte adhesion to the vasculature is significantly increased in the A2BAR-knockout mice. Exposure to an endotoxin results in augmented proinflammatory cytokine levels in A2BAR-null mice compared with control mice. Bone marrow transplantations indicated that bone marrow (and to a lesser extent vascular) A2BARs regulate these processes. Hence, we identify the A2BAR as a new critical regulator of inflammation and vascular adhesion primarily via signals from hematopoietic cells to the vasculature, focusing attention on the receptor as a therapeutic target. PMID:16823489

  20. IFN-γ prevents adenosine receptor (A2bR) upregulation to sustain the macrophage activation response

    PubMed Central

    Cohen, Heather B.; Ward, Amanda; Hamidzadeh, Kajal; Ravid, Katya; Mosser, David M.

    2015-01-01

    The priming of macrophages with IFN-γ prior to TLR stimulation results in enhanced and prolonged inflammatory cytokine production. Here, we demonstrate that following TLR stimulation, macrophages up regulate the adenosine 2b receptor (A2bR) to enhance their sensitivity to immunosuppressive extracellular adenosine. This up-regulation of A2bR leads to the induction of a macrophage with an immunoregulatory phenotype and the down regulation of inflammation. IFN-γ priming of macrophages, selectively prevents the induction of the A2bR in macrophages to mitigate sensitivity to adenosine and prevent this regulatory transition. IFN-γ-mediated A2bR blockade leads to a prolonged production of TNFα and IL-12 in response to TLR ligation. The pharmacological inhibition or the genetic deletion of the A2bR results in a hyper-inflammatory response to TLR ligation, similar to IFN-γ treatment of macrophages. Conversely, the overexpression of A2bR on macrophages blunts the IFN-γ effects and promotes the development of immunoregulatory macrophages. Thus, we propose a novel mechanism whereby IFN-γ contributes to host defense, by desensitizing macrophages to the immunoregulatory effects of adenosine. This mechanism overcomes the transient nature of TLR activation, and prolongs the anti-microbial state of the classically activated macrophage. This study may offer promising new targets to improve the clinical outcome of inflammatory diseases in which macrophage activation is dysregulated. PMID:26355158

  1. Expression of Receptors for Tetanus Toxin and Monoclonal Antibody A2B5 by Pancreatic Islet Cells

    NASA Astrophysics Data System (ADS)

    Eisenbarth, G. S.; Shimizu, K.; Bowring, M. A.; Wells, S.

    1982-08-01

    Studies of the reaction of antibody A2B5 and tetanus toxin with pancreatic islet cells, islet cell tumors, and other human amine precursor uptake and decarboxylation (APUD) tumors are described. By indirect immunofluorescence, antibody A2B5 and tetanus toxin were shown to specifically bind to the plasma membrane of human, rat, chicken, and mouse islet cells. The binding of antibody A2B5 to the cell surface of living islet cells has allowed isolation of these cells from a suspension of pancreatic cells by using a fluorescence-activated cell sorter. In studies designed to determine whether tetanus toxin and antibody A2B5 bound to the same surface antigen, A2B5 and tetanus toxin did not compete for binding to normal islet cells, a human islet cell tumor, or a rat islet cell tumor. In addition to binding to islet cell tumors, antibody A2B5 reacts with frozen sections, isolated cells, and cell lines of neural, neural crest, and APUD origin.

  2. Activation of Adenosine A2A Receptors Inhibits Neutrophil Transuroepithelial Migration ▿

    PubMed Central

    Säve, Susanne; Mohlin, Camilla; Vumma, Ravi; Persson, Katarina

    2011-01-01

    Adenosine has been identified as a significant inhibitor of inflammation by acting on adenosine A2A receptors. In this study, we examined the role of adenosine and A2A receptors in the transmigration of human neutrophils across an in vitro model of the transitional bladder urothelium. Human uroepithelial cells (UROtsa) were grown on transwell inserts; uropathogenic Escherichia coli (UPEC) and neutrophils were added to the transwell system; and the number of migrating neutrophils was evaluated. Reverse transcription-PCR (RT-PCR), immunohistochemistry, and flow cytometry were used to investigate the expression of adenosine receptors, the epithelial adhesion molecule ICAM-1, and the neutrophil integrin CD11b. Levels of proinflammatory interleukin-8 (IL-8) and phosphorylated IκBα were measured by enzyme-linked immunosorbent assays (ELISA) and Luminex assays, respectively. The neutrophils expressed all four adenosine receptor subtypes (A1, A2A, A2B, and A3 receptors), but A3 receptors were not expressed by UROtsa cells. UPEC stimulated neutrophil transuroepithelial migration, which was significantly decreased in response to the specific A2A receptor agonist CGS 21680. The inhibitory effect of CGS 21680 on neutrophil migration was reversed by the A2A receptor antagonist SCH 58261. The production of chemotactic IL-8 and the expression of the adhesion molecule ICAM-1 or CD11b were not significantly affected by CGS 21680. However, a significant decrease in the level of phosporylated IκBα was revealed in response to CGS 21680. In conclusion, UPEC infection in vitro evoked neutrophil migration through a multilayered human uroepithelium. The UPEC-evoked neutrophil transmigration decreased in response to A2A receptor activation, possibly through inhibition of NF-κB signaling pathways. PMID:21646447

  3. Shallow oceanic crust: Full waveform tomographic images of the seismic layer 2A/2B boundary

    NASA Astrophysics Data System (ADS)

    Christeson, Gail L.; Morgan, Joanna V.; Warner, Michael R.

    2012-05-01

    We present results of full-waveform tomographic inversions of four profiles acquired over young intermediate- and fast spreading rate oceanic crust. The mean velocity-depth functions from our study include a 0.25-0.30 km-thick low-velocity, low-gradient region beneath the seafloor overlying a 0.24-0.28-km-thick high-gradient region; together these regions compose seismic layer 2A. Mean layer 2A interval velocities are 3.0-3.2 km/s. The mean depth to the layer 2A/2B boundary is 0.49-0.54 km, and mean velocities within the upper 0.25 km of layer 2B are 4.7-4.9 km/s. Previous velocity analyses of the study areas using 1-D ray tracing underestimate the thickness of the high-gradient region at the base of layer 2A. We observe differences in the waveform inversion velocity models that correspond to imaging of the layer 2A event; regions with a layer 2A event have higher velocity gradients at the base of layer 2A. Intermittent high velocities, which we interpret as massive flows, are observed in the waveform inversion velocity models at 0.05-0.10 km below the seafloor (bsf) over 10-25% of the intermediate-spreading profiles and 20-45% of the fast spreading profiles. The high-gradient region located 0.25-0.54 km bsf at the base of layer 2A may be associated with an increased prevalence of massive flows, the first appearance of dikes (lava-dike transition zone), or with increased crack sealing by hydrothermal products. The upper portion of layer 2B, which begins at 0.49-0.54 km bsf, may correspond to sheeted dikes or the top of the transition zone of lavas and dikes.

  4. Adenosine A2B receptor modulates intestinal barrier function under hypoxic and ischemia/reperfusion conditions

    PubMed Central

    Yang, Yang; Qiu, Yuan; Wang, Wensheng; Xiao, Weidong; Liang, Hongyin; Zhang, Chaojun; Yang, Hanwenbo; Teitelbaum, Daniel H; Sun, Li-Hua; Yang, Hua

    2014-01-01

    Background: Intestinal barrier function failure from ischemia/reperfusion (I/R) and acute hypoxia has been implicated as a critical determinant in the predisposition to intestinal inflammation and a number of inflammatory disorders. Here, we identified the role of Adenosine A2B receptor (A2BAR) in the regulation of intestinal barrier function under I/R and acute hypoxic conditions. Methods: C57BL/6J mice were used, and were randomized into three groups: Sham, I/R, IR+PSB1115 (a specific A2BAR antagonist) groups. After surgery, the small bowel was harvested for immunohistochemical staining, RNA and protein content, and intestinal permeability analyses. Using an epithelial cell culture model, we investigated the influence of hypoxia on the epithelial function, and the role of A2BAR in the expressions of tight junction and epithelial permeability. The expressions of Claudin-1, occludin and ZO-1 were detected by RT-PCR and Western-Blot. Epithelial barrier function was assessed with transepithelial resistance (TER). Results and conclusions: The A2BAR antagonist, PSB1115, significantly increased tight junction protein expression after intestinal I/R or acute hypoxia conditions. PSB1115 also attenuated the disrupted distribution of TJ proteins. Furthermore, inhibition of A2BAR attenuated the decrease in TER induced by I/R or acute hypoxic conditions, and maintained intestinal barrier function. Antagonism of A2BAR activity improves intestinal epithelial structure and barrier function in a mouse model of intestinal I/R and a cell model of acute hypoxia. These findings support a potentially destructive role for A2BAR under intestinal I/R and acute hypoxic conditions. PMID:24966910

  5. Comparison of human recombinant adenosine A2B receptor function assessed by Fluo-3-AM fluorometry and microphysiometry.

    PubMed

    Patel, H; Porter, R H P; Palmer, A M; Croucher, M J

    2003-02-01

    1. The aim of this study was to establish the utility of a fluorometric imaging plate reader (FLIPR) assay to assess human adenosine A(2B) receptor function by characterizing its receptor pharmacology and comparing this profile to that obtained using a microphysiometer. 2. FLIPR was used, in conjunction with a Ca(2+)-sensitive dye (Fluo-3-AM), to measure rapid rises in intracellular calcium in a Chinese Hamster Ovary (CHO-K1) cell line stably transfected with both the human A(2B) receptor and a promiscuous G(alpha16) protein. Microphysiometry was used to measure rapid changes in the rate of extracellular acidification in a Human Embryonic Kidney (HEK-293) cell line also stably transfected with human A(2B) receptor. 3. Activation of A(2B) receptors by various ligands caused a concentration-dependent increase in both the intracellular calcium concentration and the extracellular acidification rate in the cells tested, with a similar rank order of potency for agonists: NECA > N(6)-Benzyl NECA > adenosine > or = R-PIA > CPA > S-PIA > CHA > CGS 21680. No comparable effects were observed in the non-transfected control cell lines. 4. The rank order of potency of the agonists examined was the same in all studies, whereas absolute potency and efficacy varied. Thus, all compounds exhibited greater potency in FLIPR than the microphysiometer and the efficacies obtained with CHO-K1 + G(alpha16) + A(2B) cell line and FLIPR were greater than those obtained with HEK-293 + A(2B) cell line in the microphysiometer. 5. ZM-241385 was the most potent of a range of adenosine antagonists tested with a pA(2) of 8.0 in both the FLIPR and microphysiometer assays. 6. In conclusion, the profile of the responses to both A(2B) receptor agonists and antagonists in FLIPR were similar to those obtained by the microphysiometer, although both potency and efficacy values were higher in the FLIPR assay. With this caveat in mind, this study shows that FLIPR coupled with a cell line transfected with both

  6. Inhibitory effects of benzodiazepines on the adenosine A(2B) receptor mediated secretion of interleukin-8 in human mast cells.

    PubMed

    Hoffmann, Kristina; Xifró, Rosa Altarcheh; Hartweg, Julia Lisa; Spitzlei, Petra; Meis, Kirsten; Molderings, Gerhard J; von Kügelgen, Ivar

    2013-01-30

    The activation of adenosine A(2B) receptors in human mast cells causes pro-inflammatory responses such as the secretion of interleukin-8. There is evidence for an inhibitory effect of benzodiazepines on mast cell mediated symptoms in patients with systemic mast cell activation disease. Therefore, we investigated the effects of benzodiazepines on adenosine A(2B) receptor mediated interleukin-8 production in human mast cell leukaemia (HMC1) cells by an enzyme linked immunosorbent assay. The adenosine analogue N-ethylcarboxamidoadenosine (NECA, 0.3-3 μM) increased interleukin-8 production about 5-fold above baseline. This effect was attenuated by the adenosine A(2B) receptor antagonist MRS1754 (N-(4-cyanophenyl)-2-{4-(2,3,6,7-tetrahydro-2,6-dioxo-1,3-dipropyl-1H-purin-8-yl)phenoxy}-acetamide) 1 μM. In addition, diazepam, 4'-chlorodiazepam and flunitrazepam (1-30 μM) markedly reduced NECA-induced interleukin-8 production in that order of potency, whereas clonazepam showed only a modest inhibition. The inhibitory effect of diazepam was not altered by flumazenil 10 μM or PK11195 (1-(2-chlorophenyl)-N-methyl-N-(1-methylpropyl)-3-isoquinolinecarboxamide) 10 μM. Diazepam attenuated the NECA-induced expression of mRNA encoding for interleukin-8. Moreover, diazepam and flunitrazepam reduced the increasing effects of NECA on cAMP-response element- and nuclear factor of activated t-cells-driven luciferase reporter gene activities in HMC1 cells. Neither diazepam nor flunitrazepam affected NECA-induced increases in cellular cAMP levels in CHO Flp-In cells stably expressing recombinant human adenosine A(2B) receptors, excluding a direct action of benzodiazepines on human adenosine A(2B) receptors. In conclusion, this is the first study showing an inhibitory action of benzodiazepines on adenosine A(2B) receptor mediated interleukin-8 production in human mast (HMC1) cells. The rank order of potency indicates the involvement of an atypical benzodiazepine binding site.

  7. Simple multiplex RT-PCR for identifying common fusion BCR-ABL transcript types and evaluation of molecular response of the a2b2 and a2b3 transcripts to Imatinib resistance in north Indian chronic myeloid leukemia patients.

    PubMed

    Mir, Rashid; Ahmad, I; Javid, J; Zuberi, M; Yadav, P; Shazia, R; Masroor, M; Guru, S; Ray, P C; Gupta, N; Saxena, A

    2015-01-01

    Chronic myeloid leukemia (CML) is characterized by the Philadelphia chromosome, an abnormally shortened chromosome 22. It is the result of a reciprocal translocation of chromosomes 9 and 22, creating BCR-ABL fusion transcripts, b3a2, b2a2, and e1a2. The aim of our study was to determine the type of BCR-ABL fusion transcripts for molecular diagnosis and investigate the frequency of BCR-ABL fusion transcripts in CML patients by multiplex RT-PCR in CML. A single reaction with multiple primers multiplex PCR was used to detect and investigate the type and frequency in 200 CML patients among which 116, 33, and 51 were in CP, AP, and BC phase, respectively. The study included 200 CML patients, among whom breakpoints in b3a2, b2a2 transcripts were detected in 68% and 24%, respectively, while 8% of the patients showed both b3a2/b2a2. A statistically significant difference was seen between frequency of BCR-ABL fusion transcripts and gender (P = 0.03), molecular response (P = 0.04), and hematological response (P = 0.05). However, there was no correlation found between frequencies of BCR-/ABL fusion transcripts and other clinicopathological parameters like age, type of therapy, thrombocytopenia, and white blood cell count. Multiplex reverse transcriptase-polymerase chain reaction is useful and saves time in the detection of BCR-ABL variants; the occurrence of these transcripts associated with CML can assist in prognosis and treatment of disease.

  8. Characterization of [125I]ZM 241385 binding to adenosine A2A receptors in the pineal of sheep brain.

    PubMed

    Yan, X; Koos, B J; Kruger, L; Linden, J; Murray, T F

    2006-06-22

    Adenosine is a ubiquitous neuromodulator and homeostatic regulator that exerts its physiologic actions through activation of A(1), A(2A), A(2B) and A(3) adenosine receptor subtypes. In the central nervous system, adenosine's action in neurons is manifested in its modulation of tonic inhibitory control. Adenosine released in the brain during hypoxia has critical depressant effects on breathing in fetal and newborn mammals, an action suggested to be mediated by A(2A) receptors in the posteromedial thalamus. In an effort to more accurately define the spatial distribution of adenosine A(2A) receptors in fetal sheep diencephalon, we have used a receptor autoradiographic technique utilizing an iodinated radioligand [(125)I]ZM 241385, which has greater sensitivity and resolution than the tritiated compound. The distribution of ligand binding sites in the fetal sheep diencephalon indicated that the highest levels of binding were in select thalamic nuclei, including those implicated in hypoxic depression of fetal breathing, and the pineal. Given the high density of labeled A(2A) receptors in the pineal, these sites were characterized more fully in homogenate radioligand binding assays. These data indicate that [(125)I]ZM 241385 binding sites display a pharmacological signature consistent with that of adenosine A(2A) receptors and are expressed at similar levels in fetal, lamb and adult ovine brain. The adenosine A(2A) receptor pharmacologic signature of the [(125)I]ZM 241385 binding site in pineal cell membranes generalized to the site characterized in membranes derived from other portions of the lamb thalamus, including the sector involved in hypoxic inhibition of fetal breathing. These results have important implications for the functional roles of adenosine A(2A) receptors in the thalamus and pineal of sheep brain.

  9. Modulation of murine dendritic cell function by adenine nucleotides and adenosine: involvement of the A(2B) receptor.

    PubMed

    Ben Addi, Abduelhakem; Lefort, Anne; Hua, Xiaoyang; Libert, Frédérick; Communi, Didier; Ledent, Catherine; Macours, Pascale; Tilley, Stephen L; Boeynaems, Jean-Marie; Robaye, Bernard

    2008-06-01

    Adenosine triphosphate has previously been shown to induce semi-mature human monocyte-derived dendritic cells (DC). These are characterized by the up-regulation of co-stimulatory molecules, the inhibition of IL-12 and the up-regulation of some genes involved in immune tolerance, such as thrombospondin-1 and indoleamine 2,3-dioxygenase. The actions of adenosine triphosphate are mediated by the P2Y(11) receptor; since there is no functional P2Y(11) gene in the murine genome, we investigated the action of adenine nucleotides on murine DC. Adenosine 5'-(3-thiotriphosphate) and adenosine inhibited the production of IL-12p70 by bone marrow-derived DC (BMDC). These inhibitions were relieved by 8-p-sulfophenyltheophylline, an adenosine receptor antagonist. The use of selective ligands and A(2B) (-/-) BMDC indicated the involvement of the A(2B) receptor. A microarray experiment, confirmed by quantitative PCR, showed that, in presence of LPS, 5'-(N-ethylcarboxamido) adenosine (NECA, the most potent A(2B) receptor agonist) regulated the expression of several genes: arginase I and II, thrombospondin-1 and vascular endothelial growth factor were up-regulated whereas CCL2 and CCL12 were down-regulated. We further showed that NECA, in combination with LPS, increased the arginase I enzymatic activity. In conclusion, the described actions of adenine nucleotides on BMDC are mediated by their degradation product, adenosine, acting on the A(2B) receptor, and will possibly lead to an impairment of Th1 response or tolerance.

  10. Clinical relevance of Helicobacter pylori babA2 and babA2/B in Costa Rica and Japan

    PubMed Central

    Con, Sergio A; Takeuchi, Hiroaki; Nishioka, Mitsuaki; Morimoto, Norihito; Sugiura, Tetsuro; Yasuda, Nobufumi; Con-Wong, Reinaldo

    2010-01-01

    AIM: To evaluate the prevalence of Helicobacter pylori (H. pylori) babA2, babB and a recombinant gene between babA2 and babB (babA2/B), and their role in the development of atrophic gastritis in Costa Rican and Japanese clinical isolates. METHODS: A total of 95 continuous H. pylori-positive Costa Rican (41 males and 54 females; mean age, 50.65 years; SD, ± 13.04 years) and 95 continuous H. pylori-positive Japanese (50 males and 45 females; mean age, 63.43; SD, ± 13.21 years) patients underwent upper endoscopy from October 2005 to July 2006. They were enrolled for the polymerase chain reaction (PCR)-based genotyping of the H. pylori babA2, babB and babA2/B genes. Statistical analysis was performed using the χ2 test and the Fisher’s exact probability test and multivariate analysis was performed by logistic regression adjusting for gender and age. P < 0.05 was regarded as statistically significant. RESULTS: The PCR-based genotyping of 95 Costa Rican and 95 Japanese isolates showed a higher prevalence of babA2 in Japan (96.8%) than in Costa Rica (73.7%), while that of babA2/B was higher in Costa Rica (11.6%) than in Japan (1.1%). In Costa Rican isolates only, babA2 was significantly associated with atrophic gastritis (P = 0.01). CONCLUSION: These results suggest that the status of babA2 and babA2/B shows geographic differences, and that babA2 has clinical relevance in Costa Rica. PMID:20101774

  11. Oxides of Nitrogen Emissions from the Testing of TF41-A-2B Engines at Naval Air Station, Lemoore, California

    DTIC Science & Technology

    1987-11-01

    for each engine test run. The procedure involves the use of a correlation coefficient which relates the weight (pounds) of NOx emissions to the...individual engines. This report establishes a correlation coefficient for the TF41-A-2B engine based on actual emissions data and the run sheets from 27...engine tests conducted in test cells at NAS Lemoore, CA. The correlation coefficient , equal to 0.01515 pounds of NOx formed per pound of fuel consumed

  12. Discovery and optimization of potent and selective functional antagonists of the human adenosine A2B receptor.

    PubMed

    Bedford, Simon T; Benwell, Karen R; Brooks, Teresa; Chen, Ijen; Comer, Mike; Dugdale, Sarah; Haymes, Tim; Jordan, Allan M; Kennett, Guy A; Knight, Anthony R; Klenke, Burkhard; LeStrat, Loic; Merrett, Angela; Misra, Anil; Lightowler, Sean; Padfield, Anthony; Poullennec, Karine; Reece, Mark; Simmonite, Heather; Wong, Melanie; Yule, Ian A

    2009-10-15

    We herein report the discovery of a novel class of antagonists of the human adenosine A2B receptor. This low molecular weight scaffold has been optimized to offer derivatives with potential utility for the alleviation of conditions associated with this receptor subtype, such as nociception, diabetes, asthma and COPD. Furthermore, preliminary pharmacokinetic analysis has revealed compounds with profiles suitable for either inhaled or systemic routes of administration.

  13. TDP-43 suppresses CGG repeat-induced neurotoxicity through interactions with HnRNP A2/B1

    PubMed Central

    He, Fang; Krans, Amy; Freibaum, Brian D.; Taylor, J. Paul; Todd, Peter K.

    2014-01-01

    Nucleotide repeat expansions can elicit neurodegeneration as RNA by sequestering specific RNA-binding proteins, preventing them from performing their normal functions. Conversely, mutations in RNA-binding proteins can trigger neurodegeneration at least partly by altering RNA metabolism. In Fragile X-associated tremor/ataxia syndrome (FXTAS), a CGG repeat expansion in the 5′UTR of the fragile X gene (FMR1) leads to progressive neurodegeneration in patients and CGG repeats in isolation elicit toxicity in Drosophila and other animal models. Here, we identify the amyotrophic lateral sclerosis (ALS)-associated RNA-binding protein TAR DNA-binding protein (TDP-43) as a suppressor of CGG repeat-induced toxicity in a Drosophila model of FXTAS. The rescue appears specific to TDP-43, as co-expression of another ALS-associated RNA-binding protein, FUS, exacerbates the toxic effects of CGG repeats. Suppression of CGG RNA toxicity was abrogated by disease-associated mutations in TDP-43. TDP-43 does not co-localize with CGG RNA foci and its ability to bind RNA is not required for rescue. TDP-43-dependent rescue does, however, require fly hnRNP A2/B1 homologues Hrb87F and Hrb98DE. Deletions in the C-terminal domain of TDP-43 that preclude interactions with hnRNP A2/B1 abolish TDP-43-dependent rescue of CGG repeat toxicity. In contrast, suppression of CGG repeat toxicity by hnRNP A2/B1 is not affected by RNAi-mediated knockdown of the fly TDP-43 orthologue, TBPH. Lastly, TDP-43 suppresses CGG repeat-triggered mis-splicing of an hnRNP A2/B1-targeted transcript. These data support a model in which TDP-43 suppresses CGG-mediated toxicity through interactions with hnRNP A2/B1 and suggest a convergence of pathogenic cascades between repeat expansion disorders and RNA-binding proteins implicated in neurodegenerative disease. PMID:24920338

  14. Involvement of NADPH oxidase in A2A adenosine receptor-mediated increase in coronary flow in isolated mouse hearts.

    PubMed

    Zhou, Zhichao; Rajamani, Uthra; Labazi, Hicham; Tilley, Stephen L; Ledent, Catherine; Teng, Bunyen; Mustafa, S Jamal

    2015-06-01

    Adenosine increases coronary flow mainly through the activation of A2A and A2B adenosine receptors (ARs). However, the mechanisms for the regulation of coronary flow are not fully understood. We previously demonstrated that adenosine-induced increase in coronary flow is in part through NADPH oxidase (Nox) activation, which is independent of activation of either A1 or A3ARs. In this study, we hypothesize that adenosine-mediated increase in coronary flow through Nox activation depends on A2A but not A2BARs. Functional studies were conducted using isolated Langendorff-perfused mouse hearts. Hydrogen peroxide (H2O2) production was measured in isolated coronary arteries from WT, A2AAR knockout (KO), and A2BAR KO mice using dichlorofluorescein immunofluorescence. Adenosine-induced concentration-dependent increase in coronary flow was attenuated by the specific Nox2 inhibitor gp91 ds-tat or reactive oxygen species (ROS) scavenger EUK134 in both WT and A2B but not A2AAR KO isolated hearts. Similarly, the A2AAR selective agonist CGS-21680-induced increase in coronary flow was significantly blunted by Nox2 inhibition in both WT and A2BAR KO, while the A2BAR selective agonist BAY 60-6583-induced increase in coronary flow was not affected by Nox2 inhibition in WT. In intact isolated coronary arteries, adenosine-induced (10 μM) increase in H2O2 formation in both WT and A2BAR KO mice was attenuated by Nox2 inhibition, whereas adenosine failed to increase H2O2 production in A2AAR KO mice. In conclusion, adenosine-induced increase in coronary flow is partially mediated by Nox2-derived H2O2, which critically depends upon the presence of A2AAR.

  15. Phylogeography of Y-chromosome haplogroup O3a2b2-N6 reveals patrilineal traces of Austronesian populations on the eastern coastal regions of Asia

    PubMed Central

    Teo, Yik-Ying; Huang, Yun-Zhi; Wang, Ling-Xiang; Yu, Ge; Saw, Woei-Yuh; Ong, Rick Twee-Hee; Lu, Yan; Zhang, Chao; Xu, Shu-Hua; Jin, Li; Li, Hui

    2017-01-01

    Austronesian diffusion is considered one of the greatest dispersals in human history; it led to the peopling of an extremely vast region, ranging from Madagascar in the Indian Ocean to Easter Island in Remote Oceania. The Y-chromosome haplogroup O3a2b*-P164(xM134), a predominant paternal lineage of Austronesian populations, is found at high frequencies in Polynesian populations. However, the internal phylogeny of this haplogroup remains poorly investigated. In this study, we analyzed -seventeen Y-chromosome sequences of haplogroup O3a2b*-P164(xM134) and generated a revised phylogenetic tree of this lineage based on 310 non-private Y-chromosome polymorphisms. We discovered that all available O3a2b*-P164(xM134) samples belong to the newly defined haplogroup O3a2b2-N6 and samples from Austronesian populations belong to the sublineage O3a2b2a2-F706. Additionally, we genotyped a series of Y-chromosome polymorphisms in a large collection of samples from China. We confirmed that the sublineage O3a2b2a2b-B451 is unique to Austronesian populations. We found that O3a2b2-N6 samples are widely distributed on the eastern coastal regions of Asia, from Korea to Vietnam. Furthermore, we propose- that the O3a2b2a2b-B451 lineage represents a genetic connection between ancestors of Austronesian populations and ancient populations in North China, where foxtail millet was domesticated about 11,000 years ago. The large number of newly defined Y-chromosome polymorphisms and the revised phylogenetic tree of O3a2b2-N6 will be helpful to explore the origin of proto-Austronesians and the early diffusion process of Austronesian populations. PMID:28380021

  16. Phylogeography of Y-chromosome haplogroup O3a2b2-N6 reveals patrilineal traces of Austronesian populations on the eastern coastal regions of Asia.

    PubMed

    Wei, Lan-Hai; Yan, Shi; Teo, Yik-Ying; Huang, Yun-Zhi; Wang, Ling-Xiang; Yu, Ge; Saw, Woei-Yuh; Ong, Rick Twee-Hee; Lu, Yan; Zhang, Chao; Xu, Shu-Hua; Jin, Li; Li, Hui

    2017-01-01

    Austronesian diffusion is considered one of the greatest dispersals in human history; it led to the peopling of an extremely vast region, ranging from Madagascar in the Indian Ocean to Easter Island in Remote Oceania. The Y-chromosome haplogroup O3a2b*-P164(xM134), a predominant paternal lineage of Austronesian populations, is found at high frequencies in Polynesian populations. However, the internal phylogeny of this haplogroup remains poorly investigated. In this study, we analyzed -seventeen Y-chromosome sequences of haplogroup O3a2b*-P164(xM134) and generated a revised phylogenetic tree of this lineage based on 310 non-private Y-chromosome polymorphisms. We discovered that all available O3a2b*-P164(xM134) samples belong to the newly defined haplogroup O3a2b2-N6 and samples from Austronesian populations belong to the sublineage O3a2b2a2-F706. Additionally, we genotyped a series of Y-chromosome polymorphisms in a large collection of samples from China. We confirmed that the sublineage O3a2b2a2b-B451 is unique to Austronesian populations. We found that O3a2b2-N6 samples are widely distributed on the eastern coastal regions of Asia, from Korea to Vietnam. Furthermore, we propose- that the O3a2b2a2b-B451 lineage represents a genetic connection between ancestors of Austronesian populations and ancient populations in North China, where foxtail millet was domesticated about 11,000 years ago. The large number of newly defined Y-chromosome polymorphisms and the revised phylogenetic tree of O3a2b2-N6 will be helpful to explore the origin of proto-Austronesians and the early diffusion process of Austronesian populations.

  17. Leishmania amazonensis impairs DC function by inhibiting CD40 expression via A2B adenosine receptor activation.

    PubMed

    Figueiredo, Amanda B; Serafim, Tiago D; Marques-da-Silva, Eduardo A; Meyer-Fernandes, José R; Afonso, Luís C C

    2012-05-01

    Dendritic cells (DCs) play an essential role in the modulation of immune responses and several studies have evaluated the interactions between Leishmania parasites and DCs. While extracellular ATP exhibits proinflammatory properties, adenosine is an important anti-inflammatory mediator. Here we investigated the effects of Leishmania infection on DC responses and the participation of purinergic signalling in this process. Bone marrow-derived dendritic cells (BMDCs) from C57BL/6J mice infected with Leishmania amazonensis, Leishmania braziliensis or Leishmania major metacyclic promastigotes showed decreased major histocompatibility complex (MHC) class II and CD86 expression and increased ectonucleotidase expression as compared with uninfected cells. In addition, L. amazonensis-infected DCs, which had lower CD40 expression, exhibited a decreased ability to induce T-cell proliferation. The presence of MRS1754, a highly selective A(2B) adenosine receptor antagonist at the time of infection increased MHC class II, CD86 and CD40 expression in L. amazonensis-infected DCs and restored the ability of the infected DCs to induce T-cell proliferation. Similar results were obtained through the inhibition of extracellular ATP hydrolysis using suramin. In conclusion, we propose that A(2B) receptor activation may be used by L. amazonensis to inhibit DC function and evade the immune response. © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  18. Purification and characterization of Yersinia enterocolitica and Yersinia pestis LcrV-cholera toxin A(2)/B chimeras.

    PubMed

    Tinker, Juliette K; Davis, Chadwick T; Arlian, Britni M

    2010-11-01

    Yersinia pestis is a virulent human pathogen and potential biological weapon. Despite a long history of research on this organism, there is no licensed vaccine to protect against pneumonic forms of Y. pestis disease. In the present study, plasmids were constructed to express cholera toxin A(2)/B chimeric molecules containing the LcrV protective antigen from Yersinia enterocolitica and Y. pestis. These chimeras were expressed and purified to high yields from the supernatant of transformed Escherichia coli. Western and GM(1) ELISA assays were used to characterize the composition, receptor-binding and relative stability of the LcrV-CTA(2)/B chimera in comparison to cholera toxin. In addition, we investigated the ability of the Y. pestis LcrV-CTA(2)/B chimera to bind to and internalize into cultured epithelial cells and macrophages by confocal microscopy. These studies indicate that the uptake and trafficking of the LcrV antigen from the chimera is comparable to the trafficking of native toxin. Together these findings report that stable, receptor-binding, non-toxic LcrV-cholera toxin A(2)/B chimeras can be expressed at high levels in E. coli and purified from the supernatant. In addition, the internalization of antigen in vitro reported here supports the development of these molecules as novel mucosal vaccine candidates. Copyright 2010 Elsevier Inc. All rights reserved.

  19. Purification and characterization of Yersinia enterocolitica and Yersinia pestis LcrV–cholera toxin A2/B chimeras

    PubMed Central

    Davis, Chadwick T.; Arlian, Britni M.

    2010-01-01

    Yersinia pestis is a virulent human pathogen and potential biological weapon. Despite a long history of research on this organism, there is no licensed vaccine to protect against pneumonic forms of Y. pestis disease. In the present study, plasmids were constructed to express cholera toxin A2/B chimeric molecules containing the LcrV protective antigen from Y. enterocolitica and Y. pestis. These chimeras were expressed and purified to high yields from the supernatant of transformed E. coli. Western and GM1 ELISA assays were used to characterize the composition, receptor-binding and relative stability of the LcrV-CTA2/B chimera in comparison to cholera toxin. In addition, we investigated the ability of the Y. pestis LcrV-CTA2/B chimera to bind to and internalize into cultured epithelial cells and macrophages by confocal microscopy. These studies indicate that the uptake and trafficking of the LcrV antigen from the chimera is comparable to the trafficking of native toxin. Together these findings report that stable, receptor-binding, non-toxic LcrV-cholera toxin A2/B chimeras can be expressed at high levels in E. coli and purified from the supernatant. In addition, the internalization of antigen in vitro reported here supports the development of these molecules as novel mucosal vaccine candidates. PMID:20438844

  20. A novel conjugated agent between dopamine and an A2A adenosine receptor antagonist as a potential anti-Parkinson multitarget approach.

    PubMed

    Dalpiaz, Alessandro; Cacciari, Barbara; Vicentini, Chiara Beatrice; Bortolotti, Fabrizio; Spalluto, Giampiero; Federico, Stephanie; Pavan, Barbara; Vincenzi, Fabrizio; Borea, Pier Andrea; Varani, Katia

    2012-03-05

    We propose a potential antiparkinsonian prodrug DP-L-A(2A)ANT (2) obtained by amidic conjugation of dopamine (1) via a succinic spacer to a new triazolo-triazine A(2A) adenosine receptor (AR) antagonist A(2A)ANT (3). The affinity of 2 and its hydrolysis products-1, 3, dopamine-linker DP-L (4) and A(2A)ANT-linker L-A(2A)ANT (5)-was evaluated for hA(1), hA(2A), hA(2B) and hA(3) ARs and rat striatum A(2A)ARs or D(2) receptors. The hydrolysis patterns of 2, 4 and 5 and the stabilities of 1 and 3 were evaluated by HPLC analysis in human whole blood and rat brain homogenates. High hA(2A) affinity was shown by compounds 2 (K(i) = 7.32 ± 0.65 nM), 3 (K(i) = 35 ± 3 nM) and 5 (K(i) = 72 ± 5 nM), whose affinity values were similar in rat striatum. These compounds were not able to change dopamine affinity for D(2) receptors but counteracted the CGS 21680-induced reduction of dopamine affinity. DP-L (4) was inactive on adenosine and dopaminergic receptors. As for stability studies, compounds 4 and 5 were not degraded in incubation media. In human blood, the prodrug 2 was hydrolyzed (half-life = 2.73 ± 0.23 h) mainly on the amidic bound coupling the A(2A)ANT (3), whereas in rat brain homogenates the prodrug 2 was hydrolyzed (half-life > eight hours) exclusively on the amidic bound coupling dopamine, allowing its controlled release and increasing its poor stability as characterized by half-life = 22.5 ± 1.5 min.

  1. Test wells SF-1A, 1B, 1C, and SF-2A, 2B, 2C, Santa Fe County, New Mexico

    USGS Publications Warehouse

    Hart, D.L.

    1989-01-01

    Two well nests, SF-1 and SF-2, were drilled in Santa Fe County, New Mexico, to monitor the hydraulic head within selected zones of the aquifer. Each well nest consists of three piezometers of shallow, middle, and deep completion within the aquifer. Each set of wells was drilled to a depth of about 2,000 ft before actual construction of the piezometers. Each piezometer was completed using either 5 or 10 ft of wire-wrapped screen. These piezometers were constructed as part of a larger ongoing program with the Santa Fe Metropolitan Water Board and New Mexico State Engineer Office to establish a regional observation-well network and to define better the groundwater flow system in the vicinity of Santa Fe and Santa Fe well fields. (USGS)

  2. Increased non-rapid eye movement sleep by cocaine withdrawal: possible involvement of A2A receptors.

    PubMed

    Yang, Shu-Long; Han, Jin-Yi; Kim, Yun-Bae; Nam, Sang-Yoon; Song, Sukgil; Hong, Jin Tae; Oh, Ki-Wan

    2011-02-01

    This study attempted to clarify whether cocaine withdrawal altered sleep architecture and the role of adenosine receptors in this process. Cocaine (20 mg/kg) was administered subcutaneously once per day for 7 days to rat implanted with sleep/wake recording electrode. Polygraphic signs of undisturbed sleep/wake activities were recorded for 24 h before cocaine administration (basal recording as control); withdrawal-day 1 (after 1 day of repeated cocaine administration), withdrawal-day 8 (after 8 days of repeated cocaine administration), and withdrawal-day 14 (after 14 days of repeated cocaine administration), respectively. On cocaine withdrawal-day 1, wakefulness was significantly increased, total sleep was decreased, non-rapid eye movement sleep was markedly reduced, and rapid eye movement sleep was enhanced. Sleep/wake cycles were also increased on cocaine withdrawal day 1. However, non-rapid eye movement sleep was increased on withdrawal-day 8 and 14, whereas rapid eye movement sleep was decreased and no significant changes were observed in the total sleep and sleep/wake cycles during these periods. Adenosine A(2A) receptors expression was increased on withdrawal-day 8 and 14, whereas A(1) receptors levels were reduced after 14 days of withdrawal and the A(2B) receptors remained unchanged. Our findings suggest that alterations of sleep and sleep architecture during cocaine subacute and subchronic withdrawals after repeated cocaine administration may be partially involved in A(2A) receptors over-expression in the rat hypothalamus.

  3. An A2B Adenosine Receptor Agonist Promotes Th17 Autoimmune Responses in Experimental Autoimmune Uveitis (EAU) via Dendritic Cell Activation.

    PubMed

    Chen, Mingjiazi; Liang, Dongchun; Zuo, Aijun; Shao, Hui; Kaplan, Henry J; Sun, Deming

    2015-01-01

    We have recently reported that, although adenosine receptor (AR) agonists have a suppressive effect on Th1 autoreactive T cells, their effect on Th17 autoreactive T cells and γδ T cells is stimulatory and this effect is mainly mediated via A2A adenosine receptors (A2ARs). In this study, we further demonstrate that treatment of C57BL/6 (B6) mice with a selective A2B adenosine receptor (A2BR) agonist greatly enhanced the development of experimental autoimmune uveitis (EAU), whereas treatment with an A2BR antagonist significantly ameliorated severity of EAU. The A2BR agonist-treated mice showed augmented Th17, but not Th1, responses. Mechanistic studies showed that the A2BR agonist-induced enhancement of the Th17 response was significantly lower when TCR-δ-/- mice received the same treatment and that transfer of γδ T cells into TCR-δ-/- mice partially restored this effect. We also showed that dendritic cells (DCs) from A2BR agonist-treated mice showed a significantly increased ability to activate γδ T cells and Th17 autoreactive T cells. Thus, our previous studies have shown that, in EAU, activated γδ T cells possess greatly increased ability to enhance Th17 autoimmune responses. In the present study, we showed that exposure of DCs to A2BR agonist facilitated γδ T cell activation, leading to augmented Th17 responses and progressive EAU development. Our results further support our previous finding that AR agonists have distinct effects on Th1 and Th17 autoimmune responses.

  4. An A2B Adenosine Receptor Agonist Promotes Th17 Autoimmune Responses in Experimental Autoimmune Uveitis (EAU) via Dendritic Cell Activation

    PubMed Central

    Chen, Mingjiazi; Liang, Dongchun; Zuo, Aijun; Shao, Hui; Kaplan, Henry J.; Sun, Deming

    2015-01-01

    We have recently reported that, although adenosine receptor (AR) agonists have a suppressive effect on Th1 autoreactive T cells, their effect on Th17 autoreactive T cells and γδ T cells is stimulatory and this effect is mainly mediated via A2A adenosine receptors (A2ARs). In this study, we further demonstrate that treatment of C57BL/6 (B6) mice with a selective A2B adenosine receptor (A2BR) agonist greatly enhanced the development of experimental autoimmune uveitis (EAU), whereas treatment with an A2BR antagonist significantly ameliorated severity of EAU. The A2BR agonist-treated mice showed augmented Th17, but not Th1, responses. Mechanistic studies showed that the A2BR agonist-induced enhancement of the Th17 response was significantly lower when TCR-δ-/- mice received the same treatment and that transfer of γδ T cells into TCR-δ-/- mice partially restored this effect. We also showed that dendritic cells (DCs) from A2BR agonist-treated mice showed a significantly increased ability to activate γδ T cells and Th17 autoreactive T cells. Thus, our previous studies have shown that, in EAU, activated γδ T cells possess greatly increased ability to enhance Th17 autoimmune responses. In the present study, we showed that exposure of DCs to A2BR agonist facilitated γδ T cell activation, leading to augmented Th17 responses and progressive EAU development. Our results further support our previous finding that AR agonists have distinct effects on Th1 and Th17 autoimmune responses. PMID:26147733

  5. Synthesis, photophysical, electrochemical and electrochemiluminescence properties of A2B2 zinc porphyrins: the effect of π-extended conjugation.

    PubMed

    Galván-Miranda, Elizabeth K; Castro-Cruz, Hiram M; Arturo Arias-Orea, J; Iurlo, Matteo; Valenti, Giovanni; Marcaccio, Massimo; Macías-Ruvalcaba, Norma A

    2016-06-01

    The synthesis of two A2B2 porphyrins, {5,15-bis-[4-(octyloxy)phenyl]-porphyrinato}zinc(ii) () and {5,15-bis-(carbazol-3-yl-ethynyl)-10,20-bis-[4-(octyloxy)phenyl]-porphinato}-zinc(ii) (), is reported. Their photophysical properties were studied by steady-state absorption and emission. Substituting the carbazolylethynyl moieties at two of the meso positions results in a large bathochromic shift of all the absorption bands, a notable increase in the absorption coefficient of the Q(0,0) band, and higher fluorescence quantum yield compared to porphyrin , with two unsubstituted meso positions. Cyclic voltammetry and digital simulation show that electrogenerated radical ions of are more stable than those of . The lack of substituents at the meso positions of leads to dimerization reactions of the radical cation. Despite this, the annihilation reaction of and produces very similar electrogenerated chemiluminescence (ECL) intensity. Spectroelectrochemical experiments demonstrate that the electroreduction of leads to a strong absorption band that might quench the ECL.

  6. a Nonthermal Model for Catalytic Surface Reaction of the Type A2+B2→2AB:

    NASA Astrophysics Data System (ADS)

    Khan, K. M.; Ahmad, W.; Iqbal, K.

    The kinetics of irreversible dimer-dimer reaction of the type A2+B2→2AB has already been studied through Monte Carlo simulation via a model based on Langmuir-Hinshelwood (thermal) mechanism. The results of this study are well known. There is single transition point (yC) at yB=0.5 (where yB is partial pressure of B2 dimer in gas phase), which separates the two poisoned states from each other. Here, we have studied this reaction on the basis of a nonthermal model, which involves the precursor motion of B2 molecule. The most interesting feature of this model is that it yields a steady reactive window. The phase diagram is similar to the ZGB model. The reactive window is separated by continuous and discontinuous irreversible phase transitions. The width of the reactive window depends upon the mobility of the precursors. The dependence of production rate on partial pressure of B2 is shown by simple mathematical equations in our model. Some interesting results are observed when reaction between precursors and chemisorbed B atoms is considered.

  7. Oxides of nitrogen emissions from the testing of Tf41-A-2B engines at Naval Air Station, Lemoore, California

    SciTech Connect

    Not Available

    1987-11-01

    NOx are air pollutants from the testing of gas turbine engines. Out-of-airframe engine testing is regulated by air pollution control agencies which require NOx emissions data on applications for permits to construct and operate engine test facilities. Aside from continuous emissions monitoring, current methods of determining NOx emissions from test cells depend on the availability of accurate records of engine operational data. This degree of record keeping is excessive given the difficult conditions under which engine testing is normally conducted. To avoid excessive record keeping, the Aircraft Environmental Support Office recommends a simple procedure for determining NOx emissions. Its use depends only on accurate records of fuel usage for each engine test run. The procedure involves the use of a correlation coefficient which relates the weight (pounds) of NOx emissions to the weight (pounds) of fuel consumed during engine testing. The coefficient is characteristic of a given engine type, demonstrating little variation among individual engines. This report establishes a correlation coefficient for the TF41-A-2B engine based on actual emissions data and the run sheets from 27 engine tests conducted in test cells at NAS Lemoore, CA. The correlation coefficient, equal to 0.01515 pounds of NOx formed per pound of fuel consumed, determined NOx emissions to within 1% of actual values. Analysis of the statistical validity of the coefficient supports its use as a reliable procedure.

  8. Targeted Segment Transfer from Rye Chromosome 2R to Wheat Chromosomes 2A, 2B, and 7B.

    PubMed

    Ren, Tianheng; Li, Zhi; Yan, Benju; Tan, Feiquan; Tang, Zongxiang; Fu, Shulan; Yang, Manyu; Ren, Zhenglong

    2017-01-01

    Increased chromosome instability was induced by a rye (Secale cereale L.) monosomic 2R chromosome into wheat (Triticum aestivum L.). Centromere breakage and telomere dysfunction result in high rates of chromosome aberrations, including breakages, fissions, fusions, deletions, and translocations. Plants with target traits were sequentially selected to produce a breeding population, from which 3 translocation lines with target traits have been selected. In these lines, wheat chromosomes 2A, 2B, and 7B recombined with segments of the rye chromosome arm 2RL. This was detected by FISH analysis using repeat sequences pSc119.2, pAs1 and genomic DNA of rye together as probes. The translocation chromosomes in these lines were named as 2ASMR, 2BSMR, and 7BSMR. The small segments that were transferred into wheat consisted of pSc119.2 repeats and other chromatin regions that conferred resistance to stripe rust and expressed target traits. These translocation lines were highly resistant to stripe rust, and expressed several typical traits that were associated with chromosome arm 2RL, which are better than those of its wheat parent, disomic addition, and substitution lines that show agronomic characteristics. The integration of molecular methods and conventional techniques to improve wheat breeding schemes are discussed. © 2017 S. Karger AG, Basel.

  9. Epithelial-specific A2B adenosine receptor signaling protects the colonic epithelial barrier during acute colitis

    PubMed Central

    Aherne, CM; Saeedi, B; Collins, CB; Masterson, JC; McNamee, EN; Perrenoud, L; Rapp, CR; Curtis, VF; Bayless, A; Fletcher, A; Glover, LE; Evans, CM; Jedlicka, P; Furuta, GT; de Zoeten, EF; Colgan, SP; Eltzschig, HK

    2015-01-01

    Central to inflammatory bowel disease (IBD) pathogenesis is loss of mucosal barrier function. Emerging evidence implicates extracellular adenosine signaling in attenuating mucosal inflammation. We hypothesized that adenosine-mediated protection from intestinal barrier dysfunction involves tissue-specific signaling through the A2B adenosine receptor (Adora2b) at the intestinal mucosal surface. To address this hypothesis, we combined pharmacologic studies and studies in mice with global or tissue-specific deletion of the Adora2b receptor. Adora2b−/− mice experienced a significantly heightened severity of colitis, associated with a more acute onset of disease and loss of intestinal epithelial barrier function. Comparison of mice with Adora2b deletion on vascular endothelial cells (Adora2bfl/flVeCadCre+) or intestinal epithelia (Adora2bfl/flVillinCre+) revealed a selective role for epithelial Adora2b signaling in attenuating colonic inflammation. In vitro studies with Adora2b knockdown in intestinal epithelial cultures or pharmacologic studies highlighted Adora2b-driven phosphorylation of vasodilator-stimulated phosphoprotein (VASP) as a specific barrier repair response. Similarly, in vivo studies in genetic mouse models or treatment studies with an Adora2b agonist (BAY 60-6583) recapitulate these findings. Taken together, our results suggest that intestinal epithelial Adora2b signaling provides protection during intestinal inflammation via enhancing mucosal barrier responses. PMID:25850656

  10. Excess adenosine in murine penile erectile tissues contributes to priapism via A2B adenosine receptor signaling

    PubMed Central

    Mi, Tiejuan; Abbasi, Shahrzad; Zhang, Hong; Uray, Karen; Chunn, Janci L.; Xia, Ling Wei; Molina, Jose G.; Weisbrodt, Norman W.; Kellems, Rodney E.; Blackburn, Michael R.; Xia, Yang

    2008-01-01

    Priapism, abnormally prolonged penile erection in the absence of sexual excitation, is associated with ischemia-mediated erectile tissue damage and subsequent erectile dysfunction. It is common among males with sickle cell disease (SCD), and SCD transgenic mice are an accepted model of the disorder. Current strategies to manage priapism suffer from a poor fundamental understanding of the molecular mechanisms underlying the disorder. Here we report that mice lacking adenosine deaminase (ADA), an enzyme necessary for the breakdown of adenosine, displayed unexpected priapic activity. ADA enzyme therapy successfully corrected the priapic activity both in vivo and in vitro, suggesting that it was dependent on elevated adenosine levels. Further genetic and pharmacologic evidence demonstrated that A2B adenosine receptor–mediated (A2BR-mediated) cAMP and cGMP induction was required for elevated adenosine–induced prolonged penile erection. Finally, priapic activity in SCD transgenic mice was also caused by elevated adenosine levels and A2BR activation. Thus, we have shown that excessive adenosine accumulation in the penis contributes to priapism through increased A2BR signaling in both Ada–/– and SCD transgenic mice. These findings provide insight regarding the molecular basis of priapism and suggest that strategies to either reduce adenosine or block A2BR activation may prove beneficial in the treatment of this disorder. PMID:18340377

  11. Elevated adenosine signaling via adenosine A2B receptor induces normal and sickle erythrocyte sphingosine kinase 1 activity.

    PubMed

    Sun, Kaiqi; Zhang, Yujin; Bogdanov, Mikhail V; Wu, Hongyu; Song, Anren; Li, Jessica; Dowhan, William; Idowu, Modupe; Juneja, Harinder S; Molina, Jose G; Blackburn, Michael R; Kellems, Rodney E; Xia, Yang

    2015-03-05

    Erythrocyte possesses high sphingosine kinase 1 (SphK1) activity and is the major cell type supplying plasma sphingosine-1-phosphate, a signaling lipid regulating multiple physiological and pathological functions. Recent studies revealed that erythrocyte SphK1 activity is upregulated in sickle cell disease (SCD) and contributes to sickling and disease progression. However, how erythrocyte SphK1 activity is regulated remains unknown. Here we report that adenosine induces SphK1 activity in human and mouse sickle and normal erythrocytes in vitro. Next, using 4 adenosine receptor-deficient mice and pharmacological approaches, we determined that the A2B adenosine receptor (ADORA2B) is essential for adenosine-induced SphK1 activity in human and mouse normal and sickle erythrocytes in vitro. Subsequently, we provide in vivo genetic evidence that adenosine deaminase (ADA) deficiency leads to excess plasma adenosine and elevated erythrocyte SphK1 activity. Lowering adenosine by ADA enzyme therapy or genetic deletion of ADORA2B significantly reduced excess adenosine-induced erythrocyte SphK1 activity in ADA-deficient mice. Finally, we revealed that protein kinase A-mediated extracellular signal-regulated kinase 1/2 activation functioning downstream of ADORA2B underlies adenosine-induced erythrocyte SphK1 activity. Overall, our findings reveal a novel signaling network regulating erythrocyte SphK1 and highlight innovative mechanisms regulating SphK1 activity in normal and SCD.

  12. Purinergic A2b Receptor Activation by Extracellular Cues Affects Positioning of the Centrosome and Nucleus and Causes Reduced Cell Migration.

    PubMed

    Ou, Young; Chan, Gordon; Zuo, Jeremy; Rattner, Jerome B; van der Hoorn, Frans A

    2016-07-15

    The tight, relative positioning of the nucleus and centrosome in mammalian cells is important for the regulation of cell migration. Under pathophysiological conditions, the purinergic A2b receptor can regulate cell motility, but the underlying mechanism remains unknown. Expression of A2b, normally low, is increased in tissues experiencing adverse physiological conditions, including hypoxia and inflammation. ATP is released from such cells. We investigated whether extracellular cues can regulate centrosome-nucleus positioning and cell migration. We discovered that hypoxia as well as extracellular ATP cause a reversible increase in the distance between the centrosome and nucleus and reduced cell motility. We uncovered the underlying pathway: both treatments act through the A2b receptor and specifically activate the Epac1/RapGef3 pathway. We show that cells lacking A2b do not respond in this manner to hypoxia or ATP but transfection of A2b restores this response, that Epac1 is critically involved, and that Rap1B is important for the relative positioning of the centrosome and nucleus. Our results represent, to our knowledge, the first report demonstrating that pathophysiological conditions can impact the distance between the centrosome and nucleus. Furthermore, we identify the A2b receptor as a central player in this process. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  13. Purinergic A2b Receptor Activation by Extracellular Cues Affects Positioning of the Centrosome and Nucleus and Causes Reduced Cell Migration*

    PubMed Central

    Ou, Young; Chan, Gordon; Zuo, Jeremy; Rattner, Jerome B.; van der Hoorn, Frans A.

    2016-01-01

    The tight, relative positioning of the nucleus and centrosome in mammalian cells is important for the regulation of cell migration. Under pathophysiological conditions, the purinergic A2b receptor can regulate cell motility, but the underlying mechanism remains unknown. Expression of A2b, normally low, is increased in tissues experiencing adverse physiological conditions, including hypoxia and inflammation. ATP is released from such cells. We investigated whether extracellular cues can regulate centrosome-nucleus positioning and cell migration. We discovered that hypoxia as well as extracellular ATP cause a reversible increase in the distance between the centrosome and nucleus and reduced cell motility. We uncovered the underlying pathway: both treatments act through the A2b receptor and specifically activate the Epac1/RapGef3 pathway. We show that cells lacking A2b do not respond in this manner to hypoxia or ATP but transfection of A2b restores this response, that Epac1 is critically involved, and that Rap1B is important for the relative positioning of the centrosome and nucleus. Our results represent, to our knowledge, the first report demonstrating that pathophysiological conditions can impact the distance between the centrosome and nucleus. Furthermore, we identify the A2b receptor as a central player in this process. PMID:27226580

  14. Involvement of 5-HT(2A/2B/2C) receptors on memory formation: simple agonism, antagonism, or inverse agonism?

    PubMed

    Meneses, Alfredo

    2002-12-01

    1. The 5-HT2 receptors subdivision into the 5-HT(2A/2B/2C) subtypes along with the advent of the selective antagonists has allowed a more detailed investigation on the role and therapeutic significance of these subtypes in cognitive functions. The present study further analyzed the 5-HT2 receptors role on memory consolidation. 2. The SB-200646 (a selective 5-HT(2B/2C) receptor antagonist) and LY215840 (a nonselective 5-HT(2/7) receptor antagonist) posttraining administration had no effect on an autoshaped memory consolidation. However, both drugs significantly and differentially antagonized the memory impairments induced by 1-(3-chlorophenyl)piperazine (mCPP), 1-naphtyl-piperazine (1-NP), mesulergine, or N-(3-trifluoromethylphenyl) piperazine (TFMPP). 3. In contrast, SB-200646 failed to modify the facilitatory procognitive effect produced by (+/-)-2.5-dimethoxy-4-iodoamphetamine (DOI) or ketanserin, which were sensitive to MDL100907 (a selective 5-HT2A receptor antagonist) and to a LY215840 high dose. 4. Finally, SB-200646 reversed the learning deficit induced by dizocilpine, but not that by scopolamine: while SB-200646 and MDL100907 coadministration reversed memory deficits induced by both drugs. 5. It is suggested that 5-HT(2B/2C) receptors might be involved on memory formation probably mediating a suppressive or constraining action. Whether the drug-induced memory impairments in this study are explained by simple agonism, antagonism, or inverse agonism at 5-HT2 receptors remains unclear at this time. 6. Notably, the 5-HT2 receptor subtypes blockade may provide some benefit to reverse poor memory consolidation conditions associated with decreasedcholinergic, glutamatergic, and/or serotonergic neurotransmission.

  15. High-pressure behavior of A2B2O7 pyrochlore (A=Eu, Dy; B=Ti, Zr)

    NASA Astrophysics Data System (ADS)

    Rittman, Dylan R.; Turner, Katlyn M.; Park, Sulgiye; Fuentes, Antonio F.; Yan, Jinyuan; Ewing, Rodney C.; Mao, Wendy L.

    2017-01-01

    In situ high-pressure X-ray diffraction and Raman spectroscopy were used to determine the influence of composition on the high-pressure behavior of A2B2O7 pyrochlore (A = Eu, Dy; B = Ti, Zr) up to ˜50 GPa. Based on X-ray diffraction results, all compositions transformed to the high-pressure cotunnite structure. The B-site cation species had a larger effect on the transition pressure than the A-site cation species, with the onset of the phase transformation occurring at ˜41 GPa for B = Ti and ˜16 GPa B = Zr. However, the A-site cation affected the kinetics of the phase transformation, with the transformation for compositions with the smaller ionic radii, i.e., A = Dy, proceeding faster than those with a larger ionic radii, i.e., A = Eu. These results were consistent with previous work in which the radius-ratio of the A- and B-site cations determined the energetics of disordering, and compositions with more similarly sized A- and B-site cations had a lower defect formation energy. Raman spectra revealed differences in the degree of short-range order of the different compositions. Due to the large phase fraction of cotunnite at high pressure for B = Zr compositions, Raman modes for cotunnite could be observed, with more modes recorded for A = Eu than A = Dy. These additional modes are attributed to increased short-to-medium range ordering in the initially pyrochlore structured Eu2Zr2O7 as compared with the initially defect-fluorite structured Dy2Zr2O7.

  16. Fluorescence and FTIR Spectra Analysis of Trans-A2B2-Substituted Di- and Tetra-Phenyl Porphyrins

    PubMed Central

    Şen, Pınar; Hirel, Catherine; Andraud, Chantal; Aronica, Christophe; Bretonnière, Yann; Mohammed, Abdelsalam; Ågren, Hans; Minaev, Boris; Minaeva, Valentina; Baryshnikov, Gleb; Lee, Hung-Hsun; Duboisset, Julien; Lindgren, Mikael

    2010-01-01

    A series of asymmetrically substituted free-base di- and tetra-phenylporphyrins and the associated Zn-phenylporphyrins were synthesized and studied by X-ray diffraction, NMR, infrared, electronic absorption spectra, as well as fluorescence emission spectroscopy, along with theoretical simulations of the electronic and vibration structures. The synthesis selectively afforded trans-A2B2 porphyrins, without scrambling observed, where the AA and BB were taken as donor- and acceptor-substituted phenyl groups. The combined results point to similar properties to symmetrically substituted porphyrins reported in the literature. The differences in FTIR and fluorescence were analyzed by means of detailed density functional theory (DFT) calculations. The X-ray diffraction analysis for single crystals of zinc-containing porphyrins revealed small deviations from planarity for the porphyrin core in perfect agreement with the DFT optimized structures. All calculated vibrational modes (2162 modes for all six compounds studied) were found and fully characterized and assigned to the observed FTIR spectra. The most intense IR bands are discussed in connection with the generic similarity and differences of calculated normal modes. Absorption spectra of all compounds in the UV and visible regions show the typical ethio type feature of meso-tetraarylporphyrins with a very intense Soret band and weak Q bands of decreasing intensity. In diphenyl derivatives, the presence of only two phenyl rings causes a pronounced hypsochromic shift of all bands in the absorption spectra. Time-dependent DFT calculations revealed some peculiarities in the electronic excited states structure and connected them with vibronic bands in the absorption and fluorescence spectra from associated vibrational sublevels. PMID:28883336

  17. A2B adenosine receptor blockade enhances macrophage-mediated bacterial phagocytosis and improves polymicrobial sepsis survival in mice.

    PubMed

    Belikoff, Bryan G; Hatfield, Stephen; Georgiev, Peter; Ohta, Akio; Lukashev, Dmitriy; Buras, Jon A; Remick, Daniel G; Sitkovsky, Michail

    2011-02-15

    Antimicrobial treatment strategies must improve to reduce the high mortality rates in septic patients. In noninfectious models of acute inflammation, activation of A2B adenosine receptors (A2BR) in extracellular adenosine-rich microenvironments causes immunosuppression. We examined A2BR in antibacterial responses in the cecal ligation and puncture (CLP) model of sepsis. Antagonism of A2BR significantly increased survival, enhanced bacterial phagocytosis, and decreased IL-6 and MIP-2 (a CXC chemokine) levels after CLP in outbred (ICR/CD-1) mice. During the CLP-induced septic response in A2BR knockout mice, hemodynamic parameters were improved compared with wild-type mice in addition to better survival and decreased plasma IL-6 levels. A2BR deficiency resulted in a dramatic 4-log reduction in peritoneal bacteria. The mechanism of these improvements was due to enhanced macrophage phagocytic activity without augmenting neutrophil phagocytosis of bacteria. Following ex vivo LPS stimulation, septic macrophages from A2BR knockout mice had increased IL-6 and TNF-α secretion compared with wild-type mice. A therapeutic intervention with A2BR blockade was studied by using a plasma biomarker to direct therapy to those mice predicted to die. Pharmacological blockade of A2BR even 32 h after the onset of sepsis increased survival by 65% in those mice predicted to die. Thus, even the late treatment with an A2BR antagonist significantly improved survival of mice (ICR/CD-1) that were otherwise determined to die according to plasma IL-6 levels. Our findings of enhanced bacterial clearance and host survival suggest that antagonism of A2BRs offers a therapeutic target to improve macrophage function in a late treatment protocol that improves sepsis survival.

  18. Primary structures of the heterogeneous nuclear ribonucleoprotein A2, B1, and C2 proteins: a diversity of RNA binding proteins is generated by small peptide inserts.

    PubMed Central

    Burd, C G; Swanson, M S; Görlach, M; Dreyfuss, G

    1989-01-01

    We have isolated cDNAs for the major heterogeneous nuclear ribonucleoprotein (hnRNP) A2, B1, and C2 proteins and determined their nucleotide and deduced amino acid sequences. The A2 and B1 cDNAs are identical except for a 36-nucleotide in-frame insert in B1. Similarly, the sequence of the C2 protein cDNA is related to that of C1 in that C2 contains an extra 39 in-frame nucleotides. Therefore, the B1 amino acid sequence is identical to A2 except for the insertion of 12 amino acids near its amino terminus, and C1 and C2 are also identical to each other except for an extra 13 amino acids near the middle of C2. All three proteins are members of a large family of RNA binding proteins that contain the consensus sequence-type RNA binding domain (CS-RBD). The A2 and B1 proteins have a modular structure similar to that of the hnRNP protein A1: they contain two CS-RBDs and a glycine-rich auxiliary domain at the carboxyl terminus. The CS-RBDs of A2 and B1 have approximately 80% amino acid identity with those of A1, whereas the glycine-rich auxiliary domain is considerably more divergent with less than 30% of the amino acids being identical. These findings indicate that the addition of small peptides, probably by alternative pre-mRNA splicing, generates some of the diversity apparent among hnRNP proteins. Images PMID:2557628

  19. Transcriptional regulation of heat shock proteins and ascorbate peroxidase by CtHsfA2b from African bermudagrass conferring heat tolerance in Arabidopsis

    PubMed Central

    Wang, Xiuyun; Huang, Wanlu; Yang, Zhimin; Liu, Jun; Huang, Bingru

    2016-01-01

    Heat stress transcription factor A2s (HsfA2s) are key regulators in plant response to high temperature. Our objectives were to isolate an HsfA2 gene (CtHsfA2b) from a warm-season grass species, African bermudagrass (Cynodon transvaalensis Burtt-Davy), and to determine the physiological functions and transcriptional regulation of HsfA2 for improving heat tolerance. Gene expression analysis revealed that CtHsfA2b was heat-inducible and exhibited rapid response to increasing temperature. Ectopic expression of CtHsfA2b improved heat tolerance in Arabidopsis and restored heat-sensitive defects of Arabidopsis hsfa2 mutant, which was demonstrated by higher survival rate and photosynthetic parameters, and lower electrolyte leakage in transgenic plants compared to the WT or hsfa2 mutant. CtHsfA2b transgenic plants showed elevated transcriptional regulation of several downstream genes, including those encoding ascorbate peroxidase (AtApx2) and heat shock proteins [AtHsp18.1-CI, AtHsp22.0-ER, AtHsp25.3-P and AtHsp26.5-P(r), AtHsp70b and AtHsp101-3]. CtHsfA2b was found to bind to the heat shock element (HSE) on the promoter of AtApx2 and enhanced transcriptional activity of AtApx2. These results suggested that CtHsfA2b could play positive roles in heat protection by up-regulating antioxidant defense and chaperoning mechanisms. CtHsfA2b has the potential to be used as a candidate gene to genetically modify cool-season species for improving heat tolerance. PMID:27320381

  20. PAR1 inhibition suppresses the self-renewal and growth of A2B5-defined glioma progenitor cells and their derived gliomas in vivo

    PubMed Central

    Auvergne, R; Wu, C; Connell, A; Au, S; Cornwell, A; Osipovitch, M; Benraiss, A; Dangelmajer, S; Guerrero-Cazares, H; Quinones-Hinojosa, A; Goldman, SA

    2016-01-01

    Glioblastoma (GBM) remains the most common and lethal intracranial tumor. In a comparison of gene expression by A2B5-defined tumor-initiating progenitor cells (TPCs) to glial progenitor cells derived from normal adult human brain, we found that the F2R gene encoding PAR1 was differentially overexpressed by A2B5-sorted TPCs isolated from gliomas at all stages of malignant development. In this study, we asked if PAR1 is causally associated with glioma progression. Lentiviral knockdown of PAR1 inhibited the expansion and self-renewal of human GBM-derived A2B5+ TPCs in vitro, while pharmacological inhibition of PAR 1 similarly slowed both the growth and migration of A2B5+ TPCs in culture. In addition, PAR1 silencing potently suppressed tumor expansion in vivo, and significantly prolonged the survival of mice following intracranial transplantation of human TPCs. These data strongly suggest the importance of PAR1 to the self-renewal and tumorigenicity of A2B5-defined glioma TPCs; as such, the abrogation of PAR1-dependent signaling pathways may prove a promising strategy for gliomas. PMID:26616854

  1. The effects of adenosine A2B receptor inhibition on VEGF and nitric oxide axis-mediated renal function in diabetic nephropathy.

    PubMed

    Patel, Leena; Thaker, Aswin

    2014-07-01

    Diabetic nephropathy (DN) is the most common cause of end-stage renal disease worldwide. The pathophysiologic mechanisms of diabetic nephropathy are incompletely understood but include overproduction of various growth factors and cytokines. Upregulation of vascular endothelial growth factor (VEGF) is a pathogenic event occurring in most forms of podocytopathy; however, the mechanisms that regulate this growth factor induction are not clearly identified. A2B receptors have been found to regulate VEGF expression under hypoxic environment in different tissues. One proposed hypothesis in mediating diabetic nephropathy is the modulation of VEGF-NO balance in renal tissue. We determined the role of adenosine A2B receptor in mediating VEGF overproduction and nitrite in diabetic nephropathy. The renal content of A2B receptors and VEGF was increased after 8 weeks of diabetes induction. The renal and plasma nitrite levels were also reduced in these animals. In vivo administration of A2B adenosine receptor antagonist (MRS1754) inhibited the renal over expression of VEGF and adverse renal function parameters. The antagonist administration also improved the kidney tissue nitrite levels. In conclusion, we demonstrated that VEGF induction via adenosine signaling might be the critical event in regulating VEGF-NO axis in diabetic nephropathy.

  2. Whole-Genome Sequence of Clostridium botulinum A2B3 87, a Highly Virulent Strain Involved in a Fatal Case of Foodborne Botulism in Italy

    PubMed Central

    Giordani, Francesco; Fillo, Silvia; Anselmo, Anna; Palozzi, Anna Maria; Fortunato, Antonella; Gentile, Bernardina; Pittiglio, Valentina; Spagnolo, Ferdinando; Anniballi, Fabrizio; Fiore, Alfonsina; Auricchio, Bruna; De Medici, Dario

    2015-01-01

    Here, we report the genome sequence of a rare bivalent strain of Clostridium botulinum, A2B3 87. The strain was isolated from a foodborne botulism case that occurred in Italy in 1995. The case was characterized by rapid evolution of the illness and failure of conventional treatments. PMID:25814616

  3. Novel Alexa Fluor-488 labeled antagonist of the A(2A) adenosine receptor: Application to a fluorescence polarization-based receptor binding assay.

    PubMed

    Kecskés, Miklós; Kumar, T Santhosh; Yoo, Lena; Gao, Zhan-Guo; Jacobson, Kenneth A

    2010-08-15

    Fluorescence polarization (FP) assay has many advantages over the traditional radioreceptor binding studies. We developed an A(2A) adenosine receptor (AR) FP assay using a newly synthesized fluorescent antagonist of the A(2A)AR (MRS5346), a pyrazolo[4,3-e][1,2,4]triazolo[1,5-c]pyrimidin-5-amine derivative conjugated to the fluorescent dye Alexa Fluor-488. MRS5346 displayed a K(i) value of 111+/-16nM in radioligand binding using [(3)H]CGS21680 and membranes prepared from HEK293 cells stably expressing the human A(2A)AR. In a cyclic AMP functional assay, MRS5346 was shown to be an A(2A)AR antagonist. MRS5346 did not show any effect on A(1) and A(3) ARs in binding or the A(2B)AR in a cyclic AMP assay at 10microM. Its suitability as a fluorescent tracer was indicated in an initial observation of an FP signal following A(2A)AR binding. The FP signal was optimal with 20nM MRS5346 and 150microg protein/mL HEK293 membranes. The association and dissociation kinetic parameters were readily determined using this FP assay. The K(d) value of MRS5346 calculated from kinetic parameters was 16.5+/-4.7nM. In FP competition binding experiments using MRS5346 as a tracer, K(i) values of known AR agonists and antagonists consistently agreed with K(i) values from radioligand binding. Thus, this FP assay, which eliminates using radioisotopes, appears to be appropriate for both routine receptor binding and high-throughput screening with respect to speed of analysis, displaceable signal and precision. The approach used in the present study could be generally applicable to other GPCRs.

  4. Role of adenosine A2A receptor signaling in the nicotine-evoked attenuation of reflex cardiac sympathetic control.

    PubMed

    El-Mas, Mahmoud M; El-Gowilly, Sahar M; Fouda, Mohamed A; Saad, Evan I

    2011-08-01

    Baroreflex dysfunction contributes to increased cardiovascular risk in cigarette smokers. Given the importance of adenosinergic pathways in baroreflex control, the hypothesis was tested that defective central adenosinergic modulation of cardiac autonomic activity mediates the nicotine-baroreflex interaction. Baroreflex curves relating changes in heart rate (HR) to increases or decreases in blood pressure (BP) evoked by i.v. doses (1-16μg/kg) of phenylephrine (PE) and sodium nitroprusside (SNP), respectively, were constructed in conscious rats; slopes of the curves were taken as measures of baroreflex sensitivity (BRS). Nicotine (25 and 100μg/kg i.v.) dose-dependently reduced BRS(SNP) in contrast to no effect on BRS(PE). BRS(SNP) was also attenuated after intracisternal (i.c.) administration of nicotine. Similar reductions in BRS(SNP) were observed in rats pretreated with atropine or propranolol. The combined treatment with nicotine and atropine produced additive inhibitory effects on BRS, an effect that was not demonstrated upon concurrent exposure to nicotine and propranolol. BRS(SNP) was reduced in preparations treated with i.c. 8-phenyltheophylline (8-PT, nonselective adenosine receptor antagonist), 8-(3-Chlorostyryl) caffeine (CSC, A(2A) antagonist), or VUF5574 (A(3) antagonist). In contrast, BRS(SNP) was preserved after blockade of A(1) (DPCPX) or A(2B) (alloxazine) receptors or inhibition of adenosine uptake by dipyridamole. CSC or 8-PT abrogated the BRS(SNP) depressant effect of nicotine whereas other adenosinergic antagonists were without effect. Together, nicotine preferentially impairs reflex tachycardia via disruption of adenosine A(2A) receptor-mediated facilitation of reflex cardiac sympathoexcitation. Clinically, the attenuation by nicotine of compensatory sympathoexcitation may be detrimental in conditions such as hypothalamic defense response, posture changes, and ventricular rhythms.

  5. The cellular protein hnRNP A2/B1 enhances HIV-1 transcription by unfolding LTR promoter G-quadruplexes

    PubMed Central

    Scalabrin, Matteo; Frasson, Ilaria; Ruggiero, Emanuela; Perrone, Rosalba; Tosoni, Elena; Lago, Sara; Tassinari, Martina; Palù, Giorgio; Richter, Sara N.

    2017-01-01

    G-quadruplexes are four-stranded conformations of nucleic acids that act as cellular epigenetic regulators. A dynamic G-quadruplex forming region in the HIV-1 LTR promoter represses HIV-1 transcription when in the folded conformation. This activity is enhanced by nucleolin, which induces and stabilizes the HIV-1 LTR G-quadruplexes. In this work by a combined pull-down/mass spectrometry approach, we consistently found hnRNP A2/B1 as an additional LTR-G-quadruplex interacting protein. Surface plasmon resonance confirmed G-quadruplex specificity over linear sequences and fluorescence resonance energy transfer analysis indicated that hnRNP A2/B1 is able to efficiently unfold the LTR G-quadruplexes. Evaluation of the thermal stability of the LTR G-quadruplexes in different-length oligonucleotides showed that the protein is fit to be most active in the LTR full-length environment. When hnRNP A2/B1 was silenced in cells, LTR activity decreased, indicating that the protein acts as a HIV-1 transcription activator. Our data highlight a tightly regulated control of transcription based on G-quadruplex folding/unfolding, which depends on interacting cellular proteins. These findings provide a deeper understanding of the viral transcription mechanism and may pave the way to the development of drugs effective against the integrated HIV-1, present both in actively and latently infected cells. PMID:28338097

  6. hnRNP A2/B1 interacts with influenza A viral protein NS1 and inhibits virus replication potentially through suppressing NS1 RNA/protein levels and NS1 mRNA nuclear export

    SciTech Connect

    Wang, Yimeng; Zhou, Jianhong; Du, Yuchun

    2014-01-20

    The NS1 protein of influenza viruses is a major virulence factor and exerts its function through interacting with viral/cellular RNAs and proteins. In this study, we identified heterogeneous nuclear ribonucleoprotein A2/B1 (hnRNP A2/B1) as an interacting partner of NS1 proteins by a proteomic method. Knockdown of hnRNP A2/B1 by small interfering RNA (siRNA) resulted in higher levels of NS vRNA, NS1 mRNA, and NS1 protein in the virus-infected cells. In addition, we demonstrated that hnRNP A2/B1 proteins are associated with NS1 and NS2 mRNAs and that knockdown of hnRNP A2/B1 promotes transport of NS1 mRNA from the nucleus to the cytoplasm in the infected cells. Lastly, we showed that knockdown of hnRNP A2/B1 leads to enhanced virus replication. Our results suggest that hnRNP A2/B1 plays an inhibitory role in the replication of influenza A virus in host cells potentially through suppressing NS1 RNA/protein levels and NS1 mRNA nucleocytoplasmic translocation. - Highlights: • Cellular protein hnRNP A2/B1 interacts with influenza viral protein NS1. • hnRNP A2/B1 suppresses the levels of NS1 protein, vRNA and mRNA in infected cells. • hnRNP A2/B1 protein is associated with NS1 and NS2 mRNAs. • hnRNP A2/B1 inhibits the nuclear export of NS1 mRNAs. • hnRNP A2/B1 inhibits influenza virus replication.

  7. Leishmania amazonensis-Induced cAMP Triggered by Adenosine A2B Receptor Is Important to Inhibit Dendritic Cell Activation and Evade Immune Response in Infected Mice.

    PubMed

    Figueiredo, Amanda Braga; Souza-Testasicca, Míriam Conceição; Mineo, Tiago Wilson Patriarca; Afonso, Luís Carlos Crocco

    2017-01-01

    Differently from others Leishmania species, infection by the protozoan parasite L. amazonensis is associated with a lack of antigen-specific T-cell responses. Dendritic cells (DC) are essential for the innate immune response and for directing the differentiation of T-helper lymphocytes. Previously, we showed that L. amazonensis infection impairs DC activation through the activation of adenosine A2B receptor, and here, we evaluated the intracellular events triggered by this receptor in infected cells. To this aim, bone marrow-derived DC from C57BL/6J mice were infected with metacyclic promastigotes of L. amazonensis. Our results show, for the first time, that L. amazonensis increases the production of cAMP and the phosphorylation of extracellular signal-regulated protein kinases 1/2 (ERK1/2) in infected DC by a mechanism dependent on the A2B receptor. Furthermore, L. amazonensis impairs CD40 expression and IL-12 production by DC, and the inhibition of adenylate cyclase, phosphoinositide 3-kinase (PI3K), and ERK1/2 prevent these effects. The increase of ERK1/2 phosphorylation and the inhibition of DC activation by L. amazonensis are independent of protein kinase A (PKA). In addition, C57BL/6J mice were inoculated in the ears with metacyclic promastigotes, in the presence of PSB1115, an A2B receptor antagonist. PSB1115 treatment increases the percentage of CD40(+) DC on ears and draining lymph nodes. Furthermore, this treatment reduces lesion size and tissue parasitism. Lymph node cells from treated mice produce higher levels of IFN-γ than control mice, without altering the production of IL-10. In conclusion, we suggest a new pathway used by the parasite (A2B receptor → cAMP → PI3K → ERK1/2) to suppress DC activation, which may contribute to the decrease of IFN-γ production following by the deficiency in immune response characteristic of L. amazonensis infection.

  8. MicroRNA-128b suppresses tumor growth and promotes apoptosis by targeting A2bR in gastric cancer

    SciTech Connect

    Wang, Ping; Guo, Xueyan; Zong, Wei; Song, Bin; Liu, Guisheng; He, Shuixiang

    2015-11-27

    MicroRNAs (miRNAs) play crucial roles in the development and progression of human cancers, including gastric cancer (GC). The discovery of miRNAs may provide a new and powerful tool for studying the mechanism, diagnosis, and treatment of GC. In this study, we aimed to investigate the role and mechanism of miR-128b in the development and progression of GC. Quantitative real-time PCR (qRT-PCR) was used to measure the expression level of miR-128b in GC tissues and cell lines. We found that miR-128b was significantly down-regulated in GC tissues and cell lines. In addition, over-expression of miR-128b inhibited GC cell proliferation, migration and invasion of GC cells in vitro. Gain-of-function in vitro experiments further showed that the miR-128b mimic significantly promoted GC cell apoptosis. Subsequent dual-luciferase reporter assay identified one of the proto-oncogene A2bR as direct target of miR-128b. Therefore, our results indicate that miR-128b is a proto-oncogene miRNA that can suppresses GC proliferation and migration through down-regulation of the oncogene gene A2bR. Taken together, our results indicate that miR-128b could serve as a potential diagnostic biomarker and therapeutic option for human GC in the near future. - Highlights: • The expression of MiR-128b is significantly down-regulated in GC tissues and cell lines. • Ectopic expression of miR-128b directly affects cell proliferation, migration and invasion in vitro. • Overexpression of miR-128b increases apoptosis in GC cells. • A2bR is a candidate target gene of miR-128b. • MiR-128b represses cell proliferation, migration and invasion and promotes apoptosis by targeting A2bR in GC.

  9. Excess adenosine A2B receptor signaling contributes to priapism through HIF-1α mediated reduction of PDE5 gene expression

    PubMed Central

    Ning, Chen; Wen, Jiaming; Zhang, Yujin; Dai, Yingbo; Wang, Wei; Zhang, Weiru; Qi, Lin; Grenz, Almut; Eltzschig, Holger K.; Blackburn, Michael R.; Kellems, Rodney E.; Xia, Yang

    2014-01-01

    Priapism is featured with prolonged and painful penile erection and is prevalent among males with sickle cell disease (SCD). The disorder is a dangerous urological and hematological emergency since it is associated with ischemic tissue damage and erectile disability. Here we report that phosphodiesterase-5 (PDE5) gene expression and PDE activity is significantly reduced in penile tissues of two independent priapic models: SCD mice and adenosine deaminase (ADA)-deficient mice. Moreover, using ADA enzyme therapy to reduce adenosine or a specific antagonist to block A2B adenosine receptor (ADORA2B) signaling, we successfully attenuated priapism in both ADA−/− and SCD mice by restoring penile PDE5 gene expression to normal levels. This finding led us to further discover that excess adenosine signaling via ADORA2B activation directly reduces PDE5 gene expression in a hypoxia-inducible factor-1α (HIF-1α)-dependent manner. Overall, we reveal that excess adenosine-mediated ADORA2B signaling underlies reduced penile PDE activity by decreasing PDE5 gene expression in a HIF-1α-dependent manner and provide new insight for the pathogenesis of priapism and novel therapies for the disease.—Ning, C., Wen, J., Zhang, Y., Dai, Y., Wang, W., Zhang, W., Qi, L., Grenz, A., Eltzschig, H. K., Blackburn, M. R., Kellems, R. E., Xia, Y. Excess adenosine A2B receptor signaling contributes to priapism through HIF-1α mediated reduction of PDE5 gene expression. PMID:24614760

  10. Identification of hnRNPs K, L and A2/B1 as candidate proteins involved in the nutritional regulation of mRNA splicing.

    PubMed

    Griffith, Brian N; Walsh, Callee M; Szeszel-Fedorowicz, Wioletta; Timperman, Aaron T; Salati, Lisa M

    2006-01-01

    Nutrient regulation of glucose-6-phosphate dehydrogenase (G6PD) expression occurs through changes in the rate of splicing of G6PD pre-mRNA. This posttranscriptional mechanism accounts for the 12- to 15-fold increase in G6PD expression in livers of mice that were starved and then refed a high-carbohydrate diet. Regulation of G6PD pre-mRNA splicing requires a cis-acting element in exon 12 of the pre-mRNA. Using RNA probes to exon 12 and nuclear extracts from livers of mice that were starved or refed, proteins of 60 kDa and 37 kDa were detected bound to nucleotides 65-79 of exon 12 and this binding was decreased by 50% with nuclear extracts from refed mice. The proteins were identified as hnRNPs K, L, and A2/B1 by LC-MS/MS. The decrease in binding of these proteins to exon 12 during refeeding was not accompanied by a decrease in the total amount of these proteins in total nuclear extract. HnRNPs K, L and A2/B1 have known roles in the regulation of mRNA splicing. The decrease in binding of these proteins during treatments that increase G6PD expression is consistent with a role for these proteins in the inhibition of G6PD mRNA splicing.

  11. MicroRNA signatures predict dysregulated vitamin D receptor and calcium pathways status in limb girdle muscle dystrophies (LGMD) 2A/2B.

    PubMed

    Aguennouz, M; Lo Giudice, C; Licata, N; Rodolico, C; Musumeci, O; Fanin, M; Migliorato, A; Ragusa, M; Macaione, V; Di Giorgio, R M; Angelini, C; Toscano, A

    2016-08-01

    miRNA expression profile and predicted pathways involved in selected limb-girdle muscular dystrophy (LGMD)2A/2B patients were investigated. A total of 187 miRNAs were dysregulated in all patients, with six miRNAs showing opposite regulation in LGMD2A versus LGMD2B patients. Silico analysis evidence: (1) a cluster of the dysregulated miRNAs resulted primarily involved in inflammation and calcium metabolism, and (2) two genes predicted as controlled by calcium-assigned miRNAs (Vitamin D Receptor gene and Guanine Nucleotide Binding protein beta polypeptide 1gene) showed an evident upregulation in LGMD2B patients, in accordance with miRNA levels. Our data support alterations in calcium pathway status in LGMD 2A/B, suggesting myofibre calcium imbalance as a potential therapeutic target. Copyright © 2016 John Wiley & Sons, Ltd.

  12. Synthesis and characterization of monoisomeric 1,8,15,22-substituted (A3B and A2B2) phthalocyanines and phthalocyanine-fullerene dyads.

    PubMed

    Ranta, Jenni; Kumpulainen, Tatu; Lemmetyinen, Helge; Efimov, Alexander

    2010-08-06

    Synthesis and characterization of three phthalocyanine-fullerene (Pc-C(60)) dyads, corresponding monoisomeric phthalocyanines (Pc), and building blocks, phthalonitriles, are described. Six novel bisaryl phthalonitriles were prepared by the Suzuki-Miyaura coupling reaction from trifluoromethanesulfonic acid 2,3-dicyanophenyl ester and various oxaborolanes. Two phthalonitriles were selected for the synthesis of A(3)B- and A(2)B(2)-type phthalocyanines. Phthalonitrile 4 has a bulky 3,5-di-tert-butylphenyl substituent at the alpha-phthalo position, which forces only one regioisomer to form and greatly increases the solubility of phthalocyanine. Phthalonitrile 8 has a 3-phenylpropanol side chain at the alpha-position making further modifications of the side group possible. Synthesized monoisomeric A(3)B- and A(2)B(2)-type phthalocyanines are modified by attachment of malonic residues. Finally, fullerene is covalently linked to phthalocyanine with one or two malonic bridges to produce Pc-C(60) dyads. Due to the monoisomeric structure and increased solubility of phthalocyanines, the quality of NMR spectra of the compounds is enhanced significantly, making detailed NMR analysis of the structures possible. The synthesized dyads have different orientations of phthalocyanine and fullerene, which strongly influence the electron transfer (ET) from phthalocyanine to fullerene moiety. Fluorescence quenchings of the dyads were measured in both polar and nonpolar solvents, and in all cases, the quenching was more efficient in the polar environment. As expected, most efficient fluorescence quenching was observed for dyad 20b, with two linkers and phthalocyanine and fullerene in face-to-face orientation.

  13. hnRNP A2/B1 interacts with influenza A viral protein NS1 and inhibits virus replication potentially through suppressing NS1 RNA/protein levels and NS1 mRNA nuclear export.

    PubMed

    Wang, Yimeng; Zhou, Jianhong; Du, Yuchun

    2014-01-20

    The NS1 protein of influenza viruses is a major virulence factor and exerts its function through interacting with viral/cellular RNAs and proteins. In this study, we identified heterogeneous nuclear ribonucleoprotein A2/B1 (hnRNP A2/B1) as an interacting partner of NS1 proteins by a proteomic method. Knockdown of hnRNP A2/B1 by small interfering RNA (siRNA) resulted in higher levels of NS vRNA, NS1 mRNA, and NS1 protein in the virus-infected cells. In addition, we demonstrated that hnRNP A2/B1 proteins are associated with NS1 and NS2 mRNAs and that knockdown of hnRNP A2/B1 promotes transport of NS1 mRNA from the nucleus to the cytoplasm in the infected cells. Lastly, we showed that knockdown of hnRNP A2/B1 leads to enhanced virus replication. Our results suggest that hnRNP A2/B1 plays an inhibitory role in the replication of influenza A virus in host cells potentially through suppressing NS1 RNA/protein levels and NS1 mRNA nucleocytoplasmic translocation.

  14. A2B adenosine receptor contributes to penile erection via PI3K/AKT signaling cascade-mediated eNOS activation

    PubMed Central

    Wen, Jiaming; Grenz, Almut; Zhang, Yujin; Dai, Yingbo; Kellems, Rodney E.; Blackburn, Michael R.; Eltzschig, Holger K.; Xia, Yang

    2011-01-01

    Normal penile erection is under the control of multiple factors and signaling pathways. Although adenosine signaling is implicated in normal and abnormal penile erection, the exact role and the underlying mechanism for adenosine signaling in penile physiology remain elusive. Here we report that shear stress leads to increased adenosine release from endothelial cells. Subsequently, we determined that ecto-5′-nucleotidase (CD73) is a key enzyme required for the production of elevated adenosine from ATP released by shear-stressed endothelial cells. Mechanistically, we demonstrate that shear stress-mediated elevated adenosine functions through the adenosine A2B receptor (A2BR) to activate the PI3K/AKT signaling cascade and subsequent increased endothelial nitric oxide synthase (eNOS) phosphorylation. These in vitro studies led us to discover further that adenosine was induced during sustained penile erection and contributes to PI3K/AKT activation and subsequent eNOS phosphorylation via A2BR signaling in intact animal. Finally, we demonstrate that lowering adenosine in wild-type mice or genetic deletion of A2BR in mutant mice significantly attenuated PI3K/AKT activation, eNOS phosphorylation, and subsequent impaired penile erection featured with the reduction of ratio of maximal intracavernosal pressure to systemic arterial pressure from 0.49 ± 0.03 to 0.41 ± 0.05 and 0.38 ± 0.04, respectively (both P<0.05). Overall, using biochemical, cellular, genetic, and physiological approaches, our findings reveal that adenosine is a novel molecule signaling via A2BR activation, contributing to penile erection via PI3K/AKT-dependent eNOS activation. These studies suggest that this signaling pathway may be a novel therapeutic target for erectile disorders.—Wen, J., Grenz, A., Zhang, Y., Dai, Y., Kellems, R. E., Blackburn, M. R., Eltzschig, H. K., Xia, Y. A2B adenosine receptor contributes to penile erection via PI3K/AKT signaling cascade-mediated eNOS activation. PMID

  15. Activation of Th1 and Tc1 cell adenosine A2A receptors directly inhibits IL-2 secretion in vitro and IL-2-driven expansion in vivo.

    PubMed

    Erdmann, Andreas A; Gao, Zhan-Guo; Jung, Unsu; Foley, Jason; Borenstein, Todd; Jacobson, Kenneth A; Fowler, Daniel H

    2005-06-15

    To evaluate the direct effect of adenosine on cytokine-polarized effector T cells, murine type 1 helper T cells (Th1) and type 1 cytotoxic T lymphocytes (Tc1) and Th2/Tc2 cells were generated using an antigen-presenting cell (APC)-free method. Tc1 and Tc2 cells had similar adenosine signaling, as measured by intracellular cyclic AMP (cAMP) increase upon adenosine A(2A) receptor agonism by CGS21680 (CGS). CGS greatly reduced Tc1 and Tc2 cell interleukin 2 (IL-2) and tumor necrosis factor alpha (TNF-alpha) secretion, with nominal effect on interferon gamma (IFN-gamma) secretion. Tc2 cell IL-4 and IL-5 secretion was not reduced by CGS, and IL-10 secretion was moderately reduced. Agonist-mediated inhibition of IL-2 and TNF-alpha secretion occurred via A(2A) receptors, with no involvement of A(1), A(2B), or A(3) receptors. Adenosine agonist concentrations that abrogated cytokine secretion did not inhibit Tc1 or Tc2 cell cytolytic function. Adenosine modulated effector T cells in vivo, as CGS administration reduced CD4(+)Th1 and CD8(+)Tc1 cell expansion to alloantigen and, in a separate model, reduced antigen-specific CD4(+) Th1 cell numbers. Remarkably, agonist-mediated T-cell inhibition was abrogated by in vivo IL-2 therapy. Adenosine receptor activation therefore preferentially inhibits type I cytokine secretion, most notably IL-2. Modulation of adenosine receptors may thus represent a suitable target primarily for inflammatory conditions mediated by Th1 and Tc1 cells.

  16. Bulk and mechanical properties of the Paintbrush tuff recovered from boreholes UE25 NRG-2, 2A, 2B, and 3: Data report

    SciTech Connect

    Boyd, P.J.; Martin, R.J.; Noel, J.S.; Price, R.H.

    1996-09-01

    An integral part of the licensing procedure for the potential nuclear waste repository at Yucca Mountain, Nevada, involves characterization of the in situ rheology for the design and construction of the facility and the emplacement of canisters containing radioactive waste. The data used to model the thermal and mechanical behavior of the repository and surrounding lithologies include dry and saturated bulk densities, average grain density, porosity, compressional and shear wave velocities, elastic moduli, and compressional and tensional fracture strengths. In this study, a suite of experiments was performed on cores recovered from boreholes UE25 NRG-2, 2A, 2B, and 3 drilled in support of the Exploratory Studies Facility (ESF) at Yucca Mountain. The holes penetrated the Timber Mountain tuff and two thermal/mechanical units of the Paintbrush tuff. The thermal/mechanical stratigraphy was defined by Ortiz to group rock horizons of similar properties for the purpose of simplifying modeling efforts. The relationship between the geologic stratigraphy and the thermal/mechanical stratigraphy for each borehole is presented. The tuff samples in this study have a wide range of welding characteristics (usually reflected in sample porosity), and a smaller range of mineralogy and petrology characteristics. Generally, the samples are silicic, ash-fall tuffs that exhibit large variability in their elastic and strength properties.

  17. Molecular motor KIF17 is fundamental for memory and learning via differential support of synaptic NR2A/2B levels.

    PubMed

    Yin, Xiling; Takei, Yosuke; Kido, Mizuho A; Hirokawa, Nobutaka

    2011-04-28

    Kinesin superfamily motor protein 17 (KIF17) is a candidate transporter of N-methyl-D-aspartate (NMDA) receptor subunit 2B (NR2B). Disruption of the murine kif17 gene inhibits NR2B transport, accompanied by decreased transcription of nr2b, resulting in a loss of synaptic NR2B. In kif17(-/-) hippocampal neurons, the NR2A level is also decreased because of accelerated ubiquitin-proteasome system-dependent degradation. Accordingly, NMDA receptor-mediated synaptic currents, early and late long-term potentiation, long-term depression, and CREB responses are attenuated in kif17(-/-) neurons, concomitant with a hippocampus-dependent memory impairment in knockout mice. In wild-type neurons, CREB is activated by synaptic inputs, which increase the levels of KIF17 and NR2B. Thus, KIF17 differentially maintains the levels of NR2A and NR2B, and, when synapses are stimulated, the NR2B/KIF17 complex is upregulated on demand through CREB activity. These KIF17-based mechanisms for maintaining NR2A/2B levels could underlie multiple phases of memory processes in vivo.

  18. DFT+ U study of electronic structure and Curie temperature of A2 B ReO6 (A=Sr, Ca and B=Cr, Fe)

    NASA Astrophysics Data System (ADS)

    Lee, Alex; Marianetti, Chris

    Re-based double perovskites (DPs) have attracted much attention due to their high Curie temperature (TC) and colossal magneto resistance with large potential for spintronic applications. Here we investigate the electronic and magnetic properties of the Re-based DPs A2 B ReO6 (A=Sr, Ca and B=Cr, Fe) using density functional theory + U (DFT+ U) calculations. While monoclinic Ca2CrReO6 and Ca2FeReO6 (monoclinic) are insulating within GGA+ U, tetragonal Sr2CrReO6 (a0a0c0) and Sr2FeReO6 (a0a0c-) remain metallic. We show that both on-site interaction U and octahedral tilting are critical to obtain the insulating phases. The a0a0c- -phase of Sr2CrReO6 is most stable and insulating with nonzero U, suggesting that the high quality Sr2CrReO6 film on STO substrate can be a semiconductor as reported in recent experiments. We explain that the insulator-to-metal transition (MIT) of Ca2FeReO6 at 140K is predominantly due to a structural phase transition which drives the insulating state. Curie temperatures of Re-based DPs are calculated using the classical Monte Carlo simulations based on the Heisenberg model.

  19. Excess adenosine A2B receptor signaling contributes to priapism through HIF-1α mediated reduction of PDE5 gene expression.

    PubMed

    Ning, Chen; Wen, Jiaming; Zhang, Yujin; Dai, Yingbo; Wang, Wei; Zhang, Weiru; Qi, Lin; Grenz, Almut; Eltzschig, Holger K; Blackburn, Michael R; Kellems, Rodney E; Xia, Yang

    2014-06-01

    Priapism is featured with prolonged and painful penile erection and is prevalent among males with sickle cell disease (SCD). The disorder is a dangerous urological and hematological emergency since it is associated with ischemic tissue damage and erectile disability. Here we report that phosphodiesterase-5 (PDE5) gene expression and PDE activity is significantly reduced in penile tissues of two independent priapic models: SCD mice and adenosine deaminase (ADA)-deficient mice. Moreover, using ADA enzyme therapy to reduce adenosine or a specific antagonist to block A(2B) adenosine receptor (ADORA2B) signaling, we successfully attenuated priapism in both ADA(-/-) and SCD mice by restoring penile PDE5 gene expression to normal levels. This finding led us to further discover that excess adenosine signaling via ADORA2B activation directly reduces PDE5 gene expression in a hypoxia-inducible factor-1α (HIF-1α)-dependent manner. Overall, we reveal that excess adenosine-mediated ADORA2B signaling underlies reduced penile PDE activity by decreasing PDE5 gene expression in a HIF-1α-dependent manner and provide new insight for the pathogenesis of priapism and novel therapies for the disease.

  20. Potential therapeutic interest of adenosine A2A receptors in psychiatric disorders.

    PubMed

    Cunha, Rodrigo A; Ferré, Sergi; Vaugeois, Jean-Marie; Chen, Jiang-Fan

    2008-01-01

    The interest on targeting adenosine A(2A) receptors in the realm of psychiatric diseases first arose based on their tight physical and functional interaction with dopamine D(2) receptors. However, the role of central A(2A) receptors is now viewed as much broader than just controlling D(2) receptor function. Thus, there is currently a major interest in the ability of A(2A) receptors to control synaptic plasticity at glutamatergic synapses. This is due to a combined ability of A(2A) receptors to facilitate the release of glutamate and the activation of NMDA receptors. Therefore, A(2A) receptors are now conceived as a normalizing device promoting adequate adaptive responses in neuronal circuits, a role similar to that fulfilled, in essence, by dopamine. This makes A(2A) receptors particularly attractive targets to manage psychiatric disorders since adenosine may act as go-between glutamate and dopamine, two of the key players in mood processing. Furthermore, A(2A) receptors also control glia function and brain metabolic adaptation, two other emerging mechanisms to understand abnormal processing of mood, and A(2A) receptors are important players in controlling the demise of neurodegeneration, considered an amplificatory loop in psychiatric disorders. Current data only provide an indirect confirmation of this putative role of A(2A) receptors, based on the effects of caffeine (an antagonist of both A(1) and A(2A) receptors) in psychiatric disorders. However, the introduction of A(2A) receptors antagonists in clinics as anti-parkinsonian agents is hoped to bolster our knowledge on the role of A(2A) receptors in mood disorders in the near future.

  1. Activation of the A2B adenosine receptor in B16 melanomas induces CXCL12 expression in FAP-positive tumor stromal cells, enhancing tumor progression

    PubMed Central

    Sorrentino, Claudia; Miele, Lucio; Porta, Amalia; Pinto, Aldo; Morello, Silvana

    2016-01-01

    The A2B receptor (A2BR) can mediate adenosine-induced tumor proliferation, immunosuppression and angiogenesis. Targeting the A2BR has proved to be therapeutically effective in some murine tumor models, but the mechanisms of these effects are still incompletely understood. Here, we report that pharmacologic inhibition of A2BR with PSB1115, which inhibits tumor growth, decreased the number of fibroblast activation protein (FAP)-expressing cells in tumors in a mouse model of melanoma. This effect was associated with reduced expression of fibroblast growth factor (FGF)-2. Treatment of melanoma-associated fibroblasts with the A2BR agonist Bay60-6583 enhanced CXCL12 and FGF2 expression. This effect was abrogated by PSB1115. The A2AR agonist CGS21680 did not induce CXCL12 or FGF2 expression in tumor associated fibroblasts. Similar results were obtained under hypoxic conditions in skin-derived fibroblasts, which responded to Bay60-6583 in an A2BR-dependent manner, by stimulating pERK1/2. FGF2 produced by Bay60-6583-treated fibroblasts directly enhanced the proliferation of melanoma cells. This effect could be reversed by PSB1115 or an anti-FGF2 antibody. Interestingly, melanoma growth in mice receiving Bay60-6583 was attenuated by inhibition of the CXCL12/CXCR4 pathway with AMD3100. CXCL12 and its receptor CXCR4 are involved in angiogenesis and immune-suppression. Treatment of mice with AMD3100 reduced the number of CD31+ cells induced by Bay60-6583. Conversely, CXCR4 blockade did not affect the accumulation of tumor-infiltrating MDSCs or Tregs. Together, our data reveal an important role for A2BR in stimulating FGF2 and CXCL12 expression in melanoma-associated fibroblasts. These factors contribute to create a tumor-promoting microenvironment. Our findings support the therapeutic potential of PSB1115 for melanoma. PMID:27590504

  2. Adenosine A2A Receptors and A2A Receptor Heteromers as Key Players in Striatal Function

    PubMed Central

    Ferré, Sergi; Quiroz, César; Orru, Marco; Guitart, Xavier; Navarro, Gemma; Cortés, Antonio; Casadó, Vicent; Canela, Enric I.; Lluis, Carme; Franco, Rafael

    2011-01-01

    A very significant density of adenosine A2A receptors (A2ARs) is present in the striatum, where they are preferentially localized postsynaptically in striatopallidal medium spiny neurons (MSNs). In this localization A2ARs establish reciprocal antagonistic interactions with dopamine D2 receptors (D2Rs). In one type of interaction, A2AR and D2R are forming heteromers and, by means of an allosteric interaction, A2AR counteracts D2R-mediated inhibitory modulation of the effects of NMDA receptor stimulation in the striatopallidal neuron. This interaction is probably mostly responsible for the locomotor depressant and activating effects of A2AR agonist and antagonists, respectively. The second type of interaction involves A2AR and D2R that do not form heteromers and takes place at the level of adenylyl cyclase (AC). Due to a strong tonic effect of endogenous dopamine on striatal D2R, this interaction keeps A2AR from signaling through AC. However, under conditions of dopamine depletion or with blockade of D2R, A2AR-mediated AC activation is unleashed with an increased gene expression and activity of the striatopallidal neuron and with a consequent motor depression. This interaction is probably the main mechanism responsible for the locomotor depression induced by D2R antagonists. Finally, striatal A2ARs are also localized presynaptically, in cortico-striatal glutamatergic terminals that contact the striato-nigral MSN. These presynaptic A2ARs heteromerize with A1 receptors (A1Rs) and their activation facilitates glutamate release. These three different types of A2ARs can be pharmacologically dissected by their ability to bind ligands with different affinity and can therefore provide selective targets for drug development in different basal ganglia disorders. PMID:21731559

  3. Lungs donated after circulatory death and prolonged warm ischemia are transplanted successfully after enhanced ex vivo lung perfusion using adenosine A2B receptor antagonism.

    PubMed

    Charles, Eric J; Mehaffey, J Hunter; Sharma, Ashish K; Zhao, Yunge; Stoler, Mark H; Isbell, James M; Lau, Christine L; Tribble, Curtis G; Laubach, Victor E; Kron, Irving L

    2017-04-12

    The current supply of acceptable donor lungs is not sufficient for the number of patients awaiting transplantation. We hypothesized that ex vivo lung perfusion (EVLP) with targeted drug therapy would allow successful rehabilitation and transplantation of donation after circulatory death lungs exposed to 2 hours of warm ischemia. Donor porcine lungs were procured after 2 hours of warm ischemia postcardiac arrest and subjected to 4 hours of cold preservation or EVLP. ATL802, an adenosine A2B receptor antagonist, was administered to select groups. Four groups (n = 4/group) were randomized: cold preservation (Cold), cold preservation with ATL802 during reperfusion (Cold + ATL802), EVLP (EVLP), and EVLP with ATL802 during ex vivo perfusion (EVLP + ATL802). Lungs subsequently were transplanted, reperfused, and assessed by measuring dynamic lung compliance and oxygenation capacity. EVLP + ATL802 significantly improved dynamic lung compliance compared with EVLP (25.0 ± 1.8 vs 17.0 ± 2.4 mL/cmH2O, P = .04), and compared with cold preservation (Cold: 12.2 ± 1.3, P = .004; Cold + ATL802: 10.6 ± 2.0 mL/cmH2O, P = .002). Oxygenation capacity was highest in EVLP (440.4 ± 37.0 vs Cold: 174.0 ± 61.3 mm Hg, P = .037). No differences in oxygenation or pulmonary edema were observed between EVLP and EVLP + ATL802. A significant decrease in interleukin-12 expression in tissue and bronchoalveolar lavage was identified between groups EVLP and EVLP + ATL802, along with less neutrophil infiltration. Severely injured donation after circulatory death lungs subjected to 2 hours of warm ischemia are transplanted successfully after enhanced EVLP with targeted drug therapy. Increased use of lungs after uncontrolled donor cardiac death and prolonged warm ischemia may be possible and may improve transplant wait list times and mortality. Copyright © 2017 The American Association for Thoracic Surgery. Published by Elsevier Inc. All rights reserved.

  4. KW-3902, a selective high affinity antagonist for adenosine A1 receptors.

    PubMed Central

    Nonaka, H.; Ichimura, M.; Takeda, M.; Kanda, T.; Shimada, J.; Suzuki, F.; Kase, H.

    1996-01-01

    1. We demonstrate that 8-(noradamantan-3-yl)-1,3-dipropylxanthine (KW-3902) is a very potent and selective adenosine A1 receptor antagonist, assessed by radioligand binding and cyclic AMP response in cells. 2. In rat forebrain adenosine A1 receptors labelled with [3H]-cyclohexyladenosine (CHA), KW-3902 had a Ki value of 0.19 nM, whereas it showed a Ki value of 170 nM in rat striatal A2A receptors labelled with [3H]-2-[p-(2-carboxyethyl)-phenethylamino]-5'-N-ethylcarboxamidoad enosine (CGS21680), indicating 890 fold A1 receptor selectivity versus the A2A receptor. KW-3902 at 10 microM showed no effect on recombinant rat A3 receptors expressed on CHO cells. 3. Saturation studies with [3H]-KW-3902 revealed that it bound with high affinity (Kd = 77 pM) and limited capacity (Bmax = 470 fmol mg-1 of protein) to a single class of recognition sites. A high positive correlation was observed between the pharmacological profile of adenosine ligands inhibiting the binding of [3H]-KW-3902 and that of [3H]-CHA. 4. KW-3902 showed potent A1 antagonism against the inhibition of forskolin-induced cyclic AMP accumulation in DDT1 MF-2 cells by the A1-selective agonist, cyclopentyladenosine with a dissociation constant (KB value) of 0.34 nM. KW-3902 antagonized 5'-N-ethylcarboxamidoadenosine-elicited cyclic AMP accumulation via A2B receptors with a KB value of 52 nM. 5. KW-3902 exhibited marked species-dependent differences in the binding affinities. The highest affinity was for the rat A1 receptor (ki = 0.19 nM) and these values for guinea-pig and dog A1 receptors were 1.3 and 10 nM, respectively. PMID:8732272

  5. The prevalent deep intronic c. 639+919 G>A GLA mutation causes pseudoexon activation and Fabry disease by abolishing the binding of hnRNPA1 and hnRNP A2/B1 to a splicing silencer.

    PubMed

    Palhais, Bruno; Dembic, Maja; Sabaratnam, Rugivan; Nielsen, Kira S; Doktor, Thomas Koed; Bruun, Gitte Hoffmann; Andresen, Brage Storstein

    2016-11-01

    Fabry disease is an X-linked recessive inborn disorder of the glycosphingolipid metabolism, caused by total or partial deficiency of the lysosomal α-galactosidase A enzyme due to mutations in the GLA gene. The prevalent c.639+919 G>A mutation in GLA leads to pathogenic insertion of a 57bp pseudoexon sequence from intron 4, which is responsible for the cardiac variant phenotype. In this study we investigate the splicing regulatory mechanism leading to GLA pseudoexon activation. Splicing analysis of GLA minigenes revealed that pseudoexon activation is influenced by cell-type. We demonstrate that the wild-type sequence harbors an hnRNP A1 and hnRNP A2/B1-binding exonic splicing silencer (ESS) overlapping the 5'splice site (5'ss) that prevents pseudoexon inclusion. The c.639+919 G>A mutation disrupts this ESS allowing U1 snRNP recognition of the 5'ss. We show that the wild-type GLA 5'ss motif with the ESS is also able to inhibit inclusion of an unrelated pseudoexon in the FGB gene, and that also in the FGB context inactivation of the ESS by the c.639+919 G>A mutation causes pseudoexon activation, underscoring the universal nature of the ESS. Finally, we demonstrate that splice switching oligonucleotide (SSO) mediated blocking of the pseudoexon 3'ss and 5'ss effectively restores normal GLA splicing. This indicates that SSO based splicing correction may be a therapeutic alternative in the treatment of Fabry disease.

  6. Ligands and therapeutic perspectives of adenosine A(2A) receptors.

    PubMed

    Diniz, C; Borges, F; Santana, L; Uriarte, E; Oliveira, J M A; Gonçalves, J; Fresco, P

    2008-01-01

    Adenosine A(2A) receptors are members of the G protein-coupled receptor family and mediate multiple physiological effects of adenosine, both at the central nervous system (CNS) and at peripheral tissues, by activating several pathways or interacting with other receptors or proteins. Increasing evidence relate A(2A) receptors with pharmacological stress testing, neurodegenerative disorders (such as Parkinson's disease) and inflammation, renewing the interest in these receptors, increasingly viewed as promising therapeutic targets. Series of agonists and antagonists have been developed by medicinal chemistry artwork either by structure activity relationship (SAR) or quantitative structure activity relationship (QSAR) studies. These studies have allowed identification of the structural and electrostatic requirements for high affinity A(2A) receptor binding and, therefore, contributing to the rational design of A(2A) receptor ligands. Additional rational chemical modifications of the existing A(2A) receptor ligands may further improve their affinity/selectivity. The purpose of this review is to analize and summarize aspects related to the medicinal chemistry of A(2A) receptor ligands, their present and potencial therapeutic applications by exploring the molecular structure and physiological and pathophysiological roles of A(2A) receptors.

  7. No evidence for priming response in Galleria mellonella larvae exposed to toxin protein PirA2B2 from Photorhabdus luminescens TT01: An association with the inhibition of the host cellular immunity.

    PubMed

    Wu, Gongqing; Yi, Yunhong; Sun, Jianyu; Li, Mei; Qiu, Lihong

    2015-11-17

    There is accumulating evidence that many invertebrates including insects can acquire enhanced immune protection against subsequently pathogens infection through immune priming. However, whether the toxin protein from pathogenic bacteria can induce such priming response remains unclear. Here we cloned, expressed and purified the toxin Photorhabdus insect-related proteins A2B2 (PirA2B2) from Photorhabdus luminescens TT01. We primed Galleria mellonella with sublethal dose of PirA2B2 and then challenged the larvae with viable P. luminescens TT01 at 48 h after priming. We found no evidence for immune priming in G. mellonella larvae exposed to PirA2B2. Priming the larvae with PirA2B2 did not improve their resistance in a subsequent challenge with P. luminescens TT01. Whereas a robust priming response was observed when the larvae exposed to lipopolysaccharide (LPS) extracted from P. luminescens TT01. Because the larvae primed with LPS showed significant higher resistance against P. luminescens TT01 infection than those of the PBS and BSA controls. Furthermore, we investigated the changes of the cellular immune parameters, such as hemocyte counts, phagocytic activity and encapsulation ability of the hemocytes, after priming. We found that the toxin PirA2B2 significantly decreased the cellular immunity of the larvae, whereas the LPS significantly increased them. These results indicated that the degree of priming response in G. mellonella correlated positively to the levels of cellular immune parameters, and the underlying mechanism in regulating the immune priming of invertebrates was not homologous to that of the immunological memory of vertebrates. Copyright © 2015 Elsevier Ltd. All rights reserved.

  8. Involvement of adenosine A2A receptors in depression and anxiety.

    PubMed

    Yamada, Koji; Kobayashi, Minoru; Kanda, Tomoyuki

    2014-01-01

    When administered to normal healthy patients, a nonselective adenosine A1/A2A antagonist, caffeine, tended to improve anxiety and depression at low doses and to exacerbate anxiety at high doses. Caffeine also appears to enhance anxiety-related symptoms in patients with panic disorder, and A2A receptor-deficient mice have been reported to exhibit higher anxiety-like behaviors, as well as a lower incidence of depression-like behaviors. Some selective A2A antagonists were reported to ameliorate anxiety-like behaviors in rodents, while others did not affect these behaviors. In addition, most A2A antagonists showed inhibitory effects on depression-like behaviors. The mechanisms underlying the relationship between A2A receptor antagonists and anxiety and depression remain unclear at the present time, although many studies have produced hypotheses. Given that a selective A2A receptor antagonist has recently become available for use in humans, research on the role of A2A receptors in the treatment of mental illness should progress in the near future. © 2014 Elsevier Inc. All rights reserved.

  9. Adenosine A2A receptor mediated protective effect of 2‐(6‐cyano‐1‐hexyn‐1‐yl)adenosine on retinal ischaemia/reperfusion damage in rats

    PubMed Central

    Konno, T; Sato, A; Uchibori, T; Nagai, A; Kogi, K; Nakahata, N

    2006-01-01

    Aims To determine the effect of 2‐(6‐cyano‐1‐hexyn‐1‐yl)adenosine (2‐CN‐Ado), an adenosine A2A receptor agonist, on retinal ischaemia/reperfusion damage in rats. Methods Retinal ischaemia/reperfusion damage was induced by elevating the intraocular pressure of one eye to 130 mm Hg for 60 minutes and returning it to normal. 7 days later, retinal ischaemia/reperfusion damage was histologically quantified by measuring the thickness of retinal layers. Intraocular pressure was measured by pressure transducer. Results Retinal ischaemia/reperfusion caused cell loss in the ganglion cell layer and thinning of the inner plexiform and nuclear layer. Both ocular topical and intravenous administration of 2‐CN‐Ado caused a reduction of retinal ischaemia/reperfusion damage. A selective A2A receptor antagonist, 1,3,7‐trimethyl‐8‐(3‐chlorostyryl) xanthine (CSC), but not a selective A1 receptor antagonist, 8‐cyclopentyl‐1,3‐dipropylxanthine (DPCPX), or a selective A2B receptor antagonist, alloxazine, reduced the protective effect of 2‐CN‐Ado. While ocular topical administration of 2‐CN‐Ado caused a sustained reduction of intraocular pressure, intravenous administration of 2‐CN‐Ado showed a transient ocular hypotensive effect. Conclusions These results suggest that 2‐CN‐Ado attenuates retinal ischaemia/reperfusion damage, and at least some of this protective effect of 2‐CN‐Ado might be mediated via activation of the adenosine A2A receptor. PMID:16613921

  10. Adenosine A1 and A3 receptors protect astrocytes from hypoxic damage.

    PubMed

    Björklund, Olga; Shang, Mingmei; Tonazzini, Ilaria; Daré, Elisabetta; Fredholm, Bertil B

    2008-10-31

    Brain levels of adenosine are elevated during hypoxia. Through effects on adenosine receptors (A(1), A(2A), A(2B) and A(3)) on astrocytes, adenosine can influence functions such as glutamate uptake, reactive gliosis, swelling, as well as release of neurotrophic and neurotoxic factors having an impact on the outcome of metabolic stress. We have studied the roles of these receptors in astrocytes by evaluating their susceptibility to damage induced by oxygen deprivation or exposure to the hypoxia mimic cobalt chloride (CoCl(2)). Hypoxia caused ATP breakdown and purine release, whereas CoCl(2) (0.8 mM) mainly reduced ATP by causing cell death in human D384 astrocytoma cells. Further experiments were conducted in primary astrocytes prepared from specific adenosine receptor knock-out (KO) and wild type (WT) mice. In WT cells purine release following CoCl(2) exposure was mainly due to nucleotide release, whereas hypoxia-induced intracellular ATP breakdown followed by nucleoside efflux. N-ethylcarboxamidoadenosine (NECA), an unselective adenosine receptor agonist, protected from cell death following hypoxia. Cytotoxicity was more pronounced in A(1)R KO astrocytes and tended to be higher in WT cells in the presence of the A(1) receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine (DPCPX). Genetic deletion of A(2A) receptor resulted in less prominent effects. A(3)R KO glial cells were more affected by hypoxia than WT cells. Accordingly, the A(3) receptor agonist 2-chloro-N(6)-(3-iodobenzyl)-N-methyl-5'-carbamoyladenosine (CL-IB-MECA) reduced ATP depletion caused by hypoxic conditions. It also reduced apoptosis in human astroglioma D384 cells after oxygen deprivation. In conclusion, the data point to a cytoprotective role of adenosine mediated by both A(1) and A(3) receptors in primary mouse astrocytes.

  11. A(1) and A(3) adenosine receptors inhibit LPS-induced hypoxia-inducible factor-1 accumulation in murine astrocytes.

    PubMed

    Gessi, Stefania; Merighi, Stefania; Stefanelli, Angela; Fazzi, Debora; Varani, Katia; Borea, Pier Andrea

    2013-10-01

    Adenosine (Ado) exerts neuroprotective and anti-inflammatory functions by acting through four receptor subtypes A1, A2A, A2B and A3. Astrocytes are one of its targets in the central nervous system. Hypoxia-inducible factor-1 (HIF-1), a master regulator of oxygen homeostasis, is induced after hypoxia, ischemia and inflammation and plays an important role in brain injury. HIF-1 is expressed by astrocytes, however the regulatory role played by Ado on HIF-1α modulation induced by inflammatory and hypoxic conditions has not been investigated. Primary murine astrocytes were activated with lipopolysaccharide (LPS) with or without Ado, Ado receptor agonists, antagonists and receptor silencing, before exposure to normoxia or hypoxia. HIF-1α accumulation and downstream genes regulation were determined. Ado inhibited LPS-increased HIF-1α accumulation under both normoxic and hypoxic conditions, through activation of A1 and A3 receptors. In cells incubated with the blockers of p44/42 MAPK and Akt, LPS-induced HIF-1α accumulation was significantly decreased in normoxia and hypoxia, suggesting the involvement of p44/42 MAPK and Akt in this effect and Ado inhibited kinases phosphorylation. A series of angiogenesis and metabolism related genes were modulated by hypoxia in an HIF-1 dependent way, but not further increased by LPS, with the exception of GLUT-1 and hexochinase II that were elevated by LPS only in normoxia and inhibited by Ado receptors. Instead, genes involved in inflammation, like inducible nitric-oxide synthase (iNOS) and A2B receptors, were increased by LPS in normoxia, strongly stimulated by LPS in concert with hypoxia and inhibited by Ado, through A1 and A3 receptor subtypes. In conclusion A1 and A3 receptors reduce the LPS-mediated HIF-1α accumulation in murine astrocytes, resulting in a downregulation of genes involved in inflammation and hypoxic injury, like iNOS and A2B receptors, in both normoxic and hypoxic conditions.

  12. Suppression of HPV-16 late L1 5′-splice site SD3632 by binding of hnRNP D proteins and hnRNP A2/B1 to upstream AUAGUA RNA motifs

    PubMed Central

    Li, Xiaoze; Johansson, Cecilia; Glahder, Jacob; Mossberg, Ann-Kristin; Schwartz, Stefan

    2013-01-01

    Human papillomavirus type 16 (HPV-16) 5′-splice site SD3632 is used exclusively to produce late L1 mRNAs. We identified a 34-nt splicing inhibitory element located immediately upstream of HPV-16 late 5′-splice site SD3632. Two AUAGUA motifs located in these 34 nt inhibited SD3632. Two nucleotide substitutions in each of the HPV-16 specific AUAGUA motifs alleviated splicing inhibition and induced late L1 mRNA production from episomal forms of the HPV-16 genome in primary human keratinocytes. The AUAGUA motifs bind specifically not only to the heterogeneous nuclear RNP (hnRNP) D family of RNA-binding proteins including hnRNP D/AUF, hnRNP DL and hnRNP AB but also to hnRNP A2/B1. Knock-down of these proteins induced HPV-16 late L1 mRNA expression, and overexpression of hnRNP A2/B1, hnRNP AB, hnRNP DL and the two hnRNP D isoforms hnRNP D37 and hnRNP D40 further suppressed L1 mRNA expression. This inhibition may allow HPV-16 to hide from the immune system and establish long-term persistent infections with enhanced risk at progressing to cancer. There is an inverse correlation between expression of hnRNP D proteins and hnRNP A2/B1 and HPV-16 L1 production in the cervical epithelium, as well as in cervical cancer, supporting the conclusion that hnRNP D proteins and A2/B1 inhibit HPV-16 L1 mRNA production. PMID:24013563

  13. Agonist Derived Molecular Probes for A2A Adenosine Receptors

    PubMed Central

    Jacobson, Kenneth A.; Pannell, Lewis K.; Ji, Xiao-duo; Jarvis, Michael F.; Williams, Michael; Hutchison, Alan J.; Barrington, William W.; Stiles, Gary L.

    2011-01-01

    The adenosine agonist 2-(4-(2-carboxyethyl)phenylethylamino)-5′-N-ethylcarboxamidoadenosine (CGS21680) was recently reported to be selective for the A2A adenosine receptor subtype, which mediates its hypotensive action. To investigate structurelactivity relationships at a distal site, CGS21680 was derivatized using a functionalized congener approach. The carboxylic group of CGS21680 has been esterified to form a methyl ester, which was then treated with ethylenediamine to produce an amine congener. The amine congener was an intermediate for acylation reactions, in which the reactive acyl species contained a reported group, or the precursor for such. For radioiodination, derivatives of p-hydroxyphenylpropionic, 2-thiophenylacetic, and p-aminophenylacetic acids were prepared. The latter derivative (PAPA-APEC) was iodinated electrophilically using [125I]iodide resulting in a radioligand which was used for studies of competition of binding to striatal A, adenosine receptors in bovine brain. A biotin conjugate and an aryl sulfonate were at least 350-fold selective for A, receptors. For spectroscopic detection, a derivative of the stable free radical tetramethyl-1-piperidinyloxy (TEMPO) was prepared. For irreversible inhibition of receptors, meta- and para-phenylenediisothiocyanate groups were incorporated in the analogs. We have demonstrated that binding at A2A receptors is relatively insensitive to distal structural changes at the 2-position, and we report high affinity molecular probes for receptor characterization by radioactive, spectroscopic and affinity labelling methodology. PMID:2561548

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

    PubMed Central

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

    2015-01-01

    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

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

    PubMed

    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.

  16. Neuroprotection by caffeine and A(2A) adenosine receptor inactivation in a model of Parkinson's disease.

    PubMed

    Chen, J F; Xu, K; Petzer, J P; Staal, R; Xu, Y H; Beilstein, M; Sonsalla, P K; Castagnoli, K; Castagnoli, N; Schwarzschild, M A

    2001-05-15

    Recent epidemiological studies have established an association between the common consumption of coffee or other caffeinated beverages and a reduced risk of developing Parkinson's disease (PD). To explore the possibility that caffeine helps prevent the dopaminergic deficits characteristic of PD, we investigated the effects of caffeine and the adenosine receptor subtypes through which it may act in the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) neurotoxin model of PD. Caffeine, at doses comparable to those of typical human exposure, attenuated MPTP-induced loss of striatal dopamine and dopamine transporter binding sites. The effects of caffeine were mimicked by several A(2A) antagonists (7-(2-phenylethyl)-5-amino-2-(2-furyl)-pyrazolo-[4,3-e]-1,2,4-triazolo[1,5-c]pyrimidine (SCH 58261), 3,7-dimethyl-1-propargylxanthine, and (E)-1,3-diethyl-8 (KW-6002)-(3,4-dimethoxystyryl)-7-methyl-3,7-dihydro-1H-purine-2,6-dione) (KW-6002) and by genetic inactivation of the A(2A) receptor, but not by A(1) receptor blockade with 8-cyclopentyl-1,3-dipropylxanthine, suggesting that caffeine attenuates MPTP toxicity by A(2A) receptor blockade. These data establish a potential neural basis for the inverse association of caffeine with the development of PD, and they enhance the potential of A(2A) antagonists as a novel treatment for this neurodegenerative disease.

  17. Adenosine A2A receptors are necessary and sufficient to trigger memory impairment in adult mice

    PubMed Central

    Pagnussat, N; Almeida, A S; Marques, D M; Nunes, F; Chenet, G C; Botton, P H S; Mioranzza, S; Loss, C M; Cunha, R A; Porciúncula, L O

    2015-01-01

    Background and Purpose Caffeine (a non-selective adenosine receptor antagonist) prevents memory deficits in aging and Alzheimer’s disease, an effect mimicked by adenosine A2A receptor, but not A1 receptor, antagonists. Hence, we investigated the effects of adenosine receptor agonists and antagonists on memory performance and scopolamine-induced memory impairment in mice. Experimental Approach We determined whether A2A receptors are necessary for the emergence of memory impairments induced by scopolamine and whether A2A receptor activation triggers memory deficits in naïve mice, using three tests to assess short-term memory, namely the object recognition task, inhibitory avoidance and modified Y-maze. Key Results Scopolamine (1.0 mg·kg−1, i.p.) impaired short-term memory performance in all three tests and this scopolamine-induced amnesia was prevented by the A2A receptor antagonist (SCH 58261, 0.1–1.0 mg·kg−1, i.p.) and by the A1 receptor antagonist (DPCPX, 0.2–5.0 mg·kg−1, i.p.), except in the modified Y-maze where only SCH58261 was effective. Both antagonists were devoid of effects on memory or locomotion in naïve rats. Notably, the activation of A2A receptors with CGS 21680 (0.1–0.5 mg·kg−1, i.p.) before the training session was sufficient to trigger memory impairment in the three tests in naïve mice, and this effect was prevented by SCH 58261 (1.0 mg·kg−1, i.p.). Furthermore, i.c.v. administration of CGS 21680 (50 nmol) also impaired recognition memory in the object recognition task. Conclusions and Implications These results show that A2A receptors are necessary and sufficient to trigger memory impairment and further suggest that A1 receptors might also be selectively engaged to control the cholinergic-driven memory impairment. PMID:25939452

  18. Gender-dependent associations of CDKN2A/2B, KCNJ11, POLI, SLC30A8, and TCF7L2 variants with type 2 diabetes in (North African) Tunisian Arabs.

    PubMed

    Turki, Amira; Al-Zaben, Ghadeer S; Khirallah, Moncef; Marmouch, Hela; Mahjoub, Touhami; Almawi, Wassim Y

    2014-03-01

    We investigated the impact of gender on T2DM association with confirmed susceptibility loci. CDKN2A/2B rs10811661, KCNJ11 rs5219, and TCF7L2 rs7903146 were associated with T2DM in females, while POLI rs488846 was associated with T2DM among males; the association of SLC30A8 rs13266634 and TCF7L2 rs4506565, rs12243326, and rs12255372 with T2DM was gender-independent. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

  19. Concurrent agonism of adenosine A2B and glucocorticoid receptors in human airway epithelial cells cooperatively induces genes with anti-inflammatory potential: a novel approach to treat chronic obstructive pulmonary disease.

    PubMed

    Greer, Stephanie; Page, Cara W; Joshi, Taruna; Yan, Dong; Newton, Robert; Giembycz, Mark A

    2013-09-01

    Chronic obstructive pulmonary disease (COPD) is a neutrophilic inflammatory disorder that is weakly responsive to glucocorticoids. Identification of ways to enhance the anti-inflammatory activity of glucocorticoids is, therefore, a major research objective. Adenosine receptor agonists that target the A2B-receptor subtype are efficacious in several cell-based assays and preclinical models of inflammation. Accordingly, the present study was designed to determine if a selective A2B-receptor agonist, 2-[6-amino-3,5-dicyano-4-[4-(cyclopropylmethoxy)phenyl]pyridin-2-ylsulphanyl]acetamide (Bay 60-6583), and a glucocorticoid, dexamethasone, in combination display putative anti-inflammatory activity that is superior to either drug alone. In BEAS-2B human airway epithelial cells stably transfected with cAMP-response element (CRE) and glucocorticoid response element (GRE) reporter constructs, Bay 60-6583 promoted CRE-dependent transcription and enhanced GRE-dependent transcription by an adenosine A2B-receptor-mediated mechanism that was associated with cAMP formation and abolished by an inhibitor of cAMP-dependent protein kinase. Analysis of the concentration-response relationship that described the enhancement of GRE-dependent transcription showed that Bay 60-6583 increased the magnitude of response without affecting the potency of dexamethasone. Bay 60-6583 and dexamethasone also induced a panel of genes that, collectively, could have benefit in COPD. These were categorized into genes that were induced in a positive cooperative manner (RGS2, p57(kip2)), an additive manner (TTP, BRL-1), or by Bay 60-6583 (CD200, CRISPLD2, SOCS3) or dexamethasone (GILZ) only. Thus, the gene induction "fingerprints" produced by Bay 60-6583 and dexamethasone, alone and in combination, were distinct. Collectively, through their actions on gene expression, an adenosine A2B-receptor agonist and a glucocorticoid administered together may have utility in the treatment of inflammatory disorders that

  20. Adenosine A2A receptors in ventral striatum, hypothalamus and nociceptive circuitry. Implications for drug addiction, sleep and pain

    PubMed Central

    Ferré, S.; Diamond, I.; Goldberg, S.R.; Yao, L.; Hourani, S.M.O.; Huang, Z.L.; Urade, Y.; Kitchen, I.

    2007-01-01

    Adenosine A2A receptors localized in the dorsal striatum are considered as a new target for the development of antiparkinsonian drugs. Co-administration of A2A receptor antagonists has shown a significant improvement of the effects of L-DOPA. The present review emphasizes the possible application of A2A receptor antagonists in pathological conditions other than parkinsonism, including drug addiction, sleep disorders and pain. In addition to the dorsal striatum, the ventral striatum (nucleus accumbens) contains a high density of A2A receptors, which presynaptically and postsynaptically regulate glutamatergic transmission in the cortical glutamatergic projections to the nucleus accumbens. It is currently believed that molecular adaptations of the cortico-accumbens glutamatergic synapses are involved in compulsive drug seeking and relapse. Here we review recent experimental evidence suggesting that A2A antagonists could become new therapeutic agents for drug addiction. Morphological and functional studies have identified lower levels of A2A receptors in brain areas other than the striatum, such as the ventrolateral preoptic area of the hypothalamus, where adenosine plays an important role in sleep regulation. Although initially believed to be mostly dependent on A1 receptors, here we review recent studies that demonstrate that the somnogenic effects of adenosine are largely mediated by hypothalamic A2A receptors. A2A receptor antagonists could therefore be considered as a possible treatment for narcolepsy and other sleep-related disorders. Finally, nociception is another adenosine-regulated neural function previously thought to mostly involve A1 receptors. Although there is some conflicting literature on the effects of agonists and antagonists, which may partly be due to the lack of selectivity of available drugs, the studies in A2A receptor knockout mice suggest that A2A receptor antagonists might have some therapeutic potential in pain states, in particular where

  1. Behavioural and neurochemical characterization of the adenosine A2A receptor antagonist ST1535.

    PubMed

    Galluzzo, Mariangela; Pintor, Anita; Pèzzola, Antonella; Grieco, Rosa; Borsini, Franco; Popoli, Patrizia

    2008-01-28

    ST1535 (2-butyl-9-methyl-8-(2H-1,2,3-triazol 2-yl)-9 H-purin-6-ylamine) is a novel compound showing a preferential adenosine A(2A) receptor antagonist profile. To explore the potential neuroprotective profile of this compound, we evaluated whether ST1535 prevented quinolinic acid (QA)-induced glutamate outflow in the rat striatum (a reliable index of neuroprotective activity in vivo). Microdialysis experiments were performed in naive Wistar rats. In these experiments, a behaviourally active and inactive doses of ST1535 were used. Both doses significantly prevented QA-induced glutamate outflow in the striatum. These results show that ST1535 protects towards striatal excitotoxicity, even though its reduced A(2A)/A(1) selectivity might limit its actual neuroprotective potential.

  2. Chemical Modification and Irreversible Inhibition of Striatal A2a Adenosine Receptors

    PubMed Central

    JACOBSON, KENNETH A.; STILES, GARY L.; JI, XIAO-DUO

    2012-01-01

    SUMMARY The ligand recognition site of A2a-adenosine receptors in rabbit striatal membranes was probed using non-site-directed labeling reagents and specific affinity labels. Exposure of membranes to diethylpyrocarbonate at a concentration of 2.5 mm, followed by washing, was found to inhibit the binding of [3H]CGS 21680 and [3H]xanthine amine congener to A2a receptors, by 86 and 30%, respectively. Protection from diethylpyrocarbonate inactivation by an adenosine receptor agonist, 5′-N-ethylcarboxamidoadenosine, and an antagonist, theophylline, suggested the presence of two histidyl residues on the receptor, one associated with agonist binding and the other with antagonist binding. Binding of [3H]CGS 21680 or [3H]xanthine amine congener was partially restored after incubation with 250 mm hydroxylamine, further supporting histidine as the modification site. Preincubation with disulfide-reactive reagents, dithiothreitol or sodium dithionite, at >5 mm inhibited radioligand binding, indicating the presence of essential disulfide bridges in A2a receptors, whereas the concentration of mercaptoethanol required to inhibit binding was >50 mm. A number of isothiocyanate-bearing affinity labels derived from the A2a-selective agonist 2-[(2-aminoethylamino)carbonylethylphenylethylamino]-5′-N-ethylcarboxamidoadenosine (APEC) were synthesized and found to inhibit A2a receptor binding in rabbit and bovine striatal membranes. Binding to rabbit A1 receptors was not inhibited. Preincubation with the affinity label 4-isothiocyanatophenylaminothiocarbonyl-APEC (100 nm) diminished the Bmax for [3H]CGS 21680 binding by 71%, and the Kd was unaffected, suggesting a direct modification of the ligand binding site. Reversal of 4-isothiocyanatophenylaminothiocarbonyl-APEC inhibition of [3H]CGS 21680 binding with hydroxylamine suggested that the site of modification by the isothiocyanate is a cysteine residue. A bromoacetyl derivative of APEC was ineffective as an affinity label at

  3. 4-amino-6-alkyloxy-2-alkylthiopyrimidine derivatives as novel non-nucleoside agonists for the adenosine A1 receptor.

    PubMed

    Cosimelli, Barbara; Greco, Giovanni; Laneri, Sonia; Novellino, Ettore; Sacchi, Antonia; Trincavelli, Maria Letizia; Giacomelli, Chiara; Taliani, Sabrina; Da Settimo, Federico; Martini, Claudia

    2016-11-01

    Three 4-amino-6-alkyloxy-2-alkylthiopyrimidine derivatives (4-6) were investigated as potential non-nucleoside agonists at human adenosine receptors (ARs). When tested in competition binding experiments, these compounds exhibited low micromolar affinity (Ki values comprised between 1.2 and 1.9 μm) for the A1 AR and no appreciable affinity for the A2A and A3 ARs. Evaluation of their efficacy profiles by measurement of intracellular cAMP levels revealed that 4 and 5 behave as non-nucleoside agonists of the A1 AR with EC50 values of 0.47 and 0.87 μm, respectively. No clear concentration-response curves could be instead obtained for 6, probably because this compound modulates one or more additional targets, thus masking the putative effects exerted by its activation of A1 AR. The three compounds were not able to modulate A2B AR-mediated cAMP accumulation induced by the non-selective AR agonist NECA, thus demonstrating no affinity toward this receptor. © 2016 John Wiley & Sons A/S.

  4. Identification of Novel Adenosine A2A Receptor Antagonists by Virtual Screening

    PubMed Central

    2012-01-01

    Virtual screening was performed against experimentally enabled homology models of the adenosine A2A receptor, identifying a diverse range of ligand efficient antagonists (hit rate 9%). By use of ligand docking and Biophysical Mapping (BPM), hits 1 and 5 were optimized to potent and selective lead molecules (11–13 from 5, pKI = 7.5–8.5, 13- to >100-fold selective versus adenosine A1; 14–16 from 1, pKI = 7.9–9.0, 19- to 59-fold selective). PMID:22250781

  5. The effects of clobazam treatment in rats on the expression of genes and proteins encoding glucronosyltransferase 1A/2B (UGT1A/2B) and multidrug resistance‐associated protein-2 (MRP2), and development of thyroid follicular cell hypertrophy

    SciTech Connect

    Miyawaki, Izuru Tamura, Akitoshi; Matsumoto, Izumi; Inada, Hiroshi; Kunimatsu, Takeshi; Kimura, Juki; Funabashi, Hitoshi

    2012-12-15

    Clobazam (CLB) is known to increase hepatobiliary thyroxine (T4) clearance in Sprague–Dawley (SD) rats, which results in hypothyroidism followed by thyroid follicular cell hypertrophy. However, the mechanism of the acceleration of T4-clearance has not been fully investigated. In the present study, we tried to clarify the roles of hepatic UDP-glucronosyltransferase (UGT) isoenzymes (UGT1A and UGT2B) and efflux transporter (multidrug resistance–associated protein-2; MRP2) in the CLB-induced acceleration of T4-clearance using two mutant rat strains, UGT1A-deficient mutant (Gunn) and MRP2-deficient mutant (EHBR) rats, especially focusing on thyroid morphology, levels of circulating hormones (T4 and triiodothyronine (T3)) and thyroid-stimulating hormone (TSH), and mRNA or protein expressions of UGTs (Ugt1a1, Ugt1a6, and Ugt2b1/2) and MRP2 (Mrp). CLB induced thyroid morphological changes with increases in TSH in SD and Gunn rats, but not in EHBR rats. T4 was slightly decreased in SD and Gunn rats, and T3 was decreased in Gunn rats, whereas these hormones were maintained in EHBR rats. Hepatic Ugt1a1, Ugt1a6, Ugt2b1/2, and Mrp2 mRNAs were upregulated in SD rats. In Gunn rats, UGT1A mRNAs (Ugt1a1/6) and protein levels were quite low, but UGT2B mRNAs (Ugt2b1/2) and protein were prominently upregulated. In SD and Gunn rats, MRP2 mRNA and protein were upregulated to the same degree. These results suggest that MRP2 is an important contributor in development of the thyroid cellular hypertrophy in CLB-treated rats, and that UGT1A and UGT2B work in concert with MRP2 in the presence of MRP2 function to enable the effective elimination of thyroid hormones. -- Highlights: ► Role of UGT and MRP2 in thyroid pathology was investigated in clobazam-treated rats. ► Clobazam induced thyroid cellular hypertrophy in SD and Gunn rats, but not EHBR rats. ► Hepatic Mrp2 gene and protein were upregulated in SD and Gunn rats, but not EHBR rats. ► Neither serum thyroid hormones (T3/T4

  6. Adenosine A(2A)-cannabinoid CB(1) receptor interaction: an integrative mechanism in striatal glutamatergic neurotransmission.

    PubMed

    Tebano, Maria Teresa; Martire, Alberto; Popoli, Patrizia

    2012-10-02

    The striatum is a subcortical area involved in sensorimotor, cognitive and emotional processes. Adenosine A(2A) receptors (A(2A)Rs) are highly expressed in the striatum, and their ability to establish functional and molecular interactions with many other receptors attributes to a pivotal role in the modulation and integration of striatal neurotransmission. This review will focus on the interaction between A(2A)Rs and cannabinoid CB(1) receptors (CB(1)Rs), taking it as a paradigmatic example of synaptic integration. Indeed, A(2A)Rs can exert an opposite (permissive vs. inhibitory) influence on CB1-dependent synaptic effect. These apparently irreconcilable functions could depend on a different role of pre- vs. postsynaptic A(2A)Rs, on their interaction with other receptors (namely adenosine A(1), metabotropic glutamate 5 and dopamine D2 receptors), and on whether A(2A)Rs form or not heteromers with CB(1)Rs. Besides providing a good example of the intricate pattern of events taking place in striatal synapses, the A(2A)/CB(1)R interaction proves very informative to understand the physiology of the basal ganglia and the mechanisms of related diseases. This article is part of a Special Issue entitled: Brain Integration.

  7. A covalent antagonist for the human adenosine A2A receptor.

    PubMed

    Yang, Xue; Dong, Guo; Michiels, Thomas J M; Lenselink, Eelke B; Heitman, Laura; Louvel, Julien; IJzerman, Ad P

    2016-12-03

    The structure of the human A2A adenosine receptor has been elucidated by X-ray crystallography with a high affinity non-xanthine antagonist, ZM241385, bound to it. This template molecule served as a starting point for the incorporation of reactive moieties that cause the ligand to covalently bind to the receptor. In particular, we incorporated a fluorosulfonyl moiety onto ZM241385, which yielded LUF7445 (4-((3-((7-amino-2-(furan-2-yl)-[1, 2, 4]triazolo[1,5-a][1, 3, 5]triazin-5-yl)amino)propyl)carbamoyl)benzene sulfonyl fluoride). In a radioligand binding assay, LUF7445 acted as a potent antagonist, with an apparent affinity for the hA2A receptor in the nanomolar range. Its apparent affinity increased with longer incubation time, suggesting an increasing level of covalent binding over time. An in silico A2A-structure-based docking model was used to study the binding mode of LUF7445. This led us to perform site-directed mutagenesis of the A2A receptor to probe and validate the target lysine amino acid K153 for covalent binding. Meanwhile, a functional assay combined with wash-out experiments was set up to investigate the efficacy of covalent binding of LUF7445. All these experiments led us to conclude LUF7445 is a valuable molecular tool for further investigating covalent interactions at this receptor. It may also serve as a prototype for a therapeutic approach in which a covalent antagonist may be needed to counteract prolonged and persistent presence of the endogenous ligand adenosine.

  8. Mass spectroscopy identifies the splicing-associated proteins, PSF, hnRNP H3, hnRNP A2/B1, and TLS/FUS as interacting partners of the ZNF198 protein associated with rearrangement in myeloproliferative disease

    SciTech Connect

    Kasyapa, Chitta S.; Kunapuli, Padmaja; Cowell, John K. . E-mail: John.Cowell@RoswellPark.org

    2005-09-10

    ZNF198 is fused with FGFR1 in an atypical myeloproliferative disease that results in constitutive activation of the kinase domain and mislocalization to the cytoplasm. We have used immunoprecipitation of a GFP-tagged ZNF198 combined with MALDI-TOF mass spectroscopy to identify interacting proteins. P splicing factor (PSF) was identified as one of the proteins and this interaction was confirmed by Western blotting. Other proteins identified were the spliceosomal components hnRNP A2/B1, hnRNP H3, and TLS/FUS. PSF is also known to interact with PTB, another member of the hnRNP family of proteins, and we further demonstrated that PTB interacts with ZNF198. The interaction between TLS/FUS and ZNF198 was confirmed using Western blot analysis. In 293 cells expressing the ZNF198/FGFR1 fusion protein, neither PSF nor PTB binds to the fusion protein, possibly because of their differential localization in the cell.

  9. Theoretical and experimental investigation of defect formation / migration in Gd2Ti2O7: General rule of oxide-ion migration in A2B2O7 pyrochlore

    NASA Astrophysics Data System (ADS)

    Nakamura, Kaoru; Mori, Masashi; Itoh, Takanori.; Ohnuma, Toshiharu

    2016-11-01

    We investigated the intrinsic defect formation energy and oxide-ion migration mechanism in Gd2Ti2O7 pyrochlore. It was found that the vacancy formation energy of Gd is lower than that of Ti. For the oxygen vacancy, O(48f) was found to show lower vacancy formation energy than O(8b). The formation energy of the vacancy complex showed that the Gd vacancy is accompanied with the O(48f) vacancy, which is consistent with our experiment. The migration energy of O(48f) along the <100> direction, which is dominant migration path for ionic conduction, was calculated to be 0.43 eV. On the other hand, we found that Gd vacancy increases O(48f) migration energy. For example, the migration energy of O(48f) along the <100> direction was increased to be 1.36 eV by the local compressive strain around Gd vacancy. This finding could explain our previous experimental result of decreasing conductivity with increasing Gd deficiency. Along with the oxide-ion migration mechanism in Gd2Ti2O7, O(48f) migration energies along both <100> and <110> directions for various A2B2O7 pyrochlore structures were investigated. As a general trend of oxide-ion migration in the pyrochlore structure, we propose that O(48f) migration along the <100> direction is governed by the strength of B-O bonding. On the other hand, the ratio of ionic radius B/A is proposed to determine O(48f) migration along the <110> direction in A2B2O7 pyrochlore.

  10. A1C test

    MedlinePlus

    HbA1C test; Glycated hemoglobin test; Glycohemoglobin test; Hemoglobin A1C; Diabetes - A1C; Diabetic - A1C ... gov/pubmed/26696680 . Chernecky CC, Berger BJ. Glycosylated hemoglobin (GHb, glycohemoglobin, glycated hemoglobin, HbA1a, HbA1b, HbA1c - blood. ...

  11. Postsynaptic Adenosine A2A Receptors Modulate Intrinsic Excitability of Pyramidal Cells in the Rat Basolateral Amygdala

    PubMed Central

    Rau, Andrew R.; Ariwodola, Olusegun J.

    2015-01-01

    Background: The basolateral amygdala plays a critical role in the etiology of anxiety disorders and addiction. Pyramidal neurons, the primary output cells of this region, display increased firing following exposure to stressors, and it is thought that this increase in excitability contributes to stress responsivity and the expression of anxiety-like behaviors. However, much remains unknown about the underlying mechanisms that regulate the intrinsic excitability of basolateral amygdala pyramidal neurons. Methods: Ex vivo gramicidin perforated patch recordings were conducted in current clamp mode where hyper- and depolarizing current steps were applied to basolateral amygdala pyramidal neurons to assess the effects of adenosine A2A receptor modulation on intrinsic excitability. Results: Activation of adenosine A2A receptors with the selective A2A receptor agonist CGS-21680 significantly increased the firing rate of basolateral amygdala pyramidal neurons in rat amygdala brain slices, likely via inhibition of the slow afterhyperpolarization potential. Both of these A2A receptor-mediated effects were blocked by preapplication of a selective A2A receptor antagonist (ZM-241385) or by intra-pipette infusion of a protein kinase A inhibitor, suggesting a postsynaptic locus of A2A receptors on basolateral amygdala pyramidal neurons. Interestingly, bath application of the A2A receptor antagonist alone significantly attenuated basolateral amygdala pyramidal cell firing, consistent with a role for tonic adenosine in the regulation of the intrinsic excitability of these neurons. Conclusions: Collectively, these data suggest that adenosine, via activation of A2A receptors, may directly facilitate basolateral amygdala pyramidal cell output, providing a possible balance for the recently described inhibitory effects of adenosine A1 receptor activation on glutamatergic excitation of basolateral amygdala pyramidal cells. PMID:25716780

  12. Association between A2a receptor gene polymorphisms and caffeine-induced anxiety.

    PubMed

    Alsene, Karen; Deckert, Jürgen; Sand, Philipp; de Wit, Harriet

    2003-09-01

    The adenosine receptor system, which mediates the psychoactive effects of caffeine, is also thought to be involved in the regulation of anxiety. In this study, we examined the association between variations in anxiogenic responses to caffeine and polymorphisms in the A1 and A2a adenosine receptor genes. Healthy, infrequent caffeine users (N=94) recorded their subjective mood states following a 150 mg oral dose of caffeine freebase or placebo in a double-blind study. We found a significant association between self-reported anxiety after caffeine administration and two linked polymorphisms on the A2a receptor gene, the 1976C>T and 2592C>Tins polymorphisms. Individuals with the 1976T/T and the 2592Tins/Tins genotypes reported greater increases in anxiety after caffeine administration than the other genotypic groups. The study shows that an adenosine receptor gene polymorphism that has been associated with Panic Disorder is also associated with anxiogenic responses to an acute dose of caffeine.

  13. High salt diet exacerbates vascular contraction in the absence of adenosine A2A receptor

    PubMed Central

    Pradhan, Isha; Zeldin, Darryl C.; Ledent, Catherine; Mustafa, S. Jamal; Falck, John R.; Nayeem, Mohammed A

    2014-01-01

    High salt (4%NaCl, HS) diet modulates adenosine-induced vascular response through adenosine A2A-receptor (A2AAR). Evidence suggests A2AAR stimulates cyp450-epoxygenases, leading to epoxyeicosatrienoic acids (EETs) generation. The aim of this study was to understand the vascular reactivity to HS and underlying signaling mechanism in the presence or absence of A2AAR. Therefore, we hypothesized that HS enhances adenosine-induced relaxation through EETs in A2AAR+/+, but exaggerates contraction in A2AAR−/−. Organ-bath and Western-blot experiments were conducted in HS and normal salt (NS, 0.18% NaCl)-fed A2AAR+/+ and A2AAR−/− mice aortae. HS produced concentration-dependent relaxation to non-selective adenosine analog, NECA in A2AAR+/+, whereas contraction was observed in A2AAR−/− mice and this was attenuated by A1AR antagonist (DPCPX). CGS-21680 (selective A2AAR-agonist) enhanced relaxation in HS-A2AAR+/+ vs. NS-A2AAR+/+, that was blocked by EETs antagonist (14,15-EEZE). Compared to NS, HS significantly upregulated expression of vasodilators A2AAR and cyp2c29, while vasoconstrictors A1AR and cyp4a in A2AAR+/+ were downregulated. In A2AAR−/− mice, however, HS significantly downregulated the expression of cyp2c29, while A1AR and cyp4a were upregulated compared to A2AAR+/+ mice. Hence, our data suggest that in A2AAR+/+, HS enhances A2AAR-induced relaxation through increased cyp-expoxygenases-derived EETs and decreased A1AR levels, whereas in A2AAR−/−, HS exaggerates contraction through decreased cyp-epoxygenases and increased A1AR levels. PMID:24390173

  14. S = 1/2 Square-Lattice Antiferromagnets: (CuX)LaB_2O_7 and (CuCl)A_2B_3O10 (X = Cl, Br; A = Ca, Sr; B = Nb, Ta)

    NASA Astrophysics Data System (ADS)

    Kageyama, H.; Kitano, T.; Nakanishi, R.; Yasuda, J.; Oba, N.; Baba, Y.; Nishi, M.; Ueda, Y.; Ajiro, Y.; Yoshimura, K.

    A series of magnetic compounds with the formula (CuX)LaB_2O_7 and (CuCl)A_2B_3O10 (X = Cl, Br; A = Ca, Sr; B = Nb, Ta) have been prepared through a low-temperature topochemical route starting from nonmagnetic double- (n = 2) and triple- (n = 3) layered perovskites, respectively. The magnetic susceptibility of these compounds typically exhibits a broad maximum at low temperatures, characteristic of low-dimensional antiferromagnetic compounds. However, depending on the choice of the parameters, X, A, B and n, physical quantities such as the Weiss temperature and the temperature at a maximum susceptibility vary to a great extent, which enables us to study the phase diagram of the S = 1/2 frustrated square-lattice antiferromagnets (the so-called J_1-J_2 model). In particular, (CuCl)LaNb_2O_7, possibly having a ferromagnetic J_1 and an antiferromagnetic J_2, shows a spin-liquid behavior with the spin gap of 27 K.

  15. A1C Test

    MedlinePlus

    ... AACC products and services. Advertising & Sponsorship: Policy | Opportunities Hemoglobin A1c Share this page: Was this page helpful? Also known as: A1c; HbA1c; Glycohemoglobin; Glycated Hemoglobin; Glycosylated Hemoglobin Formal name: Hemoglobin A1c Related tests: ...

  16. Allosteric interactions at adenosine A1 and A3 receptors: new insights into the role of small molecules and receptor dimerization

    PubMed Central

    Hill, Stephen J; May, Lauren T; Kellam, Barrie; Woolard, Jeanette

    2014-01-01

    The purine nucleoside adenosine is present in all cells in tightly regulated concentrations. It is released under a variety of physiological and pathophysiological conditions to facilitate protection and regeneration of tissues. Adenosine acts via specific GPCRs to either stimulate cyclic AMP formation, as exemplified by Gs-protein-coupled adenosine receptors (A2A and A2B), or inhibit AC activity, in the case of Gi/o-coupled adenosine receptors (A1 and A3). Recent advances in our understanding of GPCR structure have provided insights into the conformational changes that occur during receptor activation following binding of agonists to orthosteric (i.e. at the same binding site as an endogenous modulator) and allosteric regulators to allosteric sites (i.e. at a site that is topographically distinct from the endogenous modulator). Binding of drugs to allosteric sites may lead to changes in affinity or efficacy, and affords considerable potential for increased selectivity in new drug development. Herein, we provide an overview of the properties of selective allosteric regulators of the adenosine A1 and A3 receptors, focusing on the impact of receptor dimerization, mechanistic approaches to single-cell ligand-binding kinetics and the effects of A1- and A3-receptor allosteric modulators on in vivo pharmacology. Linked ArticlesThis article is part of a themed section on Molecular Pharmacology of GPCRs. To view the other articles in this section visit http://dx.doi.org/10.1111/bph.2014.171.issue-5 PMID:24024783

  17. Lidocaine relaxation in isolated rat aortic rings is enhanced by endothelial removal: possible role of Kv, KATP channels and A2a receptor crosstalk.

    PubMed

    Arsyad, Aryadi; Dobson, Geoffrey P

    2016-12-03

    Lidocaine is an approved local anesthetic and Class 1B antiarrhythmic with a number of ancillary properties. Our aim was to investigate lidocaine's vasoreactivity properties in intact versus denuded rat thoracic aortic rings, and the effect of inhibitors of nitric oxide (NO), prostenoids, voltage-dependent Kv and KATP channels, membrane Na(+)/K(+) pump, and A2a and A2b receptors. Aortic rings were harvested from adult male Sprague Dawley rats and equilibrated in an organ bath containing oxygenated, modified Krebs-Henseleit solution, pH 7.4, 37 °C. The rings were pre-contracted sub-maximally with 0.3 μM norepinephrine (NE), and the effect of increasing lidocaine concentrations was examined. Rings were tested for viability after each experiment with maximally dilating 100 μM papaverine. The drugs 4-aminopyridine (4-AP), glibenclamide, 5-hydroxydecanoate, ouabain, 8-(3-chlorostyryl) caffeine and PSB-0788 were examined. All drugs tested had no significant effect on basal tension. Lidocaine relaxation in intact rings was biphasic between 1 and 10 μM (Phase 1) and 10 and 1000 μM (Phase 2). Mechanical removal of the endothelium resulted in further relaxation, and at lower concentrations ring sensitivity (% relaxation per μM lidocaine) significantly increased 3.5 times compared to intact rings. The relaxing factor(s) responsible for enhancing ring relaxation did not appear to be NO- or prostacyclin-dependent, as L-NAME and indomethacin had little or no effect on intact ring relaxation. In denuded rings, lidocaine relaxation was completely abolished by Kv channel inhibition and significantly reduced by antagonists of the MitoKATP channel, and to a lesser extent the SarcKATP channel. Curiously, A2a subtype receptor antagonism significantly inhibited lidocaine relaxation above 100 μM, but not the A2b receptor. We show that lidocaine relaxation in rat thoracic aorta was biphasic and significantly enhanced by endothelial removal, which did not appear to be NO or

  18. 2-Aralkynyl and 2-heteroalkynyl derivatives of adenosine-5'-N-ethyluronamide as selective A2a adenosine receptor agonists.

    PubMed

    Cristalli, G; Camaioni, E; Vittori, S; Volpini, R; Borea, P A; Conti, A; Dionisotti, S; Ongini, E; Monopoli, A

    1995-04-28

    A series of new 2-aralkynyl and 2-heteroaralkynyl derivatives of NECA were synthesized and studied in binding and functional assays to assess their potency for the A2a compared to A1 adenosine receptors. Compounds bearing an aromatic or heteroaromatic ring, conjugated to the triple bond, showed generally weaker activity at the A2a receptor and lower selectivity (A2a vs A1) than the alkylakynyl derivatives previously reported. However, the (4-formylphenyl)-ethynyl derivative 17 showed affinity in the low nanomolar range and high selectivity (about 160-fold) for the A2a receptor. The presence of heteroatoms improved vasorelaxant activity, the 2-thiazolylethynyl derivative 30 being the most potent in the series. Introduction of methylene groups between the triple bond and the phenyl ring favored the A2a binding affinity, and the 5-phenyl-1-pentynyl derivative 24 was found to be highly potent and selective (about 180-fold) at A2a receptors. With regard to antiplatelet activity, the presence of aromatic or heteroaromatic rings decreased potency in comparison with that of NECA and of N-ethyl-1'-deoxy-1'-(6-amino-2-hexynyl-9H-purin-9-yl)-beta-D-ribofura nuronamide (HENECA). Introduction of a methylene group was effective in increasing antiaggregatory potency only when this group is linked to a heteroatom (31-35). From these data and those previously reported, the structure-activity relationships derived for the 2-alkynyl-substituted ribose uronamides would indicate that potentiation of A2a receptor affinity could be obtained by aromatic rings not conjugated with the triple bond or by heteroaromatic groups. As for A2a receptors on platelets, the presence of aromatic rings, either conjugated or unconjugated to the triple bond, is detrimental for the antiaggregatory activity. However, the introduction of polar groups alpha to the triple bond strongly increases the potency when steric hindrance is avoided. Some of the compounds included in this series retain interesting

  19. Pharmacological evidence for different populations of postsynaptic adenosine A2A receptors in the rat striatum

    PubMed Central

    Orrú, Marco; Quiroz, César; Guitart, Xavier; Ferré, Sergi

    2011-01-01

    Adenosine A2A receptors (A2ARs) are highly concentrated in the striatum. Two pharmacological different functional populations of A2ARs have been recently described based on their different affinities for the A2AR antagonist SCH-442416. This compound has high affinity for A2ARs not forming heteromers or forming heteromers with adenosine A1 receptors (A1Rs) while showing very low affinity for A2ARs forming heteromers with dopamine D2 receptors (D2Rs). It has been widely described that striatal A1R-A2AR heteromers are preferentially localized presynaptically in the glutamatergic terminals that contact GABAergic dynorphinergic neurons, and that A2AR-D2R heteromers are localized postsynaptically in GABAergic enkephalinergic neurons. In the present study we provide evidence suggesting that SCH-442416 also targets postsynaptic A2AR not forming heteromers with D2R, which are involved in the motor depressant effects induced by D2R antagonists. SCH-442416 counteracted motor depression in rats induced by the D2R antagonist raclopride at a dose that did not produce motor activation or that blocked motor depression induced by an A2AR agonist. Furthermore, we re-evaluated the recently suggested key role of cannabinoid CB1 receptors (CB1Rs) in the effects of A2AR antagonists acting at postsynaptic A2ARs. By recording locomotor activity and monitoring striatal glutamate release induced by cortical electrical stimulation in rats after administration of A2AR and CB1R antagonists, we did not find evidence for any significant role of endocannabinoids in the post- or presynaptic effects of A2AR antagonists. The present results further suggest the existence of at least two functionally and pharmacologically different populations of striatal postsynaptic A2ARs. PMID:21752341

  20. Caffeine and adenosine A(2a) receptor antagonists prevent beta-amyloid (25-35)-induced cognitive deficits in mice.

    PubMed

    Dall'Igna, Oscar P; Fett, Paulo; Gomes, Marcio W; Souza, Diogo O; Cunha, Rodrigo A; Lara, Diogo R

    2007-01-01

    Consumption of caffeine, an adenosine receptor antagonist, was found to be inversely associated with the incidence of Alzheimer's disease. Moreover, caffeine protects cultured neurons against beta-amyloid-induced toxicity, an effect mimicked by adenosine A(2A) but not A(1) receptor antagonists. We now tested if caffeine administration would prevent beta-amyloid-induced cognitive impairment in mice and if this was mimicked by A(2A) receptor blockade. One week after icv administration of the 25-35 fragment of beta-amyloid (Abeta, 3 nmol), mice displayed impaired performance in both inhibitory avoidance and spontaneous alternation tests. Prolonged treatment with caffeine (1 mg/ml) had no effect alone but prevented the Abeta-induced cognitive impairment in both tasks when associated with acute caffeine (30 mg/kg) 30 min treatment before Abeta administration. The same protective effect was observed after subchronic (4 days) treatment with daily injections of either caffeine (30 mg/kg) or the selective adenosine A(2A) receptor antagonist SCH58261 (0.5 mg/kg). This provides the first direct in vivo evidence that caffeine and A(2A) receptor antagonists afford a protection against Abeta-induced amnesia, which prompts their interest for managing Alzheimer's disease.

  1. Reduced striatal adenosine A2A receptor levels define a molecular subgroup in schizophrenia.

    PubMed

    Villar-Menéndez, Izaskun; Díaz-Sánchez, Sara; Blanch, Marta; Albasanz, José Luis; Pereira-Veiga, Thais; Monje, Alfonso; Planchat, Luis Maria; Ferrer, Isidre; Martín, Mairena; Barrachina, Marta

    2014-04-01

    Schizophrenia (SZ) is a mental disorder of unknown origin. Some scientific evidence seems to indicate that SZ is not a single disease entity, since there are patient groups with clear symptomatic, course and biomarker differences. SZ is characterized by a hyperdopaminergic state related to high dopamine D2 receptor activity. It has also been proposed that there is a hypoadenosynergic state. Adenosine is a nucleoside widely distributed in the organism with neuromodulative and neuroprotective activity in the central nervous system. In the brain, the most abundant adenosine receptors are A1R and A2AR. In the present report, we characterize the presence of both receptors in human postmortem putamens of patients suffering SZ with real time TaqMan PCR, western blotting and radioligand binding assay. We show that A1R levels remain unchanged with respect to age-matched controls, whereas nearly fifty percent of patients have reduced A2AR, at the transcriptional and translational levels. Moreover, we describe how DNA methylation plays a role in the pathological A2AR levels with the bisulfite-sequencing technique. In fact, an increase in 5-methylcytosine percentage in the 5' UTR region of ADORA2A was found in those SZ patients with reduced A2AR levels. Interestingly, there was a relationship between the A2A/β-actin ratio and motor disturbances as assessed with some items of the PANSS, AIMS and SAS scales. Therefore, there may be a subgroup of SZ patients with reduced striatal A2AR levels accompanied by an altered motor phenotype.

  2. Functions, dysfunctions and possible therapeutic relevance of adenosine A2A receptors in Huntington's disease.

    PubMed

    Popoli, Patrizia; Blum, David; Martire, Alberto; Ledent, Catherine; Ceruti, Stefania; Abbracchio, Maria P

    2007-04-01

    The aim of this review is to summarize and critically discuss the complex role played by adenosine A(2A) receptors (A(2A)Rs) in Huntington's disease (HD). Since A(2A)Rs are mainly localized on the neurons, which degenerate early in HD, and given their ability to stimulate glutamate outflow and inflammatory gliosis, it was hypothesized that they could be involved in the pathogenesis of HD, and that A(2A)R antagonists could be neuroprotective. This was further sustained by the demonstration that A(2A)Rs and underlying signaling systems undergo profound changes in cellular and animal models of HD. More recently, however, the equation A(2A) receptor blockade=neuroprotection has appeared too simplistic. First, it is now definitely clear that, besides mediating 'bad' responses (for example, stimulation of glutamate outflow and excessive glial activation), A(2A)Rs also promote 'good' responses (such as trophic and antinflammatory effects). This implies that A(2A)R blockade results either in pro-toxic or neuroprotective effects according to the mechanisms involved in a given experimental model. Second, since HD is a chronically progressive disease, the multiple mechanisms involving A(2A)Rs may play different relative roles along the degenerative process. Such different mechanisms can be influenced by A(2A)R activation or blockade in different ways, even leading to opposite outcomes depending on the time of agonist/antagonist administration. The number, and the complexity, of the possible scenarios is further increased by the influence of mutant Huntingtin on both the expression and functions of A(2A)Rs, and by the strikingly different effects mediated by A(2A)Rs expressed by different cell populations within the brain.

  3. A1C

    MedlinePlus

    A1C is a blood test for type 2 diabetes and prediabetes. It measures your average blood glucose, or blood sugar, level over the past 3 ... A1C alone or in combination with other diabetes tests to make a diagnosis. They also use the ...

  4. High-salt diet enhances mouse aortic relaxation through adenosine A2A receptor via CYP epoxygenases.

    PubMed

    Nayeem, Mohammed A; Ponnoth, Dovenia S; Boegehold, Matthew A; Zeldin, Darryl C; Falck, John R; Mustafa, S Jamal

    2009-03-01

    We hypothesize that A(2A) adenosine receptors (A(2A) AR) promote aortic relaxation in mice through cytochrome P450 (CYP)-epoxygenases and help to avoid salt sensitivity. Aortas from male mice maintained on a high-salt (HS; 7% NaCl) or normal-salt (NS; 0.45% NaCl) diet for 4-5 wks were used. Concentration-response curves (10(-11)-10(-5) M) for 5'-N-ethylcarboxamidoadenosine (NECA; a nonselective adenosine analog) and CGS 21680 (A(2A) AR agonist) were obtained with different antagonists including ZM 241385 (A(2A) AR antagonist; 10(-6) M), SCH 58261 (A(2A) AR antagonist; 10(-6) M), N(omega)-nitro-l-arginine methyl ester (l-NAME; endothelial nitric oxide synthase inhibitor; 10(-4) M) and inhibitors including methylsulfonyl-propargyloxyphenylhexanamide (MS-PPOH; CYP epoxygenases inhibitor; 10(-5)M), 14,15-epoxyeicosa-5(z)-enoic acid (14,15-EEZE; EET antagonist; 10(-5)M), dibromo-dodecenyl-methylsulfimide (DDMS; CYP4A inhibitor; 10(-5)M), and HET0016 (20-HETE inhibitor; 10(-5)M). At 10(-7) M of NECA, significant relaxation in HS (+22.58 +/- 3.12%) was observed compared with contraction in NS (-10.62 +/- 6.27%, P < 0.05). ZM 241385 changed the NECA response to contraction (P < 0.05) in HS. At 10(-7) M of CGS 21680, significant relaxation in HS (+32.04 +/- 3.08%) was observed compared with NS (+10.45 +/- 1.34%, P < 0.05). SCH 58261, l-NAME, MS-PPOH, and 14,15-EEZE changed the CGS 21680-induced relaxation to contraction (P < 0.05) in HS. Interestingly, DDMS and HET0016 changed CGS 21680 response to relaxation (P < 0.05) in NS; however, there was no significant difference found between DDMS, HET0016-treated HS and NS vs. nontreated HS group (P > 0.05). CYP2C29 protein was 55% and 74% upregulated in HS vs. NS (P < 0.05) mice aorta and kidney, respectively. CYP4A protein was 30.30% and 35.70% upregulated in NS vs. HS (P < 0.05) mice aorta and kidneys, respectively. A(1) AR was downregulated, whereas A(2A) AR was upregulated in HS compared with NS. These data suggest that HS

  5. High-salt diet enhances mouse aortic relaxation through adenosine A2A receptor via CYP epoxygenases

    PubMed Central

    Nayeem, Mohammed A.; Ponnoth, Dovenia S.; Boegehold, Matthew A.; Zeldin, Darryl C.; Falck, John R.; Mustafa, S. Jamal

    2009-01-01

    We hypothesize that A2A adenosine receptors (A2A AR) promote aortic relaxation in mice through cytochrome P450 (CYP)-epoxygenases and help to avoid salt sensitivity. Aortas from male mice maintained on a high-salt (HS; 7% NaCl) or normal-salt (NS; 0.45% NaCl) diet for 4–5 wks were used. Concentration-response curves (10−11–10−5 M) for 5′-N-ethylcarboxamidoadenosine (NECA; a nonselective adenosine analog) and CGS 21680 (A2A AR agonist) were obtained with different antagonists including ZM 241385 (A2A AR antagonist; 10−6 M), SCH 58261 (A2A AR antagonist; 10−6 M), Nω-nitro-l-arginine methyl ester (l-NAME; endothelial nitric oxide synthase inhibitor; 10−4 M) and inhibitors including methylsulfonyl-propargyloxyphenylhexanamide (MS-PPOH; CYP epoxygenases inhibitor; 10−5M), 14,15-epoxyeicosa-5(z)-enoic acid (14,15-EEZE; EET antagonist; 10−5M), dibromo-dodecenyl-methylsulfimide (DDMS; CYP4A inhibitor; 10−5M), and HET0016 (20-HETE inhibitor; 10−5M). At 10−7 M of NECA, significant relaxation in HS (+22.58 ± 3.12%) was observed compared with contraction in NS (−10.62 ± 6.27%, P < 0.05). ZM 241385 changed the NECA response to contraction (P < 0.05) in HS. At 10−7 M of CGS 21680, significant relaxation in HS (+32.04 ± 3.08%) was observed compared with NS (+10.45 ± 1.34%, P < 0.05). SCH 58261, l-NAME, MS-PPOH, and 14,15-EEZE changed the CGS 21680-induced relaxation to contraction (P < 0.05) in HS. Interestingly, DDMS and HET0016 changed CGS 21680 response to relaxation (P < 0.05) in NS; however, there was no significant difference found between DDMS, HET0016-treated HS and NS vs. nontreated HS group (P > 0.05). CYP2C29 protein was 55% and 74% upregulated in HS vs. NS (P < 0.05) mice aorta and kidney, respectively. CYP4A protein was 30.30% and 35.70% upregulated in NS vs. HS (P < 0.05) mice aorta and kidneys, respectively. A1 AR was downregulated, whereas A2A AR was upregulated in HS compared with NS. These data suggest that HS may activate CYP2

  6. Modulatory effect of the 5-HT1A agonist buspirone and the mixed non-hallucinogenic 5-HT1A/2A agonist ergotamine on psilocybin-induced psychedelic experience.

    PubMed

    Pokorny, Thomas; Preller, Katrin H; Kraehenmann, Rainer; Vollenweider, Franz X

    2016-04-01

    The mixed serotonin (5-HT) 1A/2A/2B/2C/6/7 receptor agonist psilocybin dose-dependently induces an altered state of consciousness (ASC) that is characterized by changes in sensory perception, mood, thought, and the sense of self. The psychological effects of psilocybin are primarily mediated by 5-HT2A receptor activation. However, accumulating evidence suggests that 5-HT1A or an interaction between 5-HT1A and 5-HT2A receptors may contribute to the overall effects of psilocybin. Therefore, we used a double-blind, counterbalanced, within-subject design to investigate the modulatory effects of the partial 5-HT1A agonist buspirone (20mg p.o.) and the non-hallucinogenic 5-HT2A/1A agonist ergotamine (3mg p.o.) on psilocybin-induced (170 µg/kg p.o.) psychological effects in two groups (n=19, n=17) of healthy human subjects. Psychological effects were assessed using the Altered State of Consciousness (5D-ASC) rating scale. Buspirone significantly reduced the 5D-ASC main scale score for Visionary Restructuralization (VR) (p<0.001), which was mostly driven by a reduction of the VR item cluster scores for elementary and complex visual hallucinations. Further, buspirone also reduced the main scale score for Oceanic Boundlessness (OB) including derealisation and depersonalisation phenomena at a trend level (p=0.062), whereas ergotamine did not show any effects on the psilocybin-induced 5D-ASC main scale scores. The present finding demonstrates that buspirone exerts inhibitory effects on psilocybin-induced effects, presumably via 5-HT1A receptor activation, an interaction between 5-HT1A and 5-HT2A receptors, or both. The data suggest that the modulation of 5-HT1A receptor activity may be a useful target in the treatment of visual hallucinations in different psychiatric and neurological diseases. Copyright © 2016 Elsevier B.V. and ECNP. All rights reserved.

  7. Adenosine A2A receptor antagonism and neuroprotection: mechanisms, lights, and shadows.

    PubMed

    Popoli, Patrizia; Minghetti, Luisa; Tebano, Maria Teresa; Pintor, Annita; Domenici, Maria Rosaria; Massotti, Marino

    2004-01-01

    Adenosine A2A receptor antagonists are regarded as potential neuroprotective drugs, although the mechanisms underlying their effects remain to be elucidated. In this review, quinolinic acid (QA)-induced striatal toxicity was used as a tool to investigate the mechanisms of the neuroprotective effects of A2A receptor antagonists. After having examined the effects of selective A2A receptor antagonists toward different mechanisms of QA toxicity, we conclude that (1) the effect elicited by A2A receptor blockade on QA-induced glutamate outflow may be one of the mechanisms of the neuroprotective activity of A2A receptor antagonists; (2) A2A receptor antagonists have a potentially worsening influence on QA-dependent NMDA receptor activation; and (3) the ability of A2A receptor antagonists to prevent QA-induced lipid peroxidation does not correlate with the neuroprotective effects. These results suggest that A2A receptor antagonists may have either potentially beneficial or detrimental influence in models of neurodegeneration that are mainly due to increased glutamate levels or enhanced sensitivity of NMDA receptors, respectively.

  8. Adenosine A2a receptors and O2 sensing in development

    PubMed Central

    2011-01-01

    Reduced mitochondrial oxidative phosphorylation, via activation of adenylate kinase and the resulting exponential rise in the cellular AMP/ATP ratio, appears to be a critical factor underlying O2 sensing in many chemoreceptive tissues in mammals. The elevated AMP/ATP ratio, in turn, activates key enzymes that are involved in physiologic adjustments that tend to balance ATP supply and demand. An example is the conversion of AMP to adenosine via 5′-nucleotidase and the resulting activation of adenosine A2A receptors, which are involved in acute oxygen sensing by both carotid bodies and the brain. In fetal sheep, A2A receptors associated with carotid bodies trigger hypoxic cardiovascular chemoreflexes, while central A2A receptors mediate hypoxic inhibition of breathing and rapid eye movements. A2A receptors are also involved in hypoxic regulation of fetal endocrine systems, metabolism, and vascular tone. In developing lambs, A2A receptors play virtually no role in O2 sensing by the carotid bodies, but brain A2A receptors remain critically involved in the roll-off ventilatory response to hypoxia. In adult mammals, A2A receptors have been implicated in O2 sensing by carotid glomus cells, while central A2A receptors likely blunt hypoxic hyperventilation. In conclusion, A2A receptors are crucially involved in the transduction mechanisms of O2 sensing in fetal carotid bodies and brains. Postnatally, central A2A receptors remain key mediators of hypoxic respiratory depression, but they are less critical for O2 sensing in carotid chemoreceptors, particularly in developing lambs. PMID:21677265

  9. Striatal adenosine-cannabinoid receptor interactions in rats over-expressing adenosine A2A receptors.

    PubMed

    Chiodi, Valentina; Ferrante, Antonella; Ferraro, Luca; Potenza, Rosa Luisa; Armida, Monica; Beggiato, Sarah; Pèzzola, Antonella; Bader, Michael; Fuxe, Kjell; Popoli, Patrizia; Domenici, Maria Rosaria

    2016-03-01

    Adenosine A2A receptors (A2 A Rs) and cannabinoid CB1 receptors (CB1 Rs) are highly expressed in the striatum, where they functionally interact and form A2A /CB1 heteroreceptor complexes. We investigated the effects of CB1 R stimulation in a transgenic rat strain over-expressing A2 A Rs under the control of the neural-specific enolase promoter (NSEA2A rats) and in age-matched wild-type (WT) animals. The effects of the CB1 R agonist WIN 55,212-2 (WIN) were significantly lower in NSEA2A rats than in WT animals, as demonstrated by i) electrophysiological recordings of synaptic transmission in corticostriatal slices; ii) the measurement of glutamate outflow from striatal synaptosomes and iii) in vivo experiments on locomotor activity. Moreover, while the effects of WIN were modulated by both A2 A R agonist (CGS 21680) and antagonists (ZM 241385, KW-6002 and SCH-442416) in WT animals, the A2 A R antagonists failed to influence WIN-mediated effects in NSEA2A rats. The present results demonstrate that in rats with genetic neuronal over-expression of A2 A Rs, the effects mediated by CB1 R activation in the striatum are significantly reduced, suggesting a change in the stoichiometry of A2A and CB1 receptors and providing a strategy to dissect the involvement of A2 A R forming or not forming heteromers in the modulation of striatal functions. These findings add additional evidence for the existence of an interaction between striatal A2 A Rs and CB1 Rs, playing a fundamental role in the regulation of striatal functions. We studied A2A -CB1 receptor interaction in transgenic rats over-expressing adenosine A2A receptors under the control of the neuron-specific enolase promoter (NSEA2A ). In these rats, we demonstrated a reduced effect of the CB1 receptor agonist WIN 55,212-2 in the modulation of corticostriatal synaptic transmission and locomotor activity, while CB1 receptor expression level did not change with respect to WT rats. A reduction in the expression of A2A -CB1

  10. Adenosine A2A Receptors in the Amygdala Control Synaptic Plasticity and Contextual Fear Memory.

    PubMed

    Simões, Ana Patrícia; Machado, Nuno J; Gonçalves, Nélio; Kaster, Manuella P; Simões, Ana T; Nunes, Ana; Pereira de Almeida, Luís; Goosens, Ki Ann; Rial, Daniel; Cunha, Rodrigo A

    2016-11-01

    The consumption of caffeine modulates working and reference memory through the antagonism of adenosine A2A receptors (A2ARs) controlling synaptic plasticity processes in hippocampal excitatory synapses. Fear memory essentially involves plastic changes in amygdala circuits. However, it is unknown if A2ARs in the amygdala regulate synaptic plasticity and fear memory. We report that A2ARs in the amygdala are enriched in synapses and located to glutamatergic synapses, where they selectively control synaptic plasticity rather than synaptic transmission at a major afferent pathway to the amygdala. Notably, the downregulation of A2ARs selectively in the basolateral complex of the amygdala, using a lentivirus with a silencing shRNA (small hairpin RNA targeting A2AR (shA2AR)), impaired fear acquisition as well as Pavlovian fear retrieval. This is probably associated with the upregulation and gain of function of A2ARs in the amygdala after fear acquisition. The importance of A2ARs to control fear memory was further confirmed by the ability of SCH58261 (0.1 mg/kg; A2AR antagonist), caffeine (5 mg/kg), but not DPCPX (0.5 mg/kg; A1R antagonist), treatment for 7 days before fear conditioning onwards, to attenuate the retrieval of context fear after 24-48 h and after 7-8 days. These results demonstrate that amygdala A2ARs control fear memory and the underlying process of synaptic plasticity in this brain region. This provides a neurophysiological basis for the association between A2AR polymorphisms and phobia or panic attacks in humans and prompts a therapeutic interest in A2ARs to manage fear-related pathologies.

  11. GABAergic involvement in motor effects of an adenosine A(2A) receptor agonist in mice.

    PubMed

    Khisti, R T; Chopde, C T; Abraham, E

    2000-04-03

    Adenosine A(2A) agonists are known to induce catalepsy and inhibit dopamine mediated motor hyperactivity. An antagonistic interaction between adenosine A(2A) and dopamine D(2) receptors is known to regulate GABA-mediated neurotransmission in striatopallidal neurons. Stimulation of adenosine A(2A) and dopamine D(2) receptors has been shown to increase and inhibit GABA release respectively in pallidal GABAergic neurons. However, the role of GABAergic neurotransmission in the motor effects of adenosine A(2A) receptors is not yet known. Therefore in the present study the effect of GABAergic agents on adenosine A(2A) receptor agonist (NECA- or CGS 21680) induced catalepsy and inhibition of amphetamine elicited motor hyperactivity was examined. Pretreatment with GABA, the GABA(A) agonist muscimol or the GABA(B) agonist baclofen potentiated whereas the GABA(A) antagonist bicuculline attenuated NECA- or CGS 21680-induced catalepsy. However, the GABA(B) antagonists phaclophen and delta-aminovaleric acid had no effect. Administration of NECA or CGS 21680 not only reduced spontaneous locomotor activity but also antagonized amphetamine elicited motor hyperactivity. These effects of NECA and CGS 21680 were potentiated by GABA or muscimol and antagonized by bicuculline. These findings provide behavioral evidence for the role of GABA in the motor effects of adenosine A(2A) receptor agonists. Activation of adenosine A(2A) receptors increases GABA release which could reduce dopaminergic tone and induce catalepsy or inhibit amphetamine mediated motor hyperactivity.

  12. Neuroprotection by adenosine A2A receptor blockade in experimental models of Parkinson's disease.

    PubMed

    Ikeda, Ken; Kurokawa, Masako; Aoyama, Shiro; Kuwana, Yoshihisa

    2002-01-01

    Adenosine A2A receptors are abundant in the caudate-putamen and involved in the motor control in several species. In MPTP-treated monkeys, A2A receptor-blockade with an antagonist alleviates parkinsonian symptoms without provoking dyskinesia, suggesting this receptor may offer a new target for the antisymptomatic therapy of Parkinson's disease. In the present study, a significant neuroprotective effect of A2A receptor antagonists is shown in experimental models of Parkinson's disease. Oral administration of A2A receptor antagonists protected against the loss of nigral dopaminergic neuronal cells induced by 6-hydroxydopamine in rats. A2A antagonists also prevented the functional loss of dopaminergic nerve terminals in the striatum and the ensuing gliosis caused by MPTP in mice. The neuroprotective property of A2A receptor antagonists may be exerted by altering the packaging of these neurotoxins into vesicles, thus reducing their effective intracellular concentration. We therefore conclude that the adenosine A2A receptor may provide a novel target for the long-term medication of Parkinson's disease, because blockade of this receptor exerts both acutely antisymptomatic and chronically neuroprotective activities.

  13. Structure of an agonist-bound human A2A adenosine receptor.

    PubMed

    Xu, Fei; Wu, Huixian; Katritch, Vsevolod; Han, Gye Won; Jacobson, Kenneth A; Gao, Zhan-Guo; Cherezov, Vadim; Stevens, Raymond C

    2011-04-15

    Activation of G protein-coupled receptors upon agonist binding is a critical step in the signaling cascade for this family of cell surface proteins. We report the crystal structure of the A(2A) adenosine receptor (A(2A)AR) bound to an agonist UK-432097 at 2.7 angstrom resolution. Relative to inactive, antagonist-bound A(2A)AR, the agonist-bound structure displays an outward tilt and rotation of the cytoplasmic half of helix VI, a movement of helix V, and an axial shift of helix III, resembling the changes associated with the active-state opsin structure. Additionally, a seesaw movement of helix VII and a shift of extracellular loop 3 are likely specific to A(2A)AR and its ligand. The results define the molecule UK-432097 as a "conformationally selective agonist" capable of receptor stabilization in a specific active-state configuration.

  14. Capable Reader Program: Lesson Plan Guide. Units A1; A2; A3; A4; [and] A5. Pilot Year 1979-1980, Final Edition 1980-1981.

    ERIC Educational Resources Information Center

    Casper, Donna; And Others

    Part of a curriculum series for academically gifted elementary students in the area of reading, the five lesson plan guides are intended to provide teachers with suggested activities stressing high levels of reading comprehension as well as encouraging teachers to use their own ideas. Each guide focuses on one of the following major objectives:…

  15. A dispersed fluorescence and ab initio investigation of the X~ 2B1 and A~ 2A1 electronic states of the PH2 molecule

    NASA Astrophysics Data System (ADS)

    Jakubek, Z. J.; Bunker, P. R.; Zachwieja, M.; Nakhate, S. G.; Simard, B.; Yurchenko, S. N.; Thiel, W.; Jensen, Per

    2006-03-01

    In this work, the X˜B12 and ÃA12 electronic states of the phosphino (PH2) free radical have been studied by dispersed fluorescence and ab initio methods. PH2 molecules were produced in a molecular free-jet apparatus by laser vaporizing a silicon rod in the presence of phosphine (PH3) gas diluted in helium. The laser-induced fluorescence, from the excited ÃA12 electronic state down to the ground electronic state, was dispersed and analyzed. Ten (υ1υ2υ3) vibrationally excited levels of the ground electronic state, with υ1⩽2, υ2⩽6, and υ3=0, have been observed. Ab initio potential-energy surfaces for the X˜B12 and ÃA12 electronic states have been calculated at 210 points. These two states correlate with a Πu2 state at linearity and they interact by the Renner-Teller coupling and spin-orbit coupling. Using the ab initio potential-energy surfaces with our RENNER computer program system, the vibronic structure and relative intensities of the ÃA12→X˜B12 emission band system have been calculated in order to corroborate the experimental assignments.

  16. Capable Reader Program: Lesson Plan Guide. Units A1; A2; A3; A4; [and] A5. Pilot Year 1979-1980, Final Edition 1980-1981.

    ERIC Educational Resources Information Center

    Casper, Donna; And Others

    Part of a curriculum series for academically gifted elementary students in the area of reading, the five lesson plan guides are intended to provide teachers with suggested activities stressing high levels of reading comprehension as well as encouraging teachers to use their own ideas. Each guide focuses on one of the following major objectives:…

  17. Adenosine A2A Receptor Blockade Prevents Rotenone-Induced Motor Impairment in a Rat Model of Parkinsonism

    PubMed Central

    Fathalla, Ahmed M.; Soliman, Amira M.; Ali, Mohamed H.; Moustafa, Ahmed A.

    2016-01-01

    Pharmacological studies implicate the blockade of adenosine receptorsas an effective strategy for reducing Parkinson’s disease (PD) symptoms. The objective of this study is to elucidate the possible protective effects of ZM241385 and 8-cyclopentyl-1, 3-dipropylxanthine, two selective A2A and A1 receptor antagonists, on a rotenone rat model of PD. Rats were split into four groups: vehicle control (1 ml/kg/48 h), rotenone (1.5 mg/kg/48 h, s.c.), ZM241385 (3.3 mg/kg/day, i.p) and 8-cyclopentyl-1, 3-dipropylxanthine (5 mg/kg/day, i.p). After that, animals were subjected to behavioral (stride length and grid walking) and biochemical (measuring concentration of dopamine levels using high performance liquid chromatography, HPLC). In the rotenone group, rats displayed a reduced motor activity and disturbed movement coordination in the behavioral tests and a decreased dopamine concentration as foundby HPLC. The effect of rotenone was partially prevented in the ZM241385 group, but not with 8-cyclopentyl-1,3-dipropylxanthine administration. The administration of ZM241385 improved motor function and movement coordination (partial increase of stride length and partial decrease in the number of foot slips) and an increase in dopamine concentration in the rotenone-injected rats. However, the 8-cyclopentyl-1,3-dipropylxanthine and rotenone groups were not significantly different. These results indicate that selective A2A receptor blockade by ZM241385, but not A1 receptor blockadeby 8-cyclopentyl-1,3-dipropylxanthine, may treat PD motor symptoms. This reinforces the potential use of A2A receptor antagonists as a treatment strategy for PD patients. PMID:26973484

  18. A critical evaluation of adenosine A2A receptors as potentially "druggable" targets in Huntington's disease.

    PubMed

    Popoli, Patrizia; Blum, David; Domenici, Maria Rosaria; Burnouf, Sylvie; Chern, Yijuang

    2008-01-01

    Huntington's disease (HD) is a dominantly inherited neurodegenerative disorder caused by the expansion of a polymorphic CAG trinucleotide repeat encoding a poly-glutamine tract within the Huntingtin protein. GABAergic enkephalin neurons of the basal ganglia, which show the highest levels of expression of adenosine A(2A) receptors, are the most vulnerable in HD. Such a selective neuronal vulnerability, which occurs despite ubiquitous expression of mutant and normal Huntingtin, has suggested that adenosine A(2A) receptors might play a pathogenetic role in HD. In agreement, changes in A(2A) receptor expression and signaling have been reported in various experimental models of HD. The interpretation of the functional significance of the aberrant A(2A) receptor phenotype in HD mice is however complicated by the conflicting data so far reported on the potential neuroprotective and neurodegenerative effects of these receptors in the brain, with some data suggesting a potential pathogenetic role and some other data suggesting activation of trophic or protective pathways in neurons. The same complex profile has emerged in experimental models of HD, in which both A(2A) receptor agonists and antagonists have shown beneficial effects. The main aim of this review is to critically evaluate whether adenosine A(2A) receptors may represent a suitable target to develop drugs against HD.

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

    PubMed Central

    Pedata, Felicita; Pugliese, Anna Maria; Coppi, Elisabetta; Dettori, Ilaria; Maraula, Giovanna; Cellai, Lucrezia; Melani, Alessia

    2014-01-01

    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. PMID:25165414

  20. A2aR antagonists: Next generation checkpoint blockade for cancer immunotherapy.

    PubMed

    Leone, Robert D; Lo, Ying-Chun; Powell, Jonathan D

    2015-01-01

    The last several years have witnessed exciting progress in the development of immunotherapy for the treatment of cancer. This has been due in great part to the development of so-called checkpoint blockade. That is, antibodies that block inhibitory receptors such as CTLA-4 and PD-1 and thus unleash antigen-specific immune responses against tumors. It is clear that tumors evade the immune response by usurping pathways that play a role in negatively regulating normal immune responses. In this regard, adenosine in the immune microenvironment leading to the activation of the A2a receptor has been shown to represent one such negative feedback loop. Indeed, the tumor microenvironment has relatively high concentrations of adenosine. To this end, blocking A2a receptor activation has the potential to markedly enhance anti-tumor immunity in mouse models. This review will present data demonstrating the ability of A2a receptor blockade to enhance tumor vaccines, checkpoint blockade and adoptive T cell therapy. Also, as several recent studies have demonstrated that under certain conditions A2a receptor blockade can enhance tumor progression, we will also explore the complexities of adenosine signaling in the immune response. Despite important nuances to the A2a receptor pathway that require further elucidation, studies to date strongly support the development of A2a receptor antagonists (some of which have already been tested in phase III clinical trials for Parkinson Disease) as novel modalities in the immunotherapy armamentarium.

  1. The A2a adenosine receptor modulates the reinforcement efficacy and neurotoxicity of MDMA.

    PubMed

    Ruiz-Medina, Jessica; Ledent, Catherine; Carretón, Olga; Valverde, Olga

    2011-04-01

    Adenosine is an endogenous purine nucleoside that plays a neuromodulatory role in the central nervous system. A2a adenosine receptors have been involved in reward-related processes, inflammatory phenomena and neurotoxicity reactions. In the present study, we investigated the role of A2a adenosine receptors on the acute pharmacological effects, reinforcement and neuroinflammation induced by MDMA administration. First, the acute effects of MDMA on body temperature, locomotor activity and anxiety-like responses were measured in A2a knockout mice and wild-type littermates. Second, MDMA reinforcing properties were evaluated using the intravenous self-administration paradigm. Finally, we assessed striatal astrogliosis and microgliosis as markers of MDMA neurotoxicity. Our results showed that acute MDMA produced a biphasic effect on body temperature and increased locomotor activity and anxiogenic-like responses in both genotypes. However, MDMA reinforcing properties were dramatically affected by the lack of A2a adenosine receptors. Thus, wild-type mice maintained MDMA self-administration under a fixed ratio 1 reinforcement schedule, whereas the operant response appeared completely abolished in A2a knockout mice. In addition, the MDMA neurotoxic regime produced an enhanced inflammatory response in striatum of wild-type mice, revealed by a significant increase in glial expression, whereas such activation was attenuated in mutant mice. This is the first report indicating that A2a adenosine receptors play a key role in reinforcement and neuroinflammation induced by the widely used psychostimulant.

  2. A-1 to Constellation

    NASA Technical Reports Server (NTRS)

    2006-01-01

    The A-1 Test Stand at NASA Stennis Space Center near Bay St. Louis, Miss., was the focus of a ceremony held Thursday to transition the storied facility to a new program of work: testing the J-2X engines that will power the agency's next generation spacecraft, Ares I & V. Standing before the historic structure, with a plaque commemorating the change, are (from left) SSC Center Director Richard Gilbrech; NASA Associate Administrator for Exploration Systems Scott Horowitz; and NASA Space Operations Deputy Associate Administrator for Program Integration Michael Hawes. Ares vehicles are the crew and cargo launch vehicles being developed under NASA's Constellation Program.

  3. A-1 to Constellation

    NASA Image and Video Library

    2006-11-09

    The A-1 Test Stand at NASA Stennis Space Center near Bay St. Louis, Miss., was the focus of a ceremony held Thursday to transition the storied facility to a new program of work: testing the J-2X engines that will power the agency's next generation spacecraft, Ares I & V. Standing before the historic structure, with a plaque commemorating the change, are (from left) SSC Center Director Richard Gilbrech; NASA Associate Administrator for Exploration Systems Scott Horowitz; and NASA Space Operations Deputy Associate Administrator for Program Integration Michael Hawes. Ares vehicles are the crew and cargo launch vehicles being developed under NASA's Constellation Program.

  4. Remodeling of striatal NMDA receptors by chronic A(2A) receptor blockade in Huntington's disease mice.

    PubMed

    Martire, Alberto; Ferrante, Antonella; Potenza, Rosa Luisa; Armida, Monica; Ferretti, Roberta; Pézzola, Antonella; Domenici, Maria Rosaria; Popoli, Patrizia

    2010-01-01

    Excitotoxicity plays a major role in the pathogenesis of Huntington disease (HD), a fatal neurodegenerative disorder. Adenosine A(2A) receptors (A(2A)Rs) modulate excitotoxicity and have been suggested to play a pathogenetic role in HD. The main aim of this study was to evaluate the effect of A(2A)R blockade on the expression and functions of NMDA receptors in the striatum of HD mice (R6/2). We found that 3 weeks' treatment with SCH 58261 (0.01 mg/kg/day i.p. from the 8th week of age) modified NR1 and NR2A/NR2B expression in the striatum of R6/2 (Western blotting) while had no effect on NMDA-induced toxicity in corticostriatal slices (electrophysiological experiments). In conclusion, in vivo A(2A)R blockade induced a remodeling of NMDA receptors in the striatum of HD mice. Even though the functional relevance of the above effect remains to be fully elucidated, these results add further evidence to the modulatory role of A(2A)Rs in HD.

  5. Adenosine A2A receptor and ecto-5'-nucleotidase/CD73 are upregulated in hippocampal astrocytes of human patients with mesial temporal lobe epilepsy (MTLE).

    PubMed

    Barros-Barbosa, Aurora R; Ferreirinha, Fátima; Oliveira, Ângela; Mendes, Marina; Lobo, M Graça; Santos, Agostinho; Rangel, Rui; Pelletier, Julie; Sévigny, Jean; Cordeiro, J Miguel; Correia-de-Sá, Paulo

    2016-12-01

    Refractoriness to existing medications of up to 80 % of the patients with mesial temporal lobe epilepsy (MTLE) prompts for finding new antiepileptic drug targets. The adenosine A2A receptor emerges as an interesting pharmacological target since its excitatory nature partially counteracts the dominant antiepileptic role of endogenous adenosine acting via inhibitory A1 receptors. Gain of function of the excitatory A2A receptor has been implicated in a significant number of brain pathologies commonly characterized by neuronal excitotoxicity. Here, we investigated changes in the expression and cellular localization of the A2A receptor and of the adenosine-generating enzyme, ecto-5'-nucleotidase/CD73, in the hippocampus of control individuals and MTLE human patients. Western blot analysis indicates that the A2A receptor is more abundant in the hippocampus of MTLE patients compared to control individuals. Immunoreactivity against the A2A receptor predominates in astrocytes staining positively for the glial fibrillary acidic protein (GFAP). No co-localization was observed between the A2A receptor and neuronal cell markers, like synaptotagmin 1/2 (nerve terminals) and neurofilament 200 (axon fibers). Hippocampal astrogliosis observed in MTLE patients was accompanied by a proportionate increase in A2A receptor and ecto-5'-nucleotidase/CD73 immunoreactivities. Given our data, we hypothesize that selective blockade of excessive activation of astrocytic A2A receptors and/or inhibition of surplus adenosine formation by membrane-bound ecto-5'-nucleotidase/CD73 may reduce neuronal excitability, thus providing a novel therapeutic target for drug-refractory seizures in MTLE patients.

  6. Triggering neurotrophic factor actions through adenosine A2A receptor activation: implications for neuroprotection

    PubMed Central

    Sebastião, Ana M; Ribeiro, Joaquim A

    2009-01-01

    G protein coupled receptors and tropomyosin-related kinase (Trk) receptors have distinct structure and transducing mechanisms; therefore, cross-talk among them was unexpected. Evidence has, however, accumulated showing that tonic adenosine A2A receptor activity is a required step to allow synaptic actions of neurotrophic factors, namely upon synaptic transmission at both pre- and post-synaptic level as well as upon synaptic plasticity. An enhancement of A2A receptor tonus upon ageing may partially compensate the loss of TrkB receptors, rescuing to certain degree the facilitatory action of brain derived neurotrophic factor in aged animals, which might prove particularly relevant in the prevention of neurodegeneration upon ageing. A2A receptors also trigger synaptic actions of other neurotrophic factors, such as glial derived neurotrophic factor at dopaminergic striatal nerve endings. The growing evidence that tonic adenosine A2A receptor activity is a crucial step to allow actions of neurotrophic factors in neurones will be reviewed and discussed in the light of therapeutic strategies for neurodegenerative diseases. PMID:19508402

  7. Effects of adenosine A2A receptor stimulation on cocaine-seeking behavior in rats.

    PubMed

    Bachtell, Ryan K; Self, David W

    2009-10-01

    Dopamine (DA) receptor stimulation in the nucleus accumbens (NAc) plays an important role in regulating cocaine-seeking behavior. Adenosine receptors antagonize the effects of DA receptor stimulation on intracellular signaling, neuronal output, and behavior. The goal of the present study is to determine the effects of adenosine A(2A) receptor stimulation on reinstatement of cocaine-seeking behavior in rats. Rats were trained to lever press for cocaine in daily self-administration sessions on a fixed-ratio 1 schedule for 3 weeks. After 1 week of abstinence, lever pressing was extinguished in six daily extinction sessions. We subsequently assessed the effects of the adenosine A(2A) receptor agonist, CGS 21680, on cocaine-, quinpirole (D(2) agonist)-, and cue-induced reinstatement to cocaine seeking. We also assessed the effects of CGS 21680 on sucrose seeking in rats extinguished from sucrose self-administration. Pretreatment of CGS 21680 dose-dependently blunted cocaine-induced reinstatement (15 mg/kg, i.p.). Pretreatment with CGS 21680 (0.03 mg/kg, i.p.) also attenuated quinpirole- and cue-induced reinstatement. A minimally effective dose of CGS 21680 failed to alter cocaine-induced locomotor activity or sucrose seeking. Stimulation of adenosine A(2A) receptors antagonizes reinstatement of cocaine seeking elicited by cocaine, DA D(2)-receptor stimulation, and cocaine-conditioned cues. These findings suggest that adenosine A(2A) receptor stimulation may oppose DA D(2) receptor signaling in the NAc that mediates cocaine relapse.

  8. Synthesis and SAR evaluation of 1,2,4-triazoles as A(2A) receptor antagonists.

    PubMed

    Alanine, Alexander; Anselm, Lilli; Steward, Lucinda; Thomi, Stefan; Vifian, Walter; Groaning, Michael D

    2004-02-09

    The synthesis and in vitro structure-activity relationships (SAR) of a series of triazoles as A(2A) receptor antagonists is reported. This resulted in the identification of potent, selective and permeable 1,2,4-triazoles such as 42 for further optimization and evaluation in vivo.

  9. Adenosine A2A receptors play a role in the pathogenesis of hepatic cirrhosis

    PubMed Central

    Chan, Edwin S L; Montesinos, Maria Carmen; Fernandez, Patricia; Desai, Avani; Delano, David L; Yee, Herman; Reiss, Allison B; Pillinger, Michael H; Chen, Jiang-Fan; Schwarzschild, Michael A; Friedman, Scott L; Cronstein, Bruce N

    2006-01-01

    Adenosine is a potent endogenous regulator of inflammation and tissue repair. Adenosine, which is released from injured and hypoxic tissue or in response to toxins and medications, may induce pulmonary fibrosis in mice, presumably via interaction with a specific adenosine receptor. We therefore determined whether adenosine and its receptors contribute to the pathogenesis of hepatic fibrosis. As in other tissues and cell types, adenosine is released in vitro in response to the fibrogenic stimuli ethanol (40 mg dl−1) and methotrexate (100 nM). Adenosine A2A receptors are expressed on rat and human hepatic stellate cell lines and adenosine A2A receptor occupancy promotes collagen production by these cells. Liver sections from mice treated with the hepatotoxins carbon tetrachloride (CCl4) (0.05 ml in oil, 50 : 50 v : v, subcutaneously) and thioacetamide (100 mg kg−1 in PBS, intraperitoneally) released more adenosine than those from untreated mice when cultured ex vivo. Adenosine A2A receptor-deficient, but not wild-type or A3 receptor-deficient, mice are protected from development of hepatic fibrosis following CCl4 or thioacetamide exposure. Similarly, caffeine (50 mg kg−1 day−1, po), a nonselective adenosine receptor antagonist, and ZM241385 (25 mg kg−1 bid), a more selective antagonist of the adenosine A2A receptor, diminished hepatic fibrosis in wild-type mice exposed to either CCl4 or thioacetamide. These results demonstrate that hepatic adenosine A2A receptors play an active role in the pathogenesis of hepatic fibrosis, and suggest a novel therapeutic target in the treatment and prevention of hepatic cirrhosis. PMID:16783407

  10. [Adenosine A2A receptor as a drug target for treatment of sepsis].

    PubMed

    Sivak, K V; Vasin, A V; Egorov, V V; Tsevtkov, V B; Kuzmich, N N; Savina, V A; Kiselev, O I

    2016-01-01

    Sepsis is a generalized infection accompanied by response of the body that manifests in a clinical and laboratory syndrome, namely, in the systemic inflammatory response syndrome (SIRS) from the organism to the infection. Although sepsis is a widespread and life-threatening disease, the assortment of drugs for its treatment is mostly limited by antibiotics. Therefore, the search for new cellular targets for drug therapy of sepsis is an urgent task of modern medicine and pharmacology. One of the most promising targets is the adenosine A(2A) receptor (A(2A)AR). The activation of this receptor, which is mediated by extracellular adenosine, manifests in almost all types of immune cells (lymphocytes, monocytes, macrophages, and dendritic cells) and results in reducing the severity of inflammation and reperfusion injury in various tissues. The activation of adenosine A(2A) receptor inhibits the proliferation of T cells and production of proinflammatory cytokines, which contributes to the activation of the synthesis of anti-inflammatory cytokines, thereby suppressing the systemic response. For this reason, various selective A(2A)AR agonists and antagonists may be considered to be drug candidates for sepsis pharmacotherapy. Nevertheless, they remain only efficient ligands and objects of pre-clinical and clinical trials. This review examines the molecular mechanisms of inflammatory response in sepsis and the structure and functions of A(2A)AR and its role in the pathogenesis of sepsis, as well as examples of using agonists and antagonists of this receptor for the treatment of SIRS and sepsis.

  11. 2-Amino-N-pyrimidin-4-ylacetamides as A2A receptor antagonists: 2. Reduction of hERG activity, observed species selectivity, and structure-activity relationships.

    PubMed

    Slee, Deborah H; Moorjani, Manisha; Zhang, Xiaohu; Lin, Emily; Lanier, Marion C; Chen, Yongsheng; Rueter, Jaimie K; Lechner, Sandra M; Markison, Stacy; Malany, Siobhan; Joswig, Tanya; Santos, Mark; Gross, Raymond S; Williams, John P; Castro-Palomino, Julio C; Crespo, María I; Prat, Maria; Gual, Silvia; Díaz, José-Luis; Jalali, Kayvon; Sai, Yang; Zuo, Zhiyang; Yang, Chun; Wen, Jenny; O'Brien, Zhihong; Petroski, Robert; Saunders, John

    2008-03-27

    Previously we have described a series of novel A 2A receptor antagonists with excellent water solubility. As described in the accompanying paper, the antagonists were first optimized to remove an unsubstituted furyl moiety, with the aim of avoiding the potential metabolic liabilities that can arise from the presence of an unsubstituted furan. This effort identified a series of potent and selective methylfuryl derivatives. Herein, we describe the further optimization of this series to increase potency, maintain selectivity for the human A 2A vs the human A 1 receptor, and minimize activity against the hERG channel. In addition, the observed structure-activity relationships against both the human and the rat A 2A receptor are reported.

  12. Role of Adenosine Receptor A2A in Traumatic Optic Neuropathies

    DTIC Science & Technology

    2012-12-01

    in Traumatic Optic Neuropathies ” PRINCIPAL INVESTIGATOR: Gregory I. Liou, PhD CONTRACTING ORGANIZATION: Georgia Health Sciences...Adenosine Receptor A2A in Traumatic Optic Neuropathies 5b. GRANT NUMBER W81XWH-11-2-0046 5c. PROGRAM ELEMENT NUMBER 6. AUTHOR(S) 5d. PROJECT...ABSTRACT Our goal is to develop an early therapeutic intervention before the progression of traumatic optic neuropathy (TON), a vision-threatening

  13. Analysis of Adenosine A2a Receptor Stability: Effects of Ligands and Disulfide Bonds

    PubMed Central

    O'Malley, Michelle A.; Naranjo, Andrea N.; Lazarova, Tzvetana; Robinson, Anne S.

    2010-01-01

    G protein-coupled receptors (GPCRs)1 constitute the largest family of integral membrane proteins present in all eukaryotic cells, yet relatively little information is known pertaining to their structure, folding, and stability. In this work, we describe several approaches to characterize conformational stability of the human adenosine A2a receptor (hA2aR). Thermal and chemical denaturation were not reversible, yet clear differences in the unfolding behavior were observed upon ligand binding via circular dichroism and fluorescence spectrometry. We found that the stability of hA2aR was increased upon incubation with the agonist N6-cyclohexyladenosine or the antagonist theophylline. When extracellular disulfide bonds were reduced with a chemical reducing agent, the ligand-binding activity decreased by ~40%, but reduction of these bonds did not compromise the unfolding transition observed via urea denaturation. Overall, these approaches offer a general strategy for characterizing the effect of surfactant and ligand effects on the stability of GPCRs. PMID:20853839

  14. Astrocytic adenosine receptor A2A and Gs-coupled signaling regulate memory

    PubMed Central

    Orr, Anna G.; Hsiao, Edward C.; Wang, Max M.; Ho, Kaitlyn; Kim, Daniel H.; Wang, Xin; Guo, Weikun; Kang, Jing; Yu, Gui-Qiu; Adame, Anthony; Devidze, Nino; Dubal, Dena B.; Masliah, Eliezer; Conklin, Bruce R.; Mucke, Lennart

    2014-01-01

    Astrocytes express a variety of G protein-coupled receptors and might influence cognitive functions, such as learning and memory. However, the roles of astrocytic Gs-coupled receptors in cognitive function are not known. We found that humans with Alzheimer’s disease (AD) had increased levels of the Gs-coupled adenosine receptor A2A in astrocytes. Conditional genetic removal of these receptors enhanced long-term memory in young and aging mice, and increased the levels of Arc/Arg3.1, an immediate-early gene required for long-term memory. Chemogenetic activation of astrocytic Gs-coupled signaling reduced long-term memory in mice without affecting learning. Similar to humans with AD, aging mice expressing human amyloid precursor protein (hAPP) showed increased levels of astrocytic A2A receptors. Conditional genetic removal of these receptors enhanced memory in aging hAPP mice. Together, these findings establish a regulatory role for astrocytic Gs-coupled receptors in memory and suggest that AD-linked increases in astrocytic A2A receptor levels contribute to memory loss. PMID:25622143

  15. Suppression of adenosine 2a receptor (A2aR)-mediated adenosine signaling improves disease phenotypes in a mouse model of amyotrophic lateral sclerosis

    PubMed Central

    Ng, Seng kah; Higashimori, Haruki; Tolman, Michaela; Yang, Yongjie

    2017-01-01

    Amyotrophic lateral sclerosis (ALS) is a rapidly progressing neurodegenerative disease in which the majority of upper and lower motor neurons are degenerated. Despite intensive efforts to identify drug targets and develop neuroprotective strategies, effective therapeutics for ALS remains unavailable. The identification and characterization of novel targets and pathways remain crucial in the development of ALS therapeutics. Adenosine is a major neuromodulator that actively regulates synaptic transmission. Interestingly, adenosine levels are significantly elevated in the cerebrospinal fluid (CSF) of progressing human ALS patients. In the current study, we showed that adenosine 2a receptor (A2aR), but not adenosine 1 receptor (A1R), is highly enriched in spinal (motor) neurons. A2aR expression is also selectively increased at the symptomatic onset in the spinal cords of SOD1G93A mice and end-stage human ALS spinal cords. Interestingly, we found that direct adenosine treatment is sufficient to induce embryonic stem cell-derived motor neuron (ESMN) cell death in cultures. Subsequent pharmacological inhibition and partial genetic ablation of A2aR (A2aR+/−) significantly protect ESMN from SOD1G93A+ astrocyte-induced cell death and delay disease progression of SOD1G93A mice. Taken together, our results provide compelling novel evidence that A2aR-mediated adenosine signaling contributes to the selective spinal motor neuron degeneration observed in the SOD1G93A mouse model of ALS. PMID:25779930

  16. Suppression of adenosine 2a receptor (A2aR)-mediated adenosine signaling improves disease phenotypes in a mouse model of amyotrophic lateral sclerosis.

    PubMed

    Ng, Seng Kah; Higashimori, Haruki; Tolman, Michaela; Yang, Yongjie

    2015-05-01

    Amyotrophic lateral sclerosis (ALS) is a rapidly progressing neurodegenerative disease in which the majority of upper and lower motor neurons are degenerated. Despite intensive efforts to identify drug targets and develop neuroprotective strategies, effective therapeutics for ALS remains unavailable. The identification and characterization of novel targets and pathways remain crucial in the development of ALS therapeutics. Adenosine is a major neuromodulator that actively regulates synaptic transmission. Interestingly, adenosine levels are significantly elevated in the cerebrospinal fluid (CSF) of progressing human ALS patients. In the current study, we showed that adenosine 2a receptor (A2aR), but not adenosine 1 receptor (A1R), is highly enriched in spinal (motor) neurons. A2aR expression is also selectively increased at the symptomatic onset in the spinal cords of SOD1G93A mice and end-stage human ALS spinal cords. Interestingly, we found that direct adenosine treatment is sufficient to induce embryonic stem cell-derived motor neuron (ESMN) cell death in cultures. Subsequent pharmacological inhibition and partial genetic ablation of A2aR (A2aR(+/-)) significantly protect ESMN from SOD1G93A(+) astrocyte-induced cell death and delay disease progression of SOD1G93A mice. Taken together, our results provide compelling novel evidence that A2aR-mediated adenosine signaling contributes to the selective spinal motor neuron degeneration observed in the SOD1G93A mouse model of ALS.

  17. Molecular Determinants of CGS21680 Binding to the Human Adenosine A2A Receptor.

    PubMed

    Lebon, Guillaume; Edwards, Patricia C; Leslie, Andrew G W; Tate, Christopher G

    2015-06-01

    The adenosine A2A receptor (A(2A)R) plays a key role in transmembrane signaling mediated by the endogenous agonist adenosine. Here, we describe the crystal structure of human A2AR thermostabilized in an active-like conformation bound to the selective agonist 2-[p-(2-carboxyethyl)phenylethyl-amino]-5'-N-ethylcarboxamido adenosine (CGS21680) at a resolution of 2.6 Å. Comparison of A(2A)R structures bound to either CGS21680, 5'-N-ethylcarboxamido adenosine (NECA), UK432097 [6-(2,2-diphenylethylamino)-9-[(2R,3R,4S,5S)-5-(ethylcarbamoyl)-3,4-dihydroxy-tetrahydrofuran-2-yl]-N-[2-[[1-(2-pyridyl)-4-piperidyl]carbamoylamino]ethyl]purine-2-carboxamide], or adenosine shows that the adenosine moiety of the ligands binds to the receptor in an identical fashion. However, an extension in CGS21680 compared with adenosine, the (2-carboxyethyl)phenylethylamino group, binds in an extended vestibule formed from transmembrane regions 2 and 7 (TM2 and TM7) and extracellular loops 2 and 3 (EL2 and EL3). The (2-carboxyethyl)phenylethylamino group makes van der Waals contacts with side chains of amino acid residues Glu169(EL2), His264(EL3), Leu267(7.32), and Ile274(7.39), and the amine group forms a hydrogen bond with the side chain of Ser67(2.65). Of these residues, only Ile274(7.39) is absolutely conserved across the human adenosine receptor subfamily. The major difference between the structures of A(2A)R bound to either adenosine or CGS21680 is that the binding pocket narrows at the extracellular surface when CGS21680 is bound, due to an inward tilt of TM2 in that region. This conformation is stabilized by hydrogen bonds formed by the side chain of Ser67(2.65) to CGS21680, either directly or via an ordered water molecule. Mutation of amino acid residues Ser67(2.65), Glu169(EL2), and His264(EL3), and analysis of receptor activation either in the presence or absence of ligands implicates this region in modulating the level of basal activity of A(2A)R.

  18. Clinical/pharmacological aspect of adenosine A2A receptor antagonist for dyskinesia.

    PubMed

    Kanda, Tomoyuki; Uchida, Shin-ichi

    2014-01-01

    Dopamine replacement therapy using the dopamine precursor, l-3,4-dihydroxyphenylalanine (l-DOPA), with a peripheral dopa decarboxylase inhibitor is the most effective treatment currently available for the symptoms of Parkinson's disease (PD). However, the long-term use of dopaminergic therapies for PD is often limited by the development of motor response complications, such as dyskinesia. Adenosine A2A receptors are a promising nondopaminergic target for the treatment of PD. The treatment of motor response complications involves combinations of regular and controlled release L-DOPA, perhaps with the addition of a COMT inhibitor or the use of a longer-acting dopamine agonist. However, when dyskinesia is already established, the increase in dopaminergic load produced by the addition of a dopamine agonist can result in an increase in the severity and duration of dyskinesia. Currently, there are no well-tolerated antidyskinesia agents available. Amantadine, which may exert its effects through the inhibition of N-methyl-D-aspartate (NMDA) receptors, shows some effects on established dyskinesia. Dyskinesia has a negative impact on the quality of life of patients, sometimes being more disabling than PD itself. Although some patients prefer experiencing dyskinesia than being in the OFF state and unable to move, alternative, more effective therapies are still required for severe disabling dyskinesia to afford patients an improved quality of life while in the ON state. The mechanisms causing and maintaining the dyskinesia have not been clarified. The application of a nondopaminergic approach to modify the basal ganglial activity would be helpful to better understand and treat dyskinesia. The use of an adenosine A2A receptor may provide one such approach. In this literature review, we will summarize the current knowledge from both clinical and nonclinical studies on the effects of adenosine A2A receptor blockade on dyskinesia.

  19. Studies on SF=1902 A2 A5, minor components of SF-1902 (globomycin).

    PubMed

    Omoto, S; Ogino, H; Inouye, S

    1981-11-01

    Four members of globomycin, SF-1902 A2, A3, A4a and A4b were newly isolated from the culture of Streptomyces hygroscopicus SF-1902. These minor components shared four amino acids in common and the fifth was either valine or allo-isoleucine. The fatty acid moiety varied from 3-hydroxy-2-methylheptanoic acid in A2 to 3-hydroxy-2-methylundecanoic acids in A4b. The length of alkyl chain greatly affected the antibacterial activity, and maximum activity was shown by the homologue (A5) possessing the longest alkyl chain.

  20. The first convergent total synthesis of penarolide sulfate A2, a novel α-glucosidase inhibitor.

    PubMed

    Gao, Yangguang; Shan, Qiuli; Liu, Jun; Wang, Linlin; Du, Yuguo

    2014-04-07

    Penarolide sulfate A2, a 31-membered macrolide encompassing a proline residue and three sulfate groups, was firstly synthesized in 16 linear steps with 4.8% overall yield. Three consecutive stereogenic centers in penarolide sulfate A2 were efficiently derived from natural chiral template l-arabinose. The crucial assembly reactions included Brown asymmetric allylation, olefin cross-metathesis, alkyne-epoxide coupling, and macrolactamization. The anti-yeast α-glucosidase activities of penarolide sulfate A2 and its fully desulfated derivative were examined showing IC50 values of 4.87 and 10.74 μg mL(-1), respectively.

  1. Physical origins of remarkable thermostabilization by an octuple mutation for the adenosine A2a receptor

    NASA Astrophysics Data System (ADS)

    Kajiwara, Yuta; Ogino, Takahiro; Yasuda, Satoshi; Takamuku, Yuuki; Murata, Takeshi; Kinoshita, Masahiro

    2016-07-01

    It was experimentally showed that the thermal stability of a membrane protein, the adenosine A2a receptor, was remarkably enhanced by an octuple mutation. Here we theoretically prove that the energy decrease arising from the formation of protein intramolecular hydrogen bonds and the solvent-entropy gain upon protein folding are made substantially larger by the mutation, leading to the remarkable enhancement. The solvent is formed by hydrocarbon groups constituting nonpolar chains of the lipid bilayer within a membrane. The mutation modifies geometric characteristics of the structure so that the solvent crowding can be reduced to a larger extent when the protein folds.

  2. Effect of low frequency electromagnetic fields on A2A adenosine receptors in human neutrophils

    PubMed Central

    Varani, Katia; Gessi, Stefania; Merighi, Stefania; Iannotta, Valeria; Cattabriga, Elena; Spisani, Susanna; Cadossi, Ruggero; Borea, Pier Andrea

    2002-01-01

    The present study describes the effect of low frequency, low energy, pulsing electromagnetic fields (PEMFs) on A2A adenosine receptors in human neutrophils.Saturation experiments performed using a high affinity adenosine antagonist [3H]-ZM 241385 revealed a single class of binding sites in control and in PEMF-treated human neutrophils with similar affinity (KD=1.05±0.10 and 1.08±0.12 nM, respectively). Furthermore, after 1 h of exposure to PEMFs the receptor density was statistically increased (P<0.01) (Bmax =126±10 and 215±15 fmol mg−1 protein, respectively).The effect of PEMFs was specific to the A2A adenosine receptors. This effect was also intensity, time and temperature dependent.In the adenylyl cyclase assays the A2A receptor agonists, HE-NECA and NECA, increased cyclic AMP accumulation in untreated human neutrophils with an EC50 value of 43 (40 – 47) and 255 (228 – 284) nM, respectively. The capability of HE-NECA and NECA to stimulate cyclic AMP levels in human neutrophils was increased (P<0.01) after exposure to PEMFs with an EC50 value of 10(8 – 13) and 61(52 – 71) nM, respectively.In the superoxide anion (O2−) production assays HE-NECA and NECA inhibited the generation of O2− in untreated human neutrophils, with an EC50 value of 3.6(3.1 – 4.2) and of 23(20 – 27) nM, respectively. Moreover, in PEMF-treated human neutrophils, the same compounds show an EC50 value of 1.6(1.2 – 2.1) and of 6.0(4.7 – 7.5) nM respectively.These results indicate the presence of significant alterations in the expression and in the functionality of adenosine A2A receptors in human neutrophils treated with PEMFs. PMID:11976268

  3. Effect of adenosine A(2A) receptor antagonists on L-DOPA-induced hydroxyl radical formation in rat striatum.

    PubMed

    Gołembiowska, Krystyna; Dziubina, Anna; Kowalska, Magdalena; Kamińska, Katarzyna

    2009-02-01

    A(2A) adenosine receptor antagonists have been proposed as a new therapy for Parkinson's disease (PD). Since oxidative stress plays an important role in the pathogenesis of PD, we studied the effect of the selective A(2A) adenosine receptor antagonists 8-(3-chlorostyryl)caffeine (CSC) and 4-(2-[7-amino-2-(2-furyl)[1,2,4]triazolo[2,3-a][1,3,5]triazin-5-ylamino]ethyl)phenol (ZM 241385) on L: -3,4-dihydroxyphenylalanine (L: -DOPA)-induced hydroxyl radical generation using in vivo microdialysis in the striatum of freely moving rats. L: -DOPA (100 mg/kg; in the presence of benserazide, 50 mg/kg) given acutely or repeatedly for 14 days generated a high level of hydroxyl radicals, measured by HPLC with electrochemical detection, as the product of their reaction with p-hydroxybenzoic acid (PBA). CSC (1 mg/kg) and ZM 241385 (3 mg/kg) decreased haloperidol (0.5 mg/kg)-induced catalepsy, while at low doses of 0.1 and 0.3 mg/kg, respectively, they did not display an effect. CSC (1 and 5 mg/kg) and ZM 241385 (3 and 9 mg/kg) given acutely, or CSC (1 mg/kg) and ZM 241385 (3 mg/kg) given repeatedly, increased the production of hydroxyl radicals in dialysates from rat striatum. Both acute and repeated administration of CSC (0.1 and 1 mg/kg) and ZM 241385 (3 mg/kg) decreased L: -DOPA-induced generation of hydroxyl radicals. However, a high single dose of either CSC (5 mg/kg) and ZM 241385 (9 mg/kg) markedly potentiated the effect of L: -DOPA on hydroxyl radical production. The increase in hydroxyl radical production by acute and chronic injection of CSC and ZM 241385 may be related to the increased release of dopamine (DA) and its metabolism in striatal dialysates. Similarly, increased DA release following a single high dose of CSC or ZM 241385 appears to be responsible for augmentation of L: -DOPA-induced hydroxyl radical formation. Conversely, the inhibition of L: -DOPA-induced production of hydroxyl radical by single and repeated low doses of CSC or repeated low doses of ZM

  4. Diabetes susceptibility in Mayas: Evidence for the involvement of polymorphisms in HHEX, HNF4α, KCNJ11, PPARγ, CDKN2A/2B, SLC30A8, CDC123/CAMK1D, TCF7L2, ABCA1 and SLC16A11 genes.

    PubMed

    Lara-Riegos, J C; Ortiz-López, M G; Peña-Espinoza, B I; Montúfar-Robles, I; Peña-Rico, M A; Sánchez-Pozos, K; Granados-Silvestre, M A; Menjivar, M

    2015-07-01

    Association of type 2 diabetes (T2D) with common variants in HHEX, HNF4α, KCNJ11, PPARγ, CDKN2A/2B, SLC30A8, CDC123/CAMK1D, TCF7L2, ABCA1 and SLC16A11 genes have been reported, mainly in populations of European and Asian ancestry and to a lesser extent in Latin Americans. Thus, we aimed to investigate the contribution of rs1111875 (HHEX), rs1800961 (HNF4α), rs5219 (KCNJ11), rs1801282 (PPARγ), rs10811661 (CDKN2A/2B), rs13266634 (SLC30A8), rs12779790 (CDC123/CAMK1D), rs7903146 (TCF7L2), rs9282541 (ABCA1) and rs13342692 (SLC16A11) polymorphisms in the genetic background of Maya population to associate their susceptibility to develop T2D. This is one of the first studies designed specifically to investigate the inherited component of T2D in the indigenous population of Mexico. SNPs were genotyped by allelic discrimination method in 575 unrelated Maya individuals. Two SNPs rs10811661 and rs928254 were significantly associated with T2D after adjusting for BMI; rs10811661 in a recessive and rs9282541 in a dominant model. Additionally, we found phenotypical alterations associated with genetic variants: HDL to rs9282541 and insulin to rs13342692. In conclusion, these findings support an association of genetic polymorphisms to develop T2D in Maya population.

  5. Agonist Dynamics and Conformational Selection during Microsecond Simulations of the A2A Adenosine Receptor

    PubMed Central

    Lee, Ji Young; Lyman, Edward

    2012-01-01

    The G-protein-coupled receptors (GPCRs) are a ubiquitous family of signaling proteins of exceptional pharmacological importance. The recent publication of structures of several GPCRs cocrystallized with ligands of differing activity offers a unique opportunity to gain insight into their function. To that end, we performed microsecond-timescale simulations of the A2A adenosine receptor bound to either of two agonists, adenosine or UK432097. Our data suggest that adenosine is highly dynamic when bound to A2A, in stark contrast to the case with UK432097. Remarkably, adenosine finds an alternate binding pose in which the ligand is inverted relative to the crystal structure, forming relatively stable interactions with helices I and II. Our observations suggest new experimental tests to validate our predictions and deepen our understanding of GPCR signaling. Overall, our data suggest an intriguing hypothesis: that the 100- to 1000-fold greater efficacy of UK432097 relative to adenosine arises because UK432097 stabilizes a much tighter neighborhood of active conformations, which manifests as a greater likelihood of G-protein activation per unit time. PMID:22824275

  6. A2A adenosine receptor regulates the human blood brain barrier permeability

    PubMed Central

    Kim, Do-Geun; Bynoe, Margaret S.

    2015-01-01

    The blood brain barrier (BBB) symbolically represents the gateway to the central nervous system. It is a single layer of specialized endothelial cells that coats the central nervous system (CNS) vasculature and physically separates the brain environment from the blood constituents, to maintain the homeostasis of the CNS. However, this protective measure is a hindrance to the delivery of therapeutics to treat neurological diseases. Here, we show that activation of A2A adenosine receptor (AR) with an FDA-approved agonist potently permeabilizes an in vitro primary human brain endothelial barrier (hBBB) to the passage of chemotherapeutic drugs and T cells. T cell migration under AR signaling occurs primarily by paracellular transendothelial route. Permeabilization of the hBBB is rapid, time-dependent and reversible and is mediated by morphological changes in actin-cytoskeletal reorganization induced by RhoA signaling and a potent down-regulation of Claudin-5 and VE-Cadherin. Moreover, the kinetics of BBB permeability in mice closely overlaps with the permeability kinetics of the hBBB. These data suggest that activation of A2A AR is an endogenous mechanism that may be used for CNS drug delivery in human. PMID:25262373

  7. Endothelial adenosine A2a receptor-mediated glycolysis is essential for pathological retinal angiogenesis.

    PubMed

    Liu, Zhiping; Yan, Siyuan; Wang, Jiaojiao; Xu, Yiming; Wang, Yong; Zhang, Shuya; Xu, Xizhen; Yang, Qiuhua; Zeng, Xianqiu; Zhou, Yaqi; Gu, Xuejiao; Lu, Sarah; Fu, Zhongjie; Fulton, David J; Weintraub, Neal L; Caldwell, Ruth B; Zhang, Wenbo; Wu, Chaodong; Liu, Xiao-Ling; Chen, Jiang-Fan; Ahmad, Aftab; Kaddour-Djebbar, Ismail; Al-Shabrawey, Mohamed; Li, Qinkai; Jiang, Xuejun; Sun, Ye; Sodhi, Akrit; Smith, Lois; Hong, Mei; Huo, Yuqing

    2017-09-19

    Adenosine/adenosine receptor-mediated signaling has been implicated in the development of various ischemic diseases, including ischemic retinopathies. Here, we show that the adenosine A2a receptor (ADORA2A) promotes hypoxia-inducible transcription factor-1 (HIF-1)-dependent endothelial cell glycolysis, which is crucial for pathological angiogenesis in proliferative retinopathies. Adora2a expression is markedly increased in the retina of mice with oxygen-induced retinopathy (OIR). Endothelial cell-specific, but not macrophage-specific Adora2a deletion decreases key glycolytic enzymes and reduces pathological neovascularization in the OIR mice. In human primary retinal microvascular endothelial cells, hypoxia induces the expression of ADORA2A by activating HIF-2α. ADORA2A knockdown decreases hypoxia-induced glycolytic enzyme expression, glycolytic flux, and endothelial cell proliferation, sprouting and tubule formation. Mechanistically, ADORA2A activation promotes the transcriptional induction of glycolytic enzymes via ERK- and Akt-dependent translational activation of HIF-1α protein. Taken together, these findings advance translation of ADORA2A as a therapeutic target in the treatment of proliferative retinopathies and other diseases dependent on pathological angiogenesis.Pathological angiogenesis in the retina is a major cause of blindness. Here the authors show that adenosine receptor A2A drives pathological angiogenesis in the oxygen-induced retinopathy mouse model by promoting glycolysis in endothelial cells via the ERK/Akt/HIF-1α pathway, thereby suggesting new therapeutic targets for disease treatment.

  8. In vitro metabolism studies of new adenosine A 2A receptor antagonists.

    PubMed

    Marucci, Gabriella; Finaurini, Sara; Buccioni, Michela; Lammi, Carmen; Kandhavelu, Meenakshisundaram; Volpini, Rosaria; Ricciutelli, Massimo; Angeli, Piero; Commandeur, Jan N M; Cristalli, Gloria

    2008-12-01

    Evidence, obtained in rodent and primate models of Parkinson's disease (PD) and in preliminary clinical trials, indicates that adenosine A(2A) receptor antagonists might represent a promising non-dopaminergic therapeutic tool for the treatment of PD. Recently, we have reported the biological evaluation of 8-substituted 9-ethyladenines (ANR) as new A(2A) receptor antagonists, three of which (ANR 82, ANR 94, and ANR 152) showed high efficacy in in vivo models for Parkinson's. Understanding the metabolic pathways of new drug candidates is an important aspect of drug discovery. The ANR compounds have been investigated in order to clarify their activity on rat liver microsomes, and more specifically on recombinant human cytochrome P450 2D6 (CYP2D6). The metabolites of all three compounds were detected by liquid chromatography/tandem mass spectrometry (LC-MS/MS). The results indicate that this class of 9-ethyladenines is metabolized only to a fraction of 1.5-5%. These compounds also act as potent mechanism-based inhibitors of CYP450 and in particular of human isoform CYP2D6. Kinetic-analysis of enzyme inactivation was used to describe the effect of these time-dependent inhibitors and to derive the inhibition parameters K(inact) and K(i) defined with respect to the O-demethylation of dextromethorphan.

  9. Hydration Site Thermodynamics Explain SARs for Triazolylpurines Analogues Binding to the A2A Receptor

    PubMed Central

    2010-01-01

    A series of triazolylpurine analogues show interesting and unintuitive structure−activity relationships against the A2A adenosine receptor. As the 2-substituted aliphatic group is initially increased to methyl and isopropyl, there is a decrease in potency; however, extending the substituent to n-butyl and n-pentyl results in a significant gain in potency. This trend cannot be readily explained by ligand−receptor interactions, steric effects, or differences in ligand desolvation. Here, we show that a novel method for characterizing solvent thermodynamics in protein binding sites correctly predicts the trend in binding affinity for this series based on the differential water displacement patterns. In brief, small unfavorable substituents occupy a region in the A2A adenosine receptor binding site predicted to contain stable waters, while the longer favorable substituents extend to a region that contains several unstable waters. The predicted binding energies associated with displacing water within these hydration sites correlate well with the experimental activities. PMID:24900189

  10. Beneficial effects of a novel agonist of the adenosine A2A receptor on monocrotaline-induced pulmonary hypertension in rats

    PubMed Central

    Alencar, Allan K N; Pereira, Sharlene L; Montagnoli, Tadeu L; Maia, Rodolfo C; Kümmerle, Arthur E; Landgraf, Sharon S; Caruso-Neves, Celso; Ferraz, Emanuelle B; Tesch, Roberta; Nascimento, José H M; de Sant'Anna, Carlos M R; Fraga, Carlos A M; Barreiro, Eliezer J; Sudo, Roberto T; Zapata-Sudo, Gisele

    2013-01-01

    Background and Purpose Pulmonary arterial hypertension (PAH) is characterized by enhanced pulmonary vascular resistance, right ventricular hypertrophy and increased right ventricular systolic pressure. Here, we investigated the effects of a N-acylhydrazone derivative, 3,4-dimethoxyphenyl-N-methyl-benzoylhydrazide (LASSBio-1359), on monocrotaline (MCT)-induced pulmonary hypertension in rats. Experimental Approach PAH was induced in male Wistar rats by a single i.p. injection of MCT (60 mg·kg−1) and 2 weeks later, oral LASSBio-1359 (50 mg·kg−1) or vehicle was given once daily for 14 days. Echocardiography was used to measure cardiac function and pulmonary artery dimensions, with histological assay of vascular collagen. Studies of binding to human recombinant adenosine receptors (A1, A2A, A3) and of docking with A2A receptors were also performed. Key Results MCT administration induced changes in vascular and ventricular structure and function, characteristic of PAH. These changes were reversed by treatment with LASSBio-1359. MCT also induced endothelial dysfunction in pulmonary artery, as measured by diminished relaxation of pre-contracted arterial rings, and this dysfunction was reversed by LASSBio-1359. In pulmonary artery rings from normal Wistar rats, LASSBio-1359 induced relaxation, which was decreased by the adenosine A2A receptor antagonist, ZM 241385. In adenosine receptor binding studies, LASSBio-1359 showed most affinity for the A2A receptor and in the docking analyses, binding modes of LASSBio-1359 and the A2A receptor agonist, CGS21680, were very similar. Conclusion and Implications In rats with MCT-induced PAH, structural and functional changes in heart and pulmonary artery were reversed by treatment with oral LASSBio-1359, most probably through the activation of adenosine A2A receptors. PMID:23530610

  11. Therapeutic Opportunities for Caffeine and A2A Receptor Antagonists in Retinal Diseases.

    PubMed

    Boia, Raquel; Ambrósio, António Francisco; Santiago, Ana Raquel

    2016-01-01

    Caffeine, the major component of coffee, is the most consumed psychostimulant in the world. Caffeine is an adenosine analog and acts as a nonselective adenosine receptor antagonist. The majority of the effects of caffeine are mainly mediated by the blockade of adenosine receptors, and the proved neuroprotective effects of caffeine in brain disorders have been mimicked by the blockade of adenosine A2A receptor (A2AR). A growing body of evidence demonstrates that microglia-mediated neuroinflammation plays a key role in the pathophysiology of brain and retinal diseases. Moreover, the control of microglia reactivity by blocking A2AR has been proposed to be the mechanism underlying the observed protective effects of caffeine. Hence, it is conceivable that caffeine and A2AR antagonists offer therapeutic value for the treatment of retinal diseases, mainly those involving microglia-mediated neuroinflammation. © 2016 S. Karger AG, Basel.

  12. Polydeoxyribonucleotides (PDRNs) From Skin to Musculoskeletal Tissue Regeneration via Adenosine A2A Receptor Involvement.

    PubMed

    Veronesi, Francesca; Dallari, Dante; Sabbioni, Giacomo; Carubbi, Chiara; Martini, Lucia; Fini, Milena

    2017-09-01

    Polydeoxyribonucleotides (PDRNs) are low molecular weight DNA molecules of natural origin that stimulate cell migration and growth, extracellular matrix (ECM) protein production, and reduce inflammation. Most preclinical and clinical studies on tissue regeneration with PDRNs focused on skin, and only few are about musculoskeletal tissues. Starting from an overview on skin regeneration studies, through the analysis of in vitro, in vivo, and clinical studies (1990-2016), the present review aimed at defining the effects of PDRN and their mechanisms of action in the regeneration of musculoskeletal tissues. This would also help future researches in this area. A total of 29 studies were found by PubMed and www.webofknowledge.com searches: 20 were on skin (six in vitro, six in vivo, one vitro/vivo, seven clinical studies), while the other nine regarded bone (one in vitro, two in vivo, one clinical studies), cartilage (one in vitro, one vitro/vivo, two clinical studies), or tendon (one clinical study) tissues regeneration. PDRNs improved cell growth, tissue repair, ECM proteins, physical activity, and reduced pain and inflammation, through the activation of adenosine A2A receptor. PDRNs are currently used for bone, cartilage, and tendon diseases, with a great variability regarding the PDRN dosage to be used in clinical practice, while the dosage for skin regeneration is well established. PDRNs are usually administered from a minimum of three to a maximum of five times and they act trough the activation of A2A receptor. Further studies are advisable to confirm the effectiveness of PDRNs and to standardize the PDRN dose. J. Cell. Physiol. 232: 2299-2307, 2017. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

  13. Water network perturbation in ligand binding: adenosine A(2A) antagonists as a case study.

    PubMed

    Bortolato, Andrea; Tehan, Ben G; Bodnarchuk, Michael S; Essex, Jonathan W; Mason, Jonathan S

    2013-07-22

    Recent efforts in the computational evaluation of the thermodynamic properties of water molecules have resulted in the development of promising new in silico methods to evaluate the role of water in ligand binding. These methods include WaterMap, SZMAP, GRID/CRY probe, and Grand Canonical Monte Carlo simulations. They allow the prediction of the position and relative free energy of the water molecule in the protein active site and the analysis of the perturbation of an explicit water network (WNP) as a consequence of ligand binding. We have for the first time extended these approaches toward the prediction of kinetics for small molecules and of relative free energy of binding with a focus on the perturbation of the water network and application to large diverse data sets. Our results support a qualitative correlation between the residence time of 12 related triazine adenosine A(2A) receptor antagonists and the number and position of high energy trapped solvent molecules. From a quantitative viewpoint, we successfully applied these computational techniques as an implicit solvent alternative, in linear combination with a molecular mechanics force field, to predict the relative ligand free energy of binding (WNP-MMSA). The applicability of this linear method, based on the thermodynamics additivity principle, did not extend to 375 diverse A(2A) receptor antagonists. However, a fast but effective method could be enabled by replacing the linear approach with a machine learning technique using probabilistic classification trees, which classified the binding affinity correctly for 90% of the ligands in the training set and 67% in the test set.

  14. A Tripartite Fusion, FaeG-FedF-LT192A2:B, of Enterotoxigenic Escherichia coli (ETEC) Elicits Antibodies That Neutralize Cholera Toxin, Inhibit Adherence of K88 (F4) and F18 Fimbriae, and Protect Pigs against K88ac/Heat-Labile Toxin Infection ▿

    PubMed Central

    Ruan, Xiaosai; Liu, Mei; Casey, Thomas A.; Zhang, Weiping

    2011-01-01

    Enterotoxigenic Escherichia coli (ETEC) strains expressing K88 (F4) or F18 fimbriae and heat-labile (LT) and/or heat-stable (ST) toxins are the major cause of diarrhea in young pigs. Effective vaccines inducing antiadhesin (anti-K88 and anti-F18) and antitoxin (anti-LT and anti-ST) immunity would provide broad protection to young pigs against ETEC. In this study, we genetically fused nucleotides coding for peptides from K88ac major subunit FaeG, F18 minor subunit FedF, and LT toxoid (LT192) A2 and B subunits for a tripartite adhesin-adhesin-toxoid fusion (FaeG-FedF-LT192A2:B). This fusion was used for immunizations in mice and pigs to assess the induction of antiadhesin and antitoxin antibodies. In addition, protection by the elicited antiadhesin and antitoxin antibodies against a porcine ETEC strain was evaluated in a gnotobiotic piglet challenge model. The data showed that this FaeG-FedF-LT192A2:B fusion elicited anti-K88, anti-F18, and anti-LT antibodies in immunized mice and pigs. In addition, the anti-porcine antibodies elicited neutralized cholera toxin and inhibited adherence against both K88 and F18 fimbriae. Moreover, immunized piglets were protected when challenged with ETEC strain 30302 (K88ac/LT/STb) and did not develop clinical disease. In contrast, all control nonvaccinated piglets developed severe diarrhea and dehydration after being challenged with the same ETEC strain. This study clearly demonstrated that this FaeG-FedF-LT192A2:B fusion antigen elicited antibodies that neutralized LT toxin and inhibited the adherence of K88 and F18 fimbrial E. coli strains and that this fusion could serve as an antigen for vaccines against porcine ETEC diarrhea. In addition, the adhesin-toxoid fusion approach used in this study may provide important information for developing effective vaccines against human ETEC diarrhea. PMID:21813665

  15. Functional efficacy of adenosine A2A receptor agonists is positively correlated to their receptor residence time

    PubMed Central

    Guo, Dong; Mulder-Krieger, Thea; IJzerman, Adriaan P; Heitman, Laura H

    2012-01-01

    BACKGROUND AND PURPOSE The adenosine A2A receptor belongs to the superfamily of GPCRs and is a promising therapeutic target. Traditionally, the discovery of novel agents for the A2A receptor has been guided by their affinity for the receptor. This parameter is determined under equilibrium conditions, largely ignoring the kinetic aspects of the ligand-receptor interaction. The aim of this study was to assess the binding kinetics of A2A receptor agonists and explore a possible relationship with their functional efficacy. EXPERIMENTAL APPROACH We set up, validated and optimized a kinetic radioligand binding assay (a so-called competition association assay) at the A2A receptor from which the binding kinetics of unlabelled ligands were determined. Subsequently, functional efficacies of A2A receptor agonists were determined in two different assays: a novel label-free impedance-based assay and a more traditional cAMP determination. KEY RESULTS A simplified competition association assay yielded an accurate determination of the association and dissociation rates of unlabelled A2A receptor ligands at their receptor. A correlation was observed between the receptor residence time of A2A receptor agonists and their intrinsic efficacies in both functional assays. The affinity of A2A receptor agonists was not correlated to their functional efficacy. CONCLUSIONS AND IMPLICATIONS This study indicates that the molecular basis of different agonist efficacies at the A2A receptor lies within their different residence times at this receptor. PMID:22324512

  16. Potentiation by tonic A2a-adenosine receptor activation of CGRP-facilitated [3H]-ACh release from rat motor nerve endings.

    PubMed Central

    Correia-de-Sá, P.; Ribeiro, J. A.

    1994-01-01

    1. The effect of calcitonin gene-related peptide (CGRP) on [3H]-acetylcholine ([3H]-ACh) release from motor nerve endings and its interaction with presynaptic facilitatory A2a-adenosine and nicotinic acetylcholine receptors was studied on rat phrenic nerve-hemidiaphragm preparations loaded with [3H]-choline. 2. CGRP (100-400 nM) increased electrically evoked [3H]-ACh release from phrenic nerve endings in a concentration-dependent manner. 3. The magnitude of CGRP excitation increased with the increase of the stimulation pulse duration from 40 microseconds to 1 ms, keeping the frequency, the amplitude and the train length constants. With 1 ms pulses, the evoked [3H]-ACh release was more intense than with 40 microseconds pulse duration. 4. Both the nicotinic acetylcholine receptor agonist, 1,1-dimethyl-4-phenylpiperazinium, and the A2a adenosine receptor agonist, CGS 21680C, increased evoked [3H]-ACh release, but only CGS 21680C potentiated the facilitatory effect of CGRP. This potentiation was prevented by the A2a adenosine receptor antagonist, PD 115,199. 5. Adenosine deaminase prevented the excitatory effect of CGRP (400 nM) on [3H]-ACh release. This effect was reversed by the non-hydrolysable A2a-adenosine receptor agonist, CGS 21680C. 6. The nicotinic antagonist, tubocurarine, did not significantly change, whereas the A2-adenosine receptor antagonist, PD 115,199, blocked the CGRP facilitation. The A1-adenosine receptor antagonist, 1,3-dipropyl-8-cyclopentylxanthine, potentiated the CGRP excitatory effect. 7. The results suggest that the facilitatory effect of CGRP on evoked [3H]-ACh release from rat phrenic motor nerve endings depends on the presence of endogenous adenosine which tonically activates A2a-adenosine receptors.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:8004402

  17. Adenosine A2A receptor antagonism and genetic deletion attenuate the effects of dopamine D2 antagonism on effort-based decision making in mice.

    PubMed

    Pardo, M; Lopez-Cruz, L; Valverde, O; Ledent, C; Baqi, Y; Müller, C E; Salamone, J D; Correa, M

    2012-04-01

    Brain dopamine (DA) and adenosine interact in the regulation of behavioral activation and effort-related processes. In the present studies, a T-maze task was developed in mice for the assessment of effort-related decision making. With this task, the two arms of the maze have different reinforcement densities, and a vertical barrier is positioned in the arm with the higher density (HD), presenting the animal with an effort-related challenge. Under control conditions mice prefer the HD arm, and climb the barrier to obtain the larger amount of food. The DA D(2) receptor antagonist haloperidol decreased selection of the HD arm and increased selection of the arm with the low density of reinforcement. However, the HD arm was still the preferred choice in haloperidol-treated mice trained with barriers in both arms. Pre-feeding the mice to reduce food motivation dramatically increased omissions, an effect that was distinct from the actions of haloperidol. Co-administration of theophylline, a nonselective adenosine receptor antagonist, partially reversed the effects of haloperidol. This effect seems to be mediated by the A(2A) receptor but not the A(1) receptor, since the A(2A) antagonist MSX-3, but not the A(1) antagonist CPT, dose dependently reversed the effects of haloperidol on effort-related choice and on c-Fos expression in the dorsal striatum and nucleus accumbens. In addition, adenosine A(2A) receptor knockout mice were resistant to the effects of haloperidol on effort-related choice in the maze. These results indicate that DA D(2) and adenosine A(2A) receptors interact to regulate effort-related decision making and effort expenditure in mice.

  18. Continuous adenosine A2A receptor antagonism after focal cerebral ischemia in spontaneously hypertensive rats.

    PubMed

    Fronz, Ulrike; Deten, Alexander; Baumann, Frank; Kranz, Alexander; Weidlich, Sarah; Härtig, Wolfgang; Nieber, Karen; Boltze, Johannes; Wagner, Daniel-Christoph

    2014-02-01

    Antagonism of the adenosine A2A receptor (A2AR) has been shown to elicit substantial neuroprotective properties when given immediately after cerebral ischemia. We asked whether the continuous application of a selective A2AR antagonist within a clinically relevant time window will be a feasible and effective approach to treat focal cerebral ischemia. To answer this question, we subjected 20 male spontaneously hypertensive rats to permanent middle cerebral artery occlusion and randomized them equally to a verum and a control group. Two hours after stroke onset, the animals received a subcutaneous implantation of an osmotic minipump filled with 5 mg kg(-1) day(-1) 8-(3-chlorostyryl) caffeine (CSC) or vehicle solution. The serum level of CSC was measured twice a day for three consecutive days. The infarct volume was determined at days 1 and 3 using magnetic resonance imaging. We found the serum level of CSC showing a bell-shaped curve with its maximum at 36 h. The infarct volume was not affected by continuous CSC treatment. These results suggest that delayed and continuous CSC application was not sufficient to treat acute ischemic stroke, potentially due to unfavorable hepatic elimination and metabolization of the pharmaceutical.

  19. Angiotensin II type 1/adenosine A 2A receptor oligomers: a novel target for tardive dyskinesia.

    PubMed

    Oliveira, Paulo A de; Dalton, James A R; López-Cano, Marc; Ricarte, Adrià; Morató, Xavier; Matheus, Filipe C; Cunha, Andréia S; Müller, Christa E; Takahashi, Reinaldo N; Fernández-Dueñas, Víctor; Giraldo, Jesús; Prediger, Rui D; Ciruela, Francisco

    2017-05-12

    Tardive dyskinesia (TD) is a serious motor side effect that may appear after long-term treatment with neuroleptics and mostly mediated by dopamine D2 receptors (D2Rs). Striatal D2R functioning may be finely regulated by either adenosine A2A receptor (A2AR) or angiotensin receptor type 1 (AT1R) through putative receptor heteromers. Here, we examined whether A2AR and AT1R may oligomerize in the striatum to synergistically modulate dopaminergic transmission. First, by using bioluminescence resonance energy transfer, we demonstrated a physical AT1R-A2AR interaction in cultured cells. Interestingly, by protein-protein docking and molecular dynamics simulations, we described that a stable heterotetrameric interaction may exist between AT1R and A2AR bound to antagonists (i.e. losartan and istradefylline, respectively). Accordingly, we subsequently ascertained the existence of AT1R/A2AR heteromers in the striatum by proximity ligation in situ assay. Finally, we took advantage of a TD animal model, namely the reserpine-induced vacuous chewing movement (VCM), to evaluate a novel multimodal pharmacological TD treatment approach based on targeting the AT1R/A2AR complex. Thus, reserpinized mice were co-treated with sub-effective losartan and istradefylline doses, which prompted a synergistic reduction in VCM. Overall, our results demonstrated the existence of striatal AT1R/A2AR oligomers with potential usefulness for the therapeutic management of TD.

  20. New adenosine A2A receptor antagonists: actions on Parkinson's disease models.

    PubMed

    Pinna, Annalisa; Volpini, Rosaria; Cristalli, Gloria; Morelli, Micaela

    2005-04-11

    The 8-substituted 9-ethyladenine derivatives: 8-bromo-9-ethyladenine (ANR 82), 8-ethoxy- 9-ethyladenine (ANR 94), and 8-furyl-9-ethyladenine (ANR 152) have been characterized in vitro as adenosine receptor antagonists. Adenosine is deeply involved in the control of motor behaviour and substantial evidences indicate that adenosine A(2A) receptor antagonists improve motor deficits in animal models of Parkinson's disease. On this basis, the efficacy of ANR 82, ANR 94, and ANR 152 in rat models of Parkinson's disease was evaluated. All compounds tested reversed the catalepsy induced by haloperidol. However, in unilaterally 6-hydroxydopamine-lesioned rats, only ANR 94 and ANR 152 potentiated l-dihydroxy-phenylalanine (l-DOPA) effect on turning behaviour and induced contralateral turning behaviour in rats sensitised to l-DOPA. Taken together the results of this study indicate that some 8-substituted 9-ethyladenine derivatives ameliorate motor deficits in rat models of Parkinson's disease, suggesting a potential therapeutic role of these compounds.

  1. Investigation of the conformational dynamics of the apo A2A adenosine receptor

    PubMed Central

    Caliman, Alisha D; Swift, Sara E; Wang, Yi; Miao, Yinglong; McCammon, J Andrew

    2015-01-01

    The activation/deactivation processes for G-protein coupled receptors (GPCRs) have been computationally studied for several different classes, including rhodopsin, the β2 adrenergic receptor, and the M2 muscarinic receptor. Despite determined cocrystal structures of the adenosine A2A receptor (A2AAR) in complex with antagonists, agonists and an antibody, the deactivation process of this GPCR is not completely understood. In this study, we investigate the convergence of two apo simulations, one starting with an agonist-bound conformation (PDB: 3QAK)14 and the other starting with an antagonist-bound conformation (PDB: 3EML)11. Despite the two simulations not completely converging, we were able to identify distinct intermediate steps of the deactivation process characterized by the movement of Y2887.53 in the NPxxY motif. We find that Y2887.53 contributes to the process by forming hydrogen bonds to residues in transmembrane helices 2 and 7 and losing these interactions upon full deactivation. Y1975.58 also plays a role in the process by forming a hydrogen bond only once the side chain moves from the lipid interface to the middle of the helical bundle. PMID:25761901

  2. The effect of caffeine to increase reaction time in the rat during a test of attention is mediated through antagonism of adenosine A2A receptors.

    PubMed

    Higgins, Guy A; Grzelak, Michael E; Pond, Annamarie J; Cohen-Williams, Mary E; Hodgson, Robert A; Varty, Geoffrey B

    2007-12-11

    Caffeine produces effects on cognitive function particularly relating to aspects of attention such as reaction time. Considering the plasma exposure levels following regular caffeine intake, and the affinity of caffeine for known protein targets, these effects are likely mediated by either the adenosine A(1) or A(2A) receptor. In the present studies, two rat strains [Long-Evans (LE) and CD] were trained to asymptote performance in a test of selective attention, the 5-choice serial reaction time task (5-CSRTT). Next, the effects of caffeine were compared to the selective A(2A) antagonists, SCH 412348 and KW-6002 (Istradefylline), and the A(1) antagonist, DPCPX. Further studies compared the psychostimulant effects of each drug. Finally, we tested the A(2A) agonist, CGS-21680, on 5-CSRTT performance and given the antipsychotic potential of this drug class, studied the interaction between CGS-21680 and amphetamine in this task. Caffeine (3-10mg/kg IP) increased reaction time in both LE and CD rats, with no effect on accuracy, an effect replicated by SCH 412348 (0.1-1mg/kg PO) and KW-6002 (1-3mg/kg PO), but not DPCPX (3-30 mg/kg PO). At least with SCH 412348, these effects were at doses that were not overtly psychostimulant. In contrast, CGS-21680 (0.03-0. 3mg/kg IP) slowed reaction speed and increased omissions. Interestingly, at a comparatively low dose of 0.03 mg/kg, CGS-21680 attenuated the increased premature responding produced by amphetamine (1mg/kg IP). The present results suggest that the attention-enhancing effects of caffeine are mediated through A(2A) receptor blockade, and selective A(2A) receptor antagonists may have potential as therapies for attention-related disorders. Furthermore, the improvement in response control in amphetamine-treated rats following CGS-21680 pretreatment supports the view that A(2A) agonists have potential as novel antipsychotics.

  3. Creatine, similarly to ketamine, affords antidepressant-like effects in the tail suspension test via adenosine A₁ and A2A receptor activation.

    PubMed

    Cunha, Mauricio P; Pazini, Francis L; Rosa, Julia M; Ramos-Hryb, Ana B; Oliveira, Ágatha; Kaster, Manuella P; Rodrigues, Ana Lúcia S

    2015-06-01

    The benefits of creatine supplementation have been reported in a broad range of central nervous systems diseases, including depression. A previous study from our group demonstrated that creatine produces an antidepressant-like effect in the tail suspension test (TST), a predictive model of antidepressant activity. Since depression is associated with a dysfunction of the adenosinergic system, we investigated the involvement of adenosine A1 and A2A receptors in the antidepressant-like effect of creatine in the TST. The anti-immobility effect of creatine (1 mg/kg, po) or ketamine (a fast-acting antidepressant, 1 mg/kg, ip) in the TST was prevented by pretreatment of mice with caffeine (3 mg/kg, ip, nonselective adenosine receptor antagonist), 8-cyclopentyl-1,3-dipropylxanthine (DPCPX) (2 mg/kg, ip, selective adenosine A1 receptor antagonist), and 4-(2-[7-amino-2-{2-furyl}{1,2,4}triazolo-{2,3-a}{1,3,5}triazin-5-yl-amino]ethyl)-phenol (ZM241385) (1 mg/kg, ip, selective adenosine A2A receptor antagonist). In addition, the combined administration of subeffective doses of creatine and adenosine (0.1 mg/kg, ip, nonselective adenosine receptor agonist) or inosine (0.1 mg/kg, ip, nucleoside formed by the breakdown of adenosine) reduced immobility time in the TST. Moreover, the administration of subeffective doses of creatine or ketamine combined with N-6-cyclohexyladenosine (CHA) (0.05 mg/kg, ip, selective adenosine A1 receptor agonist), N-6-[2-(3,5-dimethoxyphenyl)-2-(methylphenyl)ethyl]adenosine (DPMA) (0.1 mg/kg, ip, selective adenosine A2A receptor agonist), or dipyridamole (0.1 μg/mouse, icv, adenosine transporter inhibitor) produced a synergistic antidepressant-like effect in the TST. These results indicate that creatine, similarly to ketamine, exhibits antidepressant-like effect in the TST probably mediated by the activation of both adenosine A1 and A2A receptors, further reinforcing the potential of targeting the purinergic system to the management of mood disorders.

  4. Pre-synaptic adenosine A2A receptors control cannabinoid CB1 receptor-mediated inhibition of striatal glutamatergic neurotransmission.

    PubMed

    Martire, Alberto; Tebano, Maria Teresa; Chiodi, Valentina; Ferreira, Samira G; Cunha, Rodrigo A; Köfalvi, Attila; Popoli, Patrizia

    2011-01-01

    An interaction between adenosine A(2A) receptors (A(2A) Rs) and cannabinoid CB(1) receptors (CB(1) Rs) has been consistently reported to occur in the striatum, although the precise mechanisms are not completely understood. As both receptors control striatal glutamatergic transmission, we now probed the putative interaction between pre-synaptic CB(1) R and A(2A) R in the striatum. In extracellular field potentials recordings in corticostriatal slices from Wistar rats, A(2A) R activation by CGS21680 inhibited CB(1) R-mediated effects (depression of synaptic response and increase in paired-pulse facilitation). Moreover, in superfused rat striatal nerve terminals, A(2A) R activation prevented, while A(2A) R inhibition facilitated, the CB(1) R-mediated inhibition of 4-aminopyridine-evoked glutamate release. In summary, the present study provides converging neurochemical and electrophysiological support for the occurrence of a tight control of CB(1) R function by A(2A) Rs in glutamatergic terminals of the striatum. In view of the key role of glutamate to trigger the recruitment of striatal circuits, this pre-synaptic interaction between CB(1) R and A(2A) R may be of relevance for the pathogenesis and the treatment of neuropsychiatric disorders affecting the basal ganglia.

  5. Regulation of fear responses by striatal and extrastriatal adenosine A2A receptors in forebrain.

    PubMed

    Wei, Catherine J; Augusto, Elisabete; Gomes, Catarina A; Singer, Philipp; Wang, Yumei; Boison, Detlev; Cunha, Rodrigo A; Yee, Benjamin K; Chen, Jiang-Fan

    2014-06-01

    Adenosine A2A receptors (A2ARs) are enriched in the striatum but are also present at lower levels in the extrastriatal forebrain (i.e., hippocampus, cortex), integrating dopamine, glutamate, and brain-derived neurotrophic factor (BDNF) signaling, and are thus essential for striatal neuroplasticity and fear and anxiety behavior. We tested two brain region-specific A2AR knockout lines with A2ARs selectively deleted either in the striatum (st-A2AR KO) or the entire forebrain (striatum, hippocampus, and cortex [fb-A2AR KO]) on fear and anxiety-related responses. We also examined the effect of hippocampus-specific A2AR deletion by local injection of adeno-associated virus type 5 (AAV5)-Cre into floxed-A2AR knockout mice. Selectively deleting A2ARs in the striatum increased Pavlovian fear conditioning (both context and tone) in st-A2AR KO mice, but extending the deletion to the rest of the forebrain apparently spared context fear conditioning and attenuated tone fear conditioning in fb-A2AR KO mice. Moreover, focal deletion of hippocampal A2ARs by AAV5-Cre injection selectively attenuated context (but not tone) fear conditioning. Deletion of A2ARs in the entire forebrain in fb-A2AR KO mice also produced an anxiolytic phenotype in both the elevated plus maze and open field tests, and increased the startle response. These extrastriatal forebrain A2AR behavioral effects were associated with reduced BDNF levels in the fb-A2AR KO hippocampus. This study provides evidence that inactivation of striatal A2ARs facilitates Pavlovian fear conditioning, while inactivation of extrastriatal A2ARs in the forebrain inhibits fear conditioning and also affects anxiety-related behavior. Copyright © 2014. Published by Elsevier Inc.

  6. A2A Adenosine Receptor (A2AAR) as a Therapeutic Target in Diabetic Retinopathy

    PubMed Central

    Ibrahim, Ahmed S.; El-shishtawy, Mamdouh M.; Zhang, Wenbo; Caldwell, Ruth B.; Liou, Gregory I.

    2011-01-01

    In diabetic retinopathy (DR), abnormalities in vascular and neuronal function are closely related to the local production of inflammatory mediators whose potential source is microglia. A2A adenosine receptor (A2AAR) has been shown to possess anti-inflammatory properties that have not been studied in DR. Here, we evaluate the role of A2AAR and its underlying signaling in retinal complications associated with diabetes. Initial studies in wild-type mice revealed that the treatment with the A2AAR agonist resulted in marked decreases in hyperglycemia-induced retinal cell death and tumor necrosis factor (TNF)-α release. To further assess the role of A2AAR in DR, we studied the effects of A2AAR ablation on diabetes-induced retinal abnormalities. Diabetic A2AAR−/− mice had significantly more terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling-positive cells, TNF-α release, and intercellular adhesion molecule-1 expression compared with diabetic wild-type mice. To explore a potential mechanism by which A2AAR signaling regulates inflammation in DR, we performed additional studies using microglial cells treated with Amadori-glycated albumin, a risk factor in diabetic disorders. The results showed that activation of A2AAR attenuated Amadori-glycated albumin-induced TNF-α release in a cAMP/exchange protein directly activated by cAMP-dependent mechanism and significantly repressed the inflammatory cascade, C-Raf/extracellular signal-regulated kinase (ERK), in activated microglia. Collectively, this work provides pharmacological and genetic evidence for A2AAR signaling as a control point of cell death in DR and suggests that the retinal protective effect of A2AAR is mediated by abrogating the inflammatory response that occurs in microglia via interaction with C-Raf/ERK pathway. PMID:21514428

  7. Macrophage A2A Adenosinergic Receptor Modulates Oxygen-Induced Augmentation of Murine Lung Injury

    PubMed Central

    D’Alessio, Franco R.; Eto, Yoshiki; Chau, Eric; Avalos, Claudia; Waickman, Adam T.; Garibaldi, Brian T.; Mock, Jason R.; Files, Daniel C.; Sidhaye, Venkataramana; Polotsky, Vsevolod Y.; Powell, Jonathan; Horton, Maureen; King, Landon S.

    2013-01-01

    Acute respiratory distress syndrome (ARDS) causes significant morbidity and mortality. Exacerbating factors increasing the risk of ARDS remain unknown. Supplemental oxygen is often necessary in both mild and severe lung disease. The potential effects of supplemental oxygen may include augmentation of lung inflammation by inhibiting anti-inflammatory pathways in alveolar macrophages. We sought to determine oxygen-derived effects on the anti-inflammatory A2A adenosinergic (ADORA2A) receptor in macrophages, and the role of the ADORA2A receptor in lung injury. Wild-type (WT) and ADORA2A−/− mice received intratracheal lipopolysaccharide (IT LPS), followed 12 hours later by continuous exposure to 21% oxygen (control mice) or 60% oxygen for 1 to 3 days. We measured the phenotypic endpoints of lung injury and the alveolar macrophage inflammatory state. We tested an ADORA2A-specific agonist, CGS-21680 hydrochloride, in LPS plus oxygen-exposed WT and ADORA2A−/− mice. We determined the specific effects of myeloid ADORA2A, using chimera experiments. Compared with WT mice, ADORA2A−/− mice exposed to IT LPS and 60% oxygen demonstrated significantly more histologic lung injury, alveolar neutrophils, and protein. Macrophages from ADORA2A−/− mice exposed to LPS plus oxygen expressed higher concentrations of proinflammatory cytokines and cosignaling molecules. CGS-21680 prevented the oxygen-induced augmentation of lung injury after LPS only in WT mice. Chimera experiments demonstrated that the transfer of WT but not ADORA2A−/− bone marrow cells into irradiated ADORA2A−/− mice reduced lung injury after LPS plus oxygen, demonstrating myeloid ADORA2A protection. ADORA2A is protective against lung injury after LPS and oxygen. Oxygen after LPS increases macrophage activation to augment lung injury by inhibiting the ADORA2A pathway. PMID:23349051

  8. The Immunosuppressive Role of Adenosine A2A Receptors in Ischemia Reperfusion Injury and Islet Transplantation

    PubMed Central

    Chhabra, Preeti; Linden, Joel; Lobo, Peter; Okusa, Mark Douglas; Brayman, Kenneth Lewis

    2014-01-01

    Activation of adenosine A2A receptors (A2AR) reduces inflammation by generally inhibiting the activation of pro-inflammatory cells, decreasing endothelial adhesion molecule expression and reducing the release of proinflammatory cytokine mediators. Numerous preclinical studies using selective A2AR agonists, antagonists, A2AR knockout as well as chimeric mice have suggested the therapeutic potential of A2AR agonists for the treatment of ischemia reperfusion injury (IRI) and autoimmune diseases. This review summarizes the immunosuppressive actions of A2AR agonists in murine IRI models of liver, kidney, heart, lung and CNS, and gives details on the cellular effects of A2AR activation in neutrophils, macrophages, dendritic cells, natural killer cells, NKT cells, T effector cells and CD4+CD25+FoxP3+ T regulatory cells. This is discussed in the context of cytokine mediators involved in inflammatory cascades. Whilst the role of adenosine receptor agonists in various models of autoimmune disease has been well-documented, very little information is available regarding the role of A2AR activation in type 1 diabetes mellitus (T1DM). An overview of the pathogenesis of T1DM as well as early islet graft rejection in the immediate peri-transplantation period offers insight regarding the use of A2AR agonists as a beneficial intervention in clinical islet transplantation, promoting islet graft survival, minimizing early islet loss and reducing the number of islets required for successful transplantation, thereby increasing the availability of this procedure to a greater number of recipients. In summary, the use of A2AR agonists as a clinical intervention in IRI and as an adjunct to clinical immunesuppressive regimen in islet transplantation is highlighted. PMID:22934547

  9. Allosteric mechanisms within the adenosine A2A-dopamine D2 receptor heterotetramer.

    PubMed

    Ferré, Sergi; Bonaventura, Jordi; Tomasi, Dardo; Navarro, Gemma; Moreno, Estefanía; Cortés, Antonio; Lluís, Carme; Casadó, Vicent; Volkow, Nora D

    2016-05-01

    The structure constituted by a G protein coupled receptor (GPCR) homodimer and a G protein provides a main functional unit and oligomeric entities can be viewed as multiples of dimers. For GPCR heteromers, experimental evidence supports a tetrameric structure, comprised of two different homodimers, each able to signal with its preferred G protein. GPCR homomers and heteromers can act as the conduit of allosteric interactions between orthosteric ligands. The well-known agonist/agonist allosteric interaction in the adenosine A2A receptor (A2AR)-dopamine D2 receptor (D2R) heteromer, by which A2AR agonists decrease the affinity of D2R agonists, gave the first rationale for the use of A2AR antagonists in Parkinson's disease. We review new pharmacological findings that can be explained in the frame of a tetrameric structure of the A2AR-D2R heteromer: first, ligand-independent allosteric modulations by the D2R that result in changes of the binding properties of A2AR ligands; second, differential modulation of the intrinsic efficacy of D2R ligands for G protein-dependent and independent signaling; third, the canonical antagonistic Gs-Gi interaction within the frame of the heteromer; and fourth, the ability of A2AR antagonists, including caffeine, to also exert the same allosteric modulations of D2R ligands than A2AR agonists, while A2AR agonists and antagonists counteract each other's effects. These findings can have important clinical implications when evaluating the use of A2AR antagonists. They also call for the need of monitoring caffeine intake when evaluating the effect of D2R ligands, when used as therapeutic agents in neuropsychiatric disorders or as probes in imaging studies. This article is part of the Special Issue entitled 'Purines in Neurodegeneration and Neuroregeneration'. Published by Elsevier Ltd.

  10. Genetic polymorphism of the adenosine A2A receptor is associated with habitual caffeine consumption.

    PubMed

    Cornelis, Marilyn C; El-Sohemy, Ahmed; Campos, Hannia

    2007-07-01

    Caffeine is the most widely consumed stimulant in the world, and individual differences in response to its stimulating effects may explain some of the variability in caffeine consumption within a population. We examined whether genetic variability in caffeine metabolism [cytochrome P450 1A2 (CYP1A2) -163A-->C] or the main target of caffeine action in the nervous system [adenosine A(2A) receptor (ADORA2A) 1083C-->T] is associated with habitual caffeine consumption. Subjects (n=2735) were participants from a study of gene-diet interactions and risk of myocardial infarction who did not have a history of hypertension. Genotype frequencies were examined among persons who were categorized according to their self-reported daily caffeine intake, as assessed with a validated food-frequency questionnaire. The ADORA2A, but not the CYP1A2, genotype was associated with different amounts of caffeine intake. Compared with persons consuming <100 mg caffeine/d, the odds ratios for having the ADORA2A TT genotype were 0.74 (95% CI: 0.53, 1.03), 0.63 (95% CI: 0.48, 0.83), and 0.57 (95% CI: 0.42, 0.77) for those consuming 100-200, >200-400, and >400 mg caffeine/d, respectively. The association was more pronounced among current smokers than among nonsmokers (P for interaction = 0.07). Persons with the ADORA2A TT genotype also were significantly more likely to consume less caffeine (ie, <100 mg/d) than were carriers of the C allele [P=0.011 (nonsmokers), P=0.008 (smokers)]. Our findings show that the probability of having the ADORA2A 1083TT genotype decreases as habitual caffeine consumption increases. This observation provides a biologic basis for caffeine consumption behavior and suggests that persons with this genotype may be less vulnerable to caffeine dependence.

  11. A differential role for the adenosine A2A receptor in opiate reinforcement vs opiate-seeking behavior.

    PubMed

    Brown, Robyn Mary; Short, Jennifer Lynn; Cowen, Michael Scott; Ledent, Catherine; Lawrence, Andrew John

    2009-03-01

    The adenosine A(2A) receptor is specifically enriched in the medium spiny neurons that make up the 'indirect' output pathway from the ventral striatum, a structure known to have a crucial, integrative role in processes such as reward, motivation, and drug-seeking behavior. In the present study we investigated the impact of adenosine A(2A) receptor deletion on behavioral responses to morphine in a number of reward-related paradigms. The acute, rewarding effects of morphine were evaluated using the conditioned place preference paradigm. Operant self-administration of morphine on both fixed and progressive ratio schedules as well as cue-induced drug-seeking was assessed. In addition, the acute locomotor response to morphine as well as sensitization to morphine was evaluated. Decreased morphine self-administration and breakpoint in A(2A) knockout mice was observed. These data support a decrease in motivation to consume the drug, perhaps reflecting diminished rewarding effects of morphine in A(2A) knockout mice. In support of this finding, a place preference to morphine was not observed in A(2A) knockout mice but was present in wild-type mice. In contrast, robust cue-induced morphine-seeking behavior was exhibited by both A(2A) knockout and wild-type mice after a period of withdrawal. The acute locomotor response to morphine in the A(2A) knockout was similar to wild-type mice, yet A(2A) knockout mice did not display tolerance to chronic morphine under the present paradigm. Both genotypes display locomotor sensitization to morphine, implying a lack of a role for the A(2A) receptor in the drug-induced plasticity necessary for the development or expression of sensitization. Collectively, these data suggest a differential role for adenosine A(2A) receptors in opiate reinforcement compared to opiate-seeking.

  12. Deletion of striatal adenosine A(2A) receptor spares latent inhibition and prepulse inhibition but impairs active avoidance learning.

    PubMed

    Singer, Philipp; Wei, Catherine J; Chen, Jiang-Fan; Boison, Detlev; Yee, Benjamin K

    2013-04-01

    Following early clinical leads, the adenosine A(2A)R receptor (A(2A)R) has continued to attract attention as a potential novel target for treating schizophrenia, especially against the negative and cognitive symptoms of the disease because of A(2A)R's unique modulatory action over glutamatergic in addition to dopaminergic signaling. Through (i) the antagonistic interaction with the dopamine D(2) receptor, and (ii) the regulation of glutamate release and N-methyl-d-aspartate receptor function, striatal A(2A)R is ideally positioned to fine-tune the dopamine-glutamate balance, the disturbance of which is implicated in the pathophysiology of schizophrenia. However, the precise function of striatal A(2A)Rs in the regulation of schizophrenia-relevant behavior is poorly understood. Here, we tested the impact of conditional striatum-specific A(2A)R knockout (st-A(2A)R-KO) on latent inhibition (LI) and prepulse inhibition (PPI) - behavior that is tightly regulated by striatal dopamine and glutamate. These are two common cross-species translational tests for the assessment of selective attention and sensorimotor gating deficits reported in schizophrenia patients; and enhanced performance in these tests is associated with antipsychotic drug action. We found that neither LI nor PPI was significantly affected in st-A(2A)R-KO mice, although a deficit in active avoidance learning was identified in these animals. The latter phenotype, however, was not replicated in another form of aversive conditioning - namely, conditioned taste aversion. Hence, the present study shows that neither learned inattention (as measured by LI) nor sensory gating (as indexed by PPI) requires the integrity of striatal A(2A)Rs - a finding that may undermine the hypothesized importance of A(2A)R in the genesis and/or treatment of schizophrenia.

  13. Selective adenosine A2A receptor agonists and antagonists protect against spinal cord injury through peripheral and central effects

    PubMed Central

    2011-01-01

    Background Permanent functional deficits following spinal cord injury (SCI) arise both from mechanical injury and from secondary tissue reactions involving inflammation. Enhanced release of adenosine and glutamate soon after SCI represents a component in the sequelae that may be responsible for resulting functional deficits. The role of adenosine A2A receptor in central ischemia/trauma is still to be elucidated. In our previous studies we have demonstrated that the adenosine A2A receptor-selective agonist CGS21680, systemically administered after SCI, protects from tissue damage, locomotor dysfunction and different inflammatory readouts. In this work we studied the effect of the adenosine A2A receptor antagonist SCH58261, systemically administered after SCI, on the same parameters. We investigated the hypothesis that the main action mechanism of agonists and antagonists is at peripheral or central sites. Methods Spinal trauma was induced by extradural compression of SC exposed via a four-level T5-T8 laminectomy in mouse. Three drug-dosing protocols were utilized: a short-term systemic administration by intraperitoneal injection, a chronic administration via osmotic minipump, and direct injection into the spinal cord. Results SCH58261, systemically administered (0.01 mg/kg intraperitoneal. 1, 6 and 10 hours after SCI), reduced demyelination and levels of TNF-α, Fas-L, PAR, Bax expression and activation of JNK mitogen-activated protein kinase (MAPK) 24 hours after SCI. Chronic SCH58261 administration, by mini-osmotic pump delivery for 10 days, improved the neurological deficit up to 10 days after SCI. Adenosine A2A receptors are physiologically expressed in the spinal cord by astrocytes, microglia and oligodendrocytes. Soon after SCI (24 hours), these receptors showed enhanced expression in neurons. Both the A2A agonist and antagonist, administered intraperitoneally, reduced expression of the A2A receptor, ruling out the possibility that the neuroprotective effects

  14. A-1 Test Stand modifications

    NASA Image and Video Library

    2011-09-14

    Team members check the progress of a liquid nitrogen cold shock test on the A-1 Test Stand at Stennis Space Center on Sept. 15. The cold shock test is used to confirm the test stand's support system can withstand test conditions, when super-cold rocket engine propellant is piped. The A-1 Test Stand is preparing to conduct tests on the powerpack component of the J-2X rocket engine, beginning in early 2012.

  15. Past, present and future of A(2A) adenosine receptor antagonists in the therapy of Parkinson's disease.

    PubMed

    Armentero, Marie Therese; Pinna, Annalisa; Ferré, Sergi; Lanciego, José Luis; Müller, Christa E; Franco, Rafael

    2011-12-01

    Several selective antagonists for adenosine A(2A) receptors (A(2A)R) are currently under evaluation in clinical trials (phases I to III) to treat Parkinson's disease, and they will probably soon reach the market. The usefulness of these antagonists has been deduced from studies demonstrating functional interactions between dopamine D₂ and adenosine A(2A) receptors in the basal ganglia. At present it is believed that A(2A)R antagonists can be used in combination with the dopamine precursor L-DOPA to minimize the motor symptoms of Parkinson's patients. However, a considerable body of data indicates that in addition to ameliorating motor symptoms, adenosine A(2A)R antagonists may also prevent neurodegeneration. Despite these promising indications, one further issue must be considered in order to develop fully optimized antiparkinsonian drug therapy, namely the existence of (hetero)dimers/oligomers of G protein-coupled receptors, a topic that is currently the focus of intense debate within the scientific community. Dopamine D₂ receptors (D₂Rs) expressed in the striatum are known to form heteromers with A(2A) adenosine receptors. Thus, the development of heteromer-specific A(2A) receptor antagonists represents a promising strategy for the identification of more selective and safer drugs.

  16. Adenosine A(2A) receptor activation prevents progressive kidney fibrosis in a model of immune-associated chronic inflammation.

    PubMed

    Garcia, Gabriela E; Truong, Luan D; Chen, Jiang-Fan; Johnson, Richard J; Feng, Lili

    2011-08-01

    Crescentic glomerulonephritis (GN) in Wistar-Kyoto rats progresses to lethal kidney failure by macrophage (Mφ)-mediated mechanisms. Mφs in nephritic glomeruli express adenosine A(2A) receptors (A(2A)Rs), the activation of which suppresses inflammation. Here, we pharmacologically activated the A(2A)Rs with a selective agonist, CGS 21680, and inactivated them with a selective antagonist, ZM241385, to test the effects on established GN. When activation was delayed until antiglomerular basement membrane GN and extracellular matrix deposition were established, glomerular Mφ infiltration was reduced by 83%. There was also a marked improvement in glomerular lesion histology, as well as decreased proteinuria. A(2A)R activation significantly reduced type I, III, and IV collagen deposition, and E-cadherin expression was restored in association with a reduction of α-smooth muscle actin-positive myofibroblasts in the interstitium and glomeruli. In contrast, pharmacological inactivation of A(2A)Rs increased glomerular crescent formation, type I, III, and IV collagen expression, and enhanced E-cadherin loss. Activation of A(2A)Rs suppressed the expression of the Mφ-linked glomerular damage mediators, transforming growth factor-β, osteopontin-1, thrombospondin-1, and tissue inhibitor of metalloproteinase-1. Thus, A(2A)R activation can arrest GN and prevent progressive fibrosis in established pathological lesions.

  17. A2A-D2 receptor-receptor interaction modulates gliotransmitter release from striatal astrocyte processes.

    PubMed

    Cervetto, Chiara; Venturini, Arianna; Passalacqua, Mario; Guidolin, Diego; Genedani, Susanna; Fuxe, Kjell; Borroto-Esquela, Dasiel O; Cortelli, Pietro; Woods, Amina; Maura, Guido; Marcoli, Manuela; Agnati, Luigi F

    2017-01-01

    Evidence for striatal A2A-D2 heterodimers has led to a new perspective on molecular mechanisms involved in schizophrenia and Parkinson's disease. Despite the increasing recognition of astrocytes' participation in neuropsychiatric disease vulnerability, involvement of striatal astrocytes in A2A and D2 receptor signal transmission has never been explored. Here, we investigated the presence of D2 and A2A receptors in isolated astrocyte processes prepared from adult rat striatum by confocal imaging; the effects of receptor activation were measured on the 4-aminopyridine-evoked release of glutamate from the processes. Confocal analysis showed that A2A and D2 receptors were co-expressed on the same astrocyte processes. Evidence for A2A-D2 receptor-receptor interactions was obtained by measuring the release of the gliotransmitter glutamate: D2 receptors inhibited the glutamate release, while activation of A2A receptors, per se ineffective, abolished the effect of D2 receptor activation. The synthetic D2 peptide VLRRRRKRVN corresponding to the receptor region involved in electrostatic interaction underlying A2A-D2 heteromerization abolished the ability of the A2A receptor to antagonize the D2 receptor-mediated effect. Together, the findings are consistent with heteromerization of native striatal astrocytic A2A-D2 receptors that via allosteric receptor-receptor interactions could play a role in the control of striatal glutamatergic transmission. These new findings suggest possible new pathogenic mechanisms and/or therapeutic approaches to neuropsychiatric disorders. © 2016 International Society for Neurochemistry.

  18. Synthesis of [1,2,4]triazolo[1,5-a]pyrazines as adenosine A2A receptor antagonists.

    PubMed

    Dowling, James E; Vessels, Jeffrey T; Haque, Serajul; Chang, He Xi; van Vloten, Kurt; Kumaravel, Gnanasambandam; Engber, Thomas; Jin, Xiaowei; Phadke, Deepali; Wang, Joy; Ayyub, Eman; Petter, Russell C

    2005-11-01

    Potent and selective antagonists of the adenosine A2A receptor often contain a nitrogen-rich fused-ring heterocyclic core. Replacement of the core with an isomeric ring system has previously been shown to improve target affinity, selectivity, and in vivo activity. This paper describes the preparation, by a novel route, of A2A receptor antagonists containing the [1,2,4]triazolo[1,5-a]pyrazine nucleus, which is isomeric with the [1,2,4]triazolo[1,5-c]pyrimidine core of a series of known A2A antagonists with in vivo activity in animal models of Parkinson's disease.

  19. The adenosine A2A receptor antagonist ZM241385 enhances neuronal survival after oxygen-glucose deprivation in rat CA1 hippocampal slices

    PubMed Central

    Pugliese, AM; Traini, C; Cipriani, S; Gianfriddo, M; Mello, T; Giovannini, MG; Galli, A; Pedata, F

    2009-01-01

    Background and purpose: Activation of adenosine A2A receptors in the CA1 region of rat hippocampal slices during oxygen-glucose deprivation (OGD), a model of cerebral ischaemia, was investigated. Experimental approach: We made extracellular recordings of CA1 field excitatory postsynaptic potentials (fepsps) followed by histochemical and immunohistochemical techniques coupled to Western blots. Key results: OGD (7 or 30 min duration) elicited an irreversible loss of fepsps invariably followed by the appearance of anoxic depolarization (AD), an unambiguous sign of neuronal damage. The application of the selective adenosine A2A receptor antagonist, ZM241385 (4-(2-[7-amino-2-{2-furyl}{1,2,4}triazolo{2,3-a}{1,3,5}triazin-5-ylamino]ethyl)phenol; 100–500 nmol·L−1) prevented or delayed AD appearance induced by 7 or 30 min OGD and protected from the irreversible fepsp depression elicited by 7 min OGD. Two different selective adenosine A2A receptor antagonists, SCH58261 and SCH442416, were less effective than ZM241385 during 7 min OGD. The extent of CA1 cell injury was assessed 3 h after the end of 7 min OGD by propidium iodide. Substantial CA1 pyramidal neuronal damage occurred in untreated slices, exposed to OGD, whereas injury was significantly prevented by 100 nmol·L−1 ZM241385. Glial fibrillary acid protein (GFAP) immunostaining showed that 3 h after 7 min OGD, astrogliosis was appreciable. Western blot analysis indicated an increase in GFAP 30 kDa fragment which was significantly reduced by treatment with 100 nmol·L−1 ZM241385. Conclusions and implications: In the CA1 hippocampus, antagonism of A2A adenosine receptors by ZM241385 was protective during OGD (a model of cerebral ischaemia) by delaying AD appearance, decreasing astrocyte activation and improving neuronal survival. PMID:19422385

  20. Identification and analysis of CYP7A1, CYP17A1, CYP20A1, CYP27A1 and CYP51A1 in cynomolgus macaques.

    PubMed

    Uno, Yasuhiro; Hosaka, Shinya; Yamazaki, Hiroshi

    2014-12-01

    Cytochromes P450 (P450) are important for not only drug metabolism and toxicity, but also biosynthesis and metabolism of cholesterol and bile acids, and steroid synthesis. In cynomolgus macaques, widely used in biomedical research, we have characterized P450 cDNAs, which were isolated as expressed sequence tags of cynomolgus macaque liver. In this study, cynomolgus CYP7A1, CYP17A1, CYP20A1, CYP27A1 and CYP51A1 cDNAs were characterized by sequence analysis, phylogenetic analysis and tissue expression pattern. By sequence analysis, these five cynomolgus P450s had high sequence identities (94-99%) to the human orthologs in amino acids. By phylogenetic analysis, each cynomolgus P450 was more closely related to the human ortholog as compared with the dog or rat ortholog. By quantitative polymerase chain reaction, among the 10 tissue types, CYP7A1 and CYP17A1 mRNAs were preferentially expressed in liver and adrenal gland, respectively. Cynomolgus CYP27A1 and CYP51A1 mRNAs were most abundantly expressed in liver and testis, respectively. Cynomolgus CYP20A1 mRNA was expressed in all the tissues, including brain and liver. Tissue expression patterns of each cynomolgus P450 were generally similar to that of the human ortholog. These results suggest the molecular similarities of CYP7A1, CYP17A1, CYP20A1, CYP27A1 and CYP51A1 between cynomolgus macaques and humans.

  1. Key Modulatory Role of Presynaptic Adenosine A2A Receptors in Cortical Neurotransmission to the Striatal Direct Pathway

    PubMed Central

    Quiroz, César; Luján, Rafael; Uchigashima, Motokazu; Simoes, Ana Patrícia; Lerner, Talia N.; Borycz, Janusz; Kachroo, Anil; Canas, Paula M.; Orru, Marco; Schwarzschild, Michael A.; Rosin, Diane L.; Kreitzer, Anatol C.; Cunha, Rodrigo A.; Watanabe, Masahiko; Ferré, Sergi

    2010-01-01

    Basal ganglia processing results from a balanced activation of direct and indirect striatal efferent pathways, which are controlled by dopamine D1 and D2 receptors, respectively. Adenosine A2A receptors are considered novel anti-parkinsonian targets, based on their selective postsynaptic localization in the indirect pathway, where they modulate D2 receptor function. The present study provides evidence for the existence of an additional functionally significant segregation of A2A receptors at the presynaptic level. Using integrated anatomical, electrophysiological and biochemical approaches, we demonstrate that presynaptic A2A receptors are preferentially localized in cortical glutamatergic terminals that contact striatal neurons of the direct pathway, where they exert a selective modulation of cortico-striatal neurotransmission. Presynaptic striatal A2A receptors could provide a new target for the treatment of neuropsychiatric disorders. PMID:19936569

  2. Key modulatory role of presynaptic adenosine A2A receptors in cortical neurotransmission to the striatal direct pathway.

    PubMed

    Quiroz, César; Luján, Rafael; Uchigashima, Motokazu; Simoes, Ana Patrícia; Lerner, Talia N; Borycz, Janusz; Kachroo, Anil; Canas, Paula M; Orru, Marco; Schwarzschild, Michael A; Rosin, Diane L; Kreitzer, Anatol C; Cunha, Rodrigo A; Watanabe, Masahiko; Ferré, Sergi

    2009-11-18

    Basal ganglia processing results from a balanced activation of direct and indirect striatal efferent pathways, which are controlled by dopamine D1 and D2 receptors, respectively. Adenosine A2A receptors are considered novel antiparkinsonian targets, based on their selective postsynaptic localization in the indirect pathway, where they modulate D2 receptor function. The present study provides evidence for the existence of an additional, functionally significant, segregation of A2A receptors at the presynaptic level. Using integrated anatomical, electrophysiological, and biochemical approaches, we demonstrate that presynaptic A2A receptors are preferentially localized in cortical glutamatergic terminals that contact striatal neurons of the direct pathway, where they exert a selective modulation of corticostriatal neurotransmission. Presynaptic striatal A2A receptors could provide a new target for the treatment of neuropsychiatric disorders.

  3. Adenosine A2A receptor blockade differentially influences excitotoxic mechanisms at pre- and postsynaptic sites in the rat striatum.

    PubMed

    Tebano, Maria Teresa; Pintor, Annita; Frank, Claudio; Domenici, Maria Rosaria; Martire, Alberto; Pepponi, Rita; Potenza, Rosa Luisa; Grieco, Rosa; Popoli, Patrizia

    2004-07-01

    Adenosine A(2A) receptor antagonists are being regarded as potential neuroprotective drugs, although the mechanisms underlying their effects need to be better studied. The aim of this work was to investigate further the mechanism of the neuroprotective action of A(2A) receptor antagonists in models of pre- and postsynaptic excitotoxicity. In microdialysis studies, the intrastriatal perfusion of the A(2A) receptor antagonist ZM 241385 (5 and 50 nM) significantly reduced, in an inversely dose-dependent way, the raise in glutamate outflow induced by 5 mM quinolinic acid (QA). In rat corticostriatal slices, ZM 241385 (30-100 nM) significantly reduced 4-aminopyridine (4-AP)-induced paired-pulse inhibition (PPI; an index of neurotransmitter release), whereas it worsened the depression of field potential amplitude elicited by N-methyl-D-aspartate (NMDA; 12.5 and 50 microM). The A(2A) antagonist SCH 58261 (30 nM) mimicked the effects of ZM 241385, whereas the A(2A) agonist CGS 21680 (100 nM) showed a protective influence toward 50 microM NMDA. In rat striatal neurons, 50 nM ZM 241385 did not affect the increase in [Ca(2+)](i) or the release of lactate dehydrogenase (LDH) induced by 100 and 300 microM NMDA, respectively. The ability of ZM 241385 to prevent QA-induced glutamate outflow and 4-AP-induced effects confirms that A(2A) receptor antagonists have inhibitory effects on neurotransmitter release, whereas the results obtained toward NMDA-induced effects suggest that A(2A) receptor blockade does not reduce, or even amplifies, excitotoxic mechanisms due to direct NMDA receptor stimulation. This indicates that the neuroprotective potential of A(2A) antagonists may be evident mainly in models of neurodegeneration in which presynaptic mechanisms play a major role.

  4. The GS Protein-coupled A2a Adenosine Receptor Controls T Cell Help in the Germinal Center.

    PubMed

    Abbott, Robert K; Silva, Murillo; Labuda, Jasmine; Thayer, Molly; Cain, Derek W; Philbrook, Phaethon; Sethumadhavan, Shalini; Hatfield, Stephen; Ohta, Akio; Sitkovsky, Michail

    2017-01-27

    T follicular helper (TFH) cells have been shown to be critically required for the germinal center (GC) reaction where B cells undergo class switch recombination and clonal selection to generate high affinity neutralizing antibodies. However, detailed knowledge of the physiological cues within the GC microenvironment that regulate T cell help is limited. The cAMP-elevating, Gs protein-coupled A2a adenosine receptor (A2aR) is an evolutionarily conserved receptor that limits and redirects cellular immunity. However, the role of A2aR in humoral immunity and B cell differentiation is unknown. We hypothesized that the hypoxic microenvironment within the GC facilitates an extracellular adenosine-rich milieu, which serves to limit TFH frequency and function, and also promotes immunosuppressive T follicular regulatory cells (TFR). In support of this hypothesis, we found that following immunization, mice lacking A2aR (A2aRKO) exhibited a significant expansion of T follicular cells, as well as increases in TFH to TFR ratio, GC T cell frequency, GC B cell frequency, and class switching of GC B cells to IgG1. Transfer of CD4 T cells from A2aRKO or wild type donors into T cell-deficient hosts revealed that these increases were largely T cell-intrinsic. Finally, injection of A2aR agonist, CGS21680, following immunization suppressed T follicular differentiation, GC B cell frequency, and class switching of GC B cells to IgG1. Taken together, these observations point to a previously unappreciated role of GS protein-coupled A2aR in regulating humoral immunity, which may be pharmacologically targeted during vaccination or pathological states in which GC-derived autoantibodies contribute to the pathology.

  5. Sinomenine protects against lipopolysaccharide-induced acute lung injury in mice via adenosine A(2A) receptor signaling.

    PubMed

    Li, Jun; Zhao, Li; He, Xie; Zeng, Yi-Jun; Dai, Shuang-Shuang

    2013-01-01

    Sinomenine (SIN) is a bioactive alkaloid extracted from the Chinese medicinal plant Sinomenium acutum, which is widely used in the clinical treatment of rheumatoid arthritis (RA). However, its role in acute lung injury (ALI) is unclear. In this study, we investigate the role of SIN in lipopolysaccharide (LPS)-induced ALI in mice. After ALI, lung water content and histological signs of pulmonary injury were attenuated, whereas the PaO2/FIO2 (P/F) ratios were elevated significantly in the mice pretreated with SIN. Additionally, SIN markedly inhibited inflammatory cytokine TNF-α and IL-1β expression levels as well as neutrophil infiltration in the lung tissues of the mice. Microarray analysis and real-time PCR showed that SIN treatment upregulated adenosine A(2A) receptor (A(2A)R) expression, and the protective effect of SIN was abolished in A(2A)R knockout mice. Further investigation in isolated mouse neutrophils confirmed the upregulation of A(2A)R by SIN and showed that A(2A)R-cAMP-PKA signaling was involved in the anti-inflammatory effect of SIN. Taken together, these findings demonstrate an A(2A)R-associated anti-inflammatory effect and the protective role of SIN in ALI, which suggests a potential novel approach to treat ALI.

  6. Hermes A-1 Test Rockets

    NASA Technical Reports Server (NTRS)

    1950-01-01

    The first Hermes A-1 test rocket was fired at White Sand Proving Ground (WSPG). Hermes was a modified V-2 German rocket, utilizing the German aerodynamic configuration; however, internally it was a completely new design. Although it did not result in an operational vehicle, the information that was gathered in the process contributed directly to the development of the Redstone rocket.

  7. Insilico study of the A(2A)R-D (2)R kinetics and interfacial contact surface for heteromerization.

    PubMed

    Prakash, Amresh; Luthra, Pratibha Mehta

    2012-10-01

    G-protein-coupled receptors (GPCRs) are cell surface receptors. The dynamic property of receptor-receptor interactions in GPCRs modulates the kinetics of G-protein signaling and stability. In the present work, the structural and dynamic study of A(2A)R-D(2)R interactions was carried to acquire the understanding of the A(2A)R-D(2)R receptor activation and deactivation process, facilitating the design of novel drugs and therapeutic target for Parkinson's disease. The structure-based features (Alpha, Beta, SurfAlpha, and SurfBeta; GapIndex, Leakiness and Gap Volume) and slow mode model (ENM) facilitated the prediction of kinetics (K (off), K (on), and K (d)) of A(2A)R-D(2)R interactions. The results demonstrated the correlation coefficient 0.294 for K (d) and K (on) and the correlation coefficient 0.635 for K (d) and K (off), and indicated stable interfacial contacts in the formation of heterodimer. The coulombic interaction involving the C-terminal tails of the A(2A)R and intracellular loops (ICLs) of D(2)R led to the formation of interfacial contacts between A(2A)R-D(2)R. The properties of structural dynamics, ENM and KFC server-based hot-spot analysis illustrated the stoichiometry of A(2A)R-D(2)R contact interfaces as dimer. The propensity of amino acid residues involved in A(2A)R-D(2)R interaction revealed the presence of positively (R, H and K) and negatively (E and D) charged structural motif of TMs and ICL3 of A(2A)R and D(2)R at interface of dimer contact. Essentially, in silico structural and dynamic study of A(2A)R-D(2)R interactions will provide the basic understanding of the A(2A)R-D(2)R interfacial contact surface for activation and deactivation processes, and could be used as constructive model to recognize the protein-protein interactions in receptor assimilations.

  8. Novel 8-(furan-2-yl)-3-benzyl thiazolo [5,4-e][1,2,4] triazolo [1,5-c] pyrimidine-2(3H)-thione as selective adenosine A(2A) receptor antagonist.

    PubMed

    Barodia, Sandeep Kumar; Mishra, Chandra Bhushan; Prakash, Amresh; Senthil Kumar, J B; Kumari, Namrata; Luthra, Pratibha Mehta

    2011-01-13

    Adenosine A(2A) receptor (A(2A)R) antagonists have emerged as potential drug candidates to alleviate progression and symptoms of Parkinson's disease (PD), and reduce the dopaminergic side effects. The synthesis of novel compound 8-(furan-2-yl)-3-benzyl thiazolo [5,4-e][1,2,4] triazolo [1,5-c] pyrimidine-2-(3H)-thione (BTTP) was carried out to evaluate the potential of BTTP as A(2A)R antagonist using SCH58261, a standard A(2A)R antagonist. The strong interaction of BTTP with A(2A)R (ΔG=-12.46kcal/mol and K(i)=0.6nM) in silico analysis was confirmed by radioligand receptor binding studies showing high affinity (K(i)=0.004nM) and selectivity with A(2A)R (A(2A)/A(1)=1155-fold). The effect of CGS21680 (selective A(2A)R agonist) induced cAMP concentration (0.1pmol/ml) in HEK293 cells was antagonized with BTTP (0.065pmol/ml) and SCH58261 (0.075pmol/ml). Furthermore, BTTP pre-treated (5, 10 and 20mg/kg) haloperidol-induced mice demonstrated significant attenuation in catalepsy and akinesia. BTTP induced elevation in the striatal dopamine concentration (2.90μM/mg of tissue) was comparable to SCH58261 (2.92μM/mg of tissue) at the dose of 10mg/kg. The results firmly articulate that BTTP possesses potential A(2A)R antagonist activity and can be further explored for the treatment of PD. Copyright © 2010 Elsevier Ireland Ltd. All rights reserved.

  9. Synergistic Up-Regulation of Vascular Endothelial Growth Factor Expression in Murine Macrophages by Adenosine A2A Receptor Agonists and Endotoxin

    PubMed Central

    Leibovich, Samuel Joseph; Chen, Jiang-Fan; Pinhal-Enfield, Grace; Belem, Paula C.; Elson, Genie; Rosania, Anthony; Ramanathan, Madhuri; Montesinos, Carmen; Jacobson, Marlene; Schwarzschild, Michael A.; Fink, J. Stephen; Cronstein, Bruce

    2002-01-01

    Under normoxic conditions, macrophages from C57BL mice produce low levels of vascular endothelial growth factor (VEGF). Hypoxia stimulates VEGF expression by ∼500%; interferon-γ (IFN-γ) with endotoxin [lipopolysaccharide (LPS)] also stimulates VEGF expression by ∼50 to 150% in an inducible nitric oxide synthase (iNOS)-dependent manner. Treatment of normoxic macrophages with 5′-N-ethyl-carboxamido-adenosine (NECA), a nonselective adenosine A2 receptor agonist, or with 2-[p-(2-carboxyethyl)-phenylethyl amino]-5′-N-ethyl-carboxamido-adenosine (CGS21680), a specific adenosine A2A receptor agonist, modestly increases VEGF expression, whereas 2-chloro-N6-cyclopentyl adenosine (CCPA), an adenosine A1 agonist, does not. Treatment with LPS (0 to 1000 ng/ml), or with IFN-γ (0 to 300 U/ml), does not affect VEGF expression. In the presence of LPS (EC50 < 10 ng/ml), but not of IFN-γ, both NECA and CGS21680 synergistically up-regulate VEGF expression by as much as 10-fold. This VEGF is biologically active in vivo in the rat corneal bioassay of angiogenesis. Inhibitors of iNOS do not affect this synergistic induction of VEGF, and macrophages from iNOS−/− mice produce similar levels of VEGF as wild-type mice, indicating that NO does not play a role in this induction. Under hypoxic conditions, VEGF expression is slightly increased by adenosine receptor agonists but adenosine A2 or A1 receptor antagonists 3,7-dimethyl-1-propargyl xanthine (DMPX), ZM241385, and 8-cyclopentyl-1,3-dipropylxanthine (DCPCX) do not modulate VEGF expression. VEGF expression is also not reduced in hypoxic macrophages from A3−/− and A2A−/− mice. Thus, VEGF expression by hypoxic macrophages does not seem to depend on endogenously released or exogenous adenosine. VEGF expression is strongly up-regulated by LPS/NECA in macrophages from A3−/− but not A2A−/− mice, confirming the role of adenosine A2A receptors in this pathway. LPS with NECA strongly up-regulates VEGF expression by

  10. GDNF control of the glutamatergic cortico-striatal pathway requires tonic activation of adenosine A2A Receptors

    PubMed Central

    Gomes, Catarina A.R.V.; Simões, Patrícia F.; Canas, Paula M.; Quiroz, César; Sebastião, Ana M.; Ferré, Sergi; Cunha, Rodrigo A.; Ribeiro, Joaquim A.

    2009-01-01

    Glial cell line-derived neurotrophic factor (GDNF) affords neuroprotection in Parkinson’s disease in accordance with its ability to bolster nigrostriatal innervation. We previously found that GDNF facilitates dopamine release in a manner dependent on adenosine A2A receptor activation. Since motor dysfunction also involves modifications of striatal glutamatergic innervation, we now tested if GDNF and its receptor system, Ret (rearranged during transfection) and GFRα1 (GDNF family receptor alpha 1) controlled the cortico-striatal glutamatergic pathway in an A2A receptor-dependent manner. GDNF (10 ng/ml) enhanced (by ≈13%) glutamate release from rat striatal nerve endings, an effect potentiated (up to ≈ 30%) by the A2A receptor agonist CGS 21680 (10 nM) and prevented by the A2A receptor antagonist, SCH 58261 (50 nM). Triple immunocytochemical studies revealed that Ret and GFRα1 were located in 50% of rat striatal glutamatergic terminals (immunopositive for vesicular glutamate transporters-1/2), where they were found to be co-located with A2A receptors. Activation of the glutamatergic system upon in vivo electrical stimulation of the rat cortico-striatal input induced striatal Ret phosphoprylation that was prevented by pre-treatment with the A2A receptor antagonist, MSX-3 (3 mg/kg). The results provide the first functional and morphological evidence that GDNF controls cortico-striatal glutamatergic pathways in a manner largely dependent on the co-activation of adenosine A2A receptors. PMID:19141075

  11. Adenosine A2A receptors enable the synaptic effects of cannabinoid CB1 receptors in the rodent striatum.

    PubMed

    Tebano, Maria Teresa; Martire, Alberto; Chiodi, Valentina; Pepponi, Rita; Ferrante, Antonella; Domenici, Maria Rosaria; Frank, Claudio; Chen, Jiang-Fan; Ledent, Catherine; Popoli, Patrizia

    2009-09-01

    Adenosine A(2A), cannabinoid CB(1) and metabotropic glutamate 5 (mGlu(5)) receptors are all highly expressed in the striatum. The aim of the present work was to investigate whether, and by which mechanisms, the above receptors interact in the regulation of striatal synaptic transmission. By extracellular field potentials (FPs) recordings in corticostriatal slices, we demonstrated that the ability of the selective type 1 cannabinoid receptor (CB(1)R) agonist WIN55,212-2 to depress synaptic transmission was prevented by the pharmacological blockade or the genetic inactivation of A(2A)Rs. Such a permissive effect of A(2A)Rs towards CB(1)Rs does not seem to occur pre-synaptically as the ability of WIN55,212-2 to increase the R2/R1 ratio under a protocol of paired-pulse stimulation was not modified by ZM241385. Furthermore, the effects of WIN55,212-2 were reduced in slices from mice lacking post-synaptic striatal A(2A)Rs. The selective mGlu(5)R agonist (RS)-2-chloro-5-hydroxyphenylglycine (CHPG) potentiated the synaptic effects of WIN55,212-2, and such a potentiation was abolished by A(2A)R blockade. Unlike the synaptic effects, the ability of WIN55,212-2 to prevent NMDA-induced toxicity was not influenced by ZM241385. Altogether, these results show that the state of activation of A(2A)Rs regulates the synaptic effects of CB(1)Rs and that A(2A)Rs may control CB(1) effects also indirectly, namely through mGlu(5)Rs.

  12. The importance of the adenosine A(2A) receptor-dopamine D(2) receptor interaction in drug addiction.

    PubMed

    Filip, M; Zaniewska, M; Frankowska, M; Wydra, K; Fuxe, K

    2012-01-01

    Drug addiction is a serious brain disorder with somatic, psychological, psychiatric, socio-economic and legal implications in the developed world. Illegal (e.g., psychostimulants, opioids, cannabinoids) and legal (alcohol, nicotine) drugs of abuse create a complex behavioral pattern composed of drug intake, withdrawal, seeking and relapse. One of the hallmarks of drugs that are abused by humans is that they have different mechanisms of action to increase dopamine (DA) neurotransmission within the mesolimbic circuitry of the brain and indirectly activate DA receptors. Among the DA receptors, D(2) receptors are linked to drug abuse and addiction because their function has been proven to be correlated with drug reinforcement and relapses. The recognition that D(2) receptors exist not only as homomers but also can form heteromers, such as with the adenosine (A)(2A) receptor, that are pharmacologically and functionally distinct from their constituent receptors, has significantly expanded the range of potential drug targets and provided new avenues for drug design in the search for novel drug addiction therapies. The aim of this review is to bring current focus on A(2A) receptors, their physiology and pharmacology in the central nervous system, and to discuss the therapeutic relevance of these receptors to drug addiction. We concentrate on the contribution of A(2A) receptors to the effects of different classes of drugs of abuse examined in preclinical behavioral experiments carried out with pharmacological and genetic tools. The consequences of chronic drug treatment on A(2A) receptor-assigned functions in preclinical studies are also presented. Finally, the neurochemical mechanism of the interaction between A(2A) receptors and drugs of abuse in the context of the heteromeric A(2A)-D(2) receptor complex is discussed. Taken together, a significant amount of experimental analyses provide evidence that targeting A(2A) receptors may offer innovative translational strategies

  13. Stabilizing interactions between D666-S1787 and T657-Y1792 at the A2-A3 interface support factor VIIIa stability in the blood clotting pathway.

    PubMed

    Monaghan, M; Wakabayashi, H; Griffiths, A E; Fay, P J

    2016-05-01

    Essentials Factor VIIIa (FVIIIa) is unstable due to loss of A2; D666 and Y1792 contribute to its stability. We conducted a study to identify the interactions made at these residues at the A2-A3 interface. We present evidence for stabilizing interactions between D666-S1787 and T657-Y1792 in FVIIIa. A D666C/S1788C variant with a disulfide A2-A3 linkage has a FVIIIa decay rate that is 1% of wild-type. Background Factor (F)VIIIa activity and stability depends on the non-covalent association of the A2 subunit with the A1/A3C1C2 dimer, but the interactions that contribute to A2 association are not well defined. Previous work had shown that D666A and Y1792F mutations at the A2-A3 interface resulted in increased FVIIIa decay, suggesting that the residues were involved in bonding interactions important for FVIIIa stability. Objectives Several potential hydrogen bonding partners of D666 and Y1792 across the A2-A3 interface were selected from the low-resolution FVIII crystal structure, and we used mutagenesis and biochemical analysis to examine the bonding interactions occurring at D666 and Y1792. Methods Using a series of stability and functional analyses, we analyzed FVIII variants in which D666 and Y1792 were each swapped with the residues of potential bonding partners. Results and conclusions We present evidence for hydrogen bonds between D666 and S1787 and between Y1792 and T657 that are important for FVIIIa stability. D666S/S1787D and T657Y/Y1792T variants each displayed wild-type (WT)-like FVIIIa stability and performed like WT FVIII in a series of functional analyses, whereas D666S, S1787D, and Y1792T single variants showed increased FVIIIa decay and a T657Y variant had little FVIIIa activity. These results suggest that WT hydrogen bonds are disrupted with the single mutations but maintained in the swap variants. Furthermore, mutation of D666 and S1788 to cysteine resulted in disulfide bond formation across the A2-A3 interface, confirming the close proximity of D666

  14. Modulating the Strength and Threshold of NOTCH Oncogenic Signals by mir-181a-1/b-1

    PubMed Central

    Wang, Song; Schaffert, Steven; Gong, Xue; Yue, Sibiao; Luong, Richard; Min, Hyeyoung; Yashiro-Ohtani, Yumi; Davis, Mark; Pear, Warren; Chen, Chang-Zheng

    2012-01-01

    Oncogenes, which are essential for tumor initiation, development, and maintenance, are valuable targets for cancer therapy. However, it remains a challenge to effectively inhibit oncogene activity by targeting their downstream pathways without causing significant toxicity to normal tissues. Here we show that deletion of mir-181a-1/b-1 expression inhibits the development of Notch1 oncogene-induced T cell acute lymphoblastic leukemia (T-ALL). mir-181a-1/b-1 controls the strength and threshold of Notch activity in tumorigenesis in part by dampening multiple negative feedback regulators downstream of NOTCH and pre-T cell receptor (TCR) signaling pathways. Importantly, although Notch oncogenes utilize normal thymic progenitor cell genetic programs for tumor transformation, comparative analyses of mir-181a-1/b-1 function in normal thymocyte and tumor development demonstrate that mir-181a-1/b-1 can be specifically targeted to inhibit tumor development with little toxicity to normal development. Finally, we demonstrate that mir-181a-1/b-1, but not mir-181a-2b-2 and mir-181-c/d, controls the development of normal thymic T cells and leukemia cells. Together, these results illustrate that NOTCH oncogene activity in tumor development can be selectively inhibited by targeting the molecular networks controlled by mir-181a-1/b-1. PMID:22916024

  15. A2A Adenosine Receptor Antagonism Enhances Synaptic and Motor Effects of Cocaine via CB1 Cannabinoid Receptor Activation

    PubMed Central

    Tozzi, Alessandro; de Iure, Antonio; Marsili, Valentina; Romano, Rosaria; Tantucci, Michela; Di Filippo, Massimiliano; Costa, Cinzia; Napolitano, Francesco; Mercuri, Nicola Biagio; Borsini, Franco; Giampà, Carmen; Fusco, Francesca Romana; Picconi, Barbara; Usiello, Alessandro; Calabresi, Paolo

    2012-01-01

    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. PMID:22715379

  16. A2A receptor antagonists do not induce dyskinesias in drug-naive or L-dopa sensitized rats.

    PubMed

    Jones, N; Bleickardt, C; Mullins, D; Parker, E; Hodgson, R

    2013-09-01

    L-dopa, the precursor to dopamine, is currently the gold standard treatment for Parkinson's disease (PD). However, chronic exposure is associated with L-dopa-induced dyskinesias (LIDs), a serious side effect characterized by involuntary movements. Adenosine A2A receptor antagonists have been studied as a novel non-dopaminergic PD treatment. Because A2A receptor antagonists do not act on dopamine receptors, it has been hypothesized that they will not induce dyskinesias characteristic of L-dopa. To test this hypothesis in a rodent model, the A2A receptor antagonists SCH 412348 (3 mg/kg), vipadenant (10 mg/kg), caffeine (30 mg/kg), or istradefylline (3 mg/kg) were chronically (19-22 days) administered to Sprague Dawley rats, and dyskinetic behaviors were scored across this chronic dosing paradigm. Unlike L-dopa, there was no evidence of dyskinetic activity resulting from any of the four A2A receptor antagonists tested. When delivered to animals previously sensitized with L-dopa (6 mg/kg), SCH 412348, vipadenant, caffeine or istradefylline treatment produced no dyskinesias. When administered in combination with L-dopa (6 mg/kg), SCH 412348 (3 mg/kg) neither exacerbated nor prevented the induction of LIDs over the course of 19 days of treatment. Collectively, our data indicate that A2A receptor antagonists are likely to have a reduced dyskinetic liability relative to L-dopa but do not block dyskinesias when coadministered with L-dopa. Clinical studies are required to fully understand the dyskinesia profiles of A2A receptor antagonists.

  17. In vivo expression of Nurr1/Nr4a2a in developing retinal amacrine subtypes in zebrafish Tg(nr4a2a:eGFP) transgenics.

    PubMed

    Goodings, Liana; He, Jie; Wood, Alasdair J; Harris, William A; Currie, Peter D; Jusuf, Patricia R

    2017-06-01

    The Nuclear receptor subfamily 4 group A member 2 (Nr4a2) is crucial for the formation or maintenance of dopaminergic neurons in the central nervous system including the retina, where dopaminergic amacrine cells contribute to visual function. Little is known about which cells express Nr4a2 at which developmental stage. Furthermore, whether Nr4a2 functions in combination with other genes is poorly understood. Thus, we generated a novel transgenic to visualize Nr4a2 expression in vivo during zebrafish retinogenesis. A 4.1 kb fragment of the nr4a2a promoter was used to drive green fluorescent protein expression in this Tg(nr4a2a:eGFP) line. In situ hybridization showed that transgene expression follows endogenous RNA expression at a cellular level. Temporal expression and lineages were quantified using in vivo time-lapse imaging in embryos. Nr4a2 expressing retinal subtypes were characterized immunohistochemically. Nr4a2a:eGFP labeled multiple neuron subtypes including 24.5% of all amacrine interneurons. Nr4a2a:eGFP labels all tyrosine hydroxylase labeled dopaminergic amacrine cells, and other nondopaminergic GABAergic amacrine populations. Nr4a2a:eGFP is confined to a specific progenitor lineage identified by sequential expression of the bhlh transcription factor Atonal7 (Atoh7) and Pancreas transcription factor 1a (Ptf1a), and labels postmitotic postmigratory amacrine cells. Thus, developmental Nr4a2a expression indicates a role during late differentiation of specific amacrine interneurons. Tg(nr4a2a:eGFP) is an early marker of distinct neurons including dopaminergic amacrine cells. It can be utilized to assess consequences of gene manipulations and understand whether Nr4a2 only carries out its role in the presence of specific coexpressed genes. This will allow Nr4a2 use to be refined for regenerative approaches. © 2017 Wiley Periodicals, Inc.

  18. Cannabidiol, a non-psychotropic plant-derived cannabinoid, decreases inflammation in a murine model of acute lung injury: role for the adenosine A(2A) receptor.

    PubMed

    Ribeiro, Alison; Ferraz-de-Paula, Viviane; Pinheiro, Milena L; Vitoretti, Luana B; Mariano-Souza, Domenica P; Quinteiro-Filho, Wanderley M; Akamine, Adriana T; Almeida, Vinícius I; Quevedo, João; Dal-Pizzol, Felipe; Hallak, Jaime E; Zuardi, Antônio W; Crippa, José A; Palermo-Neto, João

    2012-03-05

    Acute lung injury is an inflammatory condition for which treatment is mainly supportive because effective therapies have not been developed. Cannabidiol, a non-psychotropic cannabinoid component of marijuana (Cannabis sativa), has potent immunosuppressive and anti-inflammatory properties. Therefore, we investigated the possible anti-inflammatory effect of cannabidiol in a murine model of acute lung injury. Analysis of total inflammatory cells and differential in bronchoalveolar lavage fluid was used to characterize leukocyte migration into the lungs; myeloperoxidase activity of lung tissue and albumin concentration in the bronchoalveolar lavage fluid were analyzed by colorimetric assays; cytokine/chemokine production in the bronchoalveolar lavage fluid was also analyzed by Cytometric Bead Arrays and Enzyme-Linked Immunosorbent Assay (ELISA). A single dose of cannabidiol (20mg/kg) administered prior to the induction of LPS (lipopolysaccharide)-induced acute lung injury decreases leukocyte (specifically neutrophil) migration into the lungs, albumin concentration in the bronchoalveolar lavage fluid, myeloperoxidase activity in the lung tissue, and production of pro-inflammatory cytokines (TNF and IL-6) and chemokines (MCP-1 and MIP-2) 1, 2, and 4days after the induction of LPS-induced acute lung injury. Additionally, adenosine A(2A) receptor is involved in the anti-inflammatory effects of cannabidiol on LPS-induced acute lung injury because ZM241385 (4-(2-[7-Amino-2-(2-furyl)[1,2,4]triazolo[2,3-a][1,3,5]triazin-5-ylamino]ethyl)phenol) (a highly selective antagonist of adenosine A(2A) receptor) abrogated all of the anti-inflammatory effects of cannabidiol previously described. Thus, we show that cannabidiol has anti-inflammatory effects in a murine model of acute lung injury and that this effect is most likely associated with an increase in the extracellular adenosine offer and signaling through adenosine A(2A) receptor.

  19. Past, present and future of A2A adenosine receptor antagonists in the therapy of Parkinson’s disease

    PubMed Central

    Armentero, Marie Therese; Pinna, Annalisa; Ferré, Sergi; Lanciego, José Luis; Müller, Christa E.; Franco, Rafael

    2011-01-01

    Several selective antagonists for adenosine A2A receptors (A2AR) are currently under evaluation in clinical trials (phases I to III) to treat Parkinson’s disease, and they will probably soon reach the market. The usefulness of these antagonists has been deduced from studies demonstrating functional interactions between dopamine D2 and adenosine A2A receptors in the basal ganglia. At present it is believed that A2AR antagonists can be used in combination with the dopamine precursor L-DOPA to minimize the motor symptoms of Parkinson’s patients. However, a considerable body of data indicates that in addition to ameliorating motor symptoms, adenosine A2AR antagonists may also prevent neurodegeneration. Despite these promising indications, one further issue must be considered in order to develop fully optimized anti-parkinsonian drug therapy, namely the existence of receptor (hetero)dimers/oligomers of G protein-coupled receptors, a topic currently the focus of intense debate within the scientific community. Dopamine D2 receptors (D2Rs) expressed in the striatum are known to form heteromers with A2A adenosine receptors. Thus, the development of heteromer-specific A2A receptor antagonists represents a promising strategy for the identification of more selective and safer drugs. PMID:21810444

  20. Incorporation of rare earth elements in titanite: Stabilization of the A2/a dimorph by creation of antiphase boundaries

    USGS Publications Warehouse

    Hughes, J.M.; Bloodaxe, E.S.; Hanchar, J.M.; Foord, E.E.

    1997-01-01

    The atomic arrangement of a natural rare-earth-rich titanite and two synthetic rare-earth-doped titanites have been refined in space group A2/a, and the atomic arrangement of an undoped P21/a synthetic titanite was also refined for comparison. Previous work has shown that titanite possesses a domain structure, with domains formed of like-displaced Ti atoms in the [100] octahedral chains. P21/a titanite results when the crystal is formed of a single domain, but as Ti-reversal sites occur in the octahedral chain the apparent A2/a structure results from the average of antiphase domains. Antiphase boundaries occur at O1, which is alternately overbonded or underbonded at the boundaries, depending on the displacement of the neighboring Ti atoms. Type 2 antiphase boundaries exist where two Ti atoms are displaced away from the intervening O1 atom and are energetically unfavorable because of underbonding of that O1 atom. However, substitution of a trivalent rare earth element in the adjacent Ca2+ site relieves that underbonding, favoring the creation of type 2 antiphase boundaries and stabilization of the A2/a dimorph. The results of high-precision crystal structure analyses demonstrate that rare earth substituents for Ca stabilize the A2/a dimorph at lower substitution levels than required for octahedral substitutions.

  1. FGF acts as a co-transmitter through Adenosine A2A receptor to regulate morphological and physiological synaptic plasticity

    PubMed Central

    Flajolet, Marc; Wang, Zhongfeng; Futter, Marie; Shen, Weixing; Nuangchamnong, Nina; Bendor, Jacob; Palaszewski, Iwona; Nairn, Angus C.; Surmeier, D. James; Greengard, Paul

    2009-01-01

    Summary Abnormalities of striatal function have been implicated in several major neurological and psychiatric disorders, including Parkinson's disease, schizophrenia, and depression. Adenosine, by activation of A2A receptors, antagonizes dopamine signaling at D2 receptors and A2A receptor antagonists have been tested as therapeutic agents for Parkinson's disease. We report here a direct physical interaction between the G protein-coupled A2A receptor and the receptor tyrosine kinase FGF receptor. Concomitant activation of these two classes of receptors, but not individual activation of either one alone, causes a robust activation of the MAPK/ERK pathway, differentiation and neurite extension of PC12 cells, spine morphogenesis in primary neuronal cultures, and cortico-striatal plasticity induced by a novel A2AR/FGFR-dependent mechanism. The discovery of a direct physical interaction between the A2A and FGF receptors and the robust physiological consequences of this association shed light on the mechanism underlying FGF functions as a co-transmitter and open new avenues for therapeutic interventions. PMID:18953346

  2. Structure-Activity Relationships of the Sustained Effects of Adenosine A2A Receptor Agonists Driven by Slow Dissociation Kinetics

    PubMed Central

    Hothersall, J. Daniel; Guo, Dong; Sarda, Sunil; Sheppard, Robert J.; Chen, Hongming; Keur, Wesley; Waring, Michael J.; IJzerman, Adriaan P.; Hill, Stephen J.; Dale, Ian L.

    2017-01-01

    The duration of action of adenosine A2A receptor (A2A) agonists is critical for their clinical efficacy, and we sought to better understand how this can be optimized. The in vitro temporal response profiles of a panel of A2A agonists were studied using cAMP assays in recombinantly (CHO) and endogenously (SH-SY5Y) expressing cells. Some agonists (e.g., 3cd; UK-432,097) but not others (e.g., 3ac; CGS-21680) demonstrated sustained wash-resistant agonism, where residual receptor activation continued after washout. The ability of an antagonist to reverse pre-established agonist responses was used as a surrogate read-out for agonist dissociation kinetics, and together with radioligand binding studies suggested a role for slow off-rate in driving sustained effects. One compound, 3ch, showed particularly marked sustained effects, with a reversal t1/2 > 6 hours and close to maximal effects that remained for at least 5 hours after washing. Based on the structure-activity relationship of these compounds, we suggest that lipophilic N6 and bulky C2 substituents can promote stable and long-lived binding events leading to sustained agonist responses, although a high compound logD is not necessary. This provides new insight into the binding interactions of these ligands and we anticipate that this information could facilitate the rational design of novel long-acting A2A agonists with improved clinical efficacy. PMID:27803241

  3. Blunted dynamics of adenosine A2A receptors is associated with increased susceptibility to Candida albicans infection in the elderly

    PubMed Central

    Rodrigues, Lisa; Miranda, Isabel M.; Andrade, Geanne M.; Mota, Marta; Cortes, Luísa; Rodrigues, Acácio G.; Cunha, Rodrigo A.; Gonçalves, Teresa

    2016-01-01

    Opportunistic gut infections and chronic inflammation, in particular due to overgrowth of Candida albicans present in the gut microbiota, are increasingly reported in the elder population. In aged, adult and young mice, we now compared the relative intestinal over-colonization by ingested C. albicans and their translocation to other organs, focusing on the role of adenosine A2A receptors that are a main stop signal of inflammation. We report that elderly mice are more prone to over-colonization by C. albicans than adult and young mice. This fungal over-growth seems to be related with higher growth rate in intestinal lumen, independent of gut tissues invasion, but resulting in higher GI tract inflammation. We observed a particularly high colonization of the stomach, with increased rate of yeast-to-hypha transition in aged mice. We found a correlation between A2A receptor density and tissue damage due to yeast infection: comparing with young and adults, aged mice have a lower gut A2A receptor density and C. albicans infection failed to increase it. In conclusion, this study shows that aged mice have a lower ability to cope with inflammation due to C. albicans over-colonization, associated with an inability to adaptively adjust adenosine A2A receptors density. PMID:27590517

  4. Caffeine consumption prevents memory impairment, neuronal damage, and adenosine A2A receptors upregulation in the hippocampus of a rat model of sporadic dementia.

    PubMed

    Espinosa, Janaína; Rocha, Andreia; Nunes, Fernanda; Costa, Marcelo S; Schein, Vanessa; Kazlauckas, Vanessa; Kalinine, Eduardo; Souza, Diogo O; Cunha, Rodrigo A; Porciúncula, Lisiane O

    2013-01-01

    Intracerebroventricular (icv) streptozotocin (STZ) administration induces pathological and behavioral alterations similar to those observed in Alzheimer's disease (AD) and is thus considered an experimental model of sporadic AD. Since caffeine (an adenosine receptor antagonist) and selective antagonists of adenosine A2A receptors modify the course of memory impairment in different amyloid-β-based experimental models of AD, we now tested the impact of caffeine on STZ-induced dementia and associated neurodegeneration in the hippocampus as well as on the expression and density of adenosine receptors. Adult male rats received a bilateral infusion of saline or STZ (3 mg/kg, icv), which triggered memory deficits after four weeks, as gauged by impaired object recognition memory. This was accompanied by a reduced NeuN immunoreactivity in the hippocampal CA1 region and an increased expression and density of adenosine A2A receptors (A2AR), but not A1R, in the hippocampus. Caffeine consumption (1 g/L in the drinking water starting 2 weeks before the STZ challenge) prevented the STZ-induced memory impairment and neurodegeneration as well as the upregulation of A2AR. These findings provide the first demonstration that caffeine prevents sporadic dementia and implicate the control of central A2AR as its likely mechanism of action.

  5. Muscle developmental defects in heterogeneous nuclear Ribonucleoprotein A1 knockout mice

    PubMed Central

    Liu, Ting-Yuan; Chen, Yu-Chia; Jong, Yuh-Jyh; Tsai, Huai-Jen; Lee, Chien-Chin; Chang, Ya-Sian; Chang, Jan-Gowth

    2017-01-01

    Heterogeneous ribonucleoprotein A1 (hnRNP A1) is crucial for regulating alternative splicing. Its integrated function within an organism has not, however, been identified. We generated hnRNP A1 knockout mice to study the role of hnRNP A1 in vivo. The knockout mice, hnRNP A1−/−, showed embryonic lethality because of muscle developmental defects. The blood pressure and heart rate of the heterozygous mice were higher than those of the wild-type mice, indicating heart function defects. We performed mouse exon arrays to study the muscle development mechanism. The processes regulated by hnRNP A1 included cell adhesion and muscle contraction. The expression levels of muscle development-related genes in hnRNP A1+/− mice were significantly different from those in wild-type mice, as detected using qRT-PCR. We further confirmed the alternative splicing patterns of muscle development-related genes including mef2c, lrrfip1, usp28 and abcc9. Alternative mRNA isoforms of these genes were increased in hnRNP A1+/− mice compared with wild-type mice. Furthermore, we revealed that the functionally similar hnRNP A2/B1 did not compensate for the expression of hnRNP A1 in organisms. In summary, our study demonstrated that hnRNP A1 plays a critical and irreplaceable role in embryonic muscle development by regulating the expression and alternative splicing of muscle-related genes. PMID:28077597

  6. A-1 Test Stand work

    NASA Image and Video Library

    2010-01-13

    Employees at NASA's John C. Stennis Space Center work to maneuver a structural steam beam into place on the A-1 Test Stand on Jan. 13. The beam was one of several needed to form the thrust takeout structure that will support a new thrust measurement system being installed on the stand for future rocket engine testing. Once lifted onto the stand, the beams had to be hoisted into place through the center of the test stand, with only two inches of clearance on each side. The new thrust measurement system represents a state-of-the-art upgrade from the equipment installed more than 40 years ago when the test stand was first constructed.

  7. The adenosine A2A receptor agonist CGS 21680 decreases ethanol self-administration in both non-dependent and dependent animals.

    PubMed

    Houchi, Hakim; Persyn, Wolfgang; Legastelois, Rémi; Naassila, Mickaël

    2013-09-01

    There is emerging evidence that the adenosinergic system might be involved in drug addiction and alcohol dependence. We have already demonstrated the involvement of A2A receptors (A2AR) in ethanol-related behaviours in mice. Here, we investigated whether the A2AR agonist CGS 21680 can reduce ethanol operant self-administration in both non-dependent and ethanol-dependent Wistar rats. To rule out a potential involvement of the A1R in the effects of CGS 21680, we also tested its effectiveness to reduce ethanol operant self-administration in both heterozygous and homozygous A1R knockout mice. Our results demonstrated that CGS 21680 (0.065, 0.095 and 0.125 mg/kg, i.p.) had a bimodal effect on 10% ethanol operant self-administration in non-dependent rats. The intermediate dose was also effective in reducing 2% sucrose self-administration. Interestingly, the intermediate dose reduced 10% ethanol self-administration in dependent animals more effectively (75% decrease) when compared with non-dependent animals (57% decrease). These results suggest that the A2AR are involved in CGS 21680 effects since the reduction of ethanol self-administration was not dependent upon the presence of A1R in mice. In conclusion, our findings demonstrated the effectiveness of the A2AR agonist CGS 21680 in a preclinical model of alcohol addiction and suggested that the adenosinergic pathway is a promising target to treat alcohol addiction.

  8. A tail of two signals: the C terminus of the A(2A)-adenosine receptor recruits alternative signaling pathways.

    PubMed

    Gsandtner, Ingrid; Freissmuth, Michael

    2006-08-01

    G protein-coupled receptors are endowed with carboxyl termini that vary greatly in length and sequence. In most instances, the distal portion of the C terminus is dispensable for G protein coupling. This is also true for the A(2A)-adenosine receptor, where the last 100 amino acids are of very modest relevance to G(s) coupling. The C terminus was originally viewed mainly as the docking site for regulatory proteins of the beta-arrestin family. These beta-arrestins bind to residues that have been phosphorylated by specialized kinases (G protein-coupled receptor kinases) and thereby initiate receptor desensitization and endocytosis. More recently, it has become clear that many additional "accessory" proteins bind to C termini of G protein-coupled receptors. The article by Sun et al. in the current issue of Molecular Pharmacology identifies translin-associated protein-X as yet another interaction partner of the A(2A) receptor; translin-associated protein allows the A(2A) receptor to impinge on the signaling mechanisms by which p53 regulates neuronal differentiation, but the underlying signaling pathways are uncharted territory. With a list of five known interaction partners, the C terminus of the A(2A) receptor becomes a crowded place. Hence, there must be rules that regulate the interaction. This allows the C terminus to act as coincidence detector and as signal integrator. Despite our ignorance about the precise mechanisms, the article has exciting implications: the gene encoding for translin-associated protein-X maps to a locus implicated in some forms of schizophrenia; A(2A) receptor agonists are candidate drugs for the treatment of schizophrenic symptoms. It is of obvious interest to explore a possible link.

  9. A new ethyladenine antagonist of adenosine A(2A) receptors: behavioral and biochemical characterization as an antiparkinsonian drug.

    PubMed

    Pinna, Annalisa; Tronci, Elisabetta; Schintu, Nicoletta; Simola, Nicola; Volpini, Rosaria; Pontis, Silvia; Cristalli, Gloria; Morelli, Micaela

    2010-03-01

    Adenosine A(2A) receptor antagonists have emerged as an attractive non-dopaminergic target in clinical trials aimed at evaluating improvement in motor deficits in Parkinson's disease (PD). Moreover, preclinical studies suggest that A(2A) receptor antagonists may slow the course of the underlying neurodegeneration of dopaminergic neurons. In this study, we evaluated the efficacy of the new adenosine A(2A) receptor antagonist 8-ethoxy-9-ethyladenine (ANR 94) in parkinsonian models of akinesia and tremor. In addition, induction of the immediate early gene zif-268, and neuroprotective and anti-inflammatory effects of ANR 94 were evaluated. ANR 94 was effective in reversing parkinsonian tremor induced by the administration of tacrine. ANR 94 also counteracted akinesia (stepping test) and sensorimotor deficits (vibrissae-elicited forelimb-placing test), as well as potentiating l-dopa-induced contralateral turning behavior in 6-hydroxydopamine (6-OHDA) lesion model of PD. Potentiation of motor behavior in 6-OHDA-lesioned rats was not associated with increased induction of the immediate early gene zif-268 in the striatum, suggesting that ANR 94 does not induce long-term plastic changes in this structure. Finally, in a subchronic 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) mouse model of PD, ANR 94 protected nigrostriatal dopaminergic neurons from degeneration and counteracted neuroinflammatory processes by contrasting astroglial (glial fibrillary acidic protein, GFAP) and microglial (CD11b) activation. A(2A) receptor antagonism represents a uniquely realistic opportunity for improving PD treatment, since A(2A) receptor antagonists offer substantial symptomatic benefits and possibly disease-modifying activity. The characterization of ANR 94 may represent a further therapeutic opportunity for the treatment of PD with this new class of drugs.

  10. A2A Adenosine Receptor Antagonism Reverts the Blood-Brain Barrier Dysfunction Induced by Sleep Restriction

    PubMed Central

    Hurtado-Alvarado, Gabriela; Domínguez-Salazar, Emilio; Velázquez-Moctezuma, Javier

    2016-01-01

    Chronic sleep restriction induces blood-brain barrier disruption and increases pro-inflammatory mediators in rodents. Those inflammatory mediators may modulate the blood-brain barrier and constitute a link between sleep loss and blood-brain barrier physiology. We propose that adenosine action on its A2A receptor may be modulating the blood-brain barrier dynamics in sleep-restricted rats. We administrated a selective A2A adenosine receptor antagonist (SCH58261) in sleep-restricted rats at the 10th day of sleep restriction and evaluated the blood-brain barrier permeability to dextrans coupled to fluorescein (FITC-dextrans) and Evans blue. In addition, we evaluated by western blot the expression of tight junction proteins (claudin-5, occludin, ZO-1), adherens junction protein (E-cadherin), A2A adenosine receptor, adenosine-synthesizing enzyme (CD73), and neuroinflammatory markers (Iba-1 and GFAP) in the cerebral cortex, hippocampus, basal nuclei and cerebellar vermis. Sleep restriction increased blood-brain barrier permeability to FITC-dextrans and Evans blue, and the effect was reverted by the administration of SCH58261 in almost all brain regions, excluding the cerebellum. Sleep restriction increased the expression of A2A adenosine receptor only in the hippocampus and basal nuclei without changing the expression of CD73 in all brain regions. Sleep restriction reduced the expression of tight junction proteins in all brain regions, except in the cerebellum; and SCH58261 restored the levels of tight junction proteins in the cortex, hippocampus and basal nuclei. Finally, sleep restriction induced GFAP and Iba-1 overexpression that was attenuated with the administration of SCH58261. These data suggest that the action of adenosine on its A2A receptor may have a crucial role in blood-brain barrier dysfunction during sleep loss probably by direct modulation of brain endothelial cell permeability or through a mechanism that involves gliosis with subsequent inflammation and

  11. An ab initio global potential-energy surface for NH2(A(2)A') and vibrational spectrum of the Renner-Teller A(2)A'-X(2)A" system.

    PubMed

    Zhou, Shulan; Li, Zheng; Xie, Daiqian; Lin, Shi Ying; Guo, Hua

    2009-05-14

    A global potential-energy surface for the first excited electronic state of NH(2)(A(2)A(')) has been constructed by three-dimensional cubic spline interpolation of more than 20,000 ab initio points, which were calculated at the multireference configuration-interaction level with the Davidson correction using the augmented correlation-consistent polarized valence quadruple-zeta basis set. The (J=0) vibrational energy levels for the ground (X(2)A(")) and excited (A(2)A(')) electronic states of NH(2) were calculated on our potential-energy surfaces with the diagonal Renner-Teller terms. The results show a good agreement with the experimental vibrational frequencies of NH(2) and its isotopomers.

  12. Oral administration of Japanese sake yeast (Saccharomyces cerevisiae sake) promotes non-rapid eye movement sleep in mice via adenosine A2A receptors.

    PubMed

    Nakamura, Yoshitaka; Midorikawa, Tatsuyuki; Monoi, Noriyuki; Kimura, Eriko; Murata-Matsuno, Ayumi; Sano, Tomomi; Oka, Kengo; Sugafuji, Toshihiro; Uchiyama, Akira; Murakoshi, Michiaki; Sugiyama, Keikichi; Nishino, Hoyoku; Urade, Yoshihiro

    2016-12-01

    We have demonstrated previously that Japanese sake yeast improves sleep quality in humans. In the present study, we examined the molecular mechanisms of sake yeast to induce sleep by monitoring locomotor activity, electromyogram and electroencephalogram in mice. Oral administration of Japanese sake yeast (100, 200, and 300 mg kg(-1) ) decreased the locomotor activity by 18, 46 and 59% and increased the amount of non-rapid eye movement (NREM) sleep by 1.5-, 2.3- and 2.4-fold (to 37 ± 6, 57 ± 8, and 60 ± 4 min from 25 ± 6 min in the vehicle-administered group, respectively) in a dose-dependent manner for 4 h after oral administration. However, Japanese sake yeast did not change the amount of rapid eye movement (REM) sleep, the electroencephalogram power density during NREM sleep or show any adverse effects, such as rebound of insomnia, during 24 h postadministration and on the next day. An intraperitoneal pretreatment with an adenosine A2A receptor-selective antagonist, ZM241385 (15 mg kg(-1) ), reduced the amount of NREM sleep of sake yeast-administered mice to the basal level, without changing basal amount of sleep. Conversely, an A1 receptor-selective antagonist, 8-cyclopentyltheophylline (10 mg kg(-1) ), did not affect the sleep-promoting effect of Japanese sake yeast. Thus, Japanese sake yeast promotes NREM sleep via activation of adenosine A2A but not A1 receptors.

  13. Effect of subtype-selective adenosine receptor antagonists on basal or haloperidol-regulated striatal function: studies of exploratory locomotion and c-Fos immunoreactivity in outbred and A(2A)R KO mice.

    PubMed

    Pardo, M; López-Cruz, L; Valverde, O; Ledent, C; Baqi, Y; Müller, C E; Salamone, J D; Correa, M

    2013-06-15

    Behavioral activation is regulated by dopamine (DA) in striatal areas. At low doses, while typical antipsychotic drugs produce psychomotor slowing, psychostimulants promote exploration. Minor stimulants such as caffeine, which act as adenosine receptor antagonists, can also potentiate behavioral activation. Striatal areas are rich in adenosine and DA receptors, and adenosine A2A receptors are mainly expressed in the striatum where they are co-localized with DA D2 receptors. Adenosine antagonists with different receptor-selectivity profiles were used to study spontaneous or haloperidol-impaired exploration and c-Fos expression in different striatal areas. Because A2A antagonists were expected to be more selective for reversing the effects of the D2 antagonist haloperidol, A2A receptor knockout (A2ARKO) mice were also assessed. CD1 and A2ARKO male mice were tested in an open field and in a running wheel. Only the A1/A2A receptor antagonist theophylline (5.0-15.0 mg/kg) and the A2A antagonist MSX-3 (2.0 mg/kg) increased spontaneous locomotion and rearing. Co-administration of theophylline (10.0-15.0 mg/kg), and MSX-3 (1.0-3.0 mg/kg) reversed haloperidol-induced suppression of locomotion. The A1 antagonist CPT was only marginally effective in reversing the effects of haloperidol. Although adenosine antagonists did not affect c-Fos expression on their own, theophylline and MSX-3, but not CPT, attenuated haloperidol induction of c-Fos expression. A2ARKO mice were resistant to the behavioral effects of haloperidol at intermediate doses (0.1 mg/kg) in the open field and in the running wheel. A2A receptors are important for regulating behavioral activation, and interact with D2 receptors in striatal areas to regulate neural processes involved in exploratory activity.

  14. Structure of the adenosine A(2A) receptor in complex with ZM241385 and the xanthines XAC and caffeine.

    PubMed

    Doré, Andrew S; Robertson, Nathan; Errey, James C; Ng, Irene; Hollenstein, Kaspar; Tehan, Ben; Hurrell, Edward; Bennett, Kirstie; Congreve, Miles; Magnani, Francesca; Tate, Christopher G; Weir, Malcolm; Marshall, Fiona H

    2011-09-07

    Methylxanthines, including caffeine and theophylline, are among the most widely consumed stimulant drugs in the world. These effects are mediated primarily via blockade of adenosine receptors. Xanthine analogs with improved properties have been developed as potential treatments for diseases such as Parkinson's disease. Here we report the structures of a thermostabilized adenosine A(2A) receptor in complex with the xanthines xanthine amine congener and caffeine, as well as the A(2A) selective inverse agonist ZM241385. The receptor is crystallized in the inactive state conformation as defined by the presence of a salt bridge known as the ionic lock. The complete third intracellular loop, responsible for G protein coupling, is visible consisting of extended helices 5 and 6. The structures provide new insight into the features that define the ligand binding pocket of the adenosine receptor for ligands of diverse chemotypes as well as the cytoplasmic regions that interact with signal transduction proteins.

  15. Adenosine is required for sustained inflammasome activation via the A2A receptor and the HIF-1α pathway

    NASA Astrophysics Data System (ADS)

    Ouyang, Xinshou; Ghani, Ayaz; Malik, Ahsan; Wilder, Tuere; Colegio, Oscar Rene; Flavell, Richard Anthony; Cronstein, Bruce Neil; Mehal, Wajahat Zafar

    2013-12-01

    Inflammasome pathways are important in chronic diseases; however, it is not known how the signalling is sustained after initiation. Inflammasome activation is dependent on stimuli such as lipopolysaccharide (LPS) and ATP that provide two distinct signals resulting in rapid production of interleukin (IL)-1β, with the lack of response to repeat stimulation. Here we report that adenosine is a key regulator of inflammasome activity, increasing the duration of the inflammatory response via the A2A receptor. Adenosine does not replace signals provided by stimuli such as LPS or ATP but sustains inflammasome activity via a cAMP/PKA/CREB/HIF-1α pathway. In the setting of the lack of IL-1β responses after previous exposure to LPS, adenosine can supersede this tolerogenic state and drive IL-1β production. These data reveal that inflammasome activity is sustained, after initial activation, by A2A receptor-mediated signalling.

  16. Adenosine A2A receptor deletion affects social behaviors and anxiety in mice: Involvement of anterior cingulate cortex and amygdala.

    PubMed

    López-Cruz, Laura; Carbó-Gas, Maria; Pardo, Marta; Bayarri, Pilar; Valverde, Olga; Ledent, Catherine; Salamone, John D; Correa, Mercè

    2017-03-15

    Blockade of adenosine A2A receptors can potentiate motivation to work for natural reinforcers such as food. Conspecific interaction is a potent natural reinforcer in social animals that can be manifested as preference for social exploration versus other sources of novel stimulation. Deficiencies in this type of motivated behavior (social withdrawal) have been seen in several pathologies such as autism and depression. However, the role of A2A receptors in motivation for social interaction has not been widely explored. Social interaction paradigms evaluate the natural preference of animals for exploring other conspecifics, and the ability to differentiate between familiar versus novel ones. Anxiety is one of the factors that can induce avoidance of social interaction. In the present study, adenosine A2A knockout (A2AKO) and wild-type (WT) mice were assessed for social and anxiety-related behaviors. c-Fos immunoreactivity was evaluated as a measure of neuronal activation in brain areas involved in different aspects of motivation and emotional processes. Although A2AKO mice showed an anxious profile, they displayed higher levels of sociability and were less sensitive to social novelty. WT mice displayed a typical pattern of social recognition 24h later, but not A2AKO mice, which explored equally both conspecifics. There were no differences between strains in aggressiveness, perseverance or social odor preferences. c-Fos immunoreactivity in A2AKO mice was higher in anterior cingulate and amygdala compared to WT mice. Thus, A2A receptors appear to be potential targets for the improvement of pathologies related to social function.

  17. Enhancement of AMPA currents and GluR1 membrane expression through PKA-coupled adenosine A(2A) receptors.

    PubMed

    Dias, Raquel B; Ribeiro, Joaquim A; Sebastião, Ana M

    2012-02-01

    Phosphorylation of glutamate α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors by Protein Kinase A (PKA) is known to regulate AMPA receptor (AMPAR) trafficking and stabilization at the postsynaptic membrane, which in turn is one of the key mechanisms by which synaptic transmission and plasticity are tuned. However, not much is known as to how Gs-coupled receptors contribute to endogenous PKA-mediated regulation of AMPA receptor function. Here we report that activation of the excitatory A(2A) adenosine receptor by 2-[4-(2-p-carboxyethyl)phenylamino]-5'-N-ethylcarboxamidoadenosine (CGS 21680, 1-30 nM) facilitates AMPA-evoked currents in CA1 pyramidal neurons, by a mechanism dependent on PKA activation, but not on protein synthesis. This modulation of AMPA currents was mimicked by forskolin (1 μM) and did not occur in stratum radiatum interneurons. Superfusion of the A(2A) receptor agonist also caused an increase in the amplitude of miniature excitatory postsynaptic currents (mEPSCs), as well as in the membrane levels of GluR1 subunits phosphorylated at the PKA site (Ser845). The impact of this increase on GluR1-containing AMPA receptor expression was evidenced by the potentiation of LTP at the CA3-CA1 synapse that followed brief activation of A(2A) receptors. We thus propose that in conditions of increased adenosine availability, A(2A) receptor activation is responsible for setting part of the endogenous GluR1 Ser-845 phosphorylation tonus and hence, the availability of the GluR1-containing AMPA receptor extrasynaptic pool for synaptic insertion and reinforcement of synaptic strength.

  18. Promotion of Wound Healing by an Agonist of Adenosine A2A Receptor Is Dependent on Tissue Plasminogen Activator.

    PubMed

    Montesinos, M Carmen; Desai-Merchant, Avani; Cronstein, Bruce N

    2015-12-01

    Impaired wound healing, as it occurs in diabetes mellitus or long-term corticoid treatment, is commonly associated with disability, diminished quality of life, and high economic costs. Selective agonists of the A2A receptor subtype of adenosine, an endogenous regulator of inflammation, promote tissue repair in animal models, both healthy and with impaired healing. Plasmin-mediated proteolysis of fibrin and other matrix proteins is essential for cell migration at sites of injury. Since adenosine A2A receptor activation increases plasminogen activator release from macrophages and mast cells, we studied the effect of a selective agonist, CGS-21680, on full-thickness excisional wound closure in wild-type, urokinase plasminogen activator (uPA)-deficient, and tissue plasminogen activator (tPA)-deficient mice. Wound closure was impaired in tPA- and uPA-deficient mice as compared with wild-type mice, and topical application of CGS-21680 significantly increased the rate at which wounds closed in wild-type mice and uPA-deficient mice, but not in tPA-deficient mice. Immunostaining of tissue sections showed that tPA was present in endothelial cells and histiocytes by day 3 post-wound and also by day 6. In contrast, uPA was more prominent in these cell types only by day 6 post-wound. Our results confirm that plasminogen activation contributes to wound repair and are consistent with the hypothesis that adenosine A2A receptor activation promotes wound closure by a mechanism that depends upon tPA, but not uPA. Moreover, our results suggest that topical adenosine A2A receptor agonists may be useful in promotion of wound closure in patients with impaired wound healing.

  19. Attenuation of gastric mucosal inflammation induced by indomethacin through activation of the A2A adenosine receptor in rats

    PubMed Central

    Koizumi, Shigeto; Otaka, Michiro; Jin, Mario; Linden, Joel; Watanabe, Sumio; Ohnishi, Hirohide

    2010-01-01

    Background Nonsteroidal anti-inflammatory drugs (NSAIDs) such as indomethacin induce gastric mucosal lesions in part by the activation of inflammatory cells and the production of proinflammatory cytokines. The activation of adenosine A2A receptors inhibits inflammation by increasing cyclic AMP in leukocytes and reducing both the production of various proinflammatory cytokines and neutrophil chemotaxis. The aim of present study was to determine whether administration of an orally active adenosine A2A receptor agonist (4-[3-[6-amino-9-(5-cyclopropylcarbamoyl-3,4-dihydroxy-tetrahydro-furan-2-yl)-9H-purin-2-yl]-prop-2-ynyl]-piperidine-1-carboxylic acid methyl ester; ATL-313) ameliorated indomethacin-induced gastric mucosal lesions in rats. Methods Gastric lesions were produced by oral gavage of indomethacin (30 mg/kg). ATL-313 (1–10 μg/kg) was given orally just before the indomethacin administration. Results The ulcer index induced by indomethacin was significantly (>50%) reduced by pretreatment with ATL-313 and this effect was blocked completely by the addition of equimolar ZM241385, a selective A2A receptor antagonist. The gastric content of myeloperoxidase (MPO) and proinflammatory cytokines was significantly reduced by 10 μg/kg ATL-313, but gastric mucosal prostaglandin 2 (PGE2) was not affected. Conclusion We conclude that ATL-313 does not inhibit the mucosal damaging effect of indomethacin, but it does block secondary injury due to stomach inflammation. A2A agonists may represent a class of new therapeutic drugs for NSAID-induced gastric ulcers. PMID:19333545

  20. Membrane omega-3 fatty acids modulate the oligomerisation kinetics of adenosine A2A and dopamine D2 receptors

    NASA Astrophysics Data System (ADS)

    Guixà-González, Ramon; Javanainen, Matti; Gómez-Soler, Maricel; Cordobilla, Begoña; Domingo, Joan Carles; Sanz, Ferran; Pastor, Manuel; Ciruela, Francisco; Martinez-Seara, Hector; Selent, Jana

    2016-01-01

    Membrane levels of docosahexaenoic acid (DHA), an essential omega-3 polyunsaturated fatty acid (ω-3 PUFA), are decreased in common neuropsychiatric disorders. DHA modulates key cell membrane properties like fluidity, thereby affecting the behaviour of transmembrane proteins like G protein-coupled receptors (GPCRs). These receptors, which have special relevance for major neuropsychiatric disorders have recently been shown to form dimers or higher order oligomers, and evidence suggests that DHA levels affect GPCR function by modulating oligomerisation. In this study, we assessed the effect of membrane DHA content on the formation of a class of protein complexes with particular relevance for brain disease: adenosine A2A and dopamine D2 receptor oligomers. Using extensive multiscale computer modelling, we find a marked propensity of DHA for interaction with both A2A and D2 receptors, which leads to an increased rate of receptor oligomerisation. Bioluminescence resonance energy transfer (BRET) experiments performed on living cells suggest that this DHA effect on the oligomerisation of A2A and D2 receptors is purely kinetic. This work reveals for the first time that membrane ω-3 PUFAs play a key role in GPCR oligomerisation kinetics, which may have important implications for neuropsychiatric conditions like schizophrenia or Parkinson’s disease.

  1. Salidroside attenuates chronic hypoxia-induced pulmonary hypertension via adenosine A2a receptor related mitochondria-dependent apoptosis pathway.

    PubMed

    Huang, Xiaoying; Zou, Lizhen; Yu, Xiaoming; Chen, Mayun; Guo, Rui; Cai, Hui; Yao, Dan; Xu, Xiaomei; Chen, Yanfan; Ding, Cheng; Cai, Xueding; Wang, Liangxing

    2015-05-01

    Pulmonary arterial hypertension (PAH) is characterized by pulmonary arterial remodeling mainly due to excess cellular proliferation and apoptosis resistance of pulmonary arterial smooth muscle cells (PASMCs). Salidroside, an active ingredient isolated from Rhodiola rosea is proposed to exert protective effects against PAH. However, the function of salidroside in PAH has not been investigated systematically and the underlying mechanisms are not clear. To investigate the effects of salidroside on PAH, the mice in chronic hypoxia model of PAH were given by an increasing concentration of salidroside (0, 16 mg/kg, 32 mg/kg, and 64 mg/kg). After salidroside treatment, the chronic hypoxia-induced right ventricular hypertrophy and pulmonary arterial remodeling were attenuated, suggesting a protective role played by salidroside in PAH. To explore the potential mechanisms, the apoptosis of PASMCs after salidroside treatment under hypoxia conditions were determined in vivo and in vitro, and also the mitochondria-dependent apoptosis factors, Bax, Bcl-2, cytochrome C, and caspase 9 were examined. The results revealed that salidroside reversed hypoxia-induced cell apoptosis resistance at least partially via a mitochondria-dependent pathway. In addition, salidroside upregulated the expression of adenosine A2a receptor (A2aR) in lung tissues of mice and in PASMCs in vitro after hypoxia exposure. Combined the evidence above, we conclude that salidroside can attenuate chronic hypoxia-induced PAH by promoting PASMCs apoptosis via an A2aR related mitochondria dependent pathway.

  2. Membrane omega-3 fatty acids modulate the oligomerisation kinetics of adenosine A2A and dopamine D2 receptors

    PubMed Central

    Guixà-González, Ramon; Javanainen, Matti; Gómez-Soler, Maricel; Cordobilla, Begoña; Domingo, Joan Carles; Sanz, Ferran; Pastor, Manuel; Ciruela, Francisco; Martinez-Seara, Hector; Selent, Jana

    2016-01-01

    Membrane levels of docosahexaenoic acid (DHA), an essential omega-3 polyunsaturated fatty acid (ω-3 PUFA), are decreased in common neuropsychiatric disorders. DHA modulates key cell membrane properties like fluidity, thereby affecting the behaviour of transmembrane proteins like G protein-coupled receptors (GPCRs). These receptors, which have special relevance for major neuropsychiatric disorders have recently been shown to form dimers or higher order oligomers, and evidence suggests that DHA levels affect GPCR function by modulating oligomerisation. In this study, we assessed the effect of membrane DHA content on the formation of a class of protein complexes with particular relevance for brain disease: adenosine A2A and dopamine D2 receptor oligomers. Using extensive multiscale computer modelling, we find a marked propensity of DHA for interaction with both A2A and D2 receptors, which leads to an increased rate of receptor oligomerisation. Bioluminescence resonance energy transfer (BRET) experiments performed on living cells suggest that this DHA effect on the oligomerisation of A2A and D2 receptors is purely kinetic. This work reveals for the first time that membrane ω-3 PUFAs play a key role in GPCR oligomerisation kinetics, which may have important implications for neuropsychiatric conditions like schizophrenia or Parkinson’s disease. PMID:26796668

  3. Adenosine through the A2A adenosine receptor increases IL-1β in the brain contributing to anxiety

    PubMed Central

    Chiu, Gabriel S.; Darmody, Patrick T.; Walsh, John P.; Moon, Morgan L.; Kwakwa, Kristin A.; Bray, Julie K.; McCusker, Robert H.; Freund, Gregory G.

    2014-01-01

    Anxiety is one of the most commonly reported psychiatric conditions, but its pathogenesis is poorly understood. Ailments associated with activation of the innate immune system, however, are increasingly linked to anxiety disorders. In adult male mice, we found that adenosine doubled caspase-1 activity in brain by a pathway reliant on ATP-sensitive potassium (KATP) channels, protein kinase A (PKA) and the A2A adenosine receptor (AR). In addition, adenosine-dependent activation of caspase-1 increased interleukin (IL)-1β in the brain by two-fold. Peripheral administration of adenosine in wild-type (WT) mice led to a 2.3-fold increase in caspase-1 activity in the amygdala and to a 33% and 42% reduction in spontaneous locomotor activity and food intake, respectively, that were not observed in caspase-1 knockout (KO), IL-1 receptor type 1 (IL-1R1) KO and A2A AR KO mice or in mice administered a caspase-1 inhibitor centrally. Finally, adenosine administration increased anxiety-like behaviors in WT mice by 28% in the open field test and by 55% in the elevated zero-maze. Caspase-1 KO mice, IL-1R1 KO mice, A2A AR KO mice and WT mice treated with the KATP channel blocker, glyburide, were resistant to adenosine-induced anxiety-like behaviors. Thus, our results indicate that adenosine can act as an anxiogenic by activating caspase-1 and increasing IL-1β in the brain. PMID:24907587

  4. Selecting an optimal number of binding site waters to improve virtual screening enrichments against the adenosine A2A receptor.

    PubMed

    Lenselink, Eelke B; Beuming, Thijs; Sherman, Woody; van Vlijmen, Herman W T; IJzerman, Adriaan P

    2014-06-23

    A major challenge in structure-based virtual screening (VS) involves the treatment of explicit water molecules during docking in order to improve the enrichment of active compounds over decoys. Here we have investigated this in the context of the adenosine A2A receptor, where water molecules have previously been shown to be important for achieving high enrichment rates with docking, and where the positions of some binding site waters are known from a high-resolution crystal structure. The effect of these waters (both their presence and orientations) on VS enrichment was assessed using a carefully curated set of 299 high affinity A2A antagonists and 17,337 decoys. We show that including certain crystal waters greatly improves VS enrichment and that optimization of water hydrogen positions is needed in order to achieve the best results. We also show that waters derived from a molecular dynamics simulation - without any knowledge of crystallographic waters - can improve enrichments to a similar degree as the crystallographic waters, which makes this strategy applicable to structures without experimental knowledge of water positions. Finally, we used decision trees to select an ensemble of structures with different water molecule positions and orientations that outperforms any single structure with water molecules. The approach presented here is validated against independent test sets of A2A receptor antagonists and decoys from the literature. In general, this water optimization strategy could be applied to any target with waters-mediated protein-ligand interactions.

  5. Interactions between CYP1A1 polymorphisms and exposure to environmental tobacco smoke in the modulation of lymphocyte bulky DNA adducts and chromosomal aberrations.

    PubMed

    Georgiadis, Panagiotis; Topinka, Jan; Vlachodimitropoulos, Dimitris; Stoikidou, Melpomeni; Gioka, Maria; Stephanou, Georgia; Autrup, Herman; Demopoulos, Nikolaos A; Katsouyanni, Klea; Sram, Radim; Kyrtopoulos, Soterios A

    2005-01-01

    CYP1A1 plays an important role in the metabolic activation of polycyclic aromatic hydrocarbons (PAH), carcinogenic components of air pollution. The influence of CYP1A1 genotype (*2A, *2B and *4) on the levels of lymphocyte bulky DNA adducts and the frequency of cells with aberrant chromosomes was assessed in 194 non-smoking subjects in whom recent exposure to environmental tobacco smoke (ETS) and airborne particulate-associated PAH were measured during two consecutive seasons (winter and summer). While CYP1A1*4 had no consistent effect on either biomarker of genetic damage, the levels of both biomarkers responded in a parallel fashion to changes in exposure/CYP1A1*2A genotype combinations during both seasons. Specifically, the levels of both biomarkers were increased in carriers of at least one CYP1A1*2A allele, as compared with CYP1A1*1 homozygotes, in subjects with ETS exposures >0.8 h/day during the previous 4 days and mean personal exposure to benzo[a]pyrene <0.9 ng/m3 during the previous 24 h (all P < 0.05). Outside these exposure limits the differential effect in CYP1A1*2A variants was lost. Although the numbers of subjects with the CYP1A1*2B polymorphism was small, the same trend appeared to be followed in this case. These effects are interpreted as resulting from differential induction of CYP1A1 expression in CYP1A1*2A and CYP1A1*2A/*2B carriers by components of ETS-polluted air at levels of exposure readily suffered by large segments of the general population and suggest that subjects with these genotypes may have increased susceptibility to the genotoxic effects of ETS.

  6. A1C Test and Diabetes

    MedlinePlus

    ... Diagnosis The A1C Test & Diabetes The A1C Test & Diabetes What is the A1C test? The A1C test ... A1C test be used to diagnose type 2 diabetes and prediabetes? Yes. In 2009, an international expert ...

  7. Blood Test: Hemoglobin A1C

    MedlinePlus

    ... TV, Video Games, and the Internet Blood Test: Hemoglobin A1c KidsHealth > For Parents > Blood Test: Hemoglobin A1c Print A A A What's in this ... de sangre: hemoglobina A1c What It Is A hemoglobin A1c (HbA1c) test is used to monitor long- ...

  8. Blood Test: Hemoglobin A1C

    MedlinePlus

    ... Your 1- to 2-Year-Old Blood Test: Hemoglobin A1c KidsHealth > For Parents > Blood Test: Hemoglobin A1c A A A What's in this article? ... de sangre: hemoglobina A1c What It Is A hemoglobin A1c (HbA1c) test is used to monitor long- ...

  9. Tetanic depression is overcome by tonic adenosine A2A receptor facilitation of L-type Ca2+ influx into rat motor nerve terminals

    PubMed Central

    Oliveira, Laura; Timóteo, M Alexandrina; Correia-de-Sá, Paulo

    2004-01-01

    Motor nerve terminals possess multiple voltage-sensitive calcium channels operating acetylcholine (ACh) release. In this study, we investigated whether facilitation of neuromuscular transmission by adenosine generated during neuronal firing was operated by Ca2+ influx via ‘prevalent’ P-type or via the recruitment of ‘silent’ L-type channels. The release of [3H]ACh from rat phrenic nerve endings decreased upon increasing the stimulation frequency of the trains (750 pulses) from 5 Hz (83 ± 4 × 103 disintegrations per minute per gram (d.p.m. g−1); n = 11) to 50 Hz (30 ± 3 × 103 d.p.m. g−1; n = 5). The P-type Ca2+ channel blocker, ω-agatoxin IVA (100 nm) reduced (by 40 ± 10%; n = 6) the release of [3H]ACh evoked by 50-Hz trains, while nifedipine (1 μm, an L-type blocker) was inactive. Tetanic depression was overcome (88 ± 6 × 103 d.p.m. g−1; n = 12) by stimulating the phrenic nerve with 50-Hz bursts (five bursts of 150 pulses, 20 s interburst interval). In these conditions, ω-agatoxin IVA (100 nm) failed to affect transmitter release, but nifedipine (1 μm) decreased [3H]ACh release by 21 ± 7% (n = 4). Inactivation of endogenous adenosine with adenosine deaminase (ADA, 0.5 U ml−1) reduced (by 54 ± 8%, n = 5) the release of [3H]ACh evoked with 50-Hz bursts. This effect was opposite to the excitatory actions of adenosine (0.5 mm), S-(p-nitrobenzyl)-6-thioinosine (5 μm, an adenosine uptake blocker) and CGS 21680C (3 nm, a selective A2A receptor agonist); as the A1 receptor agonist R-N6-phenylisopropyl adenosine (R-PIA, 300 nm) failed to affect the release of [3H]ACh, the results indicate that adenosine generated during 50-Hz bursts exerts an A2A-receptor-mediated tonus. The effects of ADA (0.5 U ml−1) and CGS 21680C (3 nm) were prevented by nifedipine (1 μm). Blocking tonic A2A receptor activation, with ADA (0.5 U ml−1) or 3,7-dimethyl-1-propargyl xanthine (10 μm, an A2A antagonist), recovered ω-agatoxin IVA (100 nm) inhibition and

  10. Modulation by salt intake of the vascular response mediated through adenosine A(2A) receptor: role of CYP epoxygenase and soluble epoxide hydrolase.

    PubMed

    Nayeem, Mohammed A; Zeldin, Darryl C; Boegehold, Matthew A; Morisseau, Christophe; Marowsky, Anne; Ponnoth, Dovenia S; Roush, Kevin P; Falck, John R

    2010-07-01

    High-salt intake can change the effect of adenosine on arterial tone in mice. The aim of this study was to clarify the mechanism by which this occurs. Using aortas from mice fed a 4% NaCl (HS) or 0.45% NaCl (NS) diet for 4-5 wks, concentration-response curves for ACh, 5'-N-ethylcarboxamidoadenosine (NECA; adenosine analog) and 2-p-(2-carboxyethyl)phenethylamino-5'-N-ethylcarboxamidoadenosine hydrochloride hydrate [CGS-21680; A(2A) adenosine receptor (A(2A) AR) agonist] were obtained with N(omega)-nitro-L-arginine methyl ester (L-NAME; nitric oxide inhibitor, 10(-4) M), methylsulfonyl-propargyloxyphenylhexanamide [MS-PPOH; a CYP (cytochrome P-450) epoxygenase blocker, 10(-5) M including CYP2J2], 12-(3-adamantan-1-yl-ureido)dodecanoic acid [AUDA; soluble epoxide hydrolase (sEH) blocker, 10(-5) M], dibromo-dodecenyl-methylsulfimide [DDMS; CYP omega-hydroxylase (CYP4A blocker), 10(-5) M], glibenclamide (K(ATP) channel blocker; 10(-5) M) and 5-hydroxydecanoate (5-HD; mitochondrial-K(ATP) channel blocker, 10(-4) M). HS dose response to ACh (10(-7) - 10(-5) M) was not different from NS (P > 0.05). Relaxation to 10(-6) M NECA was greater in the HS group (28.4 +/- 3.9%) than in the NS group (4.1 +/- 2.3%). Relaxation to 10(-6) M CGS-21680 was also greater in HS (27.9 +/- 4.5%) than in NS (4.9 +/- 2.2%). L-NAME was able to block the dose response of ACh (10(-7) - 10(-5) M) equally in both HS and NS (P > 0.05), whereas L-NAME did not block CGS-21680-induced response in HS. In HS the CGS-21680 response was greatly reduced by MS-PPOH (to 4.7 +/- 2.0%) and 5-HD (to 8.9 +/- 2.2%), and also abolished by glibenclamide (-1.0 +/- 5.9%). In NS, the CGS-21680 response was increased by AUDA (to 26.3 +/- 3.4%) and DDMS (to 27.2 +/- 3.0%). Compared with NS, HS vessels showed increased CYP2J2 and A(2A) AR expression (46 and 74% higher, respectively) but decreased sEH, CYP4A, and A(1) AR expression (75, 30, and 55% lower, respectively). These data suggest that in mice fed NS-containing diet

  11. Modulation by salt intake of the vascular response mediated through adenosine A2A receptor: role of CYP epoxygenase and soluble epoxide hydrolase

    PubMed Central

    Zeldin, Darryl C.; Boegehold, Matthew A.; Morisseau, Christophe; Marowsky, Anne; Ponnoth, Dovenia S.; Roush, Kevin P.; Falck, John R.

    2010-01-01

    High-salt intake can change the effect of adenosine on arterial tone in mice. The aim of this study was to clarify the mechanism by which this occurs. Using aortas from mice fed a 4% NaCl (HS) or 0.45% NaCl (NS) diet for 4–5 wks, concentration-response curves for ACh, 5′-N-ethylcarboxamidoadenosine (NECA; adenosine analog) and 2-p-(2-carboxyethyl)phenethylamino-5′-N-ethylcarboxamidoadenosine hydrochloride hydrate [CGS-21680; A2A adenosine receptor (A2A AR) agonist] were obtained with Nω-nitro-l-arginine methyl ester (l-NAME; nitric oxide inhibitor, 10−4 M), methylsulfonyl-propargyloxyphenylhexanamide [MS-PPOH; a CYP (cytochrome P-450) epoxygenase blocker, 10−5 M including CYP2J2], 12-(3-adamantan-1-yl-ureido)dodecanoic acid [AUDA; soluble epoxide hydrolase (sEH) blocker, 10−5 M], dibromo-dodecenyl-methylsulfimide [DDMS; CYP ω-hydroxylase (CYP4A blocker), 10−5 M], glibenclamide (KATP channel blocker; 10−5 M) and 5-hydroxydecanoate (5-HD; mitochondrial-KATP channel blocker, 10−4 M). HS dose response to ACh (10−7 − 10−5 M) was not different from NS (P > 0.05). Relaxation to 10−6 M NECA was greater in the HS group (28.4 ± 3.9%) than in the NS group (4.1 ± 2.3%). Relaxation to 10−6 M CGS-21680 was also greater in HS (27.9 ± 4.5%) than in NS (4.9 ± 2.2%). l-NAME was able to block the dose response of ACh (10−7 − 10−5 M) equally in both HS and NS (P > 0.05), whereas l-NAME did not block CGS-21680-induced response in HS. In HS the CGS-21680 response was greatly reduced by MS-PPOH (to 4.7 ± 2.0%) and 5-HD (to 8.9 ± 2.2%), and also abolished by glibenclamide (−1.0 ± 5.9%). In NS, the CGS-21680 response was increased by AUDA (to 26.3 ± 3.4%) and DDMS (to 27.2 ± 3.0%). Compared with NS, HS vessels showed increased CYP2J2 and A2A AR expression (46 and 74% higher, respectively) but decreased sEH, CYP4A, and A1 AR expression (75, 30, and 55% lower, respectively). These data suggest that in mice fed NS-containing diet, upregulation of

  12. ON THE DISTRIBUTION OF VITAMINS A1 AND A2

    PubMed Central

    Wald, George

    1939-01-01

    The distribution of vitamins A1 and A2 has been determined in the eye tissues and livers of a number of fishes. The vitamins were differentiated by means of the antimony chloride reaction, which yields with A1 a band at 615–620 mµ and with A2 a band at about 696 mµ. In the retina the presence of vitamin A1 is diagnostic of the operation of a rhodopsin, and vitamin A2 of a porphyropsin cycle. The eye tissues of all permanently marine fishes examined, except the tautog, contain vitamin A1 alone. Those of all permanently freshwater fishes possess only vitamin A2. Those of all euryhaline (potentially migratory) fishes, except possibly the alewife, contain mixtures of both vitamins A, and always predominantly that one which ordinarily is associated with the environment in which the fish is spawned. These correlations extend in part to the liver oils, but most livers contain mixtures of both vitamins A, and occasionally in proportions the reverse of those in the eye tissues. The vitamin A configuration does not depend upon environmental circumstances, but is determined genetically. The transfer from vitamin A1 to A2 metabolism appears associated phylogenetically with migration of marine teleosts into fresh water. PMID:19873110

  13. Crystal structure of the adenosine A2A receptor bound to an antagonist reveals a potential allosteric pocket

    PubMed Central

    Sun, Bingfa; Bachhawat, Priti; Chu, Matthew Ling-Hon; Wood, Martyn; Ceska, Tom; Sands, Zara A.; Mercier, Joel; Lebon, Florence; Kobilka, Tong Sun; Kobilka, Brian K.

    2017-01-01

    The adenosine A2A receptor (A2AR) has long been implicated in cardiovascular disorders. As more selective A2AR ligands are being identified, its roles in other disorders, such as Parkinson’s disease, are starting to emerge, and A2AR antagonists are important drug candidates for nondopaminergic anti-Parkinson treatment. Here we report the crystal structure of A2A receptor bound to compound 1 (Cmpd-1), a novel A2AR/N-methyl d-aspartate receptor subtype 2B (NR2B) dual antagonist and potential anti-Parkinson candidate compound, at 3.5 Å resolution. The A2A receptor with a cytochrome b562-RIL (BRIL) fusion (A2AR–BRIL) in the intracellular loop 3 (ICL3) was crystallized in detergent micelles using vapor-phase diffusion. Whereas A2AR–BRIL bound to the antagonist ZM241385 has previously been crystallized in lipidic cubic phase (LCP), structural differences in the Cmpd-1–bound A2AR–BRIL prevented formation of the lattice observed with the ZM241385–bound receptor. The crystals grew with a type II crystal lattice in contrast to the typical type I packing seen from membrane protein structures crystallized in LCP. Cmpd-1 binds in a position that overlaps with the native ligand adenosine, but its methoxyphenyl group extends to an exosite not previously observed in other A2AR structures. Structural analysis revealed that Cmpd-1 binding results in the unique conformations of two tyrosine residues, Tyr91.35 and Tyr2717.36, which are critical for the formation of the exosite. The structure reveals insights into antagonist binding that are not observed in other A2AR structures, highlighting flexibility in the binding pocket that may facilitate the development of A2AR-selective compounds for the treatment of Parkinson’s disease. PMID:28167788

  14. Induction of oral tremor in mice by the acetylcholinesterase inhibitor galantamine: Reversal with adenosine A2A antagonism.

    PubMed

    Podurgiel, Samantha J; Spencer, Tiahna; Kovner, Rotem; Baqi, Younis; Müller, Christa E; Correa, Merce; Salamone, John D

    2016-01-01

    Tremulous jaw movements (TJMs) have become a commonly used rat model of Parkinsonian tremor. TJMs can be induced by a number of neurochemical conditions that parallel those seen in human Parkinsonism, including DA depletion, DA antagonism, and cholinomimetic administration, and can be reduced by various antiparkinsonian agents. TJMs typically occur in bursts with the peak frequency in the range of 3-7.5 Hz, which is similar to the Parkinsonian tremor frequency range. While the vast majority of this work has been done using rats, current efforts have focused on extending the TJM model to mice. The aim of the present studies was to establish a mouse model of Parkinsonian resting tremor using the anticholinesterase galantamine, and to investigate the effects of adenosine A2A antagonism on galantamine-induced TJMs. Galantamine significantly induced TJMs in a dose-dependent manner (0.5, 1.0, 1.5, 2.0, 2.5 mg/kg IP). The TJMs tended to occur in bursts in the 3-7.5 Hz frequency range, with a peak frequency of approximately 6 Hz. Systemic administration of the adenosine A2A antagonist MSX-3 (2.5, 5.0, 10.0 mg/kg) significantly attenuated galantamine-induced TJMs. Co-administration of MSX-3 also altered the local frequency of galantamine-induced TJMs, decreasing the peak frequency from approximately 6 Hz to 5 Hz, though the vast majority of TJMs remained in the frequency range characteristic of Parkinsonian resting tremor. These results indicate that adenosine A2A antagonism is capable of reducing anticholinesterase-induced TJMs in mice. Extending the TJM model to mice gives researchers an additional avenue for investigating drug-induced Parkinsonism and tremorogenesis, and could be a useful addition to the study of motor abnormalities observed in mouse genetic models of Parkinsonism.

  15. Quantification of indirect pathway inhibition by the adenosine A2a antagonist SYN115 in Parkinson disease

    PubMed Central

    Black, Kevin J.; Koller, Jonathan M.; Campbell, Meghan C.; Gusnard, Debra A.; Bandak, Stephen I.

    2010-01-01

    Adenosine A2a receptor antagonists reduce symptom severity in Parkinson disease (PD) and animal models. Rodent studies support the hypothesis that A2a antagonists produce this benefit by reducing the inhibitory output of the basal ganglia indirect pathway. One way to test this hypothesis in humans is to quantify regional pharmacodynamic responses with cerebral blood flow (CBF) imaging. That approach has also been proposed as a tool to accelerate pharmaceutical dose-finding, but has not yet been applied in humans to drugs in development. We successfully addressed both these aims with a perfusion MRI study of the novel adenosine A2a antagonist SYN115. During a randomized, double-blind, placebo-controlled, crossover study in 21 PD patients on levodopa but no agonists, we acquired pulsed arterial spin labeling MRI at the end of each treatment period. SYN115 produced a highly significant decrease in thalamic CBF, consistent with reduced pallidothalamic inhibition via the indirect pathway. Similar decreases occurred in cortical regions whose activity decreases with increased alertness and externally-focused attention, consistent with decreased self-reported sleepiness on SYN115. Remarkably, we also derived quantitative pharmacodynamic parameters from the CBF responses to SYN115. These results suggested that the doses tested were on the low end of the effective dose range, consistent with clinical data reported separately. We conclude that (1) SYN115 enters the brain and exerts dose-dependent regional effects, (2) the most prominent of these effects is consistent with deactivation of the indirect pathway as predicted by preclinical studies; and (3) perfusion MRI can provide rapid, quantitative, clinically relevant dose-finding information for pharmaceutical development. PMID:21123574

  16. Adenosine A2A receptors and uric acid mediate protective effects of inosine against TNBS-induced colitis in rats.

    PubMed

    Rahimian, Reza; Fakhfouri, Gohar; Daneshmand, Ali; Mohammadi, Hamed; Bahremand, Arash; Rasouli, Mohammad Reza; Mousavizadeh, Kazem; Dehpour, Ahmad Reza

    2010-12-15

    Inflammatory bowel disease comprises chronic recurrent inflammation of gastrointestinal tract. This study was conducted to investigate inosine, a potent immunomodulator, in 2,4,6-trinitrobenzene sulphonic acid (TNBS)-induced chronic model of experimental colitis, and contribution of adenosine A(2A) receptors and the metabolite uric acid as possible underlying mechanisms. Experimental colitis was rendered in rats by a single colonic administration of 10 mg of TNBS. Inosine, potassium oxonate (a hepatic uricase inhibitor), SCH-442416 (a selective adenosine A(2A) receptor antagonist), inosine+potassium oxonate, or inosine+SCH-442416 were given twice daily for 7 successive days. At the end of experiment, macroscopic and histopathologic scores, colonic malondialdehyde (MDA), Tumor Necrosis Factor-alpha (TNF-α) and Interleukin-1beta (IL-1β) levels, and myeloperoxidase (MPO) activity were assessed. Plasma uric acid level was measured throughout the experiment. Both macroscopic and histological features of colonic injury were markedly ameliorated by either inosine, oxonate or inosine+oxonate. Likewise, the elevated amounts of MPO and MDA abated as well as those of TNF-α and IL-1β (P<0.05). SCH-442416 partially reversed the effect of inosine on theses markers, while inosine+oxonate showed a higher degree of protection than each treatment alone (P<.0.05). No significant difference was observed between TNBS and SCH-442416 groups. Uric acid levels were significantly higher in inosine or oxonate groups compared to control. Inosine+oxonate resulted in an even more elvelated uric acid level than each treatment alone (P<0.05). Inosine elicits notable anti-inflammatory effects on TNBS-induced colitis in rats. Uric acid and adenosine A(2A) receptors contribute to these salutary properties.

  17. Effect of adenosine A(2A) receptor antagonists and L-DOPA on hydroxyl radical, glutamate and dopamine in the striatum of 6-OHDA-treated rats.

    PubMed

    Gołembiowska, Krystyna; Dziubina, Anna

    2012-02-01

    A(2A) adenosine receptor antagonists have been proposed as a new therapy of PD. Since oxidative stress plays an important role in the pathogenesis of PD, we studied the effect of the selective A(2A) adenosine receptor antagonists 8-(-3-chlorostyryl)caffeine (CSC) and 4-(2-[7-amino-2-(2-furyl)[1,2,4]triazolo[2,3-a][1,3,5]triazin-5-ylamino]ethyl)phenol (ZM 241385) on hydroxyl radical generation, and glutamate (GLU) and dopamine (DA) extracellular level using a microdialysis in the striatum of 6-OHDA-treated rats. CSC (1 mg/kg) and ZM 241385 (3 mg/kg) given repeatedly for 14 days decreased the production of hydroxyl radical and extracellular GLU level, both enhanced by prior 6-OHDA treatment in dialysates from the rat striatum. CSC and ZM 241385 did not affect DA and its metabolites, 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanilic acid (HVA) extracellular levels in the striatum of 6-OHDA-treated rats. L-DOPA (6 mg/kg) given twice daily for two weeks in the presence of benserazide (3 mg/kg) decreased striatal hydroxyl radical and glutamate extracellular level in 6-OHDA-treated rats. At the same time, L-DOPA slightly but significantly increased the extracellular levels of DOPAC and HVA. A combined repeated administration of L-DOPA and CSC or ZM 241385 did not change the effect of L-DOPA on hydroxyl radical production and glutamate extracellular level in spite of an enhancement of extracellular DA level by CSC and elevation of extracellular level of DOPAC and HVA by ZM 241385. The data suggest that the 6-OHDA-induced damage of nigrostriatal DA-terminals is related to oxidative stress and excessive release of glutamate. Administration of L-DOPA in combination with CSC or ZM 241385, by restoring striatal DA-glutamate balance, suppressed 6-OHDA-induced overproduction of hydroxyl radical.

  18. The effect of adenosine A(2A) receptor antagonists on hydroxyl radical, dopamine, and glutamate in the striatum of rats with altered function of VMAT2.

    PubMed

    Gołembiowska, Krystyna; Dziubina, Anna

    2012-08-01

    It has been shown that a decreased vesicular monoamine transporter (VMAT2) function and the disruption of dopamine (DA) storage is an early contributor to oxidative damage of dopamine neurons in Parkinson's disease (PD). In our previous study, we demonstrated that adenosine A(2A) receptor antagonists suppressed oxidative stress in 6-hydroxydopamine-treated rats suggesting that this effect may account for neuroprotective properties of drugs. In the present study, rats were injected with reserpine (10 mg/kg sc) and 18 h later the effect of the adenosine A(2A) receptor antagonists 8-(3-chlorostyryl)caffeine (CSC) and 4-(2-[7-amino-2-(2-furyl)[1,2,4]triazolo[2,3-a][1,3,5]triazin-5-ylamino]ethyl)phenol (ZM 241385) on extracellular DA, glutamate and hydroxyl radical formation was studied in the rat striatum using in vivo microdialysis. By disrupting VMAT2 function, reserpine depleted DA stores, and increased glutamate and hydroxyl radical levels in the rat striatum. CSC (1 mg/kg) but not ZM 241385 (3 mg/kg) increased extracellular DA level and production of hydroxyl radical in reserpinised rats. Both antagonists decreased the reserpine-induced increase in extracellular glutamate. L-3,4-Dihydroxyphenylalanine (L-DOPA) (25 mg/kg) significantly enhanced extracellular DA, had no effect on reserpine-induced hydroxyl radical production and decreased extracellular glutamate concentration. CSC but not ZM 241385 given jointly with L-DOPA increased the effect of L-DOPA on extracellular DA and augmented the reserpine-induced hydroxyl radical production. CSC and ZM 241385 did not influence extracellular glutamate level, which was decreased by L-DOPA. It seems that by decreasing the MAO-dependent DA metabolism rate, CSC raised cytosolic DA and by DA autoxidation, it induced hydroxyl radical overproduction. Thus, the methylxanthine A(2A) receptor antagonists bearing properties of MAO-B inhibitor, like CSC, may cause a risk of oxidative stress resulting from dysfunctional DA storage

  19. Rotationally resolved spectroscopy of the A~ 2A1<--X~ 2B1 transition of H2S+ above the barrier to linearity using the mass-analyzed threshold ionization photofragment excitation technique

    NASA Astrophysics Data System (ADS)

    Han, Songhee; Kang, Tae Yeon; Kim, Sang Kyu

    2010-03-01

    The à A21←X˜ B21 transitions of H2S+ above the barrier to linearity have been investigated with the energy resolution high enough to identify individual rotational transition lines for the first time. The rotational cooling of the cation is achieved either by the direct ionization or mass-analyzed threshold ionization (MATI) technique employed in the vacuum-ultraviolet laser excitation of the jet-cooled H2S. Subsequent photoexcitation leads to the H2S+→H2+S+ dissociation and the S+ product yield taken as a function of the excitation energy gives the photofragment excitation (PHOFEX) spectra. The combined use of MATI and PHOFEX techniques greatly simplifies the spectrum allowing the accurate identification of the rotationally resolved bands which is otherwise a formidable task due to the intrinsic complexity of the à A21←X˜ B21 transition. Highly excited states of Ã(0,7,0), Ã(0,8,0), and Ã(0,9,0) vibronic levels with different K quantum numbers which are located above the barrier to linearity are thoroughly investigated. The bent-to-quasilinear transition of H2S+ above the barrier to linearity shows the characteristics of the Renner-Teller effect, showing the large A rotational constant and strong intensity borrowing of the highly vibrationally excited ground levels such as X˜(0,23,0) or X˜(0,24,0) in the dipole-allowed excitation. Spectroscopic parameters of term values, rotational, and spin-orbit coupling constants are precisely determined in this work, providing the most quantitative spectroscopic structure of the H2S+ to date. Quantum-state dependent photodissociation dynamics are also discussed from spectral features of PHOFEX.

  20. Adenosine A(2A) receptor gene (ADORA2A) variants may increase autistic symptoms and anxiety in autism spectrum disorder.

    PubMed

    Freitag, Christine M; Agelopoulos, Konstantin; Huy, Ellen; Rothermundt, Matthias; Krakowitzky, Petra; Meyer, Jobst; Deckert, Jürgen; von Gontard, Alexander; Hohoff, Christa

    2010-01-01

    Autism spectrum disorders (ASDs) are heterogeneous disorders presenting with increased rates of anxiety. The adenosine A(2A) receptor gene (ADORA2A) is associated with panic disorder and is located on chromosome 22q11.23. Its gene product, the adenosine A(2A) receptor, is strongly expressed in the caudate nucleus, which also is involved in ASD. As autistic symptoms are increased in individuals with 22q11.2 deletion syndrome, and large 22q11.2 deletions and duplications have been observed in ASD individuals, in this study, 98 individuals with ASD and 234 control individuals were genotyped for eight single-nucleotide polymorphisms in ADORA2A. Nominal association with the disorder was observed for rs2236624-CC, and phenotypic variability in ASD symptoms was influenced by rs3761422, rs5751876 and rs35320474. In addition, association of ADORA2A variants with anxiety was replicated for individuals with ASD. Findings point toward a possible mediating role of ADORA2A variants on phenotypic expression in ASD that need to be replicated in a larger sample.

  1. Dissecting striatal adenosine-cannabinoid receptor interactions. New clues from rats over-expressing adenosine A2A receptors.

    PubMed

    Ferré, Sergi; Sebastião, Ana Maria

    2016-03-01

    This Editorial highlights a study by Chiodi et al. () showing that the effects mediated by cannabinoid CB1 receptor (CB1R) activation in the striatum are significantly reduced in rats with neuronal over-expression of adenosine A2A receptors (A2AR). Two hypotheses are derived from that study. Hypothesis A: two subpopulations of pre-synaptic CB1R in corticostriatal glutamatergic terminals exist, one forming and another not forming heteromers with A2AR. Hypothesis B: CB1R are predominantly forming heteromers with A2AR. In the case of hypothesis A, the A2AR might be required for CB1R-A2AR heteromeric signaling, whereas non-heteromeric CB1R activity is inhibited by A2ARs. In the case of hypothesis B, up-regulation of A2ARs may perturb heteromeric stoichiometry, thus reducing CB1R functioning. In any case, pre-synaptic striatal A2AR-CB1R heteromers emerge as important targets of the effects of cannabinoids demonstrated at the neuronal and behavioral level. Read the highlighted article 'Striatal adenosine-cannabinoid receptor interactions in rats over-expressing adenosine A2A receptors' on page 907. © 2016 International Society for Neurochemistry.

  2. Expression, Purification and Crystallisation of the Adenosine A2A Receptor Bound to an Engineered Mini G Protein.

    PubMed

    Carpenter, Byron; Tate, Christopher G

    2017-04-20

    G protein-coupled receptors (GPCRs) promote cytoplasmic signalling by activating heterotrimeric G proteins in response to extracellular stimuli such as light, hormones and nucleosides. Structure determination of GPCR-G protein complexes is central to understanding the precise mechanism of signal transduction. However, these complexes are challenging targets for structural studies due to their conformationally dynamic and inherently transient nature. We recently developed an engineered G protein, mini-Gs, which addressed these problems and allowed the formation of a stable GPCR-G protein complex. Mini-Gs facilitated the structure determination of the human adenosine A2A receptor (A2AR) in its G protein-bound conformation at 3.4 Å resolution. Here, we describe a step by step protocol for the expression and purification of A2AR, and crystallisation of the A2AR-mini-Gs complex.

  3. Controlling the Dissociation of Ligands from the Adenosine A2A Receptor through Modulation of Salt Bridge Strength.

    PubMed

    Segala, Elena; Guo, Dong; Cheng, Robert K Y; Bortolato, Andrea; Deflorian, Francesca; Doré, Andrew S; Errey, James C; Heitman, Laura H; IJzerman, Adriaan P; Marshall, Fiona H; Cooke, Robert M

    2016-07-14

    The association and dissociation kinetics of ligands binding to proteins vary considerably, but the mechanisms behind this variability are poorly understood, limiting their utilization for drug discovery. This is particularly so for G protein-coupled receptors (GPCRs) where high resolution structural information is only beginning to emerge. Engineering the human A2A adenosine receptor has allowed structures to be solved in complex with the reference compound ZM241385 and four related ligands at high resolution. Differences between the structures are limited, with the most pronounced being the interaction of each ligand with a salt bridge on the extracellular side of the receptor. Mutagenesis experiments confirm the role of this salt bridge in controlling the dissociation kinetics of the ligands from the receptor, while molecular dynamics simulations demonstrate the ability of ligands to modulate salt bridge stability. These results shed light on a structural determinant of ligand dissociation kinetics and identify a means by which this property may be optimized.

  4. Binding mode similarity measures for ranking of docking poses: a case study on the adenosine A2A receptor

    NASA Astrophysics Data System (ADS)

    Anighoro, Andrew; Bajorath, Jürgen

    2016-06-01

    We report an investigation designed to explore alternative approaches for ranking of docking poses in the search for antagonists of the adenosine A2A receptor, an attractive target for structure-based virtual screening. Calculation of 3D similarity of docking poses to crystallographic ligand(s) as well as similarity of receptor-ligand interaction patterns was consistently superior to conventional scoring functions for prioritizing antagonists over decoys. Moreover, the use of crystallographic antagonists and agonists, a core fragment of an antagonist, and a model of an agonist placed into the binding site of an antagonist-bound form of the receptor resulted in a significant early enrichment of antagonists in compound rankings. Taken together, these findings showed that the use of binding modes of agonists and/or antagonists, even if they were only approximate, for similarity assessment of docking poses or comparison of interaction patterns increased the odds of identifying new active compounds over conventional scoring.

  5. Neuroprotection by caffeine in the MPTP model of parkinson's disease and its dependence on adenosine A2A receptors.

    PubMed

    Xu, K; Di Luca, D G; Orrú, M; Xu, Y; Chen, J-F; Schwarzschild, M A

    2016-05-13

    Considerable epidemiological and laboratory data have suggested that caffeine, a nonselective adenosine receptor antagonist, may protect against the underlying neurodegeneration of parkinson's disease (PD). Although both caffeine and more specific antagonists of the A2A subtype of adenosine receptor (A2AR) have been found to confer protection in animal models of PD, the dependence of caffeine's neuroprotective effects on the A2AR is not known. To definitively determine its A2AR dependence, the effect of caffeine on 1-methyl-4-phenyl-1,2,3,6 tetra-hydropyridine (MPTP) neurotoxicity was compared in wild-type (WT) and A2AR gene global knockout (A2A KO) mice, as well as in central nervous system (CNS) cell type-specific (conditional) A2AR knockout (cKO) mice that lack the receptor either in postnatal forebrain neurons or in astrocytes. In WT and in heterozygous A2AR KO mice caffeine pretreatment (25mg/kgip) significantly attenuated MPTP-induced depletion of striatal dopamine. By contrast in homozygous A2AR global KO mice caffeine had no effect on MPTP toxicity. In forebrain neuron A2AR cKO mice, caffeine lost its locomotor stimulant effect, whereas its neuroprotective effect was mostly preserved. In astrocytic A2AR cKO mice, both caffeine's locomotor stimulant and protective properties were undiminished. Taken together, these results indicate that neuroprotection by caffeine in the MPTP model of PD relies on the A2AR, although the specific cellular localization of these receptors remains to be determined.

  6. 26 CFR 1.263(a)-1 - Capital expenditures; In general.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... property (as defined in § 1.263A-1T (a)(5)(iii)), including films, sound recordings, video tapes, books, or... tangible personal property (as defined in § 1.263A-2(a)(2)), including films, sound recordings, video tapes...

  7. 18 CFR 3a.1 - Purpose.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 18 Conservation of Power and Water Resources 1 2010-04-01 2010-04-01 false Purpose. 3a.1 Section 3a.1 Conservation of Power and Water Resources FEDERAL ENERGY REGULATORY COMMISSION, DEPARTMENT OF ENERGY GENERAL RULES NATIONAL SECURITY INFORMATION General § 3a.1 Purpose. This part 3a describes the...

  8. 18 CFR 3a.1 - Purpose.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 18 Conservation of Power and Water Resources 1 2013-04-01 2013-04-01 false Purpose. 3a.1 Section 3a.1 Conservation of Power and Water Resources FEDERAL ENERGY REGULATORY COMMISSION, DEPARTMENT OF ENERGY GENERAL RULES NATIONAL SECURITY INFORMATION General § 3a.1 Purpose. This part 3a describes the...

  9. 18 CFR 3a.1 - Purpose.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 18 Conservation of Power and Water Resources 1 2012-04-01 2012-04-01 false Purpose. 3a.1 Section 3a.1 Conservation of Power and Water Resources FEDERAL ENERGY REGULATORY COMMISSION, DEPARTMENT OF ENERGY GENERAL RULES NATIONAL SECURITY INFORMATION General § 3a.1 Purpose. This part 3a describes the...

  10. 18 CFR 3a.1 - Purpose.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 18 Conservation of Power and Water Resources 1 2014-04-01 2014-04-01 false Purpose. 3a.1 Section 3a.1 Conservation of Power and Water Resources FEDERAL ENERGY REGULATORY COMMISSION, DEPARTMENT OF ENERGY GENERAL RULES NATIONAL SECURITY INFORMATION General § 3a.1 Purpose. This part 3a describes the...

  11. 14 CFR 374a.1 - Purpose.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 14 Aeronautics and Space 4 2010-01-01 2010-01-01 false Purpose. 374a.1 Section 374a.1 Aeronautics and Space OFFICE OF THE SECRETARY, DEPARTMENT OF TRANSPORTATION (AVIATION PROCEEDINGS) SPECIAL REGULATIONS EXTENSION OF CREDIT BY AIRLINES TO FEDERAL POLITICAL CANDIDATES § 374a.1 Purpose. Section 401 of...

  12. 22 CFR 3a.1 - Definitions.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 22 Foreign Relations 1 2012-04-01 2012-04-01 false Definitions. 3a.1 Section 3a.1 Foreign Relations DEPARTMENT OF STATE GENERAL ACCEPTANCE OF EMPLOYMENT FROM FOREIGN GOVERNMENTS BY MEMBERS OF THE UNIFORMED SERVICES § 3a.1 Definitions. For purposes of this part— (a) Applicant means any person who...

  13. 18 CFR 3a.1 - Purpose.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 18 Conservation of Power and Water Resources 1 2011-04-01 2011-04-01 false Purpose. 3a.1 Section 3a.1 Conservation of Power and Water Resources FEDERAL ENERGY REGULATORY COMMISSION, DEPARTMENT OF ENERGY GENERAL RULES NATIONAL SECURITY INFORMATION General § 3a.1 Purpose. This part 3a describes the...

  14. 46 CFR 147A.1 - Purpose.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... 46 Shipping 5 2012-10-01 2012-10-01 false Purpose. 147A.1 Section 147A.1 Shipping COAST GUARD, DEPARTMENT OF HOMELAND SECURITY (CONTINUED) DANGEROUS CARGOES INTERIM REGULATIONS FOR SHIPBOARD FUMIGATION General § 147A.1 Purpose. The purpose of this part is to prescribe the requirements for...

  15. 12 CFR 269a.1 - Party.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 12 Banks and Banking 4 2014-01-01 2014-01-01 false Party. 269a.1 Section 269a.1 Banks and Banking FEDERAL RESERVE SYSTEM (CONTINUED) BOARD OF GOVERNORS OF THE FEDERAL RESERVE SYSTEM (CONTINUED) DEFINITIONS § 269a.1 Party. The term Party means any person, employee, group of employees, labor organization...

  16. 12 CFR 269a.1 - Party.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 12 Banks and Banking 4 2013-01-01 2013-01-01 false Party. 269a.1 Section 269a.1 Banks and Banking FEDERAL RESERVE SYSTEM (CONTINUED) BOARD OF GOVERNORS OF THE FEDERAL RESERVE SYSTEM (CONTINUED) DEFINITIONS § 269a.1 Party. The term Party means any person, employee, group of employees, labor organization...

  17. 12 CFR 269a.1 - Party.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 12 Banks and Banking 3 2011-01-01 2011-01-01 false Party. 269a.1 Section 269a.1 Banks and Banking FEDERAL RESERVE SYSTEM (CONTINUED) BOARD OF GOVERNORS OF THE FEDERAL RESERVE SYSTEM DEFINITIONS § 269a.1 Party. The term Party means any person, employee, group of employees, labor organization, or bank as...

  18. 12 CFR 269a.1 - Party.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 12 Banks and Banking 4 2012-01-01 2012-01-01 false Party. 269a.1 Section 269a.1 Banks and Banking FEDERAL RESERVE SYSTEM (CONTINUED) BOARD OF GOVERNORS OF THE FEDERAL RESERVE SYSTEM (CONTINUED) DEFINITIONS § 269a.1 Party. The term Party means any person, employee, group of employees, labor organization...

  19. 12 CFR 269a.1 - Party.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 12 Banks and Banking 3 2010-01-01 2010-01-01 false Party. 269a.1 Section 269a.1 Banks and Banking FEDERAL RESERVE SYSTEM (CONTINUED) BOARD OF GOVERNORS OF THE FEDERAL RESERVE SYSTEM DEFINITIONS § 269a.1 Party. The term Party means any person, employee, group of employees, labor organization, or bank as...

  20. Binding of the novel nonxanthine A2A adenosine receptor antagonist [3H]SCH58261 to coronary artery membranes.

    PubMed

    Belardinelli, L; Shryock, J C; Ruble, J; Monopoli, A; Dionisotti, S; Ongini, E; Dennis, D M; Baker, S P

    1996-12-01

    This study demonstrates quantification of A2A adenosine receptors (A2AAdoRs) in membranes prepared from porcine coronary arteries, porcine striatum, and PC12 cells. Radioligand binding assays were performed using the new selective A2AAdoR antagonist radioligand [3H]-5-amino-7-(2-phenylethyl)-2-(2-furyl)-pyrazolo [4,3-epsilon]-1,2,4-triazolo[1,5-c)pyrimidine ([3H]SCH58261). Binding of the radioligand to membranes was rapid, reversible, and saturable. The densities of A2AAdoRs in membranes prepared from porcine coronary arteries, porcine striatum, and PC12 cells were 900 +/- 61, 892 +/- 35, and 959 +/- 76 fmol/mg protein, respectively. Equilibrium dissociation constants (Kd values) calculated from results of saturation binding assays were 2.19, 1.20, and 0.81 nmol/L, and Kd values calculated from results of association and dissociation assays were 2.42, 1.01, and 0.40 nmol/L for [3H]SCH58261 binding to membranes prepared from porcine coronary arteries, porcine striatum, and PC12 cells, respectively. The specific binding of [3H]SCH58261 as a percentage of total binding at a radioligand concentration equal to the Kd value was 65% to 90% in the three membrane preparations. The order of ligand potencies determined by assay of competition binding to sites in porcine coronary membranes using [3H]SCH58261, unlabeled antagonists (SCH58261, 8-(3-chlorostyryl)caffeine [CSC], and xanthine amine congener [XAC]), and unlabeled agonists ([3H]2-p-(2-carboxyethyl)-phenethylamino-5'-N-ethylcarboxamidoaden osine [CGS 21680], 2-hexynyl-5'-N-ethylcarboxamidoadenosine [HE-NECA], [3H]5'-N-ethylcarboxamidoadenosine [NECA], and R(-)N6-(2-phenylisopropyl)adenosine [R-PIA]) was SCH58261 > HE-NECA = CSC = CGS 21680 = XAC > NECA = R-PIA. The Hill coefficients of displacement by A2AAdoR ligands of [3H]SCH58261 binding were not significantly different from unity, indicating that [3H]SCH58261 bound to a group of homogeneous noninteracting sites in all membrane preparations. The order of ligand

  1. Caffeine and CSC, adenosine A2A antagonists, offer neuroprotection against 6-OHDA-induced neurotoxicity in rat mesencephalic cells.

    PubMed

    Nobre, Hélio Vitoriano; Cunha, Geanne Matos de Andrade; de Vasconcelos, Lissiana Magna; Magalhães, Hemerson Iury Ferreira; Oliveira Neto, Raimundo Nogueira; Maia, Flávio Damasceno; de Moraes, Manoel Odorico; Leal, L Kalyne A Moreira; Viana, Glauce Socorro de Barros

    2010-01-01

    In this study, the cytoprotective effects of caffeine (CAF) and 8-(3-chlorostyryl)-caffeine (CSC), A(2A) receptor antagonists, were tested against 6-OHDA-induced cytotoxicity, in rat mesencephalic cells. Both drugs significantly increased the number of viable cells, after their exposure to 6-OHDA, as measured by the MTT assay. While nitrite levels in the cells were drastically increased by 6-OHDA, their concentrations were brought toward normality after CAF or CSC, indicating that both drugs block 6-OHDA-induced oxidative stress which leads to free radicals generation. A complete blockade of 6-OHDA-induced lipid peroxidation, considered as a major source of DNA damage, was observed after cells treatment with CAF or CSC. 6-OHDA decreased the number of normal cells while increasing the number of apoptotic cells. In the CAF plus 6-OHDA group, a significant recover in the number of viable cells and a decrease in the number of apoptotic cells were seen, as compared to the group treated with 6-OHDA alone. A similar effect was observed after cells exposure to CSC in the presence of 6-OHDA. Unexpectedly, while a significant lower number of activated microglia was observed after cells exposure to CAF plus 6-OHDA, this was not the case after cells exposure to CSC under the same conditions. While CAF lowered the percentage of reactive astrocytes increased by 6-OHDA, CSC presented no effect. The effects of these drugs were also examined on the releases of myeloperoxidase (MPO), an inflammatory marker, and lactate dehydrogenase (LDH), a marker for cytotoxicity, in human neutrophils, in vitro. CSC and CAF (0.1, 1 and 10 microg/ml) produced inhibitions of the MPO release from PMA-stimulated cells, ranging from 45 to 83%. In addition, CSC and CAF (5, 50 and 100 microg/ml) did not show any cytotoxicity in the range of concentrations used, as determined by the LDH assay. All together, our results showed a strong neuroptrotection afforded by caffeine or CSC, on rat mesencephalic

  2. Imidazo[1,2-a]pyrazin-8-amine core for the design of new adenosine receptor antagonists: Structural exploration to target the A3 and A2A subtypes.

    PubMed

    Poli, Daniela; Falsini, Matteo; Varano, Flavia; Betti, Marco; Varani, Katia; Vincenzi, Fabrizio; Pugliese, Anna Maria; Pedata, Felicita; Dal Ben, Diego; Thomas, Ajiroghene; Palchetti, Ilaria; Bettazzi, Francesca; Catarzi, Daniela; Colotta, Vittoria

    2017-01-05

    The imidazo[1,2-a]pyrazine ring system has been chosen as a new decorable core skeleton for the design of novel adenosine receptor (AR) antagonists targeting either the human (h) A3 or the hA2A receptor subtype. The N(8)-(hetero)arylcarboxyamido substituted compounds 4-14 and 21-30, bearing a 6-phenyl moiety or not, respectively, show good hA3 receptor affinity and selectivity versus the other ARs. In contrast, the 8-amino-6-(hetero)aryl substituted derivatives designed for targeting the hA2A receptor subtype (compounds 31-38) and also the 6-phenyl analogues 18-20 do not bind the hA2A AR, or show hA1 or balanced hA1/hA2A AR affinity in the micromolar range. Molecular docking of the new hA3 antagonists was carried out to depict their hypothetical binding mode to our refined model of the hA3 receptor. Some derivatives were evaluated for their fluorescent potentiality and showed some fluorescent emission properties. One of the most active hA3 antagonists herein reported, i.e. the 2,6-diphenyl-8-(3-pyridoylamino)imidazo[1,2-a]pyrazine 29, tested in a rat model of cerebral ischemia, delayed the occurrence of anoxic depolarization caused by oxygen and glucose deprivation in the hippocampus and allowed disrupted synaptic activity to recover. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

  3. Allosteric interactions between agonists and antagonists within the adenosine A2A receptor-dopamine D2 receptor heterotetramer.

    PubMed

    Bonaventura, Jordi; Navarro, Gemma; Casadó-Anguera, Verònica; Azdad, Karima; Rea, William; Moreno, Estefanía; Brugarolas, Marc; Mallol, Josefa; Canela, Enric I; Lluís, Carme; Cortés, Antoni; Volkow, Nora D; Schiffmann, Serge N; Ferré, Sergi; Casadó, Vicent

    2015-07-07

    Adenosine A2A receptor (A2AR)-dopamine D2 receptor (D2R) heteromers are key modulators of striatal neuronal function. It has been suggested that the psychostimulant effects of caffeine depend on its ability to block an allosteric modulation within the A2AR-D2R heteromer, by which adenosine decreases the affinity and intrinsic efficacy of dopamine at the D2R. We describe novel unsuspected allosteric mechanisms within the heteromer by which not only A2AR agonists, but also A2AR antagonists, decrease the affinity and intrinsic efficacy of D2R agonists and the affinity of D2R antagonists. Strikingly, these allosteric modulations disappear on agonist and antagonist coadministration. This can be explained by a model that considers A2AR-D2R heteromers as heterotetramers, constituted by A2AR and D2R homodimers, as demonstrated by experiments with bioluminescence resonance energy transfer and bimolecular fluorescence and bioluminescence complementation. As predicted by the model, high concentrations of A2AR antagonists behaved as A2AR agonists and decreased D2R function in the brain.

  4. Arousal effect of caffeine depends on adenosine A2A receptors in the shell of the nucleus accumbens

    PubMed Central

    Lazarus, Michael; Shen, Hai-Ying; Cherasse, Yoan; Qu, Wei-Min; Huang, Zhi-Li; Bass, Caroline E.; Winsky-Sommerer, Raphaelle; Semba, Kazue; Fredholm, Bertil B.; Boison, Detlev; Hayaishi, Osamu; Urade, Yoshihiro; Chen, Jiang-Fan

    2011-01-01

    Caffeine, the most widely used psychoactive compound, is an adenosine receptor antagonist. It promotes wakefulness by blocking adenosine A2A receptors (A2ARs) in the brain, but the specific neurons on which caffeine acts to produce arousal have not been identified. Using selective gene deletion strategies based on the Cre/loxP technology in mice and focal RNA interference to silence the expression of A2ARs in rats by local infection with adeno-associated virus carrying short-hairpin RNA, we report that the A2ARs in the shell region of the nucleus accumbens (NAc) are responsible for the effect of caffeine on wakefulness. Caffeine-induced arousal was not affected in rats when A2ARs were focally removed from the NAc core or other A2AR-positive areas of the basal ganglia. Our observations suggest that caffeine promotes arousal by activating pathways that traditionally have been associated with motivational and motor responses in the brain. PMID:21734299

  5. Aberrant adenosine A2A receptor signaling contributes to neurodegeneration and cognitive impairments in a mouse model of synucleinopathy.

    PubMed

    Hu, Qidi; Ren, Xiangpeng; Liu, Ya; Li, Zhihui; Zhang, Liping; Chen, Xingjun; He, Chaoxiang; Chen, Jiang-Fan

    2016-09-01

    Synucleinopathy is characterized by abnormal accumulation of misfolded α-synuclein (α-Syn)-positive cytoplasmic inclusions and by neurodegeneration and cognitive impairments, but the pathogenesis mechanism of synucleinopathy remains to be defined. Using a transmission model of synucleinopathy by intracerebral injection of preformed A53T α-Syn fibrils, we investigated whether aberrant adenosine A2A receptor (A2AR) signaling contributed to pathogenesis of synucleinopathy. We demonstrated that intra-hippocampal injection of preformed mutant α-Syn fibrils triggered a striking and selective induction of A2AR expression which was closely co-localized with pSer129 α-Syn-rich inclusions in neurons and glial cells of hippocampus. Importantly, by abolishing aberrant A2AR signaling triggered by mutant α-Syn, genetic deletion of A2ARs blunted a cascade of pathological events leading to synucleinopathy, including pSer129 α-Syn-rich and p62-positive aggregates, NF-κB activation and astrogliosis, apoptotic neuronal cell death and working memory deficits without affecting motor activity. These findings define α-Syn-triggered aberrant A2AR signaling as a critical pathogenesis mechanism of synucleinopathy with dual controls of cognition and neurodegeneration by modulating α-Syn aggregates. Thus, aberrant A2AR signaling represents a useful biomarker as well as a therapeutic target of synucleinopathy. Copyright © 2016 Elsevier Inc. All rights reserved.

  6. The Role of Adenosine A2A Receptor, CYP450s, and PPARs in the Regulation of Vascular Tone

    PubMed Central

    Khayat, Maan T.

    2017-01-01

    Adenosine is an endogenous mediator involved in a myriad of physiologic functions, including vascular tone regulation. It is also implicated in some pathologic conditions. Four distinct receptor subtypes mediate the effects of adenosine, such as its role in the regulation of the vascular tone. Vascular tone regulation is a complex and continuous process which involves many mechanisms and mediators that are not fully disclosed. The vascular endothelium plays a pivotal role in regulating blood flow to and from all body organs. Also, the vascular endothelium is not merely a physical barrier; it is a complex tissue with numerous functions. Among adenosine receptors, A2A receptor subtype (A2AAR) stands out as the primary receptor responsible for the vasodilatory effects of adenosine. This review focuses on important effectors of the vascular endothelium, including adenosine, adenosine receptors, EETs (epoxyeicosatrienoic acids), HETEs (hydroxyeicosatetraenoic acids), PPARs (peroxisome proliferator-activated receptors), and KATP channels. Given the impact of vascular tone regulation in cardiovascular physiology and pathophysiology, better understanding of the mechanisms affecting it could have a significant potential for developing therapeutic agents for cardiovascular diseases. PMID:28884118

  7. Sinomenine Protects against Lipopolysaccharide-Induced Acute Lung Injury in Mice via Adenosine A2A Receptor Signaling

    PubMed Central

    Li, Jun; Zhao, Li; He, Xie; Zeng, Yi-Jun; Dai, Shuang-Shuang

    2013-01-01

    Sinomenine (SIN) is a bioactive alkaloid extracted from the Chinese medicinal plant Sinomenium acutum, which is widely used in the clinical treatment of rheumatoid arthritis (RA). However, its role in acute lung injury (ALI) is unclear. In this study, we investigate the role of SIN in lipopolysaccharide (LPS)-induced ALI in mice. After ALI, lung water content and histological signs of pulmonary injury were attenuated, whereas the PaO2/FIO2 (P/F) ratios were elevated significantly in the mice pretreated with SIN. Additionally, SIN markedly inhibited inflammatory cytokine TNF-α and IL-1β expression levels as well as neutrophil infiltration in the lung tissues of the mice. Microarray analysis and real-time PCR showed that SIN treatment upregulated adenosine A2A receptor (A2AR) expression, and the protective effect of SIN was abolished in A2AR knockout mice. Further investigation in isolated mouse neutrophils confirmed the upregulation of A2AR by SIN and showed that A2AR-cAMP-PKA signaling was involved in the anti-inflammatory effect of SIN. Taken together, these findings demonstrate an A2AR-associated anti-inflammatory effect and the protective role of SIN in ALI, which suggests a potential novel approach to treat ALI. PMID:23555007

  8. Neuroprotective potential of adenosine A2A and cannabinoid CB1 receptor antagonists in an animal model of Parkinson disease.

    PubMed

    Cerri, Silvia; Levandis, Giovanna; Ambrosi, Giulia; Montepeloso, Elena; Antoninetti, Gian Filippo; Franco, Rafael; Lanciego, José Luis; Baqi, Younis; Müller, Christa E; Pinna, Annalisa; Blandini, Fabio; Armentero, Marie Therese

    2014-05-01

    The development of nondopaminergic therapeutic strategies that may improve motor and nonmotor deficits, while possibly slowing down the neurodegenerative process and associated neuroinflammation,is a primary goal of Parkinson disease (PD) research. We investigated the neuroprotective and anti-inflammatory potential of combined and single treatment with adenosine A2A and cannabinoid CB1 receptor antagonists MSX-3 and rimonabant, respectively, in a rodent model of PD. Rats bearing a unilateral intrastriatal 6-hydroxydopamine lesion were treated chronically with MSX-3 (0.5or 1 mg/kg/d) and rimonabant (0.1 mg/kg/d) given as monotherapy or combined. The effects of the treatments to counteract dopaminergic cell death and neuroinflammation were assessed by immunohistochemistry for tyrosine hydroxylase and glial cell markers, respectively. Both rimonabant and MSX-3 (1 mg/kg/d) promoted dopaminergic neuron survival in the substantia nigra pars compacta (SNc) when given alone; this effect was weakened when the compounds were combined. Glial activation was not significantly affected by MSX-3 (1 mg/kg/d), whereas rimonabant seemed to increase astrocyte cell density in the SNc. Our findings demonstrate the neuroprotective potential of single treatments and suggest that glial cells might be involved in this protective effect. The results also indicate that the neuroprotective potential of combined therapy may not necessarily reflect or promote single-drug effects and point out that special care should be taken when considering multidrug therapies in PD.

  9. Role of adenosine A(2A) receptors in modulating synaptic functions and brain levels of BDNF: a possible key mechanism in the pathophysiology of Huntington's disease.

    PubMed

    Tebano, Maria Teresa; Martire, Alberto; Chiodi, Valentina; Ferrante, Antonella; Popoli, Patrizia

    2010-09-01

    In the last few years, accumulating evidence has shown the existence of an important cross-talk between adenosine A(2A) receptors (A(2A)Rs) and brain-derived neurotrophic factor (BDNF). Not only are A(2A)Rs involved in the mechanism of transactivation of BDNF receptor TrkB, they also modulate the effect of BDNF on synaptic transmission, playing a facilitatory and permissive role. The cAMP-PKA pathway, the main transduction system operated by A(2A)Rs, is involved in such effects. Furthermore, a basal tonus of A(2A)Rs is required to allow the regulation of BDNF physiological levels in the brain, as demonstrated by the reduced protein levels measured in A(2A)Rs KO mice. The crucial role of adenosine A(2A)Rs in the maintenance of synaptic functions and BDNF levels will be reviewed here and discussed in the light of possible implications for Huntington's disease therapy, in which a joint impairment of BDNF and A(2A)Rs seems to play a pathogenetic role.

  10. Calcium-mediated modulation of the quaternary structure and function of adenosine A2A-dopamine D2 receptor heteromers

    PubMed Central

    Ferré, Sergi; Woods, Amina S.; Navarro, Gemma; Aymerich, Marisol; Lluís, Carme; Franco, Rafael

    2009-01-01

    The adenosine A2A-dopamine D2 receptor heteromer is one of the most studied receptor heteromers. It has important implications for basal ganglia function and pathology. Recent studies using Bioluminescence and Sequential Resonance Energy Transfer techniques shed light on the role of Ca2+ in the modulation of the quaternary structure of the A2A-D2 receptor heteromer, which was found to depend on the binding of calmodulin (CaM) to the carboxy terminus of the A2A receptor in the A2A-D2 receptor heteromer. Importantly, the changes in quaternary structure correlate with changes in function. A Ca2+/CaM-dependent modulation of MAPK signaling upon agonist treatment could only be observed in cells expressing A2A-D2 receptor heteromers. These studies provide a first example of a Ca2+-mediated modulation of the quaternary structure and function of a receptor heteromer. PMID:19896897

  11. Adenosine A2A receptor agonist prevents cardiac remodeling and dysfunction in spontaneously hypertensive male rats after myocardial infarction

    PubMed Central

    da Silva, Jaqueline S; Gabriel-Costa, Daniele; Sudo, Roberto T; Wang, Hao; Groban, Leanne; Ferraz, Emanuele B; Nascimento, José Hamilton M; Fraga, Carlos Alberto M; Barreiro, Eliezer J; Zapata-Sudo, Gisele

    2017-01-01

    Background This work evaluated the hypothesis that 3,4-methylenedioxybenzoyl-2-thienylhydrazone (LASSBio-294), an agonist of adenosine A2A receptor, could be beneficial for preventing cardiac dysfunction due to hypertension associated with myocardial infarction (MI). Methods Male spontaneously hypertensive rats (SHR) were randomly divided into four groups (six animals per group): sham-operation (SHR-Sham), and myocardial infarction rats (SHR-MI) were treated orally either with vehicle or LASSBio-294 (10 and 20 mg.kg−1.d−1) for 4 weeks. Echocardiography and in vivo hemodynamic parameters measured left ventricle (LV) structure and function. Exercise tolerance was evaluated using a treadmill test. Cardiac remodeling was accessed by LV collagen deposition and tumor necrosis factor α expression. Results Early mitral inflow velocity was significantly reduced in the SHR-MI group, and there was significant recovery in a dose-dependent manner after treatment with LASSBio-294. Exercise intolerance observed in the SHR-MI group was prevented by 10 mg.kg−1.d−1 of LASS-Bio-294, and exercise tolerance exceeded that of the SHR-Sham group at 20 mg.kg−1.d−1. LV end-diastolic pressure increased after MI, and this was prevented by 10 and 20 mg.kg−1.d−1 of LASSBio-294. Sarcoplasmic reticulum Ca2+ ATPase levels were restored in a dose-dependent manner after treatment with LASSBio-294. Fibrosis and inflammatory processes were also counteracted by LASSBio-294, with reductions in LV collagen deposition and tumor necrosis factor α expression. Conclusion In summary, oral administration of LASSBio-294 after MI in a dose-dependent manner prevented the development of cardiac dysfunction, demonstrating this compound’s potential as an alternative treatment for heart failure in the setting of ischemic heart disease with superimposed chronic hypertension. PMID:28293100

  12. GPCR 3D homology models for Ligand Screening: Lessons Learned from Blind Predictions of Adenosine A2a Receptor complex

    PubMed Central

    Katritch, Vsevolod; Rueda, Manuel; Lam, Polo Chun-Hung; Yeager, Mark; Abagyan, Ruben

    2009-01-01

    Proteins of the G-protein coupled receptor (GPCR) family present numerous attractive targets for rational drug design, but also a formidable challenge for identification and conformational modeling of their 3D structure. A recently performed assessment of blind predictions of adenosine A2a receptor (AA2AR) structure in complex with ZM241385 (ZMA) antagonist provided a first example of unbiased evaluation of the current modeling algorithms on a GPCR target with ~30% sequence identity to the closest structural template. Several of the 29 groups participating in this assessment exercise (Michino et al., doi:10.1038/nrd2877) successfully predicted the overall position of the ligand ZMA in the AA2AR ligand binding pocket, however models from only three groups captured more than 40% of the ligand-receptor contacts. Here we describe two of these top performing approaches, in which all-atom models of the AA2AR were generated by homology modeling followed by ligand guided backbone ensemble receptor optimization (LiBERO). The resulting AA2AR-ZMA models, along with the best models from other groups are assessed here for their virtual ligand screening (VLS) performance on a large set of GPCR ligands. We show that ligand guided optimization was critical for improvement of both ligand-receptor contacts and VLS performance as compared to the initial raw homology models. The best blindly predicted models performed on par with the crystal structure of AA2AR in selecting known antagonists from decoys, as well as from antagonists for other adenosine subtypes and AA2AR agonists. These results suggest that despite certain inaccuracies, the optimized homology models can be useful in the drug discovery process. PMID:20063437

  13. Direct or indirect stimulation of adenosine A2A receptors enhances bone regeneration as well as bone morphogenetic protein-2.

    PubMed

    Mediero, Aránzazu; Wilder, Tuere; Perez-Aso, Miguel; Cronstein, Bruce N

    2015-04-01

    Promoting bone regeneration and repair of bone defects is a need that has not been well met to date. We have previously found that adenosine, acting via A2A receptors (A2AR) promotes wound healing and inhibits inflammatory osteolysis and hypothesized that A2AR might be a novel target to promote bone regeneration. Therefore, we determined whether direct A2AR stimulation or increasing endogenous adenosine concentrations via purine transport blockade with dipyridamole regulates bone formation. We determined whether coverage of a 3 mm trephine defect in a mouse skull with a collagen scaffold soaked in saline, bone morphogenetic protein-2 (BMP-2; 200 ng), 1 μM CGS21680 (A2AR agonist, EC50 = 160 nM), or 1 μM dipyridamole (EC50 = 32 nM) promoted bone regeneration. Microcomputed tomography examination demonstrated that CGS21680 and dipyridamole markedly enhanced bone regeneration as well as BMP-2 8 wk after surgery (60 ± 2%, 79 ± 2%, and 75 ± 1% bone regeneration, respectively, vs. 32 ± 2% in control, P < 0.001). Blockade by a selective A2AR antagonist (ZM241385, 1 μM) or deletion of A2AR abrogated the effect of CGS21680 and dipyridamole on bone regeneration. Both CGS21680 and dipyridamole treatment increased alkaline phosphatase-positive osteoblasts and diminished tartrate resistance acid phosphatase-positive osteoclasts in the defects. In vivo imaging with a fluorescent dye for new bone formation revealed a strong fluorescent signal in treated animals that was equivalent to BMP-2. In conclusion, stimulation of A2AR by specific agonists or by increasing endogenous adenosine levels stimulates new bone formation as well as BMP-2 and represents a novel approach to stimulating bone regeneration. © FASEB.

  14. Impact of purification conditions and history on A2A adenosine receptor activity: The role of CHAPS and lipids.

    PubMed

    Naranjo, Andrea N; McNeely, Patrick M; Katsaras, John; Robinson, Anne Skaja

    2016-08-01

    The adenosine A2A receptor (A2AR) is a much-studied class A G protein-coupled receptor (GPCR). For biophysical studies, A2AR is commonly purified in a detergent mixture of dodecylmaltoside (DDM), 3-(3-cholamidopropyl) dimethylammoniopropane sulfonate (CHAPS), and cholesteryl hemisuccinate (CHS). Here we studied the effects of CHAPS on the ligand binding activity and stability of wild type, full-length human A2AR. We also tested the cholesterol requirement for maintaining the active conformation of the receptor when solubilized in detergent micelles. To this end, the receptor was purified using DDM, DDM/CHAPS, or the short hydrocarbon chain lipid 1,2-dihexanoyl-sn-glycero-3-phosphocholine (DHPC, di-6:0PC). After solubilization in DDM, DDM/CHAPS, or DHPC micelles, although A2AR was found to retain its native-like fold, its binding ability was significantly compromised compared to DDM or DDM/CHAPS with CHS. It therefore appears that although cholesterol is not needed for A2AR to retain a native-like, α-helical conformation, it may be a critical component for high affinity ligand binding. Further, this result suggests that the conformational differences between the active and inactive protein may be so subtle that commonly used spectroscopic methods are unable to differentiate between the two forms, highlighting the need for activity measurements. The studies presented in this paper also underline the importance of the protein's purification history; i.e., detergents that interact with the protein during purification affect the ligand binding properties of the receptor in an irreversible manner. Copyright © 2016 Elsevier Inc. All rights reserved.

  15. Human Adenosine A2A Receptor Binds Calmodulin with High Affinity in a Calcium-Dependent Manner

    PubMed Central

    Piirainen, Henni; Hellman, Maarit; Tossavainen, Helena; Permi, Perttu; Kursula, Petri; Jaakola, Veli-Pekka

    2015-01-01

    Understanding how ligands bind to G-protein-coupled receptors and how binding changes receptor structure to affect signaling is critical for developing a complete picture of the signal transduction process. The adenosine A2A receptor (A2AR) is a particularly interesting example, as it has an exceptionally long intracellular carboxyl terminus, which is predicted to be mainly disordered. Experimental data on the structure of the A2AR C-terminus is lacking, because published structures of A2AR do not include the C-terminus. Calmodulin has been reported to bind to the A2AR C-terminus, with a possible binding site on helix 8, next to the membrane. The biological meaning of the interaction as well as its calcium dependence, thermodynamic parameters, and organization of the proteins in the complex are unclear. Here, we characterized the structure of the A2AR C-terminus and the A2AR C-terminus-calmodulin complex using different biophysical methods, including native gel and analytical gel filtration, isothermal titration calorimetry, NMR spectroscopy, and small-angle X-ray scattering. We found that the C-terminus is disordered and flexible, and it binds with high affinity (Kd = 98 nM) to calmodulin without major conformational changes in the domain. Calmodulin binds to helix 8 of the A2AR in a calcium-dependent manner that can displace binding of A2AR to lipid vesicles. We also predicted and classified putative calmodulin-binding sites in a larger group of G-protein-coupled receptors. PMID:25692595

  16. Caffeine acts through neuronal adenosine A2A receptors to prevent mood and memory dysfunction triggered by chronic stress.

    PubMed

    Kaster, Manuella P; Machado, Nuno J; Silva, Henrique B; Nunes, Ana; Ardais, Ana Paula; Santana, Magda; Baqi, Younis; Müller, Christa E; Rodrigues, Ana Lúcia S; Porciúncula, Lisiane O; Chen, Jiang Fan; Tomé, Ângelo R; Agostinho, Paula; Canas, Paula M; Cunha, Rodrigo A

    2015-06-23

    The consumption of caffeine (an adenosine receptor antagonist) correlates inversely with depression and memory deterioration, and adenosine A2A receptor (A2AR) antagonists emerge as candidate therapeutic targets because they control aberrant synaptic plasticity and afford neuroprotection. Therefore we tested the ability of A2AR to control the behavioral, electrophysiological, and neurochemical modifications caused by chronic unpredictable stress (CUS), which alters hippocampal circuits, dampens mood and memory performance, and enhances susceptibility to depression. CUS for 3 wk in adult mice induced anxiogenic and helpless-like behavior and decreased memory performance. These behavioral changes were accompanied by synaptic alterations, typified by a decrease in synaptic plasticity and a reduced density of synaptic proteins (synaptosomal-associated protein 25, syntaxin, and vesicular glutamate transporter type 1), together with an increased density of A2AR in glutamatergic terminals in the hippocampus. Except for anxiety, for which results were mixed, CUS-induced behavioral and synaptic alterations were prevented by (i) caffeine (1 g/L in the drinking water, starting 3 wk before and continued throughout CUS); (ii) the selective A2AR antagonist KW6002 (3 mg/kg, p.o.); (iii) global A2AR deletion; and (iv) selective A2AR deletion in forebrain neurons. Notably, A2AR blockade was not only prophylactic but also therapeutically efficacious, because a 3-wk treatment with the A2AR antagonist SCH58261 (0.1 mg/kg, i.p.) reversed the mood and synaptic dysfunction caused by CUS. These results herald a key role for synaptic A2AR in the control of chronic stress-induced modifications and suggest A2AR as candidate targets to alleviate the consequences of chronic stress on brain function.

  17. Caffeine acts through neuronal adenosine A2A receptors to prevent mood and memory dysfunction triggered by chronic stress

    PubMed Central

    Kaster, Manuella P.; Machado, Nuno J.; Silva, Henrique B.; Nunes, Ana; Ardais, Ana Paula; Santana, Magda; Baqi, Younis; Müller, Christa E.; Rodrigues, Ana Lúcia S.; Porciúncula, Lisiane O.; Chen, Jiang Fan; Tomé, Ângelo R.; Agostinho, Paula; Canas, Paula M.; Cunha, Rodrigo A.

    2015-01-01

    The consumption of caffeine (an adenosine receptor antagonist) correlates inversely with depression and memory deterioration, and adenosine A2A receptor (A2AR) antagonists emerge as candidate therapeutic targets because they control aberrant synaptic plasticity and afford neuroprotection. Therefore we tested the ability of A2AR to control the behavioral, electrophysiological, and neurochemical modifications caused by chronic unpredictable stress (CUS), which alters hippocampal circuits, dampens mood and memory performance, and enhances susceptibility to depression. CUS for 3 wk in adult mice induced anxiogenic and helpless-like behavior and decreased memory performance. These behavioral changes were accompanied by synaptic alterations, typified by a decrease in synaptic plasticity and a reduced density of synaptic proteins (synaptosomal-associated protein 25, syntaxin, and vesicular glutamate transporter type 1), together with an increased density of A2AR in glutamatergic terminals in the hippocampus. Except for anxiety, for which results were mixed, CUS-induced behavioral and synaptic alterations were prevented by (i) caffeine (1 g/L in the drinking water, starting 3 wk before and continued throughout CUS); (ii) the selective A2AR antagonist KW6002 (3 mg/kg, p.o.); (iii) global A2AR deletion; and (iv) selective A2AR deletion in forebrain neurons. Notably, A2AR blockade was not only prophylactic but also therapeutically efficacious, because a 3-wk treatment with the A2AR antagonist SCH58261 (0.1 mg/kg, i.p.) reversed the mood and synaptic dysfunction caused by CUS. These results herald a key role for synaptic A2AR in the control of chronic stress-induced modifications and suggest A2AR as candidate targets to alleviate the consequences of chronic stress on brain function. PMID:26056314

  18. Ex vivo Perfusion with Adenosine A2A Receptor Agonist Enhances Rehabilitation of Murine Donor Lungs after Circulatory Death

    PubMed Central

    Stone, Mathew L.; Sharma, Ashish K.; Mas, Valeria. R.; Gehrau, Ricardo C.; Mulloy, Daniel P.; Zhao, Yunge; Lau, Christine L.; Kron, Irving L.; Laubach, Victor E.

    2015-01-01

    Background Ex vivo lung perfusion (EVLP) enables assessment and rehabilitation of marginal donor lungs prior to transplantation. We previously demonstrated that adenosine A2A receptor (A2AR) agonism attenuates lung ischemia-reperfusion injury. The current study utilizes a novel murine EVLP model to test the hypothesis that A2AR agonist enhances EVLP-mediated rehabilitation of donation after circulatory death (DCD) lungs. Methods Mice underwent euthanasia and 60 min warm ischemia, and lungs were flushed with Perfadex and underwent cold static preservation (CSP, 60 min). Three groups were studied: no EVLP (CSP), EVLP with Steen solution for 60 min (EVLP), and EVLP with Steen solution supplemented with ATL1223, a selective A2AR agonist (EVLP+ATL1223). Lung function, wet/dry weight, cytokines and neutrophil numbers were measured. Microarrays were performed using the Affymetrix GeneChip Mouse Genome 430A 2.0 Array. Results EVLP significantly improved lung function versus CSP, which was further, significantly improved by EVLP+ATL1223. Lung edema, cytokines and neutrophil counts were reduced after EVLP and further, significantly reduced after EVLP+ATL1223. Gene array analysis revealed differential expression of 1,594 genes after EVLP, which comprise canonical pathways involved in inflammation and innate immunity including IL-1, IL-8, IL-6 and IL-17 signaling. Several pathways were uniquely regulated by EVLP+ATL1223 including the downregulation of genes involved in IL-1 signaling such as ADCY9, ECSIT, IRAK1, MAPK12 and TOLLIP. Conclusion EVLP modulates pro-inflammatory genes and reduces pulmonary dysfunction, edema and inflammation in DCD lungs, which are further reduced by A2AR agonism. This murine EVLP model provides a novel platform to study rehabilitative mechanisms of DCD lungs. PMID:26262504

  19. Genetic blockade of adenosine A2A receptors induces cognitive impairments and anatomical changes related to psychotic symptoms in mice.

    PubMed

    Moscoso-Castro, Maria; Gracia-Rubio, Irene; Ciruela, Francisco; Valverde, Olga

    2016-07-01

    Schizophrenia is a chronic severe mental disorder with a presumed neurodevelopmental origin, and no effective treatment. Schizophrenia is a multifactorial disease with genetic, environmental and neurochemical etiology. The main theories on the pathophysiology of this disorder include alterations in dopaminergic and glutamatergic neurotransmission in limbic and cortical areas of the brain. Early hypotheses also suggested that nucleoside adenosine is a putative affected neurotransmitter system, and clinical evidence suggests that adenosine adjuvants improve treatment outcomes, especially in poorly responsive patients. Hence, it is important to elucidate the role of the neuromodulator adenosine in the pathophysiology of schizophrenia. A2A adenosine receptor (A2AR) subtypes are expressed in brain areas controlling motivational responses and cognition, including striatum, and in lower levels in hippocampus and cerebral cortex. The aim of this study was to characterize A2AR knockout (KO) mice with complete and specific inactivation of A2AR, as an animal model for schizophrenia. We performed behavioral, anatomical and neurochemical studies to assess psychotic-like symptoms in adult male and female KO and wild-type (WT) littermates. Our results show impairments in inhibitory responses and sensory gating in A2AR KO animals. Hyperlocomotion induced by d-amphetamine and MK-801 was reduced in KO animals when compared to WT littermates. Moreover, A2AR KO animals show motor disturbances, social and cognitive alterations. Finally, behavioral impairments were associated with enlargement of brain lateral ventricles and decreased BDNF levels in the hippocampus. These data highlight the role of adenosine in the pathophysiology of schizophrenia and provide new possibilities for the therapeutic management of schizophrenia.

  20. Polydeoxyribonucleotide, an Adenosine-A2A Receptor Agonist, Preserves Blood Testis Barrier from Cadmium-Induced Injury

    PubMed Central

    Squadrito, Francesco; Micali, Antonio; Rinaldi, Mariagrazia; Irrera, Natasha; Marini, Herbert; Puzzolo, Domenico; Pisani, Antonina; Lorenzini, Cesare; Valenti, Andrea; Laurà, Rosaria; Germanà, Antonino; Bitto, Alessandra; Pizzino, Gabriele; Pallio, Giovanni; Altavilla, Domenica; Minutoli, Letteria

    2017-01-01

    Cadmium (Cd) impairs blood-testis barrier (BTB). Polydeoxyribonucleotide (PDRN), an adenosine A2A agonist, has positive effects on male reproductive system. We investigated the effects of PDRN on the morphological and functional changes induced by Cd in mice testes. Adult Swiss mice were divided into four groups: controls administered with 0.9% NaCl (1 ml/kg, i.p., daily) or with PDRN (8 mg/kg, i.p. daily), animals challenged with Cd chloride (CdCl2; 2 mg/kg, i.p, daily) and animals challenged with CdCl2 (2 mg/kg, i.p., daily) and treated with PDRN (8 mg/kg, i.p., daily). Experiments lasted 14 days. Testes were processed for biochemical, structural, and ultrastructural evaluation and hormones were assayed in serum. CdCl2 increased pERK 1/2 expression and Follicle Stimulating Hormone (FSH) and Luteinizing Hormone (LH) levels; it decreased testosterone (TE) and inhibin-B levels and induced structural damages in extratubular compartment and in seminiferous epithelium, with ultrastructural features of BTB disruption. Many TUNEL-positive germ cells were present. CdCl2 increased tubular TGF-β3 immunoreactivity and reduced claudin-11, occludin, and N-cadherin immunoreactivity. PDRN administration reduced pERK 1/2 expression, FSH, and LH levels; it increased TE and inhibin-B levels, ameliorated germinal epithelium changes and protected BTB ultrastructure. Few TUNEL-positive germ cells were present and the extratubular compartment was preserved. Furthermore, PDRN decreased TGF-β3 immunoreactivity and enhanced claudin-11, occludin, and N-cadherin immunoreactivity. We demonstrate a protective effect of PDRN on Cd-induced damages of BTB and suggest that PDRN may play an important role against Cd, particularly against its harmful effects on gametogenesis. PMID:28119612

  1. Impact of purification conditions and history on A2A adenosine receptor activity: The role of CHAPS and lipids

    DOE PAGES

    Naranjo, Andrea N.; McNeely, Patrick M.; Katsaras, John; ...

    2016-05-27

    The adenosine A2A receptor (A2AR) is a much-studied class A G protein-coupled receptor (GPCR). For biophysical studies, A2AR is commonly purified in a detergent mixture of dodecylmaltoside (DDM), 3-(3-cholamidopropyl) dimethylammoniopropane sulfonate (CHAPS), and cholesteryl hemisuccinate (CHS). Here we studied the effects of CHAPS on the ligand binding activity and stability of wild type, full-length human A2AR. We also tested the cholesterol requirement for maintaining the active conformation of the receptor when solubilized in detergent micelles. To this end, the receptor was purified using DDM, DDM/CHAPS, or the short hydrocarbon chain lipid 1,2-dihexanoyl-sn-glycero-3-phosphocholine (DHPC, di-6:0PC). After solubilization in DDM, DDM/CHAPS, ormore » DHPC micelles, although A2AR was found to retain its native-like fold, its binding ability was significantly compromised compared to DDM or DDM/CHAPS with CHS. It therefore appears that although cholesterol is not needed for A2AR to retain a native-like, α-helical conformation, it may be a critical component for high affinity ligand binding. Further, this result suggests that the conformational differences between the active and inactive protein may be so subtle that commonly used spectroscopic methods are unable to differentiate between the two forms, highlighting the need for activity measurements. Furthermore, the studies presented in this paper also underline the importance of the protein’s purification history; i.e., detergents that interact with the protein during purification affect the ligand binding properties of the receptor in an irreversible manner.« less

  2. Ex Vivo Perfusion With Adenosine A2A Receptor Agonist Enhances Rehabilitation of Murine Donor Lungs After Circulatory Death.

    PubMed

    Stone, Matthew L; Sharma, Ashish K; Mas, Valeria R; Gehrau, Ricardo C; Mulloy, Daniel P; Zhao, Yunge; Lau, Christine L; Kron, Irving L; Huerter, Mary E; Laubach, Victor E

    2015-12-01

    Ex vivo lung perfusion (EVLP) enables assessment and rehabilitation of marginal donor lungs before transplantation. We previously demonstrated that adenosine A2A receptor (A2AR) agonism attenuates lung ischemia-reperfusion injury. The current study utilizes a novel murine EVLP model to test the hypothesis that A2AR agonist enhances EVLP-mediated rehabilitation of donation after circulatory death (DCD) lungs. Mice underwent euthanasia and 60 minutes warm ischemia, and lungs were flushed with Perfadex and underwent cold static preservation (CSP, 60 minutes). Three groups were studied: no EVLP (CSP), EVLP with Steen solution for 60 minutes (EVLP), and EVLP with Steen solution supplemented with ATL1223, a selective A2AR agonist (EVLP + ATL1223). Lung function, wet/dry weight, cytokines and neutrophil numbers were measured. Microarrays were performed using the Affymetrix GeneChip Mouse Genome 430A 2.0 Array. Ex vivo lung perfusion significantly improved lung function versus CSP, which was further, significantly improved by EVLP + ATL1223. Lung edema, cytokines, and neutrophil counts were reduced after EVLP and further, significantly reduced after EVLP + ATL1223. Gene array analysis revealed differential expression of 1594 genes after EVLP, which comprise canonical pathways involved in inflammation and innate immunity including IL-1, IL-8, IL-6, and IL-17 signaling. Several pathways were uniquely regulated by EVLP + ATL1223 including the downregulation of genes involved in IL-1 signaling, such as ADCY9, ECSIT, IRAK1, MAPK12, and TOLLIP. Ex vivo lung perfusion modulates proinflammatory genes and reduces pulmonary dysfunction, edema, and inflammation in DCD lungs, which are further reduced by A2AR agonism. This murine EVLP model provides a novel platform to study rehabilitative mechanisms of DCD lungs.

  3. A(2A) adenosine receptors are differentially modulated by pharmacological treatments in rheumatoid arthritis patients and their stimulation ameliorates adjuvant-induced arthritis in rats.

    PubMed

    Vincenzi, Fabrizio; Padovan, Melissa; Targa, Martina; Corciulo, Carmen; Giacuzzo, Sarah; Merighi, Stefania; Gessi, Stefania; Govoni, Marcello; Borea, Pier Andrea; Varani, Katia

    2013-01-01

    A(2A) adenosine receptors (ARs) play a key role in the inhibition of the inflammatory process. The purpose of this study was to evaluate the modulation of A(2A)ARs in rheumatoid arthritis (RA) patients after different pharmacological treatments and to investigate the effect of A(2A)AR stimulation in a rat model of arthritis. We investigated A(2A)AR density and functionality in RA progression by using a longitudinal study in RA patients before and after methotrexate (MTX), anti-TNFα agents or rituximab treatments. A(2A)ARs were analyzed by saturation binding assays in lymphocytes from RA patients throughout the 24-month study timeframe. In an adjuvant-induced arthritis model in rats we showed the efficacy of the A(2A)AR agonist, CGS 21680 in comparison with standard therapies by means of paw volume assessment, radiographic and ultrasonographic imaging. Arthritic-associated pain was investigated in mechanical allodynia and thermal hyperalgesia tests. IL-10 release following A(2A)AR stimulation in lymphocytes from RA patients and in serum from arthritic rats was measured. In lymphocytes obtained from RA patients, the A(2A)AR up-regulation was gradually reduced in function of the treatment time and the stimulation of these receptors mediated a significant increase of IL-10 production. In the same cells, CGS 21680 did not affected cell viability and did not produced cytotoxic effects. The A(2A)AR agonist CGS 21680 was highly effective, as suggested by the marked reduction of clinical signs, in rat adjuvant-induced arthritis and associated pain. This study highlighted that A(2A)AR agonists represent a physiological-like therapeutic alternative for RA treatment as suggested by the anti-inflammatory role of A(2A)ARs in lymphocytes from RA patients. The effectiveness of A(2A)AR stimulation in a rat model of arthritis supported the role of A(2A)AR agonists as potential pharmacological treatment for RA.

  4. Chronic and acute adenosine A2A receptor blockade prevents long-term episodic memory disruption caused by acute cannabinoid CB1 receptor activation.

    PubMed

    Mouro, Francisco M; Batalha, Vânia L; Ferreira, Diana G; Coelho, Joana E; Baqi, Younis; Müller, Christa E; Lopes, Luísa V; Ribeiro, Joaquim A; Sebastião, Ana M

    2017-05-01

    Cannabinoid-mediated memory impairment is a concern in cannabinoid-based therapies. Caffeine exacerbates cannabinoid CB1 receptor (CB1R)-induced memory deficits through an adenosine A1 receptor-mediated mechanism. We now evaluated how chronic or acute blockade of adenosine A2A receptors (A2ARs) affects long-term episodic memory deficits induced by a single injection of a selective CB1R agonist. Long-term episodic memory was assessed by the novel object recognition (NOR) test. Mice received an intraperitoneal (i.p.) injection of the CB1/CB2 receptor agonist WIN 55,212-2 (1 mg/kg) immediately after the NOR training, being tested for novelty recognition 24 h later. Anxiety levels were assessed by the Elevated Plus Maze test, immediately after the NOR. Mice were also tested for exploratory behaviour at the Open Field. For chronic A2AR blockade, KW-6002 (istradefylline) (3 mg/kg/day) was administered orally for 30 days; acute blockade of A2ARs was assessed by i.p. injection of SCH 58261 (1 mg/kg) administered either together with WIN 55,212-2 or only 30 min before the NOR test phase. The involvement of CB1Rs was assessed by using the CB1R antagonist, AM251 (3 mg/kg, i.p.). WIN 55,212-2 caused a disruption in NOR, an action absent in mice also receiving AM251, KW-6002 or SCH 58261 during the encoding/consolidation phase; SCH 58251 was ineffective if present during retrieval only. No effects were detected in the Elevated Plus maze or Open Field Test. The finding that CB1R-mediated memory disruption is prevented by antagonism of adenosine A2ARs, highlights a possibility to prevent cognitive side effects when therapeutic application of CB1R drugs is desired.

  5. Guanosine may increase absence epileptic activity by means of A2A adenosine receptors in Wistar Albino Glaxo Rijswijk rats.

    PubMed

    Lakatos, Renáta Krisztina; Dobolyi, Árpád; Todorov, Mihail Ivilinov; Kékesi, Katalin A; Juhász, Gábor; Aleksza, Magdolna; Kovács, Zsolt

    2016-06-01

    The non-adenosine nucleoside guanosine (Guo) was demonstrated to decrease quinolinic acid(QA)-induced seizures, spontaneously emerged absence epileptic seizures and lipopolysaccharide(LPS)-evoked induction of absence epileptic seizures suggesting its antiepileptic potential. It was also described previously that intraperitoneal (i.p.) injection of 20 and 50mg/kg Guo decreased the number of spike-wave discharges (SWDs) in a well investigated model of human absence epilepsy, the Wistar Albino Glaxo Rijswijk (WAG/Rij) rats during 4th (20mg/kg Guo) and 3rd as well as 4th (50mg/kg Guo) measuring hours. Guanosine can potentially decrease SWD number by means of its putative receptors but absence epileptic activity changing effects of Guo by means of increased extracellular adenosine (Ado) cannot be excluded. An increase in the dose of i.p. injected Guo is limited by its low solubility in saline, therefore, we addressed in the present study whether higher doses of Guo, diluted in sodium hydroxide (NaOH) solution, have more potent antiepileptic effect in WAG/Rij rats. We confirmed that i.p. 50mg/kg Guo decreased but, surprisingly, i.p. 100mg/kg Guo enhanced the number of SWDs in WAG/Rij rats. Combined i.p. injection of a non-selective Ado receptor antagonist theophylline (5mg/kg) or a selective Ado A2A receptor (A2AR) antagonist SCH 58261 (7-(2-phenylethyl)-5-amino-2-(2-furyl)-pyrazolo-[4,3-e]-1,2,4-triazolo[1,5-c]pyrimidine) (1mg/kg) and a cyclooxygenase 1 and 2/COX-1 and COX-2 inhibitor indomethacin (10mg/kg) with 100mg/kg Guo decreased the SWD number compared to i.p. 100mg/kg Guo alone. The results suggest that i.p. 100mg/kg Guo can increase SWD number by means of the adenosinergic system.

  6. mGlu5, Dopamine D2 and Adenosine A2A Receptors in L-DOPA-induced Dyskinesias.

    PubMed

    Morin, Nicolas; Morissette, Marc; Grégoire, Laurent; Di Paolo, Thérèse

    2016-01-01

    Patients with Parkinson's disease (PD) receiving L-3,4-dihydroxyphenylalanine (L-DOPA, the gold-standard treatment for this disease) frequently develop abnormal involuntary movements, termed L-DOPA-induced dyskinesias (LID). Glutamate overactivity is well documented in PD and LID. An approach to manage LID is to add to L-DOPA specific agents to reduce dyskinesias such as metabotropic glutamate receptor (mGlu receptor) drugs. This article reviews the contribution of mGlu type 5 (mGlu5) receptors in animal models of PD. Several mGlu5 negative allosteric modulators acutely attenuate LID in 1-methyl-4-phenyl-1,2,3,6- tetrahydropyridine (MPTP) monkeys and 6-hydroxydopamine(6-OHDA)-lesioned rats. Chronic administration of mGlu5 negative allosteric modulators to MPTP monkeys and 6-OHDA rats also attenuates LID while maintaining the antiparkinsonian effect of L-DOPA. Radioligand autoradiography shows an elevation of striatal mGlu5 receptors of dyskinetic L-DOPA-treated MPTP monkeys but not in those without LID. The brain molecular correlates of the long-term effect of mGlu5 negative allosteric modulators treatments with L-DOPA attenuating development of LID was shown to extend beyond mGlu5 receptors with normalization of glutamate activity in the basal ganglia of L-DOPA-induced changes of NMDA, AMPA, mGlu2/3 receptors and VGlut2 transporter. In the basal ganglia, mGlu5 receptor negative allosteric modulators also normalize the L-DOPA-induced changes of dopamine D2receptors, their associated signaling proteins (ERK1/2 and Akt/GSK3β) and neuropeptides (preproenkephalin, preprodynorphin) as well as the adenosine A2A receptors expression. These results show in animal models of PD reduction of LID with mGlu5 negative allosteric modulation associated with normalization of glutamate, dopamine and adenosine receptors suggesting a functional link of these receptors in chronic treatment with L-DOPA.

  7. mGlu5, Dopamine D2 and Adenosine A2A Receptors in L-DOPA-induced Dyskinesias

    PubMed Central

    Morin, Nicolas; Morissette, Marc; Grégoire, Laurent; Di Paolo, Thérèse

    2016-01-01

    Patients with Parkinson’s disease (PD) receiving L-3,4-dihydroxyphenylalanine (L-DOPA, the gold-standard treatment for this disease) frequently develop abnormal involuntary movements, termed L-DOPA-induced dyskinesias (LID). Glutamate overactivity is well documented in PD and LID. An approach to manage LID is to add to L-DOPA specific agents to reduce dyskinesias such as metabotropic glutamate receptor (mGlu receptor) drugs. This article reviews the contribution of mGlu type 5 (mGlu5) receptors in animal models of PD. Several mGlu5 negative allosteric modulators acutely attenuate LID in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) monkeys and 6-hydroxydopamine(6-OHDA)-lesioned rats. Chronic administration of mGlu5 negative allosteric modulators to MPTP monkeys and 6-OHDA rats also attenuates LID while maintaining the anti-parkinsonian effect of L-DOPA. Radioligand autoradiography shows an elevation of striatal mGlu5 receptors of dyskinetic L-DOPA-treated MPTP monkeys but not in those without LID. The brain molecular correlates of the long-term effect of mGlu5 negative allosteric modulators treatments with L-DOPA attenuating development of LID was shown to extend beyond mGlu5 receptors with normalization of glutamate activity in the basal ganglia of L-DOPA-induced changes of NMDA, AMPA, mGlu2/3 receptors and VGlut2 transporter. In the basal ganglia, mGlu5 receptor negative allosteric modulators also normalize the L-DOPA-induced changes of dopamine D2 receptors, their associated signaling proteins (ERK1/2 and Akt/GSK3β) and neuropeptides (preproenkephalin, preprodynorphin) as well as the adenosine A2A receptors expression. These results show in animal models of PD reduction of LID with mGlu5 negative allosteric modulation associated with normalization of glutamate, dopamine and adenosine receptors suggesting a functional link of these receptors in chronic treatment with L-DOPA. PMID:26639458

  8. 32 CFR 169a.1 - Purpose.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

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  9. 32 CFR 169a.1 - Purpose.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

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  10. 32 CFR 168a.1 - Purpose.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

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  11. 32 CFR 169a.1 - Purpose.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

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  12. 32 CFR 168a.1 - Purpose.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

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  13. 32 CFR 169a.1 - Purpose.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

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  14. 32 CFR 237a.1 - Purpose.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

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  15. 32 CFR 168a.1 - Purpose.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

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  16. 32 CFR 242a.1 - Applicability.

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    2012-07-01

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  17. 32 CFR 168a.1 - Purpose.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

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  18. 32 CFR 242a.1 - Applicability.

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    2010-07-01

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  19. 32 CFR 242a.1 - Applicability.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

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    Code of Federal Regulations, 2014 CFR

    2014-07-01

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  1. 32 CFR 242a.1 - Applicability.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 32 National Defense 2 2014-07-01 2014-07-01 false Applicability. 242a.1 Section 242a.1 National Defense Department of Defense (Continued) OFFICE OF THE SECRETARY OF DEFENSE (CONTINUED) MISCELLANEOUS PUBLIC MEETING PROCEDURES OF THE BOARD OF REGENTS, UNIFORMED SERVICES UNIVERSITY OF THE HEALTH SCIENCES...

  2. 32 CFR 242a.1 - Applicability.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

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  3. 42 CFR 2a.1 - Applicability.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ...(a)) provides that “ he Secretary may authorize persons engaged in research on mental health... regulations in this part establish procedures under which any person engaged in research on mental health... 42 Public Health 1 2010-10-01 2010-10-01 false Applicability. 2a.1 Section 2a.1 Public...

  4. Synthesis and pharmacological characterization of a new series of 5,7-disubstituted-[1,2,4]triazolo[1,5-a][1,3,5]triazine derivatives as adenosine receptor antagonists: A preliminary inspection of ligand–receptor recognition process

    PubMed Central

    Pastorin, Giorgia; Federico, Stephanie; Paoletta, Silvia; Corradino, Marta; Cateni, Francesca; Cacciari, Barbara; Klotz, Karl-Norbert; Gao, Zhan-Guo; Jacobson, Kenneth A.; Spalluto, Giampiero; Moro, Stefano

    2011-01-01

    A new series of triazolotriazines variously substituted at the C5 and N7 (5–25) positions was synthesized and fully characterized at the four adenosine receptor (AR) subtypes. In particular, arylacetyl or arylcarbamoyl moieties were introduced at the N7 position, which enhanced affinity at the hA2B and hA3 ARs, respectively, when utilized on the pyrazolo-triazolopyrimidine nucleus as we reported in the past. In general, compounds with a free amino group at the 7 position (5, 6), showed good affinity at the rat (r) A2A AR (range 18.3–96.5 nM), while the introduction of a phenylcarbamoyl moiety at the N7 position (12, 19, 24) slightly increased the affinity at the hA3 AR (range 311–633 nM) with respect to the unsubstituted derivatives. The binding profiles of the synthesized analogues seemed to correlate with the substitutions at the C5 and N7 positions. At the hA2B AR, derivative 5, which contained a free amino group at the 7 position, was the most potent (EC50 3.42 μM) and could represent a starting point for searching new non-xanthine hA2B AR antagonists. Molecular models of the rA2A and hA3 ARs were constructed by homology to the recently reported crystallographic structure of the hA2A AR. A preliminary receptor-driven structure–activity relationship (SAR) based on the analysis of antagonist docking has been provided. PMID:20304654

  5. Up-regulation of striatal adenosine A2A receptors with iron deficiency in rats. Effects on locomotion and cortico-striatal neurotransmission

    PubMed Central

    Quiroz, César; Pearson, Virginia; Gulyani, Seema; Allen, Richard; Earley, Christopher; Ferré, Sergi

    2010-01-01

    Brain iron deficiency leads to altered dopaminergic function in experimental animals, which can provide a mechanistic explanation for iron deficiency-related human sensory-motor disorders, such as Restless Legs Syndrome (RLS). However, mechanisms linking both conditions have not been determined. Considering the strong modulation exerted by adenosine on dopamine signaling, one connection could involve changes in adenosine receptor expression or function. In the striatum, presynaptic A2A receptors are localized in glutamatergic terminals contacting GABAergic dynorphinergic neurons and their function can be analyzed by the ability of A2A receptor antagonists to block the motor output induced by cortical electrical stimulation. Postsynaptic A2A receptors are localized in the dendritic field of GABAergic enkephalinergic neurons and their function can be analyzed by studying the