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Sample records for adenosine receptor ligands

  1. Recent developments in A2B adenosine receptor ligands.

    Science.gov (United States)

    Kalla, Rao V; Zablocki, Jeff; Tabrizi, Mojgan Aghazadeh; Baraldi, Pier Giovanni

    2009-01-01

    A selective, high-affinity A(2B) adenosine receptor (AR) antagonist will be useful as a pharmacological tool to help determine the role of the A(2B)AR in inflammatory diseases and angiogenic diseases. Based on early A(2B)AR-selective ligands with nonoptimal pharmaceutical properties, such as 15 (MRS 1754: K(i)(hA(2B)) = 2 nM; K(i)(hA(1)) = 403 nM; K(i)(hA(2A)) = 503 NM, and K(i)(hA(3)) = 570 nM), several groups have discovered second-generation A(2B)AR ligands that are suitable for development. Scientists at CV Therapeutics have discovered the selective, high-affinity A(2B)AR antagonist 22, a 8-(4-pyrazolyl)-xanthine derivative, (CVT-6883, K(i)(hA(2B)) = 22 nM; K(i)(hA(1)) = 1,940 nM; K(i)(hA(2A)) = 3,280; and K(i)(hA(3)) = 1,070 nM). Compound 22 has demonstrated favorable pharmacokinetic (PK) properties (T(1/2) = 4 h and F > 35% rat), and it is a functional antagonist at the A(2B)AR(K (B) = 6 nM). In a mouse model of asthma, compound 22 demonstrated a dose-dependent efficacy supporting the role of the A(2B)AR in asthma. In two Phase I clinical trails, 22 (CVT-6883) was found to be safe, well tolerated, and suitable for once-daily dosing. Baraldi et al. have independently discovered a selective, high-affinity A(2B)AR antagonist, 30 (MRE2029F20), 8-(5-pyrazolyl)-xanthine (K(i)(hA(2B)) = 5.5 nM; K(i)(hA(1)) = 200 nM; K(i)(hA(2A), A(3)) > 1,000, that has been selected for development in conjunction with King Pharmaceuticals. Compound 30 has been demonstrated to be a functional antagonist of the A(2B)AR, and it has been radiolabeled for use in pharmacological studies. A third compound, 58 (LAS-38096), is a 2-aminopyrimidine derivative (discovered by the Almirall group) that has high A(2B)AR affinity and selectivity (K(i)(hA(2B)) = 17 nM; K(i)(hA(1)) > 1,000 nM; K(i)(hA(2A)) > 2,500; and K(i)(hA(3)) > 1,000 nM), and 58 has been moved into preclinical safety testing. A fourth selective, high-affinity A(2B)AR antagonist, 54 (OSIP339391 K(i))(hA(2B)) = 0.5 nM; K(i))(hA(1

  2. Adenosine and adenosine receptors: Newer therapeutic perspective

    Directory of Open Access Journals (Sweden)

    Manjunath S

    2009-01-01

    Full Text Available Adenosine, a purine nucleoside has been described as a ′retaliatory metabolite′ by virtue of its ability to function in an autocrine manner and to modify the activity of a range of cell types, following its extracellular accumulation during cell stress or injury. These effects are largely protective and are triggered by binding of adenosine to any of the four adenosine receptor subtypes namely A1, A2a, A2b, A3, which have been cloned in humans, and are expressed in most of the organs. Each is encoded by a separate gene and has different functions, although overlapping. For instance, both A1 and A2a receptors play a role in regulating myocardial oxygen consumption and coronary blood flow. It is a proven fact that adenosine plays pivotal role in different physiological functions, such as induction of sleep, neuroprotection and protection against oxidative stress. Until now adenosine was used for certain conditions like paroxysmal supraventricular tachycardia (PSVT and Wolff Parkinson White (WPW syndrome. Now there is a growing evidence that adenosine receptors could be promising therapeutic targets in a wide range of conditions including cardiac, pulmonary, immunological and inflammatory disorders. After more than three decades of research in medicinal chemistry, a number of selective agonists and antagonists of adenosine receptors have been discovered and some have been clinically evaluated, although none has yet received regulatory approval. So this review focuses mainly on the newer potential role of adenosine and its receptors in different clinical conditions.

  3. Adenosine Receptors and Asthma

    OpenAIRE

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

    2009-01-01

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

  4. New Pyrazolo[1',5':1,6]pyrimido[4,5-d]pyridazin-4(3H)-ones Fluoroderivatives as Human A1 Adenosine Receptor Ligands.

    Science.gov (United States)

    Graziano, Alessia; Giovannoni, Maria Paola; Cilibrizzi, Agostino; Crocetti, Letizia; Piaz, Vittorio Dal; Vergelli, Claudia; Trincavelli, Maria Letizia; Martini, Claudia; Giacomelli, Chiara

    2012-09-01

    In this paper we report the synthesis and biological evaluation of a new series of pyrazolo[1',5':1,6]pyrimido[4,5-d]pyridazin-4(3H)-ones as human A1 adenosine receptor ligands. The tricyclic scaffold was modified at position 6 and 9 by introducing small alkyl chains and substituted phenyls. The most interesting compounds showed Ki for A1 in the submicromolar range (0.105-0.244 µM) and the most interesting term (compound 4c) combined an appreciable affinity for A1 (Ki = 0.132 µM) with a good selectivity toward A2A (43% inhibition at 10 µM) and A3 (46% inhibition at 10 µM). PMID:24061322

  5. Metabolism of the A{sub 1} adenosine receptor PET ligand [{sup 18}F]CPFPX by CYP1A2: implications for bolus/infusion PET studies

    Energy Technology Data Exchange (ETDEWEB)

    Matusch, Andreas [Institute of Medicine, Research Center Juelich GmbH, D-52425 Juelich (Germany); Meyer, Philipp T. [Department of Neurology, University Hospital Aachen, D-52074 Aachen (Germany); Bier, Dirk [Institute for Neuroscience and Biophysics (INB4)-Nuclear Chemistry, Research Center Juelich GmbH, D-52425 Juelich (Germany); Holschbach, Marcus H. [Institute for Neuroscience and Biophysics (INB4)-Nuclear Chemistry, Research Center Juelich GmbH, D-52425 Juelich (Germany); Woitalla, Dirk [Neurological Department, Ruhr-University Bochum, D-44791 Bochum (Germany); Elmenhorst, David [Institute of Medicine, Research Center Juelich GmbH, D-52425 Juelich (Germany); Winz, Oliver H. [Institute of Medicine, Research Center Juelich GmbH, D-52425 Juelich (Germany); Zilles, Karl [Institute of Medicine, Research Center Juelich GmbH, D-52425 Juelich (Germany); Bauer, Andreas [Institute of Medicine, Research Center Juelich GmbH, D-52425 Juelich (Germany)]. E-mail: an.bauer@fz-juelich.de

    2006-10-15

    The A{sub 1} adenosine receptor positron emission tomography (PET) ligand 8-cyclopentyl-3-(3-[{sup 18}F]fluoropropyl)-1-propylxanthine ([{sup 18}F]CPFPX, ) undergoes a fast hepatic metabolism. An optimal design of PET quantitation approaches (e.g., bolus/infusion studies) necessitates the knowledge of factors that influence this metabolism. Metabolites of were separated by radio thin-layer chromatography. Metabolism in vivo, in pooled human liver microsomes and in recombinant human cytochrome isoenzyme preparations was studied. Dynamic PET studies using were performed on three controls and two patients, one treated with the antidepressant and inhibitor of cytochrome CYP1A2 fluvoxamine, the other suffering from liver cirrhosis. CPFPX is metabolized by cytochrome CYP1A2 with high selectivity [K {sub M}=1.1 {mu}M (95% confidence interval, or CI, 0.6-2.0 {mu}M) and V {sub max}=243 pmol min{sup -1} mg{sup -1} (95% CI, 112-373 pmol min{sup -1} mg{sup -1}) corresponding to 2.4 pmol min{sup -1} pmol{sup -1} cytochrome P-450]. This metabolism can competitively be inhibited by fluvoxamine with K {sub I}=68 nM (95% CI, 34-138 nM). At least eight compounds found in human plasma and in the CYP1A2 in vitro preparations have an identical migration pattern and account together for >90% and >80% of the respective metabolite yield. Metabolism was considerably delayed in the two patients. In conclusion, is metabolized by cytochrome CYP1A2. Its metabolism is therefore subdued to disease-related or xenobiotic-induced changes of CYP1A2 activity. The identification of the metabolic pathway of 1 allows to optimize image quantification in A{sub 1} adenosine receptor PET studies.

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

    OpenAIRE

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

    2010-01-01

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

  7. N6-(2-Hydroxyethyl)-Adenosine Exhibits Insecticidal Activity against Plutella xylostella via Adenosine Receptors

    Science.gov (United States)

    Fang, Ming; Chai, Yiqiu; Chen, Guanjv; Wang, Huidong; Huang, Bo

    2016-01-01

    The diamondback moth, Plutella xylostella, is one of the most important pests of cruciferous crops. We have earlier shown that N6-(2-hydroxyethyl)-adenosine (HEA) exhibits insecticidal activity against P. xylostella. In the present study we investigated the possible mechanism of insecticidal action of HEA on P. xylostella. HEA is a derivative of adenosine, therefore, we speculated whether it acts via P. xylostella adenosine receptor (PxAdoR). We used RNAi approach to silence PxAdoR gene and used antagonist of denosine receptor (AdoR) to study the insecticidal effect of HEA. We cloned the whole sequence of PxAdoR gene. A BLAST search using NCBI protein database showed a 61% identity with the Drosophila adenosine receptor (DmAdoR) and a 32–35% identity with human AdoR. Though the amino acids sequence of PxAdoR was different compared to other adenosine receptors, most of the amino acids that are known to be important for adenosine receptor ligand binding and signaling were present. However, only 30% binding sites key residues was similar between PxAdoR and A1R. HEA, at a dose of 1 mg/mL, was found to be lethal to the second-instar larvae of P. xylostella, and a significant reduction of mortality and growth inhibition ratio were obtained when HEA was administered to the larvae along with PxAdoR-dsRNA or antagonist of AdoR (SCH58261) for 36, 48, or 60 h. Especially at 48 h, the rate of growth inhibition of the PxAdoR knockdown group was 3.5-fold less than that of the HEA group, and the corrected mortality of SCH58261 group was reduced almost 2-fold compared with the HEA group. Our findings show that HEA may exert its insecticidal activity against P. xylostella larvae via acting on PxAdoR. PMID:27668428

  8. Internalization and desensitization of adenosine receptors.

    NARCIS (Netherlands)

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

    2008-01-01

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

  9. Mast cell adenosine receptors function: a focus on the A3 adenosine receptor and inflammation

    Directory of Open Access Journals (Sweden)

    Noam eRudich

    2012-06-01

    Full Text Available Adenosine is a metabolite, which has long been implicated in a variety of inflammatory processes. Inhaled adenosine provokes bronchoconstriction in asthmatics or chronic obstructive pulmonary disease (COPD patients, but not in non-asthmatics. This hyper responsiveness to adenosine appears to be mediated by mast cell activation. These observations have marked the receptor that mediates the bronchoconstrictor effect of adenosine on mast cells, as an attractive drug candidate. Four subtypes (A1, A2a, A2b and A3 of adenosine receptors have been cloned and shown to display distinct tissue distributions and functions. Animal models have firmly established the ultimate role of the A3 adenosine receptor (A3R in mediating hyper responsiveness to adenosine in mast cells, although the influence of the A2b adenosine receptor was confirmed as well. In contrast, studies of the A3R in humans have been controversial. In this review, we summarize data on the role of different adenosine receptors in mast cell regulation of inflammation and pathology, with a focus on the common and distinct functions of the A3R in rodent and human mast cells. The relevance of mouse studies to the human is discussed.

  10. AMPA receptor ligands

    DEFF Research Database (Denmark)

    Strømgaard, Kristian; Mellor, Ian

    2004-01-01

    polyamines are known to modulate the function of these receptors in vivo. In this study, recent developments in the medicinal chemistry of polyamine-based ligands are given, particularly focusing on the use of solid-phase synthesis (SPS) as a tool for the facile generation of libraries of polyamine toxin...

  11. Ligand-Receptor Interactions

    CERN Document Server

    Bongrand, Pierre

    2008-01-01

    The formation and dissociation of specific noncovalent interactions between a variety of macromolecules play a crucial role in the function of biological systems. During the last few years, three main lines of research led to a dramatic improvement of our understanding of these important phenomena. First, combination of genetic engineering and X ray cristallography made available a simultaneous knowledg of the precise structure and affinity of series or related ligand-receptor systems differing by a few well-defined atoms. Second, improvement of computer power and simulation techniques allowed extended exploration of the interaction of realistic macromolecules. Third, simultaneous development of a variety of techniques based on atomic force microscopy, hydrodynamic flow, biomembrane probes, optical tweezers, magnetic fields or flexible transducers yielded direct experimental information of the behavior of single ligand receptor bonds. At the same time, investigation of well defined cellular models raised the ...

  12. Adenosine modulation of [Ca2+]i in cerebellar granular cells: multiple adenosine receptors involved.

    Science.gov (United States)

    Vacas, Javier; Fernández, Mercedes; Ros, Manuel; Blanco, Pablo

    2003-12-01

    Elimination of adenosine by addition of adenosine deaminase (ADA) to the media leads to alterations in intracellular free calcium concentration ([Ca(2+)](i)) in cerebellar granular cells. Adenosine deaminase brings about increases or decreases in [Ca(2+)](i) depending on the previous activation state of the cell. These effects are dependent on the catalytic activity of adenosine deaminase, since its previous catalytic inactivation with Hg(2+) prevents the above-mentioned changes in intracellular calcium. Extracellular calcium is required for the increase in [Ca(2+)](i) promoted by ADA. This rise is insensitive to thapsigargin, but sensitive to micromolar concentrations of Ni(2+). Toxins specific for L, N and P/Q calcium channels do not overtly reduce this effect. N(6)-Cyclopentyl adenosine (CPA), an A(1) receptor agonist, produces a partial reversion of ADA effects, while CGS21680, A(2A)/A(2B) receptor agonist, slightly enhances them. Expression of A(1), A(2A), A(2B) and A(3) adenosine receptor mRNAs was detected in cerebellar granular cell cultures. These results suggest that adenosine modulate [Ca(2+)](i) in cerebellar granule cells through different adenosine receptor subtypes which, at least in part, seem to act through R-type calcium channels.

  13. 腺苷A1受体新配体YZG-404的镇静催眠作用%Sedative and hypnotic effects of a novel ligand YZG-404for adenosine A1 receptor

    Institute of Scientific and Technical Information of China (English)

    李伟; 王亚芳; 李敏; 岳正刚; 石建功; 张建军

    2011-01-01

    目的 研究新化合物YZG-404与腺苷A1受体(A1R)和腺苷A2A受体(A2AR)的亲和力及其镇静催眠作用.方法 采用放射性配体受体竞争结合实验分别测定YZG-404与腺苷A1R和腺苷A2AR的亲和力;采用开阔场实验测定其对小鼠自发活动的影响:采用协同戊巴比妥钠睡眠实验评价其镇静催眠作用.结果 YZG-404对腺苷A1R亲和力较高,K值为98.8 nmol/L,而对腺苷A2AR的亲和力较低,K值约为9828.8 nmol/L.与溶剂对照组比较,YZG-404(1.25、2,5和5 mg/kg,ig)明显抑制小鼠的自发活动,抑制率分别为26.0%、59.7%和67.1%.另外,YZG-404(1.25、2.5和5 mg/kg,ig)可以明显延长戊巴比妥钠诱导小鼠睡眠时间,延长率分别为49.7%、129.5%和126.0%,并缩短入睡潜伏期,最高缩短率为19.8%.YZG-404能提高阈下剂量戊巴比妥钠诱导小鼠入睡率,最高入睡率达80%,效果与阳性对照药地西泮相当.结论 新化合物YZG-404与腺苷A1R亲和力强,并具有强效的镇静催眠作用.%Objective To examine the affinities of YZG-404 , a novel compound, to adenosine A1 receptor (A1R) and adenosine A2A receptor (A2AR) and its sedative and hypnotic effects. Methods Radioligand binding tests were carried out for the affinity property of YZG-404 to adenosine A1R and adenosine A2AR. The influence of YZG-404 on mice spontaneous locomotor activity was investigated by open field test, and sedative and hypnotic effect of YZG-404 on sodium pentobarbital-treated mice was also evaluated. Results YZG-404 had a higher affinity to adenosine A1R than to adenosine A2AR. The values of Ki to adenosine A1R and A2AR were 98.8 and 9828.8 nmol/L, respectively. The spontaneous locomotor activity was significantly decreased by YZG-404 at test doses (1.25 , 2.5 and 5 mg/kg, ig) , and the decreasing rate was 26.0% , 59.7% and 67.1%, respectively. The duration of sleeping in sodium pentobarbital-treated mice was dose-dependently prolonged by YZG-404, which was 49.7% , 129.5% and

  14. Radiosynthesis of a novel potential adenosine A{sub 3} receptor ligand, 5-ethyl 2,4-diethyl-3-((2-[{sup 18}F]fluoroethyl)sulfanylcarbonyl)-6-phenylpyridine-5-carboxylate ([{sup 18}F]FE rate at SUPPY:2)

    Energy Technology Data Exchange (ETDEWEB)

    Haeusler, D. [Dept. of Nuclear Medicine, Medical Univ. of Vienna (Austria); Dept. of Pharmaceutical Tech. and Biopharmaceutics, Univ. of Vienna (Austria); Mitterhauser, M. [Dept. of Nuclear Medicine, Medical Univ. of Vienna (Austria); Dept. of Pharmaceutical Tech. and Biopharmaceutics, Univ. of Vienna (Austria); Hospital Pharmacy of the General Hospital of Vienna (Austria); Mien, L.K. [Dept. of Nuclear Medicine, Medical Univ. of Vienna (Austria); Dept. of Pharmaceutical Tech. and Biopharmaceutics, Univ. of Vienna (Austria); Dept. of Psychiatry and Psychotherapy, Medical Univ. of Vienna (Austria); Shanab, K.; Spreitzer, H. [Dept. of Drug and Natural Product Synthesis, Univ. of Vienna (Austria); Lanzenberger, R.R [Dept. of Psychiatry and Psychotherapy, Medical Univ. of Vienna (Austria); Schirmer, E. [Dept. of Nuclear Medicine, Medical Univ. of Vienna (Austria); Dept. of Drug and Natural Product Synthesis, Univ. of Vienna (Austria); Ungersboeck, J.; Wadsak, W. [Dept. of Nuclear Medicine, Medical Univ. of Vienna (Austria); Dept. of Inorganic Chemistry, Univ. of Vienna (Austria); Nics, L. [Dept. of Nuclear Medicine, Medical Univ. of Vienna (Austria); Dept. of Nutritional Sciences, Univ. of Vienna (Austria); Viernstein, H. [Dept. of Pharmaceutical Tech. and Biopharmaceutics, Univ. of Vienna (Austria); Dudezak, R.; Kletter, K. [Dept. of Nuclear Medicine, Medical Univ. of Vienna (Austria)

    2009-07-01

    Since, to date very limited information on the distribution and function of the adenosine A{sub 3} receptor is available, the development of suitable radioligands is needed. Recently, we introduced [{sup 18}F]FE rate at SUPPY (5-(2-[{sup 18}F]fluoroethyl) 2,4-diethyl-3-(ethylsulfanylcarbonyl)-6-phenylpyridine-5-carboxylate) as the first PET-ligand for the A3R. Regarding the metabolic profile - this class of dialkylpyridines comprises two ester functions within one molecule, one carboxylic and one thiocarboxylic - one could expect carboxylesterases significantly contributing to cleavage and degradation. Therefore, our aim was the development of [{sup 18}F]FE rate at SUPPY:2 (5-ethyl 2,4-diethyl-3-((2-[{sup 18}F]fluoroethyl)sulfanylcarbonyl)-6-phenylpyridine-5-carboxylate), the functional isomer containing the label at the thiocarboxylic moiety. For satisfactory yields in high scale radiosyntheses, a reaction temperature of 75 C has to be applied for at least 20 min using 20 mg/mL of precursor. So far, 6 complete high-scale radiosyntheses were performed. Starting from an average of 51.2 {+-} 21.8 GBq (mean{+-}SD) [{sup 18}F]fluoride, 5.8 {+-} 4.1 GBq of formulated [{sup 18}F]FE rate at SUPPY:2 (12.0{+-}5.4%, based on [{sup 18}F]fluoride, not corrected for decay) were prepared in 75 {+-} 8 min. (orig.)

  15. Radiosynthesis of a novel potential adenosine A3 receptor ligand, 5-ethyl 2,4-diethyl-3-((2-[18F]fluoroethyl)sulfanylcarbonyl) -6-phenylpyridine-5-carbox ylate ([18F]FE rate at SUPPY:2)

    International Nuclear Information System (INIS)

    Since, to date very limited information on the distribution and function of the adenosine A3 receptor is available, the development of suitable radioligands is needed. Recently, we introduced [18F]FE rate at SUPPY (5-(2-[18F]fluoroethyl) 2,4-diethyl-3-(ethylsulfanylcarbonyl)-6-phenylpyridine-5-carboxylate) as the first PET-ligand for the A3R. Regarding the metabolic profile - this class of dialkylpyridines comprises two ester functions within one molecule, one carboxylic and one thiocarboxylic - one could expect carboxylesterases significantly contributing to cleavage and degradation. Therefore, our aim was the development of [18F]FE rate at SUPPY:2 (5-ethyl 2,4-diethyl-3-((2-[18F]fluoroethyl)sulfanylcarbonyl)-6-phenylpyridine -5-carbox ylate), the functional isomer containing the label at the thiocarboxylic moiety. For satisfactory yields in high scale radiosyntheses, a reaction temperature of 75 C has to be applied for at least 20 min using 20 mg/mL of precursor. So far, 6 complete high-scale radiosyntheses were performed. Starting from an average of 51.2 ± 21.8 GBq (mean±SD) [18F]fluoride, 5.8 ± 4.1 GBq of formulated [18F]FE rate at SUPPY:2 (12.0±5.4%, based on [18F]fluoride, not corrected for decay) were prepared in 75 ± 8 min. (orig.)

  16. Determination of adenosine effects and adenosine receptors in murine corpus cavernosum.

    Science.gov (United States)

    Tostes, Rita C; Giachini, Fernanda R C; Carneiro, Fernando S; Leite, Romulo; Inscho, Edward W; Webb, R Clinton

    2007-08-01

    This study tested the hypothesis that adenosine, in murine corpora cavernosa, produces direct relaxation of smooth muscle cells and inhibition of contractile responses mediated by sympathetic nerve stimulation. Penes were excised from anesthetized male C57BL/6 mice, dissected, and cavernosal strips were mounted to record isometric force. Adenosine, 2-chloroadenosine (stable analog of adenosine), and 2-phenylaminoadenosine (CV1808) (A2(A)/A2(B) agonist) produced concentration-dependent relaxations of phenylephrine-contracted tissues. Relaxation to 2-chloroadenosine was inhibited, in a concentration-dependent manner, by 2-(2-furanyl)-7-(2-phenylethyl)-7H-pyrazolo[4,3-e][1,2,4]triazolo[1,5-c]pyrimidin-5-amine (SCH58261; A2(A) antagonist; 10(-9)-10(-6) M) and N-(4-acetylphenyl)-2-[4-(2,3,6,7-tetrahydro-2,6-dioxo-1,3-dipropyl-1H-purin-8-yl)phenoxy]acetamida (MRS1706; A2(B) antagonist; 10(-8)-10(-6) M). The combination of both antagonists abrogated 2-chloroadenosine-induced relaxation. Electrical field stimulation (EFS; 1-32 Hz) of adrenergic nerves produced frequency-dependent contractions that were inhibited by compounds that increase adenosine levels, such as 5'-iodotubercidin (adenosine kinase inhibitor), erythro-9-(2-hydroxy-3-nonyl)adenine (adenosine deaminase inhibitor), and dipyridamole (inhibitor of adenosine transport). The adenosine A1 receptor agonist N(6)-cyclopentyladenosine (C8031) right-shifted contractile responses to EFS, with a significant inhibitory effect at 10(-6) M. Blockade of adenosine A1 receptors with 8-cyclopentyl-1,3-dipropylxanthine (C101) (10(-7) M) enhanced contractile responses to EFS and eliminated the inhibitory effects of 5'-iodotubercidin. Dipyridamole and 5'-iodotubercidin had no effect on adenosine-mediated relaxation. In summary, adenosine directly relaxes cavernosal smooth muscle cells, by the activation of A2(A)/A2(B) receptor subtypes. In addition, adenosine negatively modulates sympathetic neurotransmission, by A1 receptor

  17. Distribution of adenosine receptors in human sclera fibroblasts

    OpenAIRE

    Cui, Dongmei; Trier, Klaus; Chen, Xiang; Zeng, Junwen; Yang, Xiao; Hu, Jianmin; Ge, Jian

    2008-01-01

    Purpose Systemic treatment with adenosine receptor antagonists has been reported to affect the biochemistry and ultrastructure of rabbit sclera. This study was conducted to determine whether adenosine receptors (ADORs) are present in human scleral fibroblasts (HSF). Methods Primary HSF were cultured in vitro and identified with anti-vimentin, anti-keratin, anti-desmin, and anti-S-100 antibodies. Confocal fluorescence microscopy was used to study the distribution of ADORs in the HSF cell lines...

  18. Electroacupuncture improves neuropathic pain Adenosine,adenosine 5'-triphosphate disodium and their receptors perhaps change simultaneously

    Institute of Scientific and Technical Information of China (English)

    Wen Ren; Wenzhan Tu; Songhe Jiang; Ruidong Cheng; Yaping Du

    2012-01-01

    Applying a stimulating current to acupoints through acupuncture needles-known as electroacupuncture-has the potential to produce analgesic effects in human subjects and experimental animals.When acupuncture was applied in a rat model,adenosine 5'-triphosphate disodium in the extracellular space was broken down into adenosine,which in turn inhibited pain transmission by means of an adenosine A1 receptor-dependent process.Direct injection of an adenosine A1 receptor agonist enhanced the analgesic effect of acupuncture.The analgesic effect of acupuncture appears to be mediated by activation of A1 receptors located on ascending nerves.In neuropathic pain,there is upregulation of P2X purinoceptor 3(P2X3)receptor expression in dorsal root ganglion neurons.Conversely,the onset of mechanical hyperalgesia was diminished and established hyperalgesia was significantly reversed when P2X3 receptor expression was downregulated.The pathways upon which electroacupuncture appear to act are interwoven with pain pathways,and electroacupuncture stimuli converge with impulses originating from painful areas.Electroacupuncture may act via purinergic A1 and P2X3 receptors simultaneously to induce an analgesic effect on neuropathic pain.

  19. Role of A3 adenosine receptor in diabetic neuropathy.

    Science.gov (United States)

    Yan, Heng; Zhang, Enshui; Feng, Chang; Zhao, Xin

    2016-10-01

    Neuropathy is the most common diabetic complication. Although the A1 and A2A adenosine receptors are important pharmacological targets in alleviating diabetic neuropathy, the role of the A3 adenosine receptor remains unknown. Because the A3 adenosine receptor regulates pain induced by chronic constriction injury or chemotherapy, its stimulation might also attenuate diabetic neuropathy. This study examines the effects of systemic treatment with the A3 adenosine receptor agonist 1-deoxy-1-[6-[[(3-iodophenyl)methyl]amino]-9H-purin-9-yl]-N-methyl-β-d-ribofuranuronamide (IB-MECA) on diabetic neuropathy and explores the putative mechanisms underlying its pharmacological effects. We show that IB-MECA alleviated mechanical hyperalgesia and thermal hypoalgesia in mice 2 weeks but not 4 weeks after streptozocin (STZ) treatment. Furthermore, IB-MECA prevented the reduction in sciatic motor nerve conduction velocity and sensory nerve conduction velocity in diabetic mice 2 weeks but not 4 weeks after STZ treatment. Similarly, IB-MECA inhibited the activation of nuclear factor-κB and decreased the generation of tumor necrosis factor-α in the spinal cord of mice 2 weeks but not 4 weeks after STZ treatment. These phenomena were associated with reduction of A3 adenosine receptor expression in the spinal cord after long-term diabetes. Our results suggest that the A3 adenosine receptor plays a critical role in regulating diabetic neuropathy and that reduction in A3 adenosine receptor expression/function might contribute to the progression of diabetic neuropathy. © 2016 Wiley Periodicals, Inc.

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

    Institute of Scientific and Technical Information of China (English)

    V Haktan Ozacmak; Hale Sayan

    2007-01-01

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

  1. Adenosine A1 receptor agonists inhibit trigeminovascular nociceptive transmission

    DEFF Research Database (Denmark)

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

    2002-01-01

    There is a considerable literature to suggest that adenosine A1 receptor agonists may have anti-nociceptive effects, and we sought to explore the role of adenosine A1 receptors in a model of trigeminovascular nociceptive transmission. Cats were anaesthetized (alpha-chloralose 60 mg/kg, intraperit......There is a considerable literature to suggest that adenosine A1 receptor agonists may have anti-nociceptive effects, and we sought to explore the role of adenosine A1 receptors in a model of trigeminovascular nociceptive transmission. Cats were anaesthetized (alpha-chloralose 60 mg...... from the external jugular vein to determine levels of calcitonin gene-related peptide (CGRP) release before and after drug administration. Intravenous administration of the highly selective adenosine A1 receptor agonist, GR79236 (3-100 microg/kg) had a dose-dependent inhibitory effect on SSS...... 33 +/- 2 pmol/l (n = 6) to 64 +/- 3 pmol/l, an effect substantially reduced by pre-treatment with GR79236 (30 microg/kg; P agonist, GR190178 (30-1000 microg/kg i.v.), also inhibited SSS-evoked neuronal activity in a dose-dependent fashion...

  2. The role of adenosine receptors and endogenous adenosine in citalopram-induced cardiovascular toxicity

    OpenAIRE

    Kubilay Oransay; Nil Hocaoglu; Mujgan Buyukdeligoz; Yesim Tuncok; Sule Kalkan

    2014-01-01

    Aim: We investigated the role of adenosine in citalopram-induced cardiotoxicity. Materials and Methods: Protocol 1: Rats were randomized into four groups. Sodium cromoglycate was administered to rats. Citalopram was infused after the 5% dextrose, 8-Cyclopentyl-1,3-dipropylxanthine (DPCPX; A 1 receptor antagonist), 8-(-3-chlorostyryl)-caffeine (CSC; A 2a receptor antagonist), or dimethyl sulfoxide (DMSO) administrations. Protocol 2: First group received 5% dextrose intraperitoneally 1 hour...

  3. Adenosine receptors and asthma in humans

    OpenAIRE

    Wilson, C N

    2008-01-01

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

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

    Science.gov (United States)

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

    2013-02-01

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

  5. DMPD: Shaping of monocyte and macrophage function by adenosine receptors. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 17056121 Shaping of monocyte and macrophage function by adenosine receptors. Hasko ...tml) (.csml) Show Shaping of monocyte and macrophage function by adenosine receptors. PubmedID 17056121 Titl...e Shaping of monocyte and macrophage function by adenosine receptors. Authors Has

  6. Stabilizing effects of G protein on the active conformation of adenosine A1 receptor differ depending on G protein type.

    Science.gov (United States)

    Tateyama, Michihiro; Kubo, Yoshihiro

    2016-10-01

    G protein coupled receptors (GPCRs) trigger various cellular and physiological responses upon the ligand binding. The ligand binding induces conformational change in GPCRs which allows G protein to interact with the receptor. The interaction of G protein also affects the active conformation of GPCRs. In this study, we have investigated the effects of Gαi1, Gαo and chimeric Gαqi5 on the active conformation of the adenosine A1 receptor, as each Gα showed difference in the interaction with adenosine A1 receptor. The conformational changes in the adenosine A1 receptor were detected as the agonist-induced decreases in efficiency of Förster resonance energy transfer (FRET) between fluorescent proteins (FPs) fused at the two intracellular domains of the adenosine A1 receptor. Amplitudes of the agonist-induced FRET decreases were subtle when the FP-tagged adenosine A1 receptor was expressed alone, whereas they were significantly enhanced when co-expressed with Gαi1Gβ1Gγ22 (Gi1) or Gαqi5Gβ1Gγ22 (Gqi5) but not with GαοGβ1Gγ22 (Go). The enhancement of the agonist-induced FRET decrease in the presence of Gqi5 was significantly larger than that of Gi1. Furthermore, the FRET recovery upon the agonist removal in the presence of Gqi5 was significantly slower than that of Gi1. From these results it was revealed that the agonist-bound active conformation of adenosine A1 receptor is unstable without the binding of G protein and that the stabilizing effects of G protein differ depending on the types of G protein.

  7. Radioiodinated ligands for dopamine receptors

    International Nuclear Information System (INIS)

    The dopamine receptor system is important for normal brain function; it is also the apparent action site for various neuroleptic drugs for the treatment of schizophrenia and other metal disorders. In the past few years radioiodinated ligands for single photon emission tomography (SPECT) have been successfully developed and tested in humans: [123I]TISCH for D1 dopamine receptors; [123I]IBZM, epidepride, IBF and FIDA2, four iodobenzamide derivatives, for D2/D3 dopamine receptors. In addition, [123I]β-CIT (RTI-55) and IPT, cocaine derivatives, for the dopamine reuptake site are potentially useful for diagnosis of loss of dopamine neurons. The first iodinated ligand, (R)trans-7-OH-PIPAT, for D3 dopamine receptors, was synthesized and characterized with cloned cell lines (Spodoptera frugiperda, Sf9) expressing the D2 and D3 dopamine receptors and with rat basal forebrain membrane preparations. Most of the known iodobenzamides displayed similar potency in binding to both D2 and D3 dopamine receptors expressed in the cell lines. Initial studies appear to suggest that by fine tuning the structures it may be possible to develop agents specific for D2 and D3 dopamine receptors. It is important to investigate D2/D3 selectivity for this series of potent ligands

  8. Polypharmacology of dopamine receptor ligands.

    Science.gov (United States)

    Butini, S; Nikolic, K; Kassel, S; Brückmann, H; Filipic, S; Agbaba, D; Gemma, S; Brogi, S; Brindisi, M; Campiani, G; Stark, H

    2016-07-01

    Most neurological diseases have a multifactorial nature and the number of molecular mechanisms discovered as underpinning these diseases is continuously evolving. The old concept of developing selective agents for a single target does not fit with the medical need of most neurological diseases. The development of designed multiple ligands holds great promises and appears as the next step in drug development for the treatment of these multifactorial diseases. Dopamine and its five receptor subtypes are intimately involved in numerous neurological disorders. Dopamine receptor ligands display a high degree of cross interactions with many other targets including G-protein coupled receptors, transporters, enzymes and ion channels. For brain disorders like Parkinsońs disease, schizophrenia and depression the dopaminergic system, being intertwined with many other signaling systems, plays a key role in pathogenesis and therapy. The concept of designed multiple ligands and polypharmacology, which perfectly meets the therapeutic needs for these brain disorders, is herein discussed as a general ligand-based concept while focusing on dopaminergic agents and receptor subtypes in particular. PMID:27234980

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

    OpenAIRE

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

    2012-01-01

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

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

    OpenAIRE

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

    2012-01-01

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

  11. Structure-kinetics relationships of Capadenoson derivatives as adenosine A1 receptor agonists.

    Science.gov (United States)

    Louvel, Julien; Guo, Dong; Soethoudt, Marjolein; Mocking, Tamara A M; Lenselink, Eelke B; Mulder-Krieger, Thea; Heitman, Laura H; IJzerman, Adriaan P

    2015-08-28

    We report the synthesis and biological evaluation of new derivatives of Capadenoson, a former drug candidate that was previously advanced to phase IIa clinical trials. 19 of the 20 ligands show an affinity below 100 nM at the human adenosine A1 receptor (hA1AR) and display a wide range of residence times at this target (from approx. 5 min (compound 10) up to 132 min (compound 5)). Structure-affinity and structure-kinetics relationships were established, and computational studies of a homology model of the hA1AR revealed crucial interactions for both the affinity and dissociation kinetics of this family of ligands. These results were also combined with global metrics (Ligand Efficiency, cLogP), showing the importance of binding kinetics as an additional way to better select a drug candidate amongst seemingly similar leads.

  12. Adenosine receptors and stress : Studies using methylmercury, caffeine and hypoxia

    OpenAIRE

    Björklund, Olga

    2008-01-01

    Brain development is a precisely organized process that can be disturbed by various stress factors present in the diet (e.g. exposure to xenobiotics) as well as insults such as decreased oxygen supply. The consequent adverse changes in nervous system function may not necessarily be apparent until a critical age when neurodevelopmental defects may be unmasked by a subsequent challenge. Adenosine and its receptors (AR) (A1, A2A, A2B and A3) which participate in the brain stres...

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

    Science.gov (United States)

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

    2008-01-01

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

  14. Pharmacology of the Adenosine A3 Receptor in the Vasculature and Essential Hypertension

    Science.gov (United States)

    Ho, Ming-Fen; Low, Leanne M.; Rose’Meyer, Roselyn B.

    2016-01-01

    Background Essential hypertension is considered to be a multifactorial disorder and its aetiology has yet to be clearly identified. As the adenosine receptors have a significant role in mediating vasodilation, alterations in their structures or signalling pathways may be involved in the development of hypertension. This study aimed to measure the expression of adenosine A3 receptors in a range of cardiovascular tissues and determine whether they could be altered with essential hypertension, and to functionally test responses to adenosine A3 receptor agonists in coronary blood vessels using the isolated perfused heart preparation. Methods mRNA samples from cardiovascular tissues and a range of blood vessels were collected from 10 week old male spontaneously hypertensive rats and age-gender matched Wistar rats (n = 8). The Langendorff heart perfusion preparation was used to characterise adenosine A3 receptor mediated coronary vasodilation in the rat heart. Results Adenosine A3 receptor agonists induced coronary vasodilation. The expression of adenosine A3 receptors in cardiovascular tissues was altered in a tissue-specific pattern. Specifically, down-regulation of adenosine A3 receptor expression occurred in hypertensive hearts, which might be associated with attenuated vasodilator responses observed in coronary vessels to adenosine A3 receptor agonists. Conclusions This study demonstrated alterations in the expression of adenosine A3 receptors occurred in a tissue specific mode, and reduced adenosine A3 receptor mediated coronary vasodilation in hearts from spontaneously hypertensive rats. Our findings with regard to changes in the adenosine A3 receptor in hypertensive hearts suggest that adenosine A3 receptor might play a role in the physiopathology of essential hypertension and potentially open the way to pharmacologic manipulation of vasomotor activity by the use of adenosine A3 receptor agonists. PMID:26907173

  15. Radiosynthesis of the adenosine A3 receptor ligand 5-(2-[18F]fluoroethyl) 2,4-diethyl-3-(ethylsulfanylcarbonyl)-6-phenylpyridine-5-carboxylate ([18F]FE rate at SUPPY)

    International Nuclear Information System (INIS)

    Since to date very limited information on the distribution and function of the adenosine A3 receptor is available, the development of a suitable radioligand is needed. Such a selective radioligand can then be used for quantitative autoradiography, preclinical studies in animals and subsequent human PET applications. Recently, a promising candidate compound, 5-(2-fluoroethyl) 2,4-diethyl-3-(ethylsulfanylcarbonyl)-6-phenylpyridine-5-carboxylate (FE rate at SUPPY), has been presented. The successful preparation of a suitable labelling precursor and the evaluation and optimization of the radiosynthesis of [18F]FE rate at SUPPY is presented herewith. For satisfactory yields, a reaction temperature of 75 C has to be applied for at least 20 min using 8-10 mg of precursor. Until now, 15 complete high-scale radiosyntheses were performed. Starting from an average of 51 ± 12 GBq (average ±SD; range: 30-67 GBq) [18F]fluoride, 9.4 ± 3.6 GBq of formulated [18F]FE rate at SUPPY (32.3 ± 12.4%, based on [18F]fluoride, corrected for decay) were prepared in < 105 min. (orig.)

  16. Radiosynthesis of the adenosine A{sub 3} receptor ligand 5-(2-[{sup 18}F]fluoroethyl) 2,4-diethyl-3-(ethylsulfanylcarbonyl)-6-phenylpyridine-5-carboxylate ([{sup 18}F]FE rate at SUPPY)

    Energy Technology Data Exchange (ETDEWEB)

    Wadsak, W. [Dept. of Nuclear Medicine, Medical Univ. of Vienna (Austria); Dept. of Inorganic Chemistry, Univ. of Vienna (Austria); Mien, L.K. [Dept. of Nuclear Medicine, Medical Univ. of Vienna (Austria); Dept. of Pharmaceutical Technology and Biopharmaceutics, Univ. of Vienna (Austria); Dept. of Psychiatry and Psychotherapy, Medical Univ. of Vienna (Austria); Shanab, K.; Spreitzer, H. [Dept. of Psychiatry and Psychotherapy, Medical Univ. of Vienna (Austria); Weber, K.; Schmidt, B.; Haeusler, D. [Dept. of Nuclear Medicine, Medical Univ. of Vienna (Austria); Dept. of Pharmaceutical Technology and Biopharmaceutics, Univ. of Vienna (Austria); Sindelar, K.M.; Ettlinger, D.E.; Dudczak, R.; Kletter, K. [Dept. of Nuclear Medicine, Medical Univ. of Vienna (Austria); Keppler, B.K.; Viernstein, H. [Dept. of Inorganic Chemistry, Univ. of Vienna (Austria); Mitterhauser, M. [Dept. of Nuclear Medicine, Medical Univ. of Vienna (Austria); Dept. of Pharmaceutical Technology and Biopharmaceutics, Univ. of Vienna (Austria); Hospital Pharmacy of the General Hospital of Vienna (Austria)

    2008-07-01

    Since to date very limited information on the distribution and function of the adenosine A{sub 3} receptor is available, the development of a suitable radioligand is needed. Such a selective radioligand can then be used for quantitative autoradiography, preclinical studies in animals and subsequent human PET applications. Recently, a promising candidate compound, 5-(2-fluoroethyl) 2,4-diethyl-3-(ethylsulfanylcarbonyl)-6-phenylpyridine-5-carboxylate (FE rate at SUPPY), has been presented. The successful preparation of a suitable labelling precursor and the evaluation and optimization of the radiosynthesis of [{sup 18}F]FE rate at SUPPY is presented herewith. For satisfactory yields, a reaction temperature of 75 C has to be applied for at least 20 min using 8-10 mg of precursor. Until now, 15 complete high-scale radiosyntheses were performed. Starting from an average of 51 {+-} 12 GBq (average {+-}SD; range: 30-67 GBq) [{sup 18}F]fluoride, 9.4 {+-} 3.6 GBq of formulated [{sup 18}F]FE rate at SUPPY (32.3 {+-} 12.4%, based on [{sup 18}F]fluoride, corrected for decay) were prepared in < 105 min. (orig.)

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

    Science.gov (United States)

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

    2016-05-12

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

  18. Distribution of adenosine receptors in human sclera fibroblasts

    Science.gov (United States)

    Cui, Dongmei; Trier, Klaus; Chen, Xiang; Zeng, Junwen; Yang, Xiao; Hu, Jianmin

    2008-01-01

    Purpose Systemic treatment with adenosine receptor antagonists has been reported to affect the biochemistry and ultrastructure of rabbit sclera. This study was conducted to determine whether adenosine receptors (ADORs) are present in human scleral fibroblasts (HSF). Methods Primary HSF were cultured in vitro and identified with anti-vimentin, anti-keratin, anti-desmin, and anti-S-100 antibodies. Confocal fluorescence microscopy was used to study the distribution of ADORs in the HSF cell lines and in the frozen human scleral sections. ADOR protein expression in HSF and human sclera was confirmed by western blot analysis of cell lysates. Results ADORs were expressed in both HSF and human sclera. This was confirmed by western blot. ADORA1 expression was concentrated in the nucleus. ADORA2A was concentrated mainly in one side of the cytoplasm, and ADORA2B was found both in the nucleus and the cytoplasm. ADORA3 was expressed weakly in the cytoplasm. Conclusions All four subtypes of ADOR were found in HSF and may play a role in scleral remodeling. PMID:18385786

  19. Genetically Controlled Upregulation of Adenosine A(1) Receptor Expression Enhances the Survival of Primary Cortical Neurons

    NARCIS (Netherlands)

    Serchov, Tsvetan; Atas, Hasan-Cem; Normann, Claus; van Calker, Dietrich; Biber, Knut

    2012-01-01

    Adenosine has a key endogenous neuroprotective role in the brain, predominantly mediated by the adenosine A(1) receptor (A(1)R). This has been mainly explored using pharmacological tools and/or receptor knockout mice strains. It has long been suggested that the neuroprotective effects of A(1)R are i

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

    OpenAIRE

    Pousinha, Paula Isabel Antunes, 1978-

    2012-01-01

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

  1. Modulating effect of adenosine deaminase on function of adenosine A1receptors

    Institute of Scientific and Technical Information of China (English)

    Wan-chun SUN; Yan CAO; Lei JIN; Li-zhen WANG; Fan MENG; Xing-zu ZHU

    2005-01-01

    Aim: To study the modulating effect of adenosine deaminase (ADA) on yhe adenosine A1 receptor (A1R) in HEK293 cells stably expressing the human A1R.Methods: cDNA was amplified by RT-PCR using total RNA from human embryo brain tissue as the template. The PCR products were subcloned into the plasmid pcDNA3 and cloned into the plasmid pcDNA3.1. The cloned A1R cDNA was sequenced and stably expressed in HEK293 cells. The modulating effect of adenosine deaminase on A1R was studied by using [3H]DPCPX binding assay and an intracellular calcium assay. Results: HEK293 cells stably expressing human A1R were obtained. Saturation studies showed that the KD value and Bmax value of [3H]DPCPX were 1.6±0.2 nmol/L and 1.819±0.215 nmol/g of protein respectively, in the absence of ecto-ADA respectively, and 1.3±0.2 nmol/L and 1.992±0.130 nmol/g of protein in the presence of ecto-ADA respectively, suggesting that the KD value and Bmax value of [3H]DPCPX were unaffected by ecto-ADA. In the case of [3H]DPCPX competition curves obtained from intact cells or membranes, A1R agonist CCPA/[3H]DPCPX competition curve could be fitted well to a one-site model in the absence of ecto-ADA and a two-site model in the presence of ecto ADA with a KH value of 0.74 (0.11-4.8) nmol/L (intact cells) or 1.8 (0.25-10) nmol/L (membrane) and a KL value of 0.94 (0.62-1.41) μmol/L (intact cells) or 0.77 (0.29-0.99) μmol/L (membrane). The KL value is not significantly different from the IC50 value of 0.84(0.57-1.23) μmol/L (intact cells) or 0.84 (0.63-1.12) μmol/L (membrane) obtained in the absence of ecto-ADA. Similar results were obtained from the CPA/[3H]DPCPX competition curve in the absence or presence of ecto-ADA on intact cells or membranes. Intracellular calcium assay demonstrated that the EC50 value of CPA were 10 (5-29) nmol/L and 94 (38-229) nmol/L in the presence or absence of ecto-ADA, respectively. Conclusion: A1R stably expressed in the HEK293 cells display a low affinity for agonists in

  2. Cloning, expression and pharmacological characterization of rabbit adenosine A1 and A3 receptors.

    Science.gov (United States)

    Hill, R J; Oleynek, J J; Hoth, C F; Kiron, M A; Weng, W; Wester, R T; Tracey, W R; Knight, D R; Buchholz, R A; Kennedy, S P

    1997-01-01

    The role of adenosine A1 and A3 receptors in mediating cardioprotection has been studied predominantly in rabbits, yet the pharmacological characteristics of rabbit adenosine A1 and A3 receptor subtypes are unknown. Thus, the rabbit adenosine A3 receptor was cloned and expressed, and its pharmacology was compared with that of cloned adenosine A1 receptors. Stable transfection of rabbit A1 or A3 cDNAs in Chinese hamster ovary-K1 cells resulted in high levels of expression of each of the receptors, as demonstrated by high-affinity binding of the A1/A3 adenosine receptor agonist N6-(4-amino-3-[125I]iodobenzyl)adenosine (125I-ABA). For both receptors, binding of 125I-ABA was inhibited by the GTP analog 5'-guanylimidodiphosphate, and forskolin-stimulated cyclic AMP accumulation was inhibited by the adenosine receptor agonist (R)-phenylisopropyladenosine. The rank orders of potency of adenosine receptor agonists for inhibition of 125I-ABA binding were as follows: rabbit A1, N6-cyclopentyladenosine = (R)-phenylisopropyladenosine > N-ethylcarboxamidoadenosine > or = I-ABA > or = N6-2-(4-aminophenyl) ethyladenosine > > N6-(3-iodobenzyl)adenosine-5'-N-methyluronamide > N6-(4-amino-3-benzyl)adenosine; rabbit A3, N6-(3-iodobenzyl)adenosine-5'-N-methyluronamide > or = I-ABA > > N-ethylcarboxamidoadenosine > N6-2-(4-aminophenyl) ethyladenosine = N6-cyclopentyladenosine = (R)-phenylisopropyladenosine > N6-(4-amino-3-benzyl)adenosine. The adenosine receptor antagonist rank orders were as follow: rabbit A1, 8-cyclopentyl-1,3-dipropylxanthine > 1,3- dipropyl-8-(4-acrylate)phenylxanthine > or = xanthine amine congener > > 8-(p-sulfophenyl)theophylline; rabbit A3, xanthine amine congener > 1,3-dipropyl-8-(4-acrylate)phenylxanthine > or = 8-cyclopentyl-1,3-dipropylxanthine > > 8-(p-sulfophenyl)theophylline. These observations confirm the identity of the expressed proteins as A1 and A3 receptors. The results will facilitate further in-depth studies of the roles of A1 and A3 receptors in

  3. A3 Adenosine Receptor Allosteric Modulator Induces an Anti-Inflammatory Effect: In Vivo Studies and Molecular Mechanism of Action

    Directory of Open Access Journals (Sweden)

    Shira Cohen

    2014-01-01

    Full Text Available The A3 adenosine receptor (A3AR is overexpressed in inflammatory cells and in the peripheral blood mononuclear cells of individuals with inflammatory conditions. Agonists to the A3AR are known to induce specific anti-inflammatory effects upon chronic treatment. LUF6000 is an allosteric compound known to modulate the A3AR and render the endogenous ligand adenosine to bind to the receptor with higher affinity. The advantage of allosteric modulators is their capability to target specifically areas where adenosine levels are increased such as inflammatory and tumor sites, whereas normal body cells and tissues are refractory to the allosteric modulators due to low adenosine levels. LUF6000 administration induced anti-inflammatory effect in 3 experimental animal models of rat adjuvant induced arthritis, monoiodoacetate induced osteoarthritis, and concanavalin A induced liver inflammation in mice. The molecular mechanism of action points to deregulation of signaling proteins including PI3K, IKK, IκB, Jak-2, and STAT-1, resulting in decreased levels of NF-κB, known to mediate inflammatory effects. Moreover, LUF6000 induced a slight stimulatory effect on the number of normal white blood cells and neutrophils. The anti-inflammatory effect of LUF6000, mechanism of action, and the differential effects on inflammatory and normal cells position this allosteric modulator as an attractive and unique drug candidate.

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

    Science.gov (United States)

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

    2016-06-01

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

  5. Activation of A(2) adenosine receptors dilates cortical efferent arterioles in mouse

    DEFF Research Database (Denmark)

    Al-Mashhadi, Rozh H; Skøtt, Ole; Vanhoutte, Paul M;

    2009-01-01

    that the adenosine-induced vasodilatation was inhibited by the A(2)-specific receptor blocker 3,7-dimethyl-1-propargylxanthine. In the presence of this inhibitor, adenosine failed to alter the basal vessel diameter of quiescent efferent arterioles. Using primer-specific polymerase chain reaction we found...

  6. Adenosine receptors in COPD and asymptomatic smokers : effects of smoking cessation

    NARCIS (Netherlands)

    Versluis, Mieke; ten Hacken, Nick; Postma, Dirkje; Barroso, Begona; Rutgers, Bea; Geerlings, Marie; Willemse, Brigitte; Timens, Wim; Hylkema, Machteld

    2009-01-01

    Our group has shown that 1-year smoking cessation persisted or increased airway inflammation in chronic obstructive pulmonary disease (COPD). We compared adenosine and adenosine receptor (AR) expression in COPD and asymptomatic smokers (AS) before and after 1-year smoking cessation. Sputum cytospins

  7. Adenosine, type 1 receptors: role in proximal tubule Na+ reabsorption.

    Science.gov (United States)

    Welch, W J

    2015-01-01

    Adenosine type 1 receptor (A1 -AR) antagonists induce diuresis and natriuresis in experimental animals and humans. Much of this effect is due to inhibition of A1 -ARs in the proximal tubule, which is responsible for 60-70% of the reabsorption of filtered Na(+) and fluid. Intratubular application of receptor antagonists indicates that A1 -AR mediates a portion of Na(+) uptake in PT and PT cells, via multiple transport systems, including Na(+) /H(+) exchanger-3 (NHE3), Na(+) /PO4(-) co-transporter and Na(+) -dependent glucose transporter, SGLT. Renal microperfusion and recollection studies have shown that fluid reabsorption is reduced by A1 -AR antagonists and is lower in A1 -AR KO mice, compared to WT mice. Absolute proximal reabsorption (APR) measured by free-flow micropuncture is equivocal, with studies that show either lower APR or similar APR in A1 -AR KO mice, compared to WT mice. Inhibition of A1 -ARs lowers elevated blood pressure in models of salt-sensitive hypertension, partially due to their effects in the proximal tubule. PMID:25345761

  8. Adenosine A2A receptors and A2A receptor heteromers as key players in striatal function

    Directory of Open Access Journals (Sweden)

    Sergi eFerre

    2011-06-01

    Full Text Available A very significant density of adenosine 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 striato-pallidal 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 striato-pallidal 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.

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

    Science.gov (United States)

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

    2014-09-15

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

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

    Science.gov (United States)

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

    2014-09-15

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

  11. Influence of the adenosine A1 receptor on blood pressure regulation and renin release

    DEFF Research Database (Denmark)

    Brown, Russell D.; Thorén, Peter; Steege, Andreas;

    2006-01-01

    The present study was performed to investigate the role of adenosine A1 receptors in regulating blood pressure in conscious mice. Adenosine A1-receptor knockout (A1R-/-) mice and their wild-type (A1R+/+) littermates were placed on standardized normal-salt (NS), high-salt (HS), or salt-deficient (SD...... in sodium excretion between the two genotypes on the HS diet. Even on the SD diet, A1R-/- mice had an increased sodium excretion compared with A1R+/+ mice. An abolished tubuloglomerular feedback response and reduced tubular reabsorption can account for the elevated salt excretion found in A1R-/- animals....... The elevated plasma renin concentrations found in the A1R-/- mice could also result in increased blood pressure. Our results confirm that adenosine, acting through the adenosine A1 receptor, plays an important role in regulating blood pressure, renin release, and sodium excretion....

  12. β-Nicotinamide adenine dinucleotide acts at prejunctional adenosine A1 receptors to suppress inhibitory musculomotor neurotransmission in guinea pig colon and human jejunum.

    Science.gov (United States)

    Wang, Guo-Du; Wang, Xi-Yu; Liu, Sumei; Xia, Yun; Zou, Fei; Qu, Meihua; Needleman, Bradley J; Mikami, Dean J; Wood, Jackie D

    2015-06-01

    Intracellular microelectrodes were used to record neurogenic inhibitory junction potentials in the intestinal circular muscle coat. Electrical field stimulation was used to stimulate intramural neurons and evoke contraction of the smooth musculature. Exposure to β-nicotinamide adenine dinucleotide (β-NAD) did not alter smooth muscle membrane potential in guinea pig colon or human jejunum. ATP, ADP, β-NAD, and adenosine, as well as the purinergic P2Y1 receptor antagonists MRS 2179 and MRS 2500 and the adenosine A1 receptor agonist 2-chloro-N6-cyclopentyladenosine, each suppressed inhibitory junction potentials in guinea pig and human preparations. β-NAD suppressed contractile force of twitch-like contractions evoked by electrical field stimulation in guinea pig and human preparations. P2Y1 receptor antagonists did not reverse this action. Stimulation of adenosine A1 receptors with 2-chloro-N6-cyclopentyladenosine suppressed the force of twitch contractions evoked by electrical field stimulation in like manner to the action of β-NAD. Blockade of adenosine A1 receptors with 8-cyclopentyl-1,3-dipropylxanthine suppressed the inhibitory action of β-NAD on the force of electrically evoked contractions. The results do not support an inhibitory neurotransmitter role for β-NAD at intestinal neuromuscular junctions. The data suggest that β-NAD is a ligand for the adenosine A1 receptor subtype expressed by neurons in the enteric nervous system. The influence of β-NAD on intestinal motility emerges from adenosine A1 receptor-mediated suppression of neurotransmitter release at inhibitory neuromuscular junctions.

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

    Science.gov (United States)

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

    2015-07-01

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

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

    Science.gov (United States)

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

    2015-07-01

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

  15. Rapid tolerance against focal cerebral ischemia induced by isoflurane anesthesia is attenuated by adenosine A1 receptor antagonist in rats

    Institute of Scientific and Technical Information of China (English)

    刘艳红; 熊利泽

    2003-01-01

    The brief anesthesia with isoflurane induces rapid tolerance against focal cerebral ischemia in rats and adenosine A1 receptor antagonist, DPCPX, attenuates the beneficial effect of isoflurane preconditioning.

  16. Chronic hypoxia reduces adenosine A2A receptor-mediated inhibition of calcium current in rat PC12 cells via downregulation of protein kinase A.

    Science.gov (United States)

    Kobayashi, S; Beitner-Johnson, D; Conforti, L; Millhorn, D E

    1998-10-15

    1. Adenosine has been shown to decrease Ca2+ current (ICa) and attenuate the hypoxia-induced enhancement of intracellular free Ca2+ ([Ca2+]i) in oxygen-sensitive rat phaeochromocytoma (PC12) cells. These effects are mediated via the adenosine A2A receptor and protein kinase A (PKA). The current study was undertaken to determine the effects of adenosine on Ca2+ current and hypoxia-induced change in [Ca2+]i during chronic hypoxia. 2. Whole cell patch-clamp studies revealed that the effect of adenosine on ICa was significantly reduced when PC12 cells were exposed to hypoxia (10 % O2) for 24 and 48 h. 3. Ca2+ imaging studies using fura-2 revealed that the anoxia-induced increase in [Ca2+]i was significantly enhanced when PC12 cells were exposed to 10 % O2 for up to 48 h. In contrast, the inhibitory effects of adenosine on anoxia-induced elevation of [Ca2+]i was significantly blunted in PC12 cells exposed to hypoxia for 48 h. 4. Northern blot analysis revealed that mRNA for the A2A receptor, which is the only adenosine receptor subtype expressed in PC12 cells, was significantly upregulated by hypoxia. Radioligand binding analysis with [3H]CGS21680, a selective A2A receptor ligand, showed that the number of adenosine A2A receptor binding sites was similarly increased during exposure to 10% O2 for 48 h. 5. PKA enzyme activity was significantly inhibited when PC12 cells were exposed to 10% O2 for 24 and 48 h. However, we found that hypoxia failed to induce change in adenosine- and forskolin-stimulated adenylate cyclase enzyme activity. Chronic hypoxia also did not alter the immunoreactivity level of the G protein Gsalpha, an effector of the A2 signalling pathway. 6. Whole cell patch-clamp analysis showed that the effect of 8-bromo-cAMP, an activator of PKA, on ICa was significantly attenuated during 48 h exposure to 10% O2.7. We conclude therefore that the reduced effect of adenosine on ICa and [Ca2+]i in PC12 cells exposed to chronic hypoxia is due to hypoxia

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

    OpenAIRE

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

    2010-01-01

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

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

    Science.gov (United States)

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

    2014-02-01

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

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

    OpenAIRE

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

    2014-01-01

    Selective stimulation of inhibitory A1 and facilitatory A2a adenosine receptor subtypes located in the nucleus of the solitary tract (NTS) powerfully inhibits cardiopulmonary chemoreflex (CCR) control of regional sympathetic outputs via different mechanisms: direct inhibition of glutamate release and facilitation of an inhibitory neurotransmitter release, respectively. However, it remains unknown whether adenosine naturally released into the NTS has similar inhibitory effects on the CCR as th...

  20. Adenosine enhances sweet taste through A2B receptors in the taste bud.

    Science.gov (United States)

    Dando, Robin; Dvoryanchikov, Gennady; Pereira, Elizabeth; Chaudhari, Nirupa; Roper, Stephen D

    2012-01-01

    Mammalian taste buds use ATP as a neurotransmitter. Taste Receptor (type II) cells secrete ATP via gap junction hemichannels into the narrow extracellular spaces within a taste bud. This ATP excites primary sensory afferent fibers and also stimulates neighboring taste bud cells. Here we show that extracellular ATP is enzymatically degraded to adenosine within mouse vallate taste buds and that this nucleoside acts as an autocrine neuromodulator to selectively enhance sweet taste. In Receptor cells in a lingual slice preparation, Ca(2+) mobilization evoked by focally applied artificial sweeteners was significantly enhanced by adenosine (50 μM). Adenosine had no effect on bitter or umami taste responses, and the nucleoside did not affect Presynaptic (type III) taste cells. We also used biosensor cells to measure transmitter release from isolated taste buds. Adenosine (5 μM) enhanced ATP release evoked by sweet but not bitter taste stimuli. Using single-cell reverse transcriptase (RT)-PCR on isolated vallate taste cells, we show that many Receptor cells express the adenosine receptor, Adora2b, while Presynaptic (type III) and Glial-like (type I) cells seldom do. Furthermore, Adora2b receptors are significantly associated with expression of the sweet taste receptor subunit, Tas1r2. Adenosine is generated during taste stimulation mainly by the action of the ecto-5'-nucleotidase, NT5E, and to a lesser extent, prostatic acid phosphatase. Both these ecto-nucleotidases are expressed by Presynaptic cells, as shown by single-cell RT-PCR, enzyme histochemistry, and immunofluorescence. Our findings suggest that ATP released during taste reception is degraded to adenosine to exert positive modulation particularly on sweet taste.

  1. Downregulation of adenosine and P2X receptor-mediated cardiovascular responses in heart failure rats

    DEFF Research Database (Denmark)

    Zhao, Xin; Sun, X Y; Erlinge, D;

    2000-01-01

    Neurohormonal changes in congestive heart failure (CHF) include an enhanced peripheral sympathetic nerve activity which results in increased release of noradrenaline, neuropeptide Y and ATP. To examine if such changes in CHF would modulate peripheral pre- and postsynaptic receptors of ATP and its...... effects mediated by the endothelial P2Y receptors are unaffected in CHF. Moreover, the adenosine-mediated inhibitory effects on heart rate and blood pressure were also attenuated in the CHF rats. The most important changes in adenosine and P2-receptor function induced by ischaemic CHF were the reduced...... pressor effect mediated by the P2X receptor and the increased heart rate due to an attenuated inhibitory effect of adenosine....

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

    DEFF Research Database (Denmark)

    Hayashi, M.; Inagaki, A.; Novak, Ivana;

    2016-01-01

    Adenosine modulates a wide variety of biological processes via adenosine receptors. In the exocrine pancreas, adenosine regulates transepithelial anion secretion in duct cells and is considered to play a role in acini-to-duct signaling. To identify the functional adenosine receptors and Cl− chann...

  3. 2-(1-Hexyn-1-yl)adenosine-induced intraocular hypertension is mediated via K+ channel opening through adenosine A2A receptor in rabbits.

    Science.gov (United States)

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

    2005-08-22

    The present study was performed to clarify the mechanism of change in intraocular pressure by 2-(1-hexyn-1-yl)adenosine (2-H-Ado), a selective adenosine A2 receptor agonist, in rabbits. 2-H-Ado (0.1%, 50 microl)-induced ocular hypertension (E(max): 7.7 mm Hg) was inhibited by an adenosine A2A receptor antagonist 1,3,7-trimethyl-8-(3-chlorostyryl)xanthine, ATP-sensitive K+ channel blocker glibenclamide or 5-hydroxydecanoic acid, but not by an adenosine A1 receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine, an adenosine A2B receptor antagonist alloxazine or a cyclooxygenase inhibitor indomethacin. The outflow facility induced by 2-H-Ado seems to be independent of increase in intraocular pressure or ATP-sensitive K+ channel. In contrast, the recovery rate in intraocular pressure decreased by hypertonic saline was accelerated by 2-H-Ado, and this response was dependent on ATP-sensitive K+ channel. These results suggest that 2-H-Ado-induced ocular hypertension is mediated via K+ channel opening through adenosine A2A receptor, and this is probably due to aqueous formation, but independent of change in outflow facility or prostaglandin production.

  4. 2-(1-Hexyn-1-yl)adenosine-induced intraocular hypertension is mediated via K+ channel opening through adenosine A2A receptor in rabbits.

    Science.gov (United States)

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

    2005-08-22

    The present study was performed to clarify the mechanism of change in intraocular pressure by 2-(1-hexyn-1-yl)adenosine (2-H-Ado), a selective adenosine A2 receptor agonist, in rabbits. 2-H-Ado (0.1%, 50 microl)-induced ocular hypertension (E(max): 7.7 mm Hg) was inhibited by an adenosine A2A receptor antagonist 1,3,7-trimethyl-8-(3-chlorostyryl)xanthine, ATP-sensitive K+ channel blocker glibenclamide or 5-hydroxydecanoic acid, but not by an adenosine A1 receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine, an adenosine A2B receptor antagonist alloxazine or a cyclooxygenase inhibitor indomethacin. The outflow facility induced by 2-H-Ado seems to be independent of increase in intraocular pressure or ATP-sensitive K+ channel. In contrast, the recovery rate in intraocular pressure decreased by hypertonic saline was accelerated by 2-H-Ado, and this response was dependent on ATP-sensitive K+ channel. These results suggest that 2-H-Ado-induced ocular hypertension is mediated via K+ channel opening through adenosine A2A receptor, and this is probably due to aqueous formation, but independent of change in outflow facility or prostaglandin production. PMID:16023100

  5. Ligands for Ionotropic Glutamate Receptors

    Science.gov (United States)

    Swanson, Geoffrey T.; Sakai, Ryuichi

    Marine-derived small molecules and peptides have played a central role in elaborating pharmacological specificities and neuronal functions of mammalian ionotropic glutamate receptors (iGluRs), the primary mediators of excitatory syn-aptic transmission in the central nervous system (CNS). As well, the pathological sequelae elicited by one class of compounds (the kainoids) constitute a widely-used animal model for human mesial temporal lobe epilepsy (mTLE). New and existing molecules could prove useful as lead compounds for the development of therapeutics for neuropathologies that have aberrant glutamatergic signaling as a central component. In this chapter we discuss natural source origins and pharmacological activities of those marine compounds that target ionotropic glutamate receptors.

  6. Nitrosamines as nicotinic receptor ligands.

    Science.gov (United States)

    Schuller, Hildegard M

    2007-05-30

    Nitrosamines are carcinogens formed in the mammalian organism from amine precursors contained in food, beverages, cosmetics and drugs. The potent carcinogen, NNK, and the weaker carcinogen, NNN, are nitrosamines formed from nicotine. Metabolites of the nitrosamines react with DNA to form adducts responsible for genotoxic effects. We have identified NNK as a high affinity agonist for the alpha7 nicotinic acetylcholine receptor (alpha7nAChR) whereas NNN bound with high affinity to epibatidine-sensitive nAChRs. Diethylnitrosamine (DEN) bound to both receptors but with lower affinity. High levels of the alpha7nAChR were expressed in human small cell lung cancer (SCLC) cell lines and in hamster pulmonary neuroendocrine cells (PNECs), which serve as a model for the cell of origin of human SCLC. Exposure of SCLC or PNECs to NNK or nicotine increased expression of the alpha7nAChR and caused influx of Ca(2+), activation of PKC, Raf-1, ERK1/2, and c-myc, resulting in the stimulation of cell proliferation. Signaling via the alpha7nAChR was enhanced when cells were maintained in an environment of 10-15% CO(2) similar to that in the diseased lung. Hamsters with hyperoxia-induced pulmonary fibrosis developed neuroendocrine lung carcinomas similar to human SCLC when treated with NNK, DEN, or nicotine. The development of the NNK-induced tumors was prevented by green tea or theophylline. The beta-adrenergic receptor agonist, isoproterenol or theophylline blocked NNK-induced cell proliferation in vitro. NNK and nicotine-induced hyperactivity of the alpha7nAChR/RAF/ERK1/2 pathway thus appears to play a crucial role in the development of SCLC in smokers and could be targeted for cancer prevention. PMID:17459420

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

    Directory of Open Access Journals (Sweden)

    Felicita Pedata

    2014-01-01

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

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

    Directory of Open Access Journals (Sweden)

    Shinji Kataoka

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

  9. Novel ligands for the human adenosine A1 receptor

    NARCIS (Netherlands)

    Chang, Lisa Chung Wai

    2005-01-01

    This research describes the quest to create 'super-caffeines', substances that only produce the desired effects of caffeine, and unlike caffeine, substances that should only have to be taken in measured, minute, controlled amounts to achieve these effects. Unless particular steps are taken to avoid

  10. Stimulation of adenosine receptors in the nucleus accumbens reverses the expression of cocaine sensitization and cross-sensitization to dopamine D2 receptors in rats

    OpenAIRE

    Hobson, Benjamin D.; Merritt, Kathryn E.; Bachtell, Ryan K.

    2012-01-01

    Adenosine receptors co-localize with dopamine receptors on medium spiny nucleus accumbens (NAc) neurons where they antagonize dopamine receptor activity. It remains unclear whether adenosine receptor stimulation in the NAc restores cocaine-induced enhancements in dopamine receptor sensitivity. The goal of these studies was to determine whether stimulating A1 or A2A receptors in the NAc reduces the expression of cocaine sensitization. Rats were sensitized with 7 daily treatments of cocaine (15...

  11. Glycomimetic ligands for the human asialoglycoprotein receptor.

    Science.gov (United States)

    Mamidyala, Sreeman K; Dutta, Sanjay; Chrunyk, Boris A; Préville, Cathy; Wang, Hong; Withka, Jane M; McColl, Alexander; Subashi, Timothy A; Hawrylik, Steven J; Griffor, Matthew C; Kim, Sung; Pfefferkorn, Jeffrey A; Price, David A; Menhaji-Klotz, Elnaz; Mascitti, Vincent; Finn, M G

    2012-02-01

    The asialoglycoprotein receptor (ASGPR) is a high-capacity galactose-binding receptor expressed on hepatocytes that binds its native substrates with low affinity. More potent ligands are of interest for hepatic delivery of therapeutic agents. We report several classes of galactosyl analogues with varied substitution at the anomeric, C2-, C5-, and C6-positions. Significant increases in binding affinity were noted for several trifluoromethylacetamide derivatives without covalent attachment to the protein. A variety of new ligands were obtained with affinity for ASGPR as good as or better than that of the parent N-acetylgalactosamine, showing that modification on either side of the key C3,C4-diol moiety is well tolerated, consistent with previous models of a shallow binding pocket. The galactosyl pyranose motif therefore offers many opportunities for the attachment of other functional units or payloads while retaining low-micromolar or better affinity for the ASGPR.

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

    LENUS (Irish Health Repository)

    Wakai, A

    2012-02-03

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

  13. In vivo adenosine A(2B) receptor desensitization in guinea-pig airway smooth muscle: implications for asthma.

    Science.gov (United States)

    Breschi, Maria Cristina; Blandizzi, Corrado; Fogli, Stefano; Martinelli, Cinzia; Adinolfi, Barbara; Calderone, Vincenzo; Camici, Marcella; Martinotti, Enrica; Nieri, Paola

    2007-12-01

    This study was aimed at characterizing the role of adenosine receptor subtypes in the contractility modulation of guinea-pig airway smooth muscle in normal and pathological settings. In vitro and in vivo experiments were performed by testing selective agonists and antagonists on isolated tracheal smooth muscle preparations and pulmonary inflation pressure, respectively, under normal conditions or following ovalbumin-induced allergic sensitization. In normal and sensitized animals, the adenosine A(2A)/A(2B) receptor agonist, NECA, evoked relaxing responses of isolated tracheal preparations precontracted with histamine, and such an effect was reversed by the adenosine A(2B) antagonist, MRS 1706, in the presence or in the absence of epithelium. The expression of mRNA coding for adenosine A(2B) receptors was demonstrated in tracheal specimens. In vitro desensitization with 100 microM NECA markedly reduced the relaxing effect of the agonist. In vivo NECA or adenosine administration to normal animals inhibited histamine-mediated bronchoconstriction, while these inhibitory effects no longer occurred in sensitized guinea-pigs. Adenosine plasma levels were significantly higher in sensitized than normal animals. In conclusion, our data demonstrate that: (i) adenosine A(2B) receptors are responsible for the relaxing effects of adenosine on guinea-pig airways; (ii) these receptors can undergo rapid adaptive changes that may affect airway smooth muscle responsiveness to adenosine; (iii) ovalbumin-induced sensitization promotes a reversible inactivation of adenosine A(2B) receptors which can be ascribed to homologous desensitization. These findings can be relevant to better understand adenosine functions in airways as well as mechanisms of action of asthma therapies targeting the adenosine system.

  14. Adenosine receptors in rat and human pancreatic ducts stimulate chloride transport

    DEFF Research Database (Denmark)

    Novak, Ivana; Hede, Susanne; Hansen, Mette

    2007-01-01

    Previously, we have shown that pancreatic acini release adenosine triphosphate (ATP) and ATP-handling enzymes, and pancreatic ducts express various purinergic P2 receptors. The aim of the present study was to establish whether pancreatic ducts also express adenosine receptors and whether...... these could be involved in secretory processes, which involve cystic fibrosis transmembrane regulator (CFTR) Cl(-) channels or Ca(2+)-activated Cl(-) channels and [Formula: see text] transporters. Reverse transcriptase polymerase chain reaction analysis on rat pancreatic ducts and human duct cell......) for duct cell lines. Whole-cell patch-clamp recordings on rat pancreatic ducts showed that, in about half of the recordings, adenosine depolarized the membrane voltage, and this was because of the opening of Cl(-) channels. Using a Cl(-)-sensitive fluorophore and single-cell imaging on duct cell lines...

  15. A novel chemogenomics analysis of G protein-coupled receptors (GPCRs and their ligands: a potential strategy for receptor de-orphanization

    Directory of Open Access Journals (Sweden)

    Emmerich Michael TM

    2010-06-01

    Full Text Available Abstract Background G protein-coupled receptors (GPCRs represent a family of well-characterized drug targets with significant therapeutic value. Phylogenetic classifications may help to understand the characteristics of individual GPCRs and their subtypes. Previous phylogenetic classifications were all based on the sequences of receptors, adding only minor information about the ligand binding properties of the receptors. In this work, we compare a sequence-based classification of receptors to a ligand-based classification of the same group of receptors, and evaluate the potential to use sequence relatedness as a predictor for ligand interactions thus aiding the quest for ligands of orphan receptors. Results We present a classification of GPCRs that is purely based on their ligands, complementing sequence-based phylogenetic classifications of these receptors. Targets were hierarchically classified into phylogenetic trees, for both sequence space and ligand (substructure space. The overall organization of the sequence-based tree and substructure-based tree was similar; in particular, the adenosine receptors cluster together as well as most peptide receptor subtypes (e.g. opioid, somatostatin and adrenoceptor subtypes. In ligand space, the prostanoid and cannabinoid receptors are more distant from the other targets, whereas the tachykinin receptors, the oxytocin receptor, and serotonin receptors are closer to the other targets, which is indicative for ligand promiscuity. In 93% of the receptors studied, de-orphanization of a simulated orphan receptor using the ligands of related receptors performed better than random (AUC > 0.5 and for 35% of receptors de-orphanization performance was good (AUC > 0.7. Conclusions We constructed a phylogenetic classification of GPCRs that is solely based on the ligands of these receptors. The similarities and differences with traditional sequence-based classifications were investigated: our ligand

  16. Ligand binding was acquired during evolution of nuclear receptors

    OpenAIRE

    Escriva, Hector; Safi, Rachid; Hänni, Catherine; Langlois, Marie-Claire; Saumitou-Laprade, Pierre; Stehelin, Dominique; Capron, André; Pierce, Raymond; Laudet, Vincent

    1997-01-01

    The nuclear receptor (NR) superfamily comprises, in addition to ligand-activated transcription factors, members for which no ligand has been identified to date. We demonstrate that orphan receptors are randomly distributed in the evolutionary tree and that there is no relationship between the position of a given liganded receptor in the tree and the chemical nature of its ligand. NRs are specific to metazoans, as revealed by a screen of NR-related sequences in early- and non-metazoan organism...

  17. Effect of adenosine and adenosine receptor antagonist on Müller cell potassium channel in Rat chronic ocular hypertension models.

    Science.gov (United States)

    Yang, Zijian; Huang, Ping; Liu, Xiaohong; Huang, Shouyue; Deng, Lianfu; Jin, Zhe; Xu, Shuo; Shen, Xi; Luo, Xunda; Zhong, Yisheng

    2015-01-01

    Müller cells are principal glial cells in rat retina and have attracted much attention in glaucoma studies. However, it is not clear whether adenosine and adenosine receptor (AR) antagonists play any roles in the regulation of potassium channels in Müller cells and subsequently in the promotion of glutamine synthetase (GS) and L-Glutamate/L-Aspartate Transporter (GLAST) functions. We found that chronic ocular hypertension (COH) in rat down-regulated Müller cells Kir2.1, Kir4.1, TASK-1, GS and GLAST expressions and attenuated the peak of inward potassium current. Retinal ganglion cells (RGC) count was lower in the COH rats than that in the sham operation animals. Intravitreal injection of selective A2A AR antagonist SCH442416 up-regulated Müller cell Kir4.1, TASK-1, GS and GLAST expressions and enhanced inward potassium currents compared with those in the COH rats with vehicle control. Meanwhile, the RGC count was higher following intravitreal injection of SCH442416 in the COH rats than that after vehicle injection. The fact that PKA inhibitor H-89 blocked these SCH442416 effects suggested that the PKA signaling pathway was involved in the observed ocular responses following the intravitreal SCH442416 injection. PMID:26063641

  18. Adenosine A2A receptor binding profile of two antagonists, ST1535 and KW6002: consideration on the presence of atypical adenosine A2A binding sites

    Directory of Open Access Journals (Sweden)

    Teresa Riccioni

    2010-08-01

    Full Text Available Adenosine A2A receptors seem to exist in typical (more in striatum and atypical (more in hippocampus and cortex subtypes. In the present study, we investigated the affinity of two adenosine A2A receptor antagonists, ST1535 [2 butyl -9-methyl-8-(2H-1,2,3-triazol 2-yl-9H-purin-6-xylamine] and KW6002 [(E-1,3-diethyl-8-(3,4-dimethoxystyryl-7-methyl-3,7-dihydro-1H-purine-2,6,dione] to the “typical” and “atypical” A2A binding sites. Affinity was determined by radioligand competition experiments in membranes from rat striatum and hippocampus. Displacement of the adenosine analog [3H]CGS21680 [2-p-(2-carboxyethylphenethyl-amino-5’-N-ethylcarbox-amidoadenosine] was evaluated in the absence or in the presence of either CSC [8-(3-chlorostyryl-caffeine], an adenosine A2A antagonist that pharmacologically isolates atypical binding sites, or DPCPX (8-cyclopentyl-1,3-dipropylxanthine, an adenosine A1 receptor antagonist that pharmacologically isolates typical binding site. ZM241385 [84-(2-[7-amino-2-(2-furyl [1,2,4]-triazol[2,3-a][1,3,5]triazin-5-yl amino]ethyl phenol] and SCH58261 [(5-amino-7-(β-phenylethyl-2-(8-furylpyrazolo(4,3-e-1,2,4-triazolo(1,5-c pyrimidine], two other adenosine A2A receptor antagonists, which were reported to differently bind to atypical and typical A2A receptors, were used as reference compounds. ST1535, KW6002, ZM241385 and SCH58261 displaced [3H]CGS21680 with higher affinity in striatum than in hippocampus. In hippocampus, no typical adenosine A2A binding was detected, and ST1535 was the only compound that occupied atypical A2A adenosine receptors. Present data are explained in terms of heteromeric association among adenosine A2A, A2B and A1 receptors, rather than with the presence of atypical A2A receptor subtype.

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

    Science.gov (United States)

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

    2016-07-01

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

  20. NCS-1 associates with adenosine A2A receptors and modulates receptor function

    Directory of Open Access Journals (Sweden)

    Gemma eNavarro

    2012-04-01

    Full Text Available Modulation of G protein-coupled receptor (GPCR signalling by local changes in intracellular calcium concentration is an established function of Calmodulin which is known to interact with many GPCRs. Less is known about the functional role of the closely related neuronal EF-hand Ca2+-sensor proteins that frequently associate with calmodulin targets with different functional outcome. In the present study we aimed to investigate if a target of calmodulin – the A2A adenosine receptor, is able to associate with two other neuronal calcium binding proteins, namely NCS-1 and caldendrin. Using bioluminescence resonance energy transfer and co-immunoprecipitation experiments we show the existence of A2A - NCS-1 complexes in living cells whereas caldendrin did not associate with A2A receptors under the conditions tested. Interestingly, NCS-1 binding modulated downstream A2A receptor intracellular signalling in a Ca2+-dependent manner. Taken together this study provides further evidence that neuronal Ca2+-sensor proteins play an important role in modulation of GPCR signalling.

  1. Untangling dopamine-adenosine receptor-receptor assembly in experimental parkinsonism in rats

    Directory of Open Access Journals (Sweden)

    Víctor Fernández-Dueñas

    2015-01-01

    Full Text Available Parkinson’s disease (PD is a dopaminergic-related pathology in which functioning of the basal ganglia is altered. It has been postulated that a direct receptor-receptor interaction – i.e. of dopamine D2 receptor (D2R with adenosine A2A receptor (A2AR (forming D2R-A2AR oligomers – finely regulates this brain area. Accordingly, elucidating whether the pathology prompts changes to these complexes could provide valuable information for the design of new PD therapies. Here, we first resolved a long-standing question concerning whether D2R-A2AR assembly occurs in native tissue: by means of different complementary experimental approaches (i.e. immunoelectron microscopy, proximity ligation assay and TR-FRET, we unambiguously identified native D2R-A2AR oligomers in rat striatum. Subsequently, we determined that, under pathological conditions (i.e. in a rat PD model, D2R-A2AR interaction was impaired. Collectively, these results provide definitive evidence for alteration of native D2R-A2AR oligomers in experimental parkinsonism, thus conferring the rationale for appropriate oligomer-based PD treatments.

  2. Pyrazolo Derivatives as Potent Adenosine Receptor Antagonists: An Overview on the Structure-Activity Relationships

    Directory of Open Access Journals (Sweden)

    Siew Lee Cheong

    2011-01-01

    Full Text Available In the past few decades, medicinal chemistry research towards potent and selective antagonists of human adenosine receptors (namely, A1, A2A, A2B, and A3 has been evolving rapidly. These antagonists are deemed therapeutically beneficial in several pathological conditions including neurological and renal disorders, cancer, inflammation, and glaucoma. Up to this point, many classes of compounds have been successfully synthesized and identified as potent human adenosine receptor antagonists. In this paper, an overview of the structure-activity relationship (SAR profiles of promising nonxanthine pyrazolo derivatives is reported and discussed. We have emphasized the SAR for some representative structures such as pyrazolo-[4,3-e]-1,2,4-triazolo-[1,5-c]pyrimidines; pyrazolo-[3,4-c] or -[4,3-c]quinolines; pyrazolo-[4,3-d]pyrimidinones; pyrazolo-[3,4-d]pyrimidines and pyrazolo-[1,5-a]pyridines. This overview not only clarifies the structural requirements deemed essential for affinity towards individual adenosine receptor subtypes, but it also sheds light on the rational design and optimization of existing structural templates to allow us to conceive new, more potent adenosine receptor antagonists.

  3. Therapeutic efficacy of the adenosine A1 receptor agonist N6-cyclopentyladenosine (CPA) against organophosphate intoxication

    NARCIS (Netherlands)

    Bueters, T.J.H.; Groen, B.; Danhof, M.; IJzerman, A.P.; Helden, H.P.M. van

    2002-01-01

    The objective of the present study was to investigate whether reduction of central acetylcholine (ACh) accumulation by adenosine receptor agonists could serve as a generic treatment against organophosphate (OP) poisoning. The OPs studied were tabun (O-ethyl-N-dimethylphosphoramidocyanidate), sarin (

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

    Science.gov (United States)

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

    2005-04-01

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

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

    Science.gov (United States)

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

    2005-04-01

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

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

    Science.gov (United States)

    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.

  7. A2A adenosine receptor-mediated increase in coronary flow in hyperlipidemic APOE–knockout mice

    OpenAIRE

    Teng, Bunyen

    2011-01-01

    Bunyen Teng, S Jamal MustafaDepartment of Physiology and Pharmacology and Center for Cardiovascular and Respiratory Sciences, West Virginia University, Morgantown, WV, USAAbstract: Adenosine-induced coronary vasodilation is predominantly A2A adenosine receptor (AR)-mediated, whereas A1 AR is known to negatively modulate the coronary flow (CF). However, the coronary responses to adenosine in hyperlipidemia and atherosclerosis are not well understood. Using hyperlipidemic/atherosclerotic apolip...

  8. Wound Healing Is Accelerated by Agonists of Adenosine A2 (Gα s-linked) Receptors

    OpenAIRE

    Montesinos, M. Carmen; Gadangi, Pratap; Longaker, Michael; Sung, Joanne; Levine, Jamie; Nilsen, Diana; Reibman, Joan; Min LI; Jiang, Chuan-Kui; Hirschhorn, Rochelle; Recht, Phoebe A.; Ostad, Edward; Levin, Richard I.; Cronstein, Bruce N.

    1997-01-01

    The complete healing of wounds is the final step in a highly regulated response to injury. Although many of the molecular mediators and cellular events of healing are known, their manipulation for the enhancement and acceleration of wound closure has not proven practical as yet. We and others have established that adenosine is a potent regulator of the inflammatory response, which is a component of wound healing. We now report that ligation of the Gαs-linked adenosine receptors on the cells o...

  9. GIRK channel activation via adenosine or muscarinic receptors has similar effects on rat atrial electrophysiology.

    Science.gov (United States)

    Wang, Xiaodong; Liang, Bo; Skibsbye, Lasse; Olesen, Søren-Peter; Grunnet, Morten; Jespersen, Thomas

    2013-08-01

    G protein-coupled inwardly rectifying K⁺ channels (GIRK) are important in the regulation of heart rate and atrial electrophysiology. GIRK channels are activated by G protein-coupled receptors, including muscarinic M₂ receptors and adenosine A₁ receptors. The aim of this study was to characterize and compare the electrophysiological effects of acetylcholine (ACh) and adenosine on GIRK channels in rat atria. Action potential duration at 90% repolarization (APD₉₀), effective refractory period (ERP), and resting membrane potential (RMP) were investigated in isolated rat atria by intracellular recordings. Both the adenosine analog N6-cyclopentyladenosine (CPA) and ACh profoundly shortened APD₉₀ and ERP and hyperpolarized the RMP. No additive or synergistic effect of CPA and ACh coapplication was observed. To antagonize GIRK channel activation, the specific inhibitor rTertiapin Q (TTQ) was applied. The coapplication of TTQ reversed the CPA and ACh-induced effects. When TTQ was applied without exogenous receptor activator, both APD₉₀ and ERP were prolonged and RMP was depolarized, confirming a basal activity of the GIRK current. The results reveal that activation of A₁ and M₂ receptors has a profound and equal effect on the electrophysiology in rat atrium. This effect is to a major extent mediated through GIRK channels. Furthermore, these results support the notion that atrial GIRK currents from healthy hearts have a basal component and additional activation can be mediated via at least 2 different receptor mechanisms. PMID:23609329

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

    Science.gov (United States)

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

    2015-12-01

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

  11. Structural basis for ligand recognition of incretin receptors

    DEFF Research Database (Denmark)

    Underwood, Christina Rye; Parthier, Christoph; Reedtz-Runge, Steffen

    2010-01-01

    The glucose-dependent insulinotropic polypeptide (GIP) receptor and the glucagon-like peptide-1 (GLP-1) receptor are homologous G-protein-coupled receptors (GPCRs). Incretin receptor agonists stimulate the synthesis and secretion of insulin from pancreatic β-cells and are therefore promising agents...... for the treatment of type 2 diabetes. It is well established that the N-terminal extracellular domain (ECD) of incretin receptors is important for ligand binding and ligand specificity, whereas the transmembrane domain is involved in receptor activation. Structures of the ligand-bound ECD of incretin receptors have...... appear to be the main driving force for ligand binding to the ECD of incretin receptors. Obviously, the-still missing-structures of full-length incretin receptors are required to construct a complete picture of receptor function at the molecular level. However, the progress made recently in structural...

  12. The 2.6 Angstrom Crystal Structure of a Human A[subscript 2A] Adenosine Receptor Bound to an Antagonist

    Energy Technology Data Exchange (ETDEWEB)

    Jaakola, Veli-Pekka; Griffith, Mark T.; Hanson, Michael A.; Cherezov, Vadim; Chien, Ellen Y.T.; Lane, J. Robert; IJzerman, Adriaan P.; Stevens, Raymond C. (Scripps); (Leiden/Amsterdam)

    2009-01-15

    The adenosine class of heterotrimeric guanine nucleotide-binding protein (G protein)-coupled receptors (GPCRs) mediates the important role of extracellular adenosine in many physiological processes and is antagonized by caffeine. We have determined the crystal structure of the human A{sub 2A} adenosine receptor, in complex with a high-affinity subtype-selective antagonist, ZM241385, to 2.6 angstrom resolution. Four disulfide bridges in the extracellular domain, combined with a subtle repacking of the transmembrane helices relative to the adrenergic and rhodopsin receptor structures, define a pocket distinct from that of other structurally determined GPCRs. The arrangement allows for the binding of the antagonist in an extended conformation, perpendicular to the membrane plane. The binding site highlights an integral role for the extracellular loops, together with the helical core, in ligand recognition by this class of GPCRs and suggests a role for ZM241385 in restricting the movement of a tryptophan residue important in the activation mechanism of the class A receptors.

  13. Integrin receptors and ligand-gated channels.

    Science.gov (United States)

    Morini, Raffaella; Becchetti, Andrea

    2010-01-01

    Plastic expression of different integrin subunits controls the different stages of neural development, whereas in the adult integrins regulate synaptic stability. Evidence of integrin-channel crosstalk exists for ionotropic glutamate receptors. As is often the case in other tissues, integrin engagement regulates channel activity through complex signaling pathways that often include tyrosine phosphorylation cascades. The specific pathways recruited by integrin activation depend on cerebral region and cell type. In turn, ion channels control integrin expression onto the plasma membrane and their ligand binding affinity. The most extensive studies concern the hippocampus and suggest implications for neuronal circuit plasticity. The physiological relevance of these findings depends on whether adhesion molecules, aside from determining tissue stability, contribute to synaptogenesis and the responsiveness of mature synapses, thus contributing to long-term circuit consolidation. Little evidence is available for other ligand-gated channels, with the exception of nicotinic receptors. These exert a variety of functions in neurons and non neural tissue, both in development and in the adult, by regulating cell cycle, synaptogenesis and synaptic circuit refinement. Detailed studies in epidermal keratinocytes have shed some light on the possible mechanisms through which ACh can regulate cell motility, which may be of general relevance for morphogenetic processes. As to the control of mature synapses, most results concern the integrinic control of nicotinic receptors in the neuromuscular junction. Following this lead, a few studies have addressed similar topics in adult cerebral synapses. However, pursuing and interpreting these results in the brain is especially difficult because of the complexity of the nicotinic roles and the widespread contribution of nonsynaptic, paracrine transmission. From a pathological point of view, considering the well-known contribution of both

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

    OpenAIRE

    de Mendonça, Alexandre; Ribeiro, J. A.

    1997-01-01

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

  15. The Safety of an Adenosine A(1)-Receptor Antagonist, Rolofylline, in Patients with Acute Heart Failure and Renal Impairment Findings from PROTECT

    NARCIS (Netherlands)

    Teerlink, John R.; Iragui, Vicente J.; Mohr, Jay P.; Carson, Peter E.; Hauptman, Paul J.; Lovett, David H.; Miller, Alan B.; Pina, Ileana L.; Thomson, Scott; Varosy, Paul D.; Zile, Michael R.; Cleland, John G. F.; Givertz, Michael M.; Metra, Marco; Ponikowski, Piotr; Voors, Adriaan A.; Davison, Beth A.; Cotter, Gad; Wolko, Denise; DeLucca, Paul; Salerno, Christina M.; Mansoor, George A.; Dittrich, Howard; O'Connor, Christopher M.; Massi, Barry M.

    2012-01-01

    Background: Adenosine exerts actions in multiple organ systems, and adenosine receptors are a therapeutic target in many development programmes. Objective: The aim of this analysis was to evaluate the safety of rolofylline, an adenosine A(1)-receptor antagonist, in patients with acute heart failure.

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

    International Nuclear Information System (INIS)

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

  17. Substrate coated with receptor and labelled ligand for assays

    International Nuclear Information System (INIS)

    Improvements in the procedures for assaying ligands are described. The assay consists of a polystyrene tube on which receptors are present for both the ligand to be assayed and a radioactively labelled form of the ligand. The receptors on the bottom portion of the tube are also coated with labelled ligands, thus eliminating the necessity for separate addition of the labelled ligand and sample during an assay. Examples of ligands to which this method is applicable include polypeptides, nucleotides, nucleosides and proteins. Specific examples are given in which the ligand to be assayed is digoxin, the labelled form of the ligand is 3-0-succinyl digoxyigenin tyrosine (125I) and the receptor is digoxin antibody. (U.K.)

  18. Increased accuracy of ligand sensing by receptor internalization

    CERN Document Server

    Aquino, Gerardo

    2010-01-01

    Many types of cells can sense external ligand concentrations with cell-surface receptors at extremely high accuracy. Interestingly, ligand-bound receptors are often internalized, a process also known as receptor-mediated endocytosis. While internalization is involved in a vast number of important functions for the life of a cell, it was recently also suggested to increase the accuracy of sensing ligand as the overcounting of the same ligand molecules is reduced. Here we show, by extending simple ligand-receptor models to out-of-equilibrium thermodynamics, that internalization increases the accuracy with which cells can measure ligand concentrations in the external environment. Comparison with experimental rates of real receptors demonstrates that our model has indeed biological significance.

  19. GIRK channel activation via adenosine or muscarinic receptors has similar effects on rat atrial electrophysiology

    DEFF Research Database (Denmark)

    Wang, Xiaodong; Liang, Bo; Skibsbye, Lasse;

    2013-01-01

    and compare the electrophysiological effects of acetylcholine (ACh) and adenosine on GIRK channels in rat atria. Action potential duration at 90% repolarization (APD90), effective refractory period (ERP), and resting membrane potential (RMP) were investigated in isolated rat atria by intracellular recordings....... Both the adenosine analog N6-cyclopentyladenosine (CPA) and ACh profoundly shortened APD90 and ERP and hyperpolarized the RMP. No additive or synergistic effect of CPA and ACh coapplication was observed. To antagonize GIRK channel activation, the specific inhibitor rTertiapin Q (TTQ) was applied....... The coapplication of TTQ reversed the CPA and ACh-induced effects. When TTQ was applied without exogenous receptor activator, both APD90 and ERP were prolonged and RMP was depolarized, confirming a basal activity of the GIRK current. The results reveal that activation of A1 and M2 receptors has a profound and equal...

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

    Science.gov (United States)

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

    1998-04-01

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

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

    DEFF Research Database (Denmark)

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

    2014-01-01

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

  2. Activation of NTS A(1) adenosine receptors inhibits regional sympathetic responses evoked by activation of cardiopulmonary chemoreflex.

    Science.gov (United States)

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

    2012-09-01

    Previously we have shown that adenosine operating via the A(1) receptor subtype may inhibit glutamatergic transmission in the baroreflex arc within the nucleus of the solitary tract (NTS) and differentially increase renal (RSNA), preganglionic adrenal (pre-ASNA), and lumbar (LSNA) sympathetic nerve activity (ASNA>RSNA≥LSNA). Since the cardiopulmonary chemoreflex and the arterial baroreflex are mediated via similar medullary pathways, and glutamate is a primary transmitter in both pathways, it is likely that adenosine operating via A(1) receptors in the NTS may differentially inhibit regional sympathetic responses evoked by activation of cardiopulmonary chemoreceptors. Therefore, in urethane-chloralose-anesthetized rats (n = 37) we compared regional sympathoinhibition evoked by the cardiopulmonary chemoreflex (activated with right atrial injections of serotonin 5HT(3) receptor agonist phenylbiguanide, PBG, 1-8 μg/kg) before and after selective stimulation of NTS A(1) adenosine receptors [microinjections of N(6)-cyclopentyl adenosine (CPA), 0.033-330 pmol/50 nl]. Activation of cardiopulmonary chemoreceptors evoked differential, dose-dependent sympathoinhibition (RSNA>ASNA>LSNA), and decreases in arterial pressure and heart rate. These differential sympathetic responses were uniformly attenuated in dose-dependent manner by microinjections of CPA into the NTS. Volume control (n = 11) and blockade of adenosine receptor subtypes in the NTS via 8-(p-sulfophenyl)theophylline (8-SPT, 1 nmol in 100 nl) (n = 9) did not affect the reflex responses. We conclude that activation of NTS A(1) adenosine receptors uniformly inhibits neural and cardiovascular cardiopulmonary chemoreflex responses. A(1) adenosine receptors have no tonic modulatory effect on this reflex under normal conditions. However, when adenosine is released into the NTS (i.e., during stress or severe hypotension/ischemia), it may serve as negative feedback regulator for depressor and sympathoinhibitory reflexes

  3. Death receptors and ligands in cervical carcinogenesis : an immunohistochemical study

    NARCIS (Netherlands)

    Reesink-Peters, N; Hougardy, B M T; van den Heuvel, F A J; Ten Hoor, K A; Hollema, H; Boezen, H M; de Vries, E G E; de Jong, S; van der Zee, A G J

    2005-01-01

    OBJECTIVE: Increasing imbalance between proliferation and apoptosis is important in cervical carcinogenesis. The death ligands FasL and tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) induce apoptosis by binding to their cognate cell-surface death receptors Fas or death receptor (DR)

  4. The in vivo respiratory phenotype of the adenosine A1 receptor knockout mouse.

    Science.gov (United States)

    Heitzmann, Dirk; Buehler, Philipp; Schweda, Frank; Georgieff, Michael; Warth, Richard; Thomas, Joerg

    2016-02-01

    The nucleoside adenosine has been implicated in the regulation of respiration, especially during hypoxia in the newborn. In this study the role of adenosine A1 receptors for the control of respiration was investigated in vivo. To this end, respiration of unrestrained adult and neonatal adenosine A1 receptor knockout mice (A1R(-/-)) was measured in a plethysmographic device. Under control conditions (21% O2) and mild hypoxia (12-15% O2) no difference of respiratory parameters was observed between adult wildtype (A1R(+/+)) and A1R(-/-) mice. Under more severe hypoxia (6-10% O2) A1R(+/+) mice showed, after a transient increase of respiration, a decrease of respiration frequency (fR) and tidal volume (VT) leading to a decrease of minute volume (MV). This depression of respiration during severe hypoxia was absent in A1R(-/-) mice which displayed a stimulated respiration as indicated by the enhancement of MV by some 50-60%. During hypercapnia-hyperoxia (3-10% CO2/97-90 % O2), no obvious differences in respiration of A1R(-/-) and A1R(+/+) was observed. In neonatal mice, the respiratory response to hypoxia was surprisingly similar in both genotypes. However, neonatal A1R(-/-) mice appeared to have more frequently periods of apnea during hypoxia and in the post-hypoxic control period. In conclusion, these data indicate that the adenosine A1 receptor is an important molecular component mediating hypoxic depression in adult mice and it appears to stabilize respiration of neonatal mice. PMID:26593641

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

    DEFF Research Database (Denmark)

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

    2011-01-01

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

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

    NARCIS (Netherlands)

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

    2011-01-01

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

  7. A3 Adenosine Receptors Modulate Hypoxia-inducible Factor-1a Expression in Human A375 Melanoma Cells

    Directory of Open Access Journals (Sweden)

    Stefania Merighi

    2005-10-01

    Full Text Available Hypoxia-inducible factor-1 (HIF-1 is a key regulator of genes crucial to many aspects of cancer biology. The purine nucleoside, adenosine, accumulates within many tissues under hypoxic conditions, including that of tumors. Because the levels of both HIF-1 and adenosine are elevated within the hypoxic environment of solid tumors, we investigated whether adenosine may regulate HIF-1. Here we show that, under hypoxic conditions (< 2% 02, adenosine upregulates HIF-1α protein expression in a dose-dependent and timedependent manner, exclusively through the A3 receptor subtype. The response to adenosine was generated at the cell surface because the inhibition of A3 receptor expression, by using small interfering RNA, abolished nucleoside effects. A3 receptor stimulation in hypoxia also increases angiopoietin-2 (Ang-2 protein accumulation through the induction of HIF-1α. In particular, we found that A3 receptor stimulation activates p44/p42 and p38 mitogen-activated protein kinases, which are required for A3-induced increase of HIF-1a and Ang-2. Collectively, these results suggest a cooperation between hypoxic and adenosine signals that ultimately may lead to the increase in HIF-1-mediated effects in cancer cells.

  8. A response calculus for immobilized T cell receptor ligands

    DEFF Research Database (Denmark)

    Andersen, P S; Menné, C; Mariuzza, R A;

    2001-01-01

    determine the level of T cell activation. When fitted to T cell responses against purified ligands immobilized on plastic surfaces, the 2D-affinity model adequately simulated changes in cellular activation as a result of varying ligand affinity and ligand density. These observations further demonstrated......To address the molecular mechanism of T cell receptor (TCR) signaling, we have formulated a model for T cell activation, termed the 2D-affinity model, in which the density of TCR on the T cell surface, the density of ligand on the presenting surface, and their corresponding two-dimensional affinity...... the importance of receptor cross-linking density in determining TCR signaling. Moreover, it was found that the functional two-dimensional affinity of TCR ligands was affected by the chemical composition of the ligand-presenting surface. This makes it possible that cell-bound TCR ligands, despite their low...

  9. Gene expression and function of adenosine A(2A) receptor in the rat carotid body.

    Science.gov (United States)

    Kobayashi, S; Conforti, L; Millhorn, D E

    2000-08-01

    The present study was undertaken to determine whether rat carotid bodies express adenosine (Ado) A(2A) receptors and whether this receptor is involved in the cellular response to hypoxia. Our results demonstrate that rat carotid bodies express the A(2A) and A(2B) Ado receptor mRNAs but not the A(1) or A(3) receptor mRNAs as determined by reverse transcriptase-polymerase chain reaction. In situ hybridization confirmed the expression of the A(2A) receptor mRNA. Immunohistochemical studies further showed that the A(2A) receptor is expressed in the carotid body and that it is colocalized with tyrosine hydroxylase in type I cells. Whole cell voltage-clamp studies using isolated type I cells showed that Ado inhibited the voltage-dependent Ca(2+) currents and that this inhibition was abolished by the selective A(2A) receptor antagonist ZM-241385. Ca(2+) imaging studies using fura 2 revealed that exposure to severe hypoxia induced elevation of intracellular Ca(2+) concentration ([Ca(2+)](i)) in type I cells and that extracellularly applied Ado significantly attenuated the hypoxia-induced elevation of [Ca(2+)](i). Taken together, our findings indicate that A(2A) receptors are present in type I cells and that activation of A(2A) receptors modulates Ca(2+) accumulation during hypoxia. This mechanism may play a role in regulating intracellular Ca(2+) homeostasis and cellular excitability during hypoxia. PMID:10926550

  10. Caffeine acts via A1 adenosine receptors to disrupt embryonic cardiac function.

    Directory of Open Access Journals (Sweden)

    Daniela L Buscariollo

    Full Text Available BACKGROUND: Evidence suggests that adenosine acts via cardiac A1 adenosine receptors (A1ARs to protect embryos against hypoxia. During embryogenesis, A1ARs are the dominant regulator of heart rate, and A1AR activation reduces heart rate. Adenosine action is inhibited by caffeine, which is widely consumed during pregnancy. In this study, we tested the hypothesis that caffeine influences developing embryos by altering cardiac function. METHODOLOGY/PRINCIPAL FINDINGS: Effects of caffeine and adenosine receptor-selective antagonists on heart rate were studied in vitro using whole murine embryos at E9.5 and isolated hearts at E12.5. Embryos were examined in room air (21% O(2 or hypoxic (2% O(2 conditions. Hypoxia decreased heart rates of E9.5 embryos by 15.8% and in E12.5 isolated hearts by 27.1%. In room air, caffeine (200 µM had no effect on E9.5 heart rates; however, caffeine increased heart rates at E12.5 by 37.7%. Caffeine abolished hypoxia-mediated bradycardia at E9.5 and blunted hypoxia-mediated bradycardia at E12.5. Real-time PCR analysis of RNA from isolated E9.5 and E12.5 hearts showed that A1AR and A2aAR genes were expressed at both ages. Treatment with adenosine receptor-selective antagonists revealed that SCH-58261 (A2aAR-specific antagonist had no affects on heart function, whereas DPCPX (A1AR-specific antagonist had effects similar to caffeine treatment at E9.5 and E12.5. At E12.5, embryonic hearts lacking A1AR expression (A1AR-/- had elevated heart rates compared to A1AR+/- littermates, A1AR-/- heart rates failed to decrease to levels comparable to those of controls. Caffeine did not significantly affect heart rates of A1AR-/- embryos. CONCLUSIONS/SIGNIFICANCE: These data show that caffeine alters embryonic cardiac function and disrupts the normal cardiac response to hypoxia through blockade of A1AR action. Our results raise concern for caffeine exposure during embryogenesis, particularly in pregnancies with increased risk of

  11. Stimulation of NTS A1 adenosine receptors differentially resets baroreflex control of regional sympathetic outputs.

    Science.gov (United States)

    Scislo, Tadeusz J; Ichinose, Tomoko K; O'Leary, Donal S

    2008-01-01

    Previously we showed that pressor and differential regional sympathoexcitatory responses (adrenal > renal >/= lumbar) evoked by stimulation of A(1) adenosine receptors located in the nucleus of the solitary tract (NTS) were attenuated/abolished by baroreceptor denervation or blockade of glutamatergic transmission in the NTS, suggesting A(1) receptor-elicited inhibition of glutamatergic transmission in baroreflex pathways. Therefore we tested the hypothesis that stimulation of NTS A(1) adenosine receptors differentially inhibits/resets baroreflex responses of preganglionic adrenal (pre-ASNA), renal (RSNA), and lumbar (LSNA) sympathetic nerve activity. In urethane-chloralose-anesthetized male Sprague-Dawley rats (n = 65) we compared baroreflex-response curves (iv nitroprusside and phenylephrine) evoked before and after bilateral microinjections into the NTS of A(1) adenosine receptor agonist (N(6)-cyclopentyladenosine, CPA; 0.033-330 pmol/50 nl). CPA evoked typical dose-dependent pressor and differential sympathoexcitatory responses and similarly shifted baroreflex curves for pre-ASNA, RSNA, and LSNA toward higher mean arterial pressure (MAP) in a dose-dependent manner; the maximal shifts were 52.6 +/- 2.8, 48.0 +/- 3.6, and 56.8 +/- 6.7 mmHg for pre-ASNA, RSNA, and LSNA, respectively. These shifts were not a result of simple baroreceptor resetting because they were two to three times greater than respective increases in baseline MAP evoked by CPA. Baroreflex curves for pre-ASNA were additionally shifted upward: the maximal increases of upper and lower plateaus were 41.8 +/- 16.4% and 45.3 +/- 8.7%, respectively. Maximal gain (%/mmHg) measured before vs. after CPA increased for pre-ASNA (3.0 +/- 0.6 vs. 4.9 +/- 1.3), decreased for RSNA (4.1 +/- 0.6 vs. 2.3 +/- 0.3), and remained unaltered for LSNA (2.1 +/- 0.2 vs. 2.0 +/- 0.1). Vehicle control did not alter the baroreflex curves. We conclude that the activation of NTS A(1) adenosine receptors differentially inhibits

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

    OpenAIRE

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

    2015-01-01

    This study was designed to investigate whether reduced adenosine formation linked to deficits in extracellular ATP hydrolysis by NTPDases contributes to detrusor neuromodulatory changes associated with bladder outlet obstruction in men with benign prostatic hyperplasia (BPH). The kinetics of ATP catabolism and adenosine formation as well as the role of P1 receptor agonists on muscle tension and nerve-evoked [3H]ACh release were evaluated in mucosal-denuded detrusor strips from BPH patients (n...

  13. Nucleoside-Derived Antagonists to A3 Adenosine Receptors Lower Mouse Intraocular Pressure and Act across Species

    OpenAIRE

    Wang, Zhao; Do, Chi Wai; Avila, Marcel Y.; Peterson-Yantorno, Kim; Stone, Richard A.; Gao, Zhan-Guo; Joshi, Bhalchandra; Besada, Pedro; Jeong, Lak Shin; Jacobson, Kenneth A.; Civan, Mortimer M.

    2009-01-01

    The purpose of the study was to determine whether novel, selective antagonists of human A3 adenosine receptors (ARs) derived from the A3-selective agonist Cl-IB-MECA lower intraocular pressure (IOP) and act across species. IOP was measured invasively with a micropipette by the Servo-Null Micropipette System (SNMS) and by non-invasive pneumotonometry during topical drug application. Antagonist efficacy was also assayed by measuring inhibition of adenosine-triggered shrinkage of native bovine n...

  14. Polyamidoamine (PAMAM) Dendrimer Conjugates of Clickable Agonists of the A3 Adenosine Receptor and Coactivation of the P2Y14 Receptor by a Tethered Nucleotide

    Energy Technology Data Exchange (ETDEWEB)

    Tosh, Dilip, K. [National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health; Yoo, Lena S. [National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health; Chinn, Moshe [National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health; Hong, Kunlun [ORNL; Kilbey, II, S Michael [ORNL; Barrett, Matthew O. [University of North Carolina School of Medicine; Fricks, Ingrid P. [University of North Carolina School of Medicine; Harden, T. Kendall [University of North Carolina School of Medicine; Jacobson, Kenneth A. [National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health

    2010-01-01

    We previously synthesized a series of potent and selective A{sub 3} adenosine receptor (AR) agonists (North-methanocarba nucleoside 5{prime}-uronamides) containing dialkyne groups on extended adenine C2 substituents. We coupled the distal alkyne of a 2-octadiynyl nucleoside by Cu(I)-catalyzed 'click' chemistry to azide-derivatized G4 (fourth-generation) PAMAM dendrimers to form triazoles. A{sub 3}AR activation was preserved in these multivalent conjugates, which bound with apparent Ki of 0.1-0.3 nM. They were substituted with nucleoside moieties, solely or in combination with water-solubilizing carboxylic acid groups derived from hexynoic acid. A comparison with various amide-linked dendrimers showed that triazole-linked conjugates displayed selectivity and enhanced A{sub 3}AR affinity. We prepared a PAMAM dendrimer containing equiproportioned peripheral azido and amino groups for conjugation of multiple ligands. A bifunctional conjugate activated both A{sub 3} and P2Y{sub 14} receptors (via amide-linked uridine-5{prime}-diphosphoglucuronic acid), with selectivity in comparison to other ARs and P2Y receptors. This is the first example of targeting two different GPCRs with the same dendrimer conjugate, which is intended for activation of heteromeric GPCR aggregates. Synergistic effects of activating multiple GPCRs with a single dendrimer conjugate might be useful in disease treatment.

  15. Cordycepin Increases Nonrapid Eye Movement Sleep via Adenosine Receptors in Rats

    Directory of Open Access Journals (Sweden)

    Zhenzhen Hu

    2013-01-01

    Full Text Available Cordycepin (3′-deoxyadenosine is a naturally occurring adenosine analogue and one of the bioactive constituents isolated from Cordyceps militaris/Cordyceps sinensis, species of the fungal genus Cordyceps. It has traditionally been a prized Chinese folk medicine for the human well-being. Because of similarity of chemical structure of adenosine, cordycepin has been focused on the diverse effects of the central nervous systems (CNSs, like sleep regulation. Therefore, this study was undertaken to know whether cordycepin increases the natural sleep in rats, and its effect is mediated by adenosine receptors (ARs. Sleep was recorded using electroencephalogram (EEG for 4 hours after oral administration of cordycepin in rats. Sleep architecture and EEG power spectra were analyzed. Cordycepin reduced sleep-wake cycles and increased nonrapid eye movement (NREM sleep. Interestingly, cordycepin increased θ (theta waves power density during NREM sleep. In addition, the protein levels of AR subtypes (A1, A2A, and A2B were increased after the administration of cordycepin, especially in the rat hypothalamus which plays an important role in sleep regulation. Therefore, we suggest that cordycepin increases theta waves power density during NREM sleep via nonspecific AR in rats. In addition, this experiment can provide basic evidence that cordycepin may be helpful for sleep-disturbed subjects.

  16. [Mast cells, their adenosine receptors and reactive oxygen species in chronic inflammatory pathologies of childhood].

    Science.gov (United States)

    Renke, Joanna; Popadiuk, Stefan; Wozniak, Michał; Szlagatys-Sidorkiewicz, Agnieszka; Hansdorfer-Korzon, Rita

    2006-01-01

    Mast cells were described by Erhlich at the end of XIX-th century. Their role was deeply investigated in asthma and allergy. The massive degranulation of mast cells in allergy can lead to anaphylactic shock. Recently, mast cells have been recognized again as a very interesting topic for investigation, due to their possible role in chronic inflammation. Moreover, through adenosine receptors, mast cells can be activated or inactivated. That is why these cells are regarded as a potential target of new drugs. It has been reported, that mast cells generate intracellular reactive oxygen species (ROS) in response to stimulation with divergent physiologically relevant stimulants. The intensification of ROS production may be measured by the level of carbonyl groups, as a marker of protein peroxidation. However, the role of mast cells in other than asthma diseases with chronic inflammation needs further investigation. It was found out that the information about mast cell distribution in colonic mucosa may serve as help in differentiation between inflammatory bowel disease and collagenous colitis. Moreover, its accumulation in focal active gastritis was confirmed in patients with Crohn's disease. An important role in regulation of inflammatory process seems to be reserved for adenosine receptors present on mastocytes. The activation of mast cells through the adenosine receptor is connected with 11-8 release, which stimulate the migration of leukocytes and oxidation reactions. The detection of mast cells in tissues should not be limited only to the simple histologic examination. It should be completed by the detection of products of degranulation, e.g. tryptase. This is the way to find out their actual function and state of activation. PMID:17203808

  17. Activation of NTS A2a adenosine receptors differentially resets baroreflex control of renal vs. adrenal sympathetic nerve activity.

    Science.gov (United States)

    Ichinose, Tomoko K; O'Leary, Donal S; Scislo, Tadeusz J

    2009-04-01

    The role of nucleus of solitary tract (NTS) A(2a) adenosine receptors in baroreflex mechanisms is controversial. Stimulation of these receptors releases glutamate within the NTS and elicits baroreflex-like decreases in mean arterial pressure (MAP), heart rate (HR), and renal sympathetic nerve activity (RSNA), whereas inhibition of these receptors attenuates HR baroreflex responses. In contrast, stimulation of NTS A(2a) adenosine receptors increases preganglionic adrenal sympathetic nerve activity (pre-ASNA), and the depressor and sympathoinhibitory responses are not markedly affected by sinoaortic denervation and blockade of NTS glutamatergic transmission. To elucidate the role of NTS A(2a) adenosine receptors in baroreflex function, we compared full baroreflex stimulus-response curves for HR, RSNA, and pre-ASNA (intravenous nitroprusside/phenylephrine) before and after bilateral NTS microinjections of selective adenosine A(2a) receptor agonist (CGS-21680; 2.0, 20 pmol/50 nl), selective A(2a) receptor antagonist (ZM-241385; 40 pmol/100 nl), and nonselective A(1) + A(2a) receptor antagonist (8-SPT; 1 nmol/100 nl) in urethane/alpha-chloralose anesthetized rats. Activation of A(2a) receptors decreased the range, upper plateau, and gain of baroreflex-response curves for RSNA, whereas these parameters all increased for pre-ASNA, consistent with direct effects of the agonist on regional sympathetic activity. However, no resetting of baroreflex-response curves along the MAP axis occurred despite the marked decreases in baseline MAP. The antagonists had no marked effects on baseline variables or baroreflex-response functions. We conclude that the activation of NTS A(2a) adenosine receptors differentially alters baroreflex control of HR, RSNA, and pre-ASNA mostly via non-baroreflex mechanism(s), and these receptors have virtually no tonic action on baroreflex control of these sympathetic outputs.

  18. Sliding tethered ligands add topological interactions to the toolbox of ligand-receptor design

    Science.gov (United States)

    Bauer, Martin; Kékicheff, Patrick; Iss, Jean; Fajolles, Christophe; Charitat, Thierry; Daillant, Jean; Marques, Carlos M.

    2015-09-01

    Adhesion in the biological realm is mediated by specific lock-and-key interactions between ligand-receptor pairs. These complementary moieties are ubiquitously anchored to substrates by tethers that control the interaction range and the mobility of the ligands and receptors, thus tuning the kinetics and strength of the binding events. Here we add sliding anchoring to the toolbox of ligand-receptor design by developing a family of tethered ligands for which the spacer can slide at the anchoring point. Our results show that this additional sliding degree of freedom changes the nature of the adhesive contact by extending the spatial range over which binding may sustain a significant force. By introducing sliding tethered ligands with self-regulating length, this work paves the way for the development of versatile and reusable bio-adhesive substrates with potential applications for drug delivery and tissue engineering.

  19. Ligand-specific conformational changes in the alpha1 glycine receptor ligand-binding domain

    DEFF Research Database (Denmark)

    Pless, Stephan Alexander; Lynch, Joseph W

    2009-01-01

    indicate that channel opening is accompanied by conformational rearrangements in both beta-sheets. In an attempt to resolve ligand-dependent movements in the ligand-binding domain, we employed voltage-clamp fluorometry on alpha1 glycine receptors to compare changes mediated by the agonist, glycine...... in the inner beta-sheet and pre-M1 domain that may be important for activation, desensitization, or both. In contrast, most labeled residues in loops C and F yielded fluorescence changes identical in magnitude for glycine and strychnine. A notable exception was H201C in loop C. This labeled residue responded...... differently to glycine and strychnine, thus underlining the importance of loop C in ligand discrimination. These results provide an important step toward mapping the domains crucial for ligand discrimination in the ligand-binding domain of glycine receptors and possibly other Cys loop receptors....

  20. Adenosine receptors located in the NTS contribute to renal sympathoinhibition during hypotensive phase of severe hemorrhage in anesthetized rats.

    Science.gov (United States)

    Scislo, Tadeusz J; O'Leary, Donal S

    2006-11-01

    Stimulation of nucleus of the solitary tract (NTS) A(2a)-adenosine receptors elicits cardiovascular responses quite similar to those observed with rapid, severe hemorrhage, including bradycardia, hypotension, and inhibition of renal but activation of preganglionic adrenal sympathetic nerve activity (RSNA and pre-ASNA, respectively). Because adenosine levels in the central nervous system increase during severe hemorrhage, we investigated to what extent these responses to hemorrhage may be due to activation of NTS adenosine receptors. In urethane- and alpha-chloralose-anesthetized male Sprague-Dawley rats, rapid hemorrhage was performed before and after bilateral nonselective or selective blockade of NTS adenosine-receptor subtypes [A(1)- and A(2a)-adenosine-receptor antagonist 8-(p-sulfophenyl)theophylline (1 nmol/100 nl) and A(2a)-receptor antagonist ZM-241385 (40 pmol/100 nl)]. The nonselective blockade reversed the response in RSNA (-21.0 +/- 9.6 Delta% vs. +7.3 +/- 5.7 Delta%) (where Delta% is averaged percent change from baseline) and attenuated the average heart rate response (change of -14.8 +/- 4.8 vs. -4.4 +/- 3.4 beats/min). The selective blockade attenuated the RSNA response (-30.4 +/- 5.2 Delta% vs. -11.1 +/- 7.7 Delta%) and tended to attenuate heart rate response (change of -27.5 +/- 5.3 vs. -15.8 +/- 8.2 beats/min). Microinjection of vehicle (100 nl) had no significant effect on the responses. The hemorrhage-induced increases in pre-ASNA remained unchanged with either adenosine-receptor antagonist. We conclude that adenosine operating in the NTS via A(2a) and possibly A(1) receptors may contribute to posthemorrhagic sympathoinhibition of RSNA but not to the sympathoactivation of pre-ASNA. The differential effects of NTS adenosine receptors on RSNA vs. pre-ASNA responses to hemorrhage supports the hypothesis that these receptors are differentially located/expressed on NTS neurons/synaptic terminals controlling different sympathetic outputs.

  1. Cerebral A{sub 1} adenosine receptors (A{sub 1}AR) in liver cirrhosis

    Energy Technology Data Exchange (ETDEWEB)

    Boy, Christian [Research Centre Juelich, Brain Imaging Centre West, Institute of Medicine, Juelich (Germany); University Hospital Essen, Department of Nuclear Medicine, Essen (Germany); Meyer, Philipp T. [Research Centre Juelich, Brain Imaging Centre West, Institute of Medicine, Juelich (Germany); University Hospital Aachen, Department of Nuclear Medicine, Aachen (Germany); Kircheis, Gerald; Haussinger, Dieter [University of Duesseldorf, Clinic for Gastroenterology, Hepatology and Infectiology, Duesseldorf (Germany); Holschbach, Marcus H.; Coenen, Heinz H. [Research Centre Juelich, Institute of Nuclear Chemistry, Juelich (Germany); Herzog, Hans; Elmenhorst, David [Research Centre Juelich, Brain Imaging Centre West, Institute of Medicine, Juelich (Germany); Kaiser, Hans J. [University Hospital Aachen, Department of Nuclear Medicine, Aachen (Germany); Zilles, Karl [Research Centre Juelich, Brain Imaging Centre West, Institute of Medicine, Juelich (Germany); C. and O. Vogt Institute of Brain Research, Duesseldorf (Germany); Bauer, Andreas [Research Centre Juelich, Brain Imaging Centre West, Institute of Medicine, Juelich (Germany); University of Duesseldorf, Department of Neurology, Duesseldorf (Germany)

    2008-03-15

    The cerebral mechanisms underlying hepatic encephalopathy (HE) are poorly understood. Adenosine, a neuromodulator that pre- and postsynaptically modulates neuronal excitability and release of classical neurotransmitters via A{sub 1} adenosine receptors (A{sub 1}AR), is likely to be involved. The present study investigates changes of cerebral A{sub 1}AR binding in cirrhotic patients by means of positron emission tomography (PET) and [{sup 18}F]CPFPX, a novel selective A{sub 1}AR antagonist. PET was performed in cirrhotic patients (n = 10) and healthy volunteers (n = 10). Quantification of in vivo receptor density was done by Logan's non-invasive graphical analysis (pons as reference region). The outcome parameter was the apparent binding potential (aBP, proportional to B{sub max}/K{sub D}). Cortical and subcortical regions showed lower A{sub 1}AR binding in cirrhotic patients than in controls. The aBP changes reached statistical significance vs healthy controls (p < 0.05, U test with Bonferroni-Holm adjustment for multiple comparisons) in cingulate cortex (-50.0%), precentral gyrus (-40.9%), postcentral gyrus (-38.6%), insular cortex (-38.6%), thalamus (-32.9%), parietal cortex (-31.7%), frontal cortex (-28.6), lateral temporal cortex (-28.2%), orbitofrontal cortex (-27.9%), occipital cortex (-24.6), putamen (-22.7%) and mesial temporal lobe (-22.4%). Regional cerebral adenosinergic neuromodulation is heterogeneously altered in cirrhotic patients. The decrease of cerebral A{sub 1}AR binding may further aggravate neurotransmitter imbalance at the synaptic cleft in cirrhosis and hepatic encephalopathy. Different pathomechanisms may account for these alterations including decrease of A{sub 1}AR density or affinity, as well as blockade of the A{sub 1}AR by endogenous adenosine or exogenous xanthines. (orig.)

  2. Insight into the binding mode and the structural features of the pyrimidine derivatives as human A2A adenosine receptor antagonists.

    Science.gov (United States)

    Zhang, Lihui; Liu, Tianjun; Wang, Xia; Wang, Jinan; Li, Guohui; Li, Yan; Yang, Ling; Wang, Yonghua

    2014-01-01

    The interaction of 278 monocyclic and bicyclic pyrimidine derivatives with human A2A adenosine receptor (AR) was investigated by employing molecular dynamics, thermodynamic analysis and three-dimensional quantitative structure-activity relationship (3D-QSAR) approaches. The binding analysis reveals that the pyrimidine derivatives are anchored in TM2, 3, 5, 6 and 7 of A2A AR by the aromatic stacking and hydrogen bonding interactions. The key residues involving Phe168, Glu169, and Asn253 stabilize the monocyclic and bicyclic cores of inhibitors. The thermodynamic analysis by molecular mechanics/Poisson Boltzmann surface area (MM-PBSA) approach also confirms the reasonableness of the binding modes. In addition, the ligand-/receptor-based comparative molecular similarity indices analysis (CoMSIA) models of high statistical significance were generated and the resulting contour maps correlate well with the structural features of the antagonists essential for high A2A AR affinity. A minor/bulky group with negative charge at C2/C6 of pyrimidine ring respectively enhances the activity for all these pyrimidine derivatives. Particularly, the higher electron density of the ring in the bicyclic derivatives, the more potent the antagonists. The obatined results might be helpful in rational design of novel candidate of A2A adenosine receptor antagonist for treatment of Parkinson's disease. PMID:23665268

  3. Biotechnological Fluorescent Ligands of the Bradykinin B1 Receptor: Protein Ligands for a Peptide Receptor.

    Directory of Open Access Journals (Sweden)

    Xavier Charest-Morin

    Full Text Available The bradykinin (BK B1 receptor (B1R is a peculiar G protein coupled receptor that is strongly regulated to the point of being inducible in immunopathology. Limited clinical evidence suggests that its expression in peripheral blood mononuclear cells is a biomarker of active inflammatory states. In an effort to develop a novel imaging/diagnostic tool, we report the rational design and testing of a fusion protein that is a ligand of the human B1R but not likely to label peptidases. This ligand is composed of a fluorescent protein (FP (enhanced green FP [EGFP] or mCherry prolonged at its N-terminus by a spacer peptide and a classical peptide agonist or antagonist (des-Arg9-BK, [Leu8]des-Arg9-BK, respectively. The design of the spacer-ligand joint peptide was validated by a competition assay for [3H]Lys-des-Arg9-BK binding to the human B1R applied to 4 synthetic peptides of 18 or 19 residues. The labeling of B1R-expressing cells with EGFP or mCherry fused with 7 of such peptides was performed in parallel (microscopy. Both assays indicated that the best design was FP-(Asn-Glyn-Lys-des-Arg9-BK; n = 15 was superior to n = 5, suggesting benefits from minimizing steric hindrance between the FP and the receptor. Cell labeling concerned mostly plasma membranes and was inhibited by a B1R antagonist. EGFP-(Asn-Gly15-Lys-des-Arg9-BK competed for the binding of [3H]Lys-des-Arg9-BK to human recombinant B1R, being only 10-fold less potent than the unlabeled form of Lys-des-Arg9-BK to do so. The fusion protein did not label HEK 293a cells expressing recombinant human BK B2 receptors or angiotensin converting enzyme. This study identifies a modular C-terminal sequence that can be adapted to protein cargoes, conferring high affinity for the BK B1R, with possible applications in diagnostic cytofluorometry, histology and drug delivery (e.g., in oncology.

  4. Cross sectional PET study of cerebral adenosine A1 receptors in premanifest and manifest Huntington's disease

    International Nuclear Information System (INIS)

    To study cerebral adenosine receptors (AR) in premanifest and manifest stages of Huntington's disease (HD). We quantified the cerebral binding potential (BPND) of the A1AR in carriers of the HD CAG trinucleotide repeat expansion using the radioligand [18 F]CPFPX and PET. Four groups were investigated: (i) premanifest individuals far (preHD-A; n = 7) or (ii) near (preHD-B; n = 6) to the predicted symptom onset, (iii) manifest HD patients (n = 8), and (iv) controls (n = 36). Cerebral A1AR values of preHD-A subjects were generally higher than those of controls (by up to 31 %, p 1AR BPND was observed to the levels of controls in preHD-B and undercutting controls in manifest HD by down to 25 %, p 1AR BPND and years to onset. Before onset of HD, the assumed annual rates of change of A1AR density were -1.2 % in the caudatus, -1.7 % in the thalamus and -3.4 % in the amygdala, while the corresponding volume losses amounted to 0.6 %, 0.1 % and 0.2 %, respectively. Adenosine receptors switch from supra to subnormal levels during phenoconversion of HD. This differential regulation may play a role in the pathophysiology of altered energy metabolism. (orig.)

  5. CF102 an A3 Adenosine Receptor Agonist Mediates Anti-Tumor and Anti-Inflammatory Effects in the Liver

    OpenAIRE

    Cohen, S.; Stemmer, S M; ZOZULYA, G.; Ochaion, A.; PATOKA, R.; Barer, F.; BAR-YEHUDA, S.; RATH-WOLFSON, L.; Jacobson, K. A.; Fishman, P

    2011-01-01

    The Gi protein-associated A3 adenosine receptor (A3AR) is a member of the adenosine receptor family. Selective agonists at the A3AR, such as CF101 and CF102 were found to induce anti-inflammatory and anti-cancer effects. In this study, we examined the differential effect of CF102 in pathological conditions of the liver. The anti-inflammatory protective effect of CF101 was tested in a model of liver inflammation induced by Concanavalin A (Con. A) and the anti-cancer effect of CF102 was examine...

  6. The Effects of Nucleus Accumbens μ-opioid and Adenosine 2A Receptor Stimulation and Blockade on Instrumental Learning

    OpenAIRE

    Clissold, Kara A.; Pratt, Wayne E.

    2014-01-01

    Prior research has shown that glutamate and dopamine receptors in the nucleus accumbens (NAcc) core are critical for the learning of an instrumental response for food reinforcement. It has also been demonstrated that μ-opioid and adenosine A2A receptors within the NAcc impact feeding and motivational processes. In these experiments, we examined the potential roles of NAcc μ-opioid and A2A receptors on instrumental learning and performance. Sprague-Dawley rats were food restricted and trained ...

  7. Preparation and first evaluation of [18F]FE-SUPPY: a new PET tracer for the adenosine A3 receptor

    International Nuclear Information System (INIS)

    Introduction: Changes of the adenosine A3 receptor subtype (A3AR) expression have been shown in a variety of pathologies, especially neurological and affective disorders, cardiac diseases and oncological and inflammation processes. Recently, 5-(2-fluoroethyl) 2,4-diethyl-3-(ethylsulfanylcarbonyl)-6-phenylpyridine-5-carboxylate (FE-SUPPY) was presented as a high-affinity ligand for the A3AR with good selectivity. Our aims were the development of a suitable labeling precursor, the establishment of a reliable radiosynthesis for the fluorine-18-labeled analogue [18F]FE-SUPPY and a first evaluation of [18F]FE-SUPPY in rats. Methods: [18F]FE-SUPPY was prepared in a feasible and reliable manner by radiofluorination of the corresponding tosylated precursor. Biodistribution was carried out in rats, and organs were removed and counted. Autoradiography was performed on rat brain slices in the presence or absence of 2-Cl-IB-MECA. Results: Overall yields and radiochemical purity were sufficient for further preclinical and clinical applications. The uptake pattern of [18F]FE-SUPPY found in rats mainly followed the described mRNA distribution pattern of the A3AR. Specific uptake in brain was demonstrated by blocking with a selective A3AR agonist. Conclusion: We conclude that [18F]FE-SUPPY has the potential to serve as the first positron emission tomography tracer for the A3AR

  8. The imidazoline receptors and ligands in pain modulation

    Directory of Open Access Journals (Sweden)

    Nurcan Bektas

    2015-01-01

    Full Text Available Pain is an unpleasant experience and effects daily routine negatively. Although there are various drugs, many of them are not entirely successful in relieving pain, since pain modulation is a complex process involving numerous mediators and receptors. Therefore, it is a rational approach to identify the factors involved in the complex process and develop new agents that act on these pain producing mechanisms. In this respect, the involvement of the imidazoline receptors in pain modulation has drawn attention in recent years. In this review, it is aimed to focus on the imidazoline receptors and their ligands which contribute to the pain modulation. It is demonstrated that imidazoline-2 (I2 receptors are steady new drug targets for analgesics. Even if the mechanism of I2receptor is not well known in the modulation of pain, it is known that it plays a role in tonic and chronic pain but not in acute phasic pain. Moreover, the I2receptor ligands increase the analgesic effects of opioids in both acute and chronic pain and prevent the development of opioid tolerance. So, they are valuable for the chronic pain treatment and also therapeutic coadjuvants in the management of chronic pain with opiate drugs due to the attenuation of opioid tolerance and addiction. Thus, the use of the ligands which bind to the imidazoline receptors is an effective strategy for relieving pain. This educational forum exhibits the role of imidazoline receptors and ligands in pain process by utilizing experimental studies.

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

    Directory of Open Access Journals (Sweden)

    Alessandro Tozzi

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

  10. Functional expression of adenosine A2A and A3 receptors in the mouse dendritic cell line XS-106.

    Science.gov (United States)

    Dickenson, John M; Reeder, Steve; Rees, Bob; Alexander, Steve; Kendall, Dave

    2003-08-01

    There is increasing evidence to suggest that adenosine receptors can modulate the function of cells involved in the immune system. For example, human dendritic cells derived from blood monocytes have recently been described to express functional adenosine A1, A2A and A3 receptors. Therefore, in the present study, we have investigated whether the recently established murine dendritic cell line XS-106 expresses functional adenosine receptors. The selective adenosine A3 receptor agonist 1-[2-chloro-6[[(3-iodophenyl)methyl]amino]-9H-purin-9-yl]-1-deoxy-N-methyl-beta-D-ribofuranuronamide (2-Cl-IB-MECA) inhibited forskolin-mediated [3H]cyclic AMP accumulation and stimulated concentration-dependent increases in p42/p44 mitogen-activated protein kinase (MAPK) phosphorylation. The selective adenosine A2A receptor agonist 4-[2-[[-6-amino-9-(N-ethyl-beta-D-ribofuranuronamidosyl)-9H-purin-2-yl]amino]ethyl]benzene-propanoic acid (CGS 21680) stimulated a robust increase in [3H]cyclic AMP accumulation and p42/p44 MAPK phosphorylation. In contrast, the selective adenosine A1 receptor agonist CPA (N6-cyclopentyladenosine) did not inhibit forskolin-mediated [3H]cyclic AMP accumulation or stimulate increases in p42/p44 MAPK phosphorylation. These observations suggest that XS-106 cells express functional adenosine A2A and A3 receptors. The non-selective adenosine receptor agonist 5'-N-ethylcarboxamidoadenosine (NECA) inhibited lipopolysaccharide-induced tumour necrosis factor-alpha (TNF-alpha) release from XS-106 cells in a concentration-dependent fashion. Furthermore, treatment with Cl-IB-MECA (1 microM) or CGS 21680 (1 microM) alone produced a partial inhibition of lipopolysaccharide-induced TNF-alpha release (when compared to NECA), whereas a combination of both agonists resulted in the inhibition of TNF-alpha release comparable to that observed with NECA alone. Treatment of cells with the adenosine A2A receptor selective antagonists 4-(2-[7-amino-2-(2-furyl)[1,2,4]triazolo[2,3-a

  11. Adenosine is required for sustained inflammasome activation via the A2A receptor and the HIF-1α pathway

    Science.gov (United States)

    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.

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

    OpenAIRE

    Cunha, Rodrigo A.

    2008-01-01

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

  13. Overexpression of adenosine A2A receptors in rats: effects on depression, locomotion and anxiety

    Directory of Open Access Journals (Sweden)

    Joana E Coelho

    2014-06-01

    Full Text Available Adenosine A2A receptors (A2AR are a sub-type of receptors enriched in basal ganglia, activated by the neuromodulator adenosine, which interact with dopamine D2 receptors. Although this reciprocal antagonistic interaction is well established in motor function, the outcome in dopamine-related behaviors remains uncertain, in particular in depression and anxiety. We have demonstrated an upsurge of A2AR associated to aging and chronic stress. Furthermore, Alzheimer’s disease patients present A2AR accumulation in cortical areas together with depressive signs. We now tested the impact of overexpressing A2AR in forebrain neurons on dopamine related behavior, namely depression. Adult male rats overexpressing human A2AR under the control of CaMKII promoter [Tg(CaMKII-hA2AR] and aged-matched wild-types (WT of the same strain (Sprague-Dawley were studied. The forced swimming test (FST, sucrose preference test (SPT and the open-field test (OFT were performed to evaluate behavioral despair, anhedonia, locomotion and anxiety. Tg(CaMKII-hA2AR animals spent more time floating and less time swimming in the FST and presented a decreased sucrose preference at 48h in the SPT. They also covered higher distances in the OFT and spent more time in the central zone than the WT. The results indicate that Tg(CaMKII-hA2AR rats exhibit depressive-like behavior, hyperlocomotion and altered exploratory behavior. This A2AR overexpression may explain the depressive signs found in aging, chronic stress and Alzheimer’s disease.

  14. Kinetics of Receptor-Ligand Interactions in Immune Responses

    Institute of Scientific and Technical Information of China (English)

    Mian Long; Shouqin Lü; Ganyun Sun

    2006-01-01

    Receptor-ligand interactions in blood flow are crucial to initiate the biological processes as inflammatory cascade,platelet thrombosis, as well as tumor metastasis. To mediate cell adhesions, the interacting receptors and ligands must be anchored onto two apposing surfaces of two cells or a cell and a substratum, i.e., the two-dimensional (2D) binding, which is different from the binding of a soluble ligand in fluid phase to a receptor, i.e., three-dimensional (3D) binding. While numerous works have been focused on 3D kinetics of receptor-ligand interactions in immune systems, 2D kinetics and its regulations have less been understood, since no theoretical framework and experimental assays have been established until 1993. Not only does the molecular structure dominate 2D binding kinetics, but the shear force in blood flow also regulates cell adhesions mediated by interacting receptors and ligands. Here we provided the overview of current progresses in 2D bindings and regulations. Relevant issues of theoretical frameworks, experimental measurements, kinetic rates and binding affinities, and force regulations,were discussed.

  15. Ultraslow Water-Mediated Transmembrane Interactions Regulate the Activation of A2A Adenosine Receptor.

    Science.gov (United States)

    Lee, Yoonji; Kim, Songmi; Choi, Sun; Hyeon, Changbong

    2016-09-20

    Water molecules inside a G-protein coupled receptor (GPCR) have recently been spotlighted in a series of crystal structures. To decipher the dynamics and functional roles of internal water molecules in GPCR activity, we studied the A2A adenosine receptor using microsecond molecular-dynamics simulations. Our study finds that the amount of water flux across the transmembrane (TM) domain varies depending on the receptor state, and that the water molecules of the TM channel in the active state flow three times more slowly than those in the inactive state. Depending on the location in solvent-protein interface as well as the receptor state, the average residence time of water in each residue varies from ∼O(10(2)) ps to ∼O(10(2)) ns. Especially, water molecules, exhibiting ultraslow relaxation (∼O(10(2)) ns) in the active state, are found around the microswitch residues that are considered activity hotspots for GPCR function. A continuous allosteric network spanning the TM domain, arising from water-mediated contacts, is unique in the active state, underscoring the importance of slow water molecules in the activation of GPCRs. PMID:27653477

  16. Synthesis and Properties of a New Water-Soluble Prodrug of the Adenosine A2A Receptor Antagonist MSX-2

    Directory of Open Access Journals (Sweden)

    Christa E. Müller

    2008-02-01

    Full Text Available The compound L-valine-3-{8-[(E-2-[3-methoxyphenylethenyl]-7-methyl-1-propargylxanthine-3-yl}propyl ester hydrochloride (MSX-4 was synthesized as an aminoacid ester prodrug of the adenosine A2A receptor antagonist MSX-2. It was found to bestable in artificial gastric acid, but readily cleaved by pig liver esterase.

  17. Role of Adenosine Receptor(s) in the Control of Vascular Tone in the Mouse Pudendal Artery.

    Science.gov (United States)

    Labazi, Hicham; Tilley, Stephen L; Ledent, Catherine; Mustafa, S Jamal

    2016-03-01

    Activation of adenosine receptors (ARs) has been implicated in the modulation of renal and cardiovascular systems, as well as erectile functions. Recent studies suggest that adenosine-mediated regulation of erectile function is mainly mediated through A2BAR activation. However, no studies have been conducted to determine the contribution of AR subtype in the regulation of the vascular tone of the pudendal artery (PA), the major artery supplying and controlling blood flow to the penis. Our aim was to characterize the contribution of AR subtypes and identify signaling mechanisms involved in adenosine-mediated vascular tone regulation in the PA. We used a DMT wire myograph for muscle tension measurements in isolated PAs from wild-type, A2AAR knockout, A2BAR knockout, and A2A/A2BAR double-knockout mice. Real-time reverse transcription-polymerase chain reaction was used to determine the expression of the AR subtypes. Data from our pharmacologic and genetic approaches suggest that AR activation-mediated vasodilation in the PA is mediated by both the A2AAR and A2BAR, whereas neither the A1AR nor A3AR play a role in vascular tone regulation of the PA. In addition, we showed that A2AAR- and A2BAR-mediated vasorelaxation requires activation of nitric oxide and potassium channels; however, only the A2AAR-mediated response requires protein kinase A activation. Our data are complemented by mRNA expression showing the expression of all AR subtypes with the exception of the A3AR. AR signaling in the PA may play an important role in mediating erection and represent a promising therapeutic option for the treatment of erectile dysfunction. PMID:26718241

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

    Science.gov (United States)

    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.

  19. Activation of adenosine receptor potentiates the anticonvulsant effect of phenytoin against amygdala kindled seizures.

    Science.gov (United States)

    Sun, Zhen; Zhong, Xiao-Ling; Zong, Yu; Wu, Zhong-Chen; Zhang, Qun; Yu, Jin-Tai; Tan, Lan

    2015-01-01

    Drug resistance in epilepsy is considered as a complicated and multifactorial problem. Poor penetration of antiepileptic drugs (AEDs) across blood-brain barrier (BBB) into the brain, which results in insufficient level of the drugs at the targeted brain region, has been discussed as one mechanism contributing to pharmacoresistance of epilepsies. Therefore, modulating permeability of BBB is the effective treatment strategy since it facilitates the entry of AEDs into the central nervous system (CNS). Recently, signaling through receptors for the adenosine has been identified as a potent modulator of BBB permeability. This paper aimed to investigate the effects of auxiliary application of adenosine receptor (AR) agonist on amygdala-kindled seizures in adult male Wistar rats. When fully kindled seizures were achieved by daily electrical stimulation of the amygdala, rats were randomly divided into three groups: control, phenytoin, and phenytoin (PHT)+5'-N-ethylcarboxamidoadenosine (NECA) groups. NECA (0.08 mg/kg, i.v.) was applied to the PHT+NECA group after the administration of PHT (75 mg/kg, i.p. on the first day; 50mg/kg, i.p. on the following 9 days). Intravenous infusion of NECA resulted in a significant increase in brain PHT levels as compared with the PHT treatment alone. On the other hand, the auxiliary application of NECA dramatically decreased the frequency of generalized seizures and seizure stage, shortened duration of afterdischarge and generalized seizures, as well as the elevated the afterdischarge threshold and generalized seizures threshold. Our study demonstrated that auxiliary application of AR agonist enhanced brain antiepileptic drug levels and strengthened the anticonvulsant properties of PHT against amygdala kindled seizures.

  20. A/sub 1/ and A/sub 2/ adenosine receptor regulation of erythropoietin production

    Energy Technology Data Exchange (ETDEWEB)

    Ueno, M.; Brookins, J.; Beckman, B.; Fisher, J.W.

    1988-01-01

    The effects of adenosine (ADE) and ADE agonists on erythropoietin (Ep) production were determined using percent (%) /sup 59/Fe incorporation in red cells of exhypoxic polycythemic mice. The hemisulfate salt of ADE produced a significant increase in % /sup 59/Fe incorporation in response to hypoxia in concentrations of 400 to 1600 nmol/kg/day. 5'-N-ethyl-carboxamideadenosine (NECA), a selective A/sub 2/ receptor agonist, increased radioiron incorporation in a dose-dependent manner. In contrast, N/sup 6/-cyclohexyladenosine (CHA), a selective A/sub 1/ receptor agonist, did not affect radioiron incorporation in concentrations up to 1600 nmol/kg/day. Albuterol, a beta 2-adrenergic agonist, enhanced % /sup 59/Fe incorporation in polycythemic mice and low doses of CHA, which were not effective alone on % /sup 59/Fe incorporation in polycythemic mice exposed to hypoxia, inhibited the enhancement in radioiron induced by albuterol plus hypoxia. Theophylline, a well-known antagonist of ADE receptors, blocked the ADE and NECA enhancement in radioiron incorporation at a dose of theophylline alone which produced only a slight enhancement of % /sup 59/Fe incorporation.

  1. Triazoloquinazolines as Human A3 Adenosine Receptor Antagonists: A QSAR Study

    Directory of Open Access Journals (Sweden)

    Dae-Sil Lee

    2006-11-01

    Full Text Available Multiple linear regression analysis was performed on the quantitative structure-activity relationships (QSAR of the triazoloquinazoline adenosine antagonists for human A3receptors. The data set used for the QSAR analysis encompassed the activities of 33triazoloquinazoline derivatives and 72 physicochemical descriptors. A template moleculewas derived using the known molecular structure for one of the compounds when bound tothe human A2B receptor, in which the amide bond was in a cis-conformation. All the testcompounds were aligned to the template molecule. In order to identify a reasonable QSARequation to describe the data set, we developed a multiple linear regression program thatexamined every possible combination of descriptors. The QSAR equation derived from thisanalysis indicates that the spatial and electronic effects is greater than that of hydrophobiceffects in binding of the antagonists to the human A3 receptor. It also predicts that a largesterimol length parameter is advantageous to activity, whereas large sterimol widthparameters and fractional positive partial surface areas are nonadvatageous.

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

    OpenAIRE

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

    2012-01-01

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

  3. Adenosine and the adenosine A2A receptor agonist, CGS21680, upregulate CD39 and CD73 expression through E2F-1 and CREB in regulatory T cells isolated from septic mice.

    Science.gov (United States)

    Bao, Rui; Shui, Xianqi; Hou, Jiong; Li, Jinbao; Deng, Xiaoming; Zhu, Xiaoyan; Yang, Tao

    2016-09-01

    The number of regulatory T cells (Treg cells) and the expression of ectonucleoside triphosphate diphosphohydrolase 1 (ENTPD1; also known as CD39) and 5'-ectonucleotidase (NT5E; also known as CD73) on the Treg cell surface are increased during sepsis. In this study, to determine the factors leading to the high expression of CD39 and CD73, and the regulation of the CD39/CD73/adenosine pathway in Treg cells under septic conditions, we constructed a mouse model of sepsis and separated the Treg cells using a flow cytometer. The Treg cells isolated from the peritoneal lavage and splenocytes of the mice were treated with adenosine or the specific adenosine A2A receptor agonist, CGS21680, and were transfected with specific siRNA targeting E2F transcription factor 1 (E2F-1) or cyclic adenosine monophosphate (cAMP) response element-binding protein (CREB), which are predicted transcription regulatory factors of CD39 or CD73. The regulatory relationships among these factors were then determined by western blot analysis and dual-luciferase reporter assay. In addition, changes in adenosine metabolism were measured in the treated cells. The results revealed that adenosine and CGS21680 significantly upregulated CD39 and CD73 expression (PTreg cell surface during sepsis. Adenosine and its A2A receptor agonist served as the signal transducer factors of the CD39/CD73/adenosine pathway, accelerating adenosine generation. Our study may benefit further research on adenosine metabolism for the treatment of sepsis. PMID:27430240

  4. Fluorescent ligands for studying neuropeptide receptors by confocal microscopy

    Directory of Open Access Journals (Sweden)

    Beaudet A.

    1998-01-01

    Full Text Available This paper reviews the use of confocal microscopy as it pertains to the identification of G-protein coupled receptors and the study of their dynamic properties in cell cultures and in mammalian brain following their tagging with specific fluorescent ligands. Principles that should guide the choice of suitable ligands and fluorophores are discussed. Examples are provided from the work carried out in the authors' laboratory using custom synthetized fluoresceinylated or BODIPY-tagged bioactive peptides. The results show that confocal microscopic detection of specifically bound fluorescent ligands permits high resolution appraisal of neuropeptide receptor distribution both in cell culture and in brain sections. Within the framework of time course experiments, it also allows for a dynamic assessment of the internalization and subsequent intracellular trafficking of bound fluorescent molecules. Thus, it was found that neurotensin, somatostatin and mu- and delta-selective opioid peptides are internalized in a receptor-dependent fashion and according to receptor-specific patterns into their target cells. In the case of neurotensin, this internalization process was found to be clathrin-mediated, to proceed through classical endosomal pathways and, in neurons, to result in a mobilization of newly formed endosomes from neural processes to nerve cell bodies and from the periphery of cell bodies towards the perinuclear zone. These mechanisms are likely to play an important role for ligand inactivation, receptor regulation and perhaps also transmembrane signaling.

  5. Syntheses of oxysterol receptors'(LXRs) ligands

    Institute of Scientific and Technical Information of China (English)

    2003-01-01

    The LXRs' agonist, 24S,25-epoxycholesterol 1, was synthesized stereoselectively (100% d.e.) in 56% overall yield from methyl hyodeoxycholanate 4 in 9 steps with desmosterol acetate 11 as the key intermediate and the modified Sharpless asymmetric dihydroxylation as the key step. The LXR? subtype selective agonist 5α,6α:24S,25-diepoxycho- lesterol 2 and the novel LXRs' ligand 5β,6β:24S,25-diepo- xycholesterol 3 were also synthesized from 1.

  6. Stimulation of NTS A1 adenosine receptors evokes counteracting effects on hindlimb vasculature.

    Science.gov (United States)

    McClure, Joseph M; O'Leary, Donal S; Scislo, Tadeusz J

    2005-12-01

    Our previous studies concluded that stimulation of the nucleus of the solitary tract (NTS) A2a receptors evokes preferential hindlimb vasodilation mainly via inducing increases in preganglionic sympathetic nerve activity (pre-ASNA) directed to the adrenal medulla. This increase in pre-ASNA causes the release of epinephrine and subsequent activation of beta-adrenergic receptors that are preferentially located in the skeletal muscle vasculature. Selective activation of NTS A1 adenosine receptors evokes variable, mostly pressor effects and increases pre-ASNA, as well as lumbar sympathetic activity, which is directed to the hindlimb. These counteracting factors may have opposite effects on the hindlimb vasculature resulting in mixed vascular responses. Therefore, in chloralose-urethane-anesthetized rats, we evaluated the contribution of vasodilator versus vasoconstrictor effects of stimulation of NTS A1 receptors on the hindlimb vasculature. We compared the changes in iliac vascular conductance evoked by microinejctions into the NTS of the selective A1 receptor agonist N6-cyclopentyladenosine (330 pmol in 50 nl volume) in intact animals with the responses evoked after beta-adrenergic blockade, bilateral adrenalectomy, bilateral lumbar sympathectomy, and combined adrenalectomy + lumbar sympathectomy. In intact animals, stimulation of NTS A1 receptors evoked variable effects: increases and decreases in mean arterial pressure and iliac conductance with prevailing pressor and vasoconstrictor effects. Peripheral beta-adrenergic receptor blockade and bilateral adrenalectomy eliminated the depressor component of the responses, markedly potentiated iliac vasoconstriction, and tended to increase the pressor responses. Lumbar sympathectomy tended to decrease the pressor and vasoconstrictor responses. After bilateral adrenalectomy plus lumbar sympathectomy, a marked vasoconstriction in iliac vascular bed still persisted, suggesting that the vasoconstrictor component of the

  7. Inside job: ligand-receptor pharmacology beneath the plasma membrane

    Science.gov (United States)

    Babcock, Joseph J; Li, Min

    2013-01-01

    Most drugs acting on the cell surface receptors are membrane permeable and thus able to engage their target proteins in different subcellular compartments. However, these drugs' effects on cell surface receptors have historically been studied on the plasma membrane alone. Increasing evidence suggests that small molecules may also modulate their targeted receptors through membrane trafficking or organelle-localized signaling inside the cell. These additional modes of interaction have been reported for functionally diverse ligands of GPCRs, ion channels, and transporters. Such intracellular drug-target engagements affect cell surface expression. Concurrent intracellular and cell surface signaling may also increase the complexity and therapeutic opportunities of small molecule modulation. Here we discuss examples of ligand-receptor interactions that are present in both intra- and extracellular sites, and the potential therapeutic opportunities presented by this phenomenon. PMID:23685953

  8. Inside job: ligand-receptor pharmacology beneath the plasma membrane

    Institute of Scientific and Technical Information of China (English)

    Joseph J BABCOCK; Min LI

    2013-01-01

    Most drugs acting on the cell surface receptors are membrane permeable and thus able to engage their target proteins in different subcellular compartments.However,these drugs' effects on cell surface receptors have historically been studied on the plasma membrane alone.Increasing evidence suggests that small molecules may also modulate their targeted receptors through membrane trafficking or organelle-localized signaling inside the cell.These additional modes of interaction have been reported for functionally diverse ligands of GPCRs,ion channels,and transporters.Such intracellular drug-target engagements affect cell surface expression.Concurrent intracellular and cell surface signaling may also increase the complexity and therapeutic opportunities of small molecule modulation.Here we discuss examples of ligand-receptor interactions that are present in both intra- and extracellular sites,and the potential therapeutic opportunities presented by this phenomenon.

  9. Novel retinoic acid receptor ligands in Xenopus embryos.

    OpenAIRE

    Blumberg, B; Bolado, J; Derguini, F; Craig, A G; Moreno, T A; Chakravarti, D; Heyman, R A; Buck, J.; Evans, R M

    1996-01-01

    Retinoids are a large family of natural and synthetic compounds related to vitamin A that have pleiotropic effects on body physiology, reproduction, immunity, and embryonic development. The diverse activities of retinoids are primarily mediated by two families of nuclear retinoic acid receptors, the RARs and RXRs. Retinoic acids are thought to be the only natural ligands for these receptors and are widely assumed to be the active principle of vitamin A. However, during an unbiased, bioactivit...

  10. The Quintiles Prize Lecture 2004. The identification of the adenosine A2B receptor as a novel therapeutic target in asthma.

    Science.gov (United States)

    Holgate, Stephen T

    2005-08-01

    Adenosine is a powerful bronchoconstrictor of asthmatic, but not normal, airways. In vitro studies on isolated human mast cells and basophils revealed that adenosine and selective analogues augmented inflammatory mediator release from mast cells by stimulating A(2) receptors. Pharmacological blockade of mast cell mediator release in vivo also attenuated adenosine-induced bronchoconstriction, as did theophylline, by adenosine A(2) receptor antagonism. Further in vitro studies revealed that the asthmatic response to adenosine is likely to be mediated via the A(2B) subtype which is selectively antagonised by enprofylline. Studies in animal models, especially mice, have shown a close synergistic interaction between adenosine, Th2 and airway remodelling responses. The recent description of A(2B) receptors on human airway smooth muscle cells that mediate cytokine and chemokine release and induce differentiation of fibroblasts into myofibroblasts strengthens the view that adenosine maybe more than an inflammatory mediator in asthma but also participates in airway wall remodelling in this disease. These data have provided a firm basis for developing adenosine A(2B) receptor antagonists as a new therapeutic approach to this disease. PMID:15980878

  11. The Quintiles Prize Lecture 2004: The identification of the adenosine A2B receptor as a novel therapeutic target in asthma

    Science.gov (United States)

    Holgate, Stephen T

    2005-01-01

    Adenosine is a powerful bronchoconstrictor of asthmatic, but not normal, airways. In vitro studies on isolated human mast cells and basophils revealed that adenosine and selective analogues augmented inflammatory mediator release from mast cells by stimulating A2 receptors. Pharmacological blockade of mast cell mediator release in vivo also attenuated adenosine-induced bronchoconstriction, as did theophylline, by adenosine A2 receptor antagonism. Further in vitro studies revealed that the asthmatic response to adenosine is likely to be mediated via the A2B subtype which is selectively antagonised by enprofylline. Studies in animal models, especially mice, have shown a close synergistic interaction between adenosine, Th2 and airway remodelling responses. The recent description of A2B receptors on human airway smooth muscle cells that mediate cytokine and chemokine release and induce differentiation of fibroblasts into myofibroblasts strengthens the view that adenosine maybe more than an inflammatory mediator in asthma but also participates in airway wall remodelling in this disease. These data have provided a firm basis for developing adenosine A2B receptor antagonists as a new therapeutic approach to this disease. PMID:15980878

  12. The Quintiles Prize Lecture 2004. The identification of the adenosine A2B receptor as a novel therapeutic target in asthma.

    Science.gov (United States)

    Holgate, Stephen T

    2005-08-01

    Adenosine is a powerful bronchoconstrictor of asthmatic, but not normal, airways. In vitro studies on isolated human mast cells and basophils revealed that adenosine and selective analogues augmented inflammatory mediator release from mast cells by stimulating A(2) receptors. Pharmacological blockade of mast cell mediator release in vivo also attenuated adenosine-induced bronchoconstriction, as did theophylline, by adenosine A(2) receptor antagonism. Further in vitro studies revealed that the asthmatic response to adenosine is likely to be mediated via the A(2B) subtype which is selectively antagonised by enprofylline. Studies in animal models, especially mice, have shown a close synergistic interaction between adenosine, Th2 and airway remodelling responses. The recent description of A(2B) receptors on human airway smooth muscle cells that mediate cytokine and chemokine release and induce differentiation of fibroblasts into myofibroblasts strengthens the view that adenosine maybe more than an inflammatory mediator in asthma but also participates in airway wall remodelling in this disease. These data have provided a firm basis for developing adenosine A(2B) receptor antagonists as a new therapeutic approach to this disease.

  13. Plasma membrane Ca2+ pumping plays a prominent role in adenosine A(1) receptor mediated changes in [Ca2+](i) in DDT1 MF-2 cells

    NARCIS (Netherlands)

    Sipma, H; Fredholm, BB; DenHertog, A; Nelemans, A

    1996-01-01

    Adenosine A(1) receptor mediated formation of inositol 1,4,5-trisphosphate (Ins(1,4,5)P-3) and accumulation of cytoplasmic Ca2+ ([Ca2+](i)) were investigated in DDT1 MF-2 smooth muscle cells. A strong reduction of the adenosine and N-6-cyclopentyladenosine (CPA) induced rise in [Ca2+](i) was observe

  14. Evaluation of iodinated and brominated [{sup 11}C]styrylxanthine derivatives as in vivo radioligands mapping adenosine A{sub 2A} receptor in the central nervous system

    Energy Technology Data Exchange (ETDEWEB)

    Ishiwata, Kiichi; Wang, Wei-Fang; Ishii, Shin-ichi; Senda, Michio [Tokyo Metropolitan Inst. of Gerontology (Japan). Positron Medical Center; Shimada, Junichi; Harakawa, Hiroyuki; Kiyosawa, Motohiro; Suzuki, Fumio

    2000-08-01

    In vivo assessment of the adenosine A{sub 2A} receptors localized in the striatum by PET or SPECT offers us a new diagnostic tool for neurological disorders. In the present study, we evaluated the potential of iodinated and brominated styrylxanthine derivatives labeled with {sup 11}C as an in vivo probe. [7-Methyl-{sup 11}C]-(E)-3,7-dimethyl-8-(3-iodostyryl)-1-propargylxanthine ([{sup 11}C]IS-DMPX) and [7-methyl-{sup 11}C]-(E)-8-(3-bromostyryl)-3,7-dimethyl-1-propargylxanthine ([{sup 11}C]BS-DMPX) were prepared by the {sup 11}C-methylation of corresponding 7-demethyl derivatives. An in vitro membrane binding study showed a high affinity (Ki values) of the two ligands for A{sub 2A} receptor: 8.9 nM for IS-DMPX and 7.7 nM for BS-DMPX, and a high A{sub 2A}/A{sub 1} selectivity: >1100 for IS-DMPX and 300 for BS-DMPX. In mice, [{sup 11}C]IS-DMPX and [{sup 11}C]BS-DMPX were taken up slightly more in the striatum than in the reference regions such as the cortex and cerebellum. The uptake ratios of striatum to cortex and striatum to cerebellum gradually increased but were very small: 1.6-1.7 for the striatum-to-cortex ratio and 1.2 for the striatum-to-cerebellum ratio at 60 min postinjection. The uptake by these three regions was reduced by co-injection of an excess amount of carrier or an A{sub 2A} antagonist KF17837, but not by an A{sub 1} antagonist KF15372. The blocking effects in the three regions were greater for [{sup 11}C]BS-DMPX (32-57%) than for [{sup 11}C]IS-DMPX (6-29%). Ex vivo autoradiography confirmed that the two ligands were slightly concentrated in the striatum. [{sup 11}C]BS-DMPX showed more selective affinity for adenosine A{sub 2A} receptors than [{sup 11}C]IS-DMPX, but these results have shown that the two tracers were not suitable as in vivo ligands because of low selectivity for the striatal A{sub 2A} receptors and a high nonspecific binding. (author)

  15. Striatal pre- and postsynaptic profile of adenosine A(2A receptor antagonists.

    Directory of Open Access Journals (Sweden)

    Marco Orru

    Full Text Available Striatal adenosine A(2A receptors (A(2ARs are highly expressed in medium spiny neurons (MSNs of the indirect efferent pathway, where they heteromerize with dopamine D(2 receptors (D(2Rs. A(2ARs are also localized presynaptically in cortico-striatal glutamatergic terminals contacting MSNs of the direct efferent pathway, where they heteromerize with adenosine A(1 receptors (A(1Rs. It has been hypothesized that postsynaptic A(2AR antagonists should be useful in Parkinson's disease, while presynaptic A(2AR antagonists could be beneficial in dyskinetic disorders, such as Huntington's disease, obsessive-compulsive disorders and drug addiction. The aim or this work was to determine whether selective A(2AR antagonists may be subdivided according to a preferential pre- versus postsynaptic mechanism of action. The potency at blocking the motor output and striatal glutamate release induced by cortical electrical stimulation and the potency at inducing locomotor activation were used as in vivo measures of pre- and postsynaptic activities, respectively. SCH-442416 and KW-6002 showed a significant preferential pre- and postsynaptic profile, respectively, while the other tested compounds (MSX-2, SCH-420814, ZM-241385 and SCH-58261 showed no clear preference. Radioligand-binding experiments were performed in cells expressing A(2AR-D(2R and A(1R-A(2AR heteromers to determine possible differences in the affinity of these compounds for different A(2AR heteromers. Heteromerization played a key role in the presynaptic profile of SCH-442416, since it bound with much less affinity to A(2AR when co-expressed with D(2R than with A(1R. KW-6002 showed the best relative affinity for A(2AR co-expressed with D(2R than co-expressed with A(1R, which can at least partially explain the postsynaptic profile of this compound. Also, the in vitro pharmacological profile of MSX-2, SCH-420814, ZM-241385 and SCH-58261 was is in accordance with their mixed pre- and postsynaptic profile

  16. Aryl hydrocarbon receptor ligands in cancer: friend and foe.

    Science.gov (United States)

    Murray, Iain A; Patterson, Andrew D; Perdew, Gary H

    2014-12-01

    The aryl hydrocarbon receptor (AHR) is a ligand-activated transcription factor that is best known for mediating the toxicity and tumour-promoting properties of the carcinogen 2,3,7,8-tetrachlorodibenzo-p-dioxin, commonly referred to as ‘dioxin’. AHR influences the major stages of tumorigenesis — initiation, promotion, progression and metastasis — and physiologically relevant AHR ligands are often formed during disease states or during heightened innate and adaptive immune responses. Interestingly, ligand specificity and affinity vary between rodents and humans. Studies of aggressive tumours and tumour cell lines show increased levels of AHR and constitutive localization of this receptor in the nucleus. This suggests that the AHR is chronically activated in tumours, thus facilitating tumour progression. This Review discusses the role of AHR in tumorigenesis and the potential for therapeutic modulation of its activity in tumours. PMID:25568920

  17. Competitive antagonism of AMPA receptors by ligands of different classes

    DEFF Research Database (Denmark)

    Hogner, Anders; Greenwood, Jeremy R; Liljefors, Tommy;

    2003-01-01

    Ionotropic glutamate receptors (iGluRs) constitute a family of ligand-gated ion channels that are essential for mediating fast synaptic transmission in the central nervous system. This study presents a high-resolution X-ray structure of the competitive antagonist (S)-2-amino-3-[5-tert-butyl-3-(ph...

  18. Modeling of ligand binding to dopamine D2 receptor

    Directory of Open Access Journals (Sweden)

    Ostopovici-Halip Liliana

    2014-01-01

    Full Text Available The dopaminic receptors have been for long time the major targets for developing new small molecules with high affinity and selectivity to treat psychiatric disorders, neurodegeneration, drug abuse, and other therapeutic areas. In the absence of a 3D structure for the human D2 dopamine (HDD2 receptor, the efforts for discovery and design of new potential drugs rely on comparative models generation, docking and pharmacophore development studies. To get a better understanding of the HDD2 receptor binding site and the ligand-receptor interactions a homology model of HDD2 receptor based on the X-ray structure of β2-adrenergic receptor has been built and used to dock a set of partial agonists of HDD2 receptor. The main characteristics of the binding mode for the HDD2 partial agonists set are given by the ligand particular folding and a complex network of contacts represented by stacking interactions, salt bridge and hydrogen bond formation. The characterization of the partial agonist binding mode at HDD2 receptor provide the needed information to generate pharmacophore models which represent essential information in the future virtual screening studies in order to identify new potential HDD2 partial agonists.

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

    Directory of Open Access Journals (Sweden)

    Esposito Emanuela

    2011-04-01

    Full Text Available Abstract 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

  20. Computational study of the binding modes of caffeine to the adenosine A2A receptor.

    Science.gov (United States)

    Liu, Yuli; Burger, Steven K; Ayers, Paul W; Vöhringer-Martinez, Esteban

    2011-12-01

    Using the recently solved crystal structure of the human adenosine A(2A) receptor, we applied MM/PBSA to compare the binding modes of caffeine with those of the high-affinity selective antagonist ZM241385. MD simulations were performed in the environment of the lipid membrane bilayer. Four low-energy binding modes of caffeine-A(2A) were found, all of which had similar energies. Assuming an equal contribution of each binding mode of caffeine, the computed binding free energy difference between caffeine and ZM241385 is -2.4 kcal/mol, which compares favorably with the experimental value, -3.6 kcal/mol. The configurational entropy contribution of -0.9 kcal/mol from multiple binding modes of caffeine helps explain how a small molecule like caffeine can compete with a significantly larger molecule, ZM241385, which can form many more interactions with the receptor. We also performed residue-wise energy decomposition and found that Phe168, Leu249, and Ile274 contribute most significantly to the binding modes of caffeine and ZM241385. PMID:21970461

  1. The effects of nucleus accumbens μ-opioid and adenosine 2A receptor stimulation and blockade on instrumental learning.

    Science.gov (United States)

    Clissold, Kara A; Pratt, Wayne E

    2014-11-01

    Prior research has shown that glutamate and dopamine receptors in the nucleus accumbens (NAcc) core are critical for the learning of an instrumental response for food reinforcement. It has also been demonstrated that μ-opioid and adenosine A2A receptors within the NAcc impact feeding and motivational processes. In these experiments, we examined the potential roles of NAcc μ-opioid and A2A receptors on instrumental learning and performance. Sprague-Dawley rats were food restricted and trained to lever press following daily intra-accumbens injections of the A2A receptor agonist CGS 21680 (at 0.0, 6.0, or 24.0ng/side), the A2A antagonist pro-drug MSX-3 (at 0.0, 1.0, or 3.0μg/side), the μ-opioid agonist DAMGO (at 0.0, 0.025, or 0.025μg/side), or the opioid receptor antagonist naltrexone (at 0.0, 2.0 or 20.0μg/side). After five days, rats continued training without drug injections until lever pressing rates stabilized, and were then tested with a final drug test to assess potential performance effects. Stimulation, but not inhibition, of NAcc adenosine A2A receptors depressed lever pressing during learning and performance tests, but did not impact lever pressing on non-drug days. Both μ-opioid receptor stimulation and blockade inhibited learning of the lever-press response, though only naltrexone treatment caused impairments in lever-pressing after the task had been learned. The effect of A2A receptor stimulation on learning and performance were consistent with known effects of adenosine on effort-related processes, whereas the pattern of lever presses, magazine approaches, and pellet consumption following opioid receptor manipulations suggested that their effects may have been driven by drug-induced shifts in the incentive value of the sugar reinforcer. PMID:25101542

  2. Capacity of Diffusion-based Molecular Communication with Ligand Receptors

    CERN Document Server

    Einolghozati, Arash; Fekri, Faramarz

    2012-01-01

    A diffusion-based molecular communication system has two major components: the diffusion in the medium, and the ligand-reception. Information bits, encoded in the time variations of the concentration of molecules, are conveyed to the receiver front through the molecular diffusion in the medium. The receiver, in turn, measures the concentration of the molecules in its vicinity in order to retrieve the information. This is done via ligand-reception process. In this paper, we develop models to study the constraints imposed by the concentration sensing at the receiver side and derive the maximum rate by which a ligand-receiver can receive information. Therefore, the overall capacity of the diffusion channel with the ligand receptors can be obtained by combining the results presented in this paper with our previous work on the achievable information rate of molecular communication over the diffusion channel.

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

    Science.gov (United States)

    de Mendonça, A; Ribeiro, J A

    1997-08-01

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

  4. Renal effects of the novel selective adenosine A1 receptor blocker SLV329 in experimental liver cirrhosis in rats.

    Directory of Open Access Journals (Sweden)

    Berthold Hocher

    Full Text Available Liver cirrhosis is often complicated by an impaired renal excretion of water and sodium. Diuretics tend to further deteriorate renal function. It is unknown whether chronic selective adenosine A(1 receptor blockade, via inhibition of the hepatorenal reflex and the tubuloglomerular feedback, might exert diuretic and natriuretic effects without a reduction of the glomerular filtration rate. In healthy animals intravenous treatment with the novel A(1 receptor antagonist SLV329 resulted in a strong dose-dependent diuretic (up to 3.4-fold and natriuretic (up to 13.5-fold effect without affecting creatinine clearance. Male Wistar rats with thioacetamide-induced liver cirrhosis received SLV329, vehicle or furosemide for 12 weeks. The creatinine clearance of cirrhotic animals decreased significantly (-36.5%, p<0.05, especially in those receiving furosemide (-41.9%, p<0.01. SLV329 was able to prevent this decline of creatinine clearance. Mortality was significantly lower in cirrhotic animals treated with SLV329 in comparison to animals treated with furosemide (17% vs. 54%, p<0.05. SLV329 did not relevantly influence the degree of liver fibrosis, kidney histology or expression of hepatic or renal adenosine receptors. In conclusion, chronic treatment with SLV329 prevented the decrease of creatinine clearance in a rat model of liver cirrhosis. Further studies will have to establish whether adenosine A(1 receptor antagonists are clinically beneficial at different stages of liver cirrhosis.

  5. Evolution of ligand specificity in vertebrate corticosteroid receptors

    Directory of Open Access Journals (Sweden)

    Deitcher David L

    2011-01-01

    Full Text Available Abstract Background Corticosteroid receptors include mineralocorticoid (MR and glucocorticoid (GR receptors. Teleost fishes have a single MR and duplicate GRs that show variable sensitivities to mineralocorticoids and glucocorticoids. How these receptors compare functionally to tetrapod MR and GR, and the evolutionary significance of maintaining two GRs, remains unclear. Results We used up to seven steroids (including aldosterone, cortisol and 11-deoxycorticosterone [DOC] to compare the ligand specificity of the ligand binding domains of corticosteroid receptors between a mammal (Mus musculus and the midshipman fish (Porichthys notatus, a teleost model for steroid regulation of neural and behavioral plasticity. Variation in mineralocorticoid sensitivity was considered in a broader phylogenetic context by examining the aldosterone sensitivity of MR and GRs from the distantly related daffodil cichlid (Neolamprologus pulcher, another teleost model for neurobehavioral plasticity. Both teleost species had a single MR and duplicate GRs. All MRs were sensitive to DOC, consistent with the hypothesis that DOC was the initial ligand of the ancestral MR. Variation in GR steroid-specificity corresponds to nine identified amino acid residue substitutions rather than phylogenetic relationships based on receptor sequences. Conclusion The mineralocorticoid sensitivity of duplicate GRs in teleosts is highly labile in the context of their evolutionary phylogeny, a property that likely led to neo-functionalization and maintenance of two GRs.

  6. Hide and seek: a comparative autoradiographic in vitro investigation of the adenosine A3 receptor

    Energy Technology Data Exchange (ETDEWEB)

    Haeusler, D.; Fuchshuber, F.; Girschele, F.; Hacker, M.; Wadsak, W.; Mitterhauser, Markus [Medical University of Vienna, Department of Biomedical Imaging and Image-guided Therapy, Vienna (Austria); Grassinger, L. [University of Applied Sciences Wiener Neustadt, Department of Biomedical Analytics, Wiener Neustadt (Austria); Hoerleinsberger, W.J. [Medical University of Vienna, Department of Biomedical Imaging and Image-guided Therapy, Vienna (Austria); University of Vienna, Cognitive Science Research Platform, Vienna (Austria); Hoeftberger, R.; Leisser, I. [Medical University of Vienna, Institute of Neurology, Vienna (Austria); Shanab, K.; Spreitzer, H. [University of Vienna, Department of Drug and Natural Product Synthesis, Vienna (Austria); Gerdenitsch, W. [Medical University of Vienna, Institute of Biomedicinal Research, Vienna (Austria)

    2015-05-01

    Since the adenosine A3 receptor (A3R) is considered to be of high clinical importance in the diagnosis and treatment of ischaemic conditions (heart and brain), glaucoma, asthma, arthritis, cancer and inflammation, a suitable and selective A3R PET tracer such as [{sup 18}F]FE rate at SUPPY would be of high clinical value for clinicians as well as patients. A3R was discovered in the late 1990s, but there is still little known regarding its distribution in the CNS and periphery. Hence, in autoradiographic experiments the distribution of A3R in human brain and rat tissues was investigated and the specific binding of the A3R antagonist FE rate at SUPPY and MRS1523 compared. Immunohistochemical staining (IHC) experiments were also performed to validate the autoradiographic findings. For autoradiographic competition experiments human post-mortem brain and rat tissues were incubated with [{sup 125}I]AB-MECA and highly selective compounds to block the other adenosine receptor subtypes. Additionally, IHC was performed with an A3 antibody. Specific A3R binding of MRS1523 and FE rate at SUPPY was found in all rat peripheral tissues examined with the highest amounts in the spleen (44.0 % and 46.4 %), lung (44.5 % and 45.0 %), heart (39.9 % and 42.9 %) and testes (27.4 % and 29.5 %, respectively). Low amounts of A3R were found in rat brain tissues (5.9 % and 5.6 %, respectively) and human brain tissues (thalamus 8.0 % and 9.1 %, putamen 7.8 % and 8.2 %, cerebellum 6.0 % and 7.8 %, hippocampus 5.7 % and 5.6 %, caudate nucleus 4.9 % and 6.4 %, cortex 4.9 % and 6.3 %, respectively). The outcome of the A3 antibody staining experiments complemented the results of the autoradiographic experiments. The presence of A3R protein was verified in central and peripheral tissues by autoradiography and IHC. The specificity and selectivity of FE rate at SUPPY was confirmed by direct comparison with MRS1523, providing further evidence that [{sup 18}F]FE rate at SUPPY may be a suitable A3 PET

  7. Hide and seek: a comparative autoradiographic in vitro investigation of the adenosine A3 receptor

    International Nuclear Information System (INIS)

    Since the adenosine A3 receptor (A3R) is considered to be of high clinical importance in the diagnosis and treatment of ischaemic conditions (heart and brain), glaucoma, asthma, arthritis, cancer and inflammation, a suitable and selective A3R PET tracer such as [18F]FE rate at SUPPY would be of high clinical value for clinicians as well as patients. A3R was discovered in the late 1990s, but there is still little known regarding its distribution in the CNS and periphery. Hence, in autoradiographic experiments the distribution of A3R in human brain and rat tissues was investigated and the specific binding of the A3R antagonist FE rate at SUPPY and MRS1523 compared. Immunohistochemical staining (IHC) experiments were also performed to validate the autoradiographic findings. For autoradiographic competition experiments human post-mortem brain and rat tissues were incubated with [125I]AB-MECA and highly selective compounds to block the other adenosine receptor subtypes. Additionally, IHC was performed with an A3 antibody. Specific A3R binding of MRS1523 and FE rate at SUPPY was found in all rat peripheral tissues examined with the highest amounts in the spleen (44.0 % and 46.4 %), lung (44.5 % and 45.0 %), heart (39.9 % and 42.9 %) and testes (27.4 % and 29.5 %, respectively). Low amounts of A3R were found in rat brain tissues (5.9 % and 5.6 %, respectively) and human brain tissues (thalamus 8.0 % and 9.1 %, putamen 7.8 % and 8.2 %, cerebellum 6.0 % and 7.8 %, hippocampus 5.7 % and 5.6 %, caudate nucleus 4.9 % and 6.4 %, cortex 4.9 % and 6.3 %, respectively). The outcome of the A3 antibody staining experiments complemented the results of the autoradiographic experiments. The presence of A3R protein was verified in central and peripheral tissues by autoradiography and IHC. The specificity and selectivity of FE rate at SUPPY was confirmed by direct comparison with MRS1523, providing further evidence that [18F]FE rate at SUPPY may be a suitable A3 PET tracer for use in

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

    Science.gov (United States)

    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.

  9. DMPD: Endogenous ligands of Toll-like receptors. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 15178705 Endogenous ligands of Toll-like receptors. Tsan MF, Gao B. J Leukoc Biol. ...2004 Sep;76(3):514-9. Epub 2004 Jun 3. (.png) (.svg) (.html) (.csml) Show Endogenous ligands of Toll-like re...ceptors. PubmedID 15178705 Title Endogenous ligands of Toll-like receptors. Authors Tsan MF, Gao B. Publicat

  10. Preclinical studies on [{sup 11}C]MPDX for mapping adenosine A{sub 1} receptors by positron emission tomography

    Energy Technology Data Exchange (ETDEWEB)

    Ishiwata, Kiichi; Kimura, Yuichi; Oda, Keiichi; Kawamura, Kazunori; Ishii, Kenji; Senda, Michio [Tokyo Metropolitan Inst. of Gerontology (Japan). Positron Medical Center; Nariai, Tadashi; Wakabayashi, Shinichi [Tokyo Medical and Dental Univ. (Japan). School of Medicine; Shimada, Junichi [Kyowa Hakko Kogyo Co. Ltd., Tokyo (Japan). Pharmaceutical Research Inst.

    2002-09-01

    In previous in vivo studies with mice, rats and cats, we have demonstrated that [{sup 11}C]MPDX ([1-methyl-{sup 11}C]8-dicyclopropylmethyl-1-methyl-3-propylxanthine) is a potential radioligand for mapping adenosine A{sub 1} receptors of the brain by positron emission tomography (PET). In the present study, we performed a preclinical study. The radiation absorbed-dose by [{sup 11}C]MPDX in humans estimated from the tissue distribution in mice was low enough for clinical use, and the acute toxicity and mutagenicity of MPDX were not found. The monkey brain was clearly visualized by PET with [{sup 11}C]MPDX. We have concluded that [{sup 11}C]MPDX is suitable for mapping adenosine A{sub 1} receptors in the human brain by PET. (author)

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

    OpenAIRE

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

    2012-01-01

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

  12. Adenosine A2B receptor-mediated leukemia inhibitory factor release from astrocytes protects cortical neurons against excitotoxicity

    Directory of Open Access Journals (Sweden)

    Moidunny Shamsudheen

    2012-08-01

    Full Text Available Abstract Background Neuroprotective and neurotrophic properties of leukemia inhibitory factor (LIF have been widely reported. In the central nervous system (CNS, astrocytes are the major source for LIF, expression of which is enhanced following disturbances leading to neuronal damage. How astrocytic LIF expression is regulated, however, has remained an unanswered question. Since neuronal stress is associated with production of extracellular adenosine, we investigated whether LIF expression in astrocytes was mediated through adenosine receptor signaling. Methods Mouse cortical neuronal and astrocyte cultures from wild-type and adenosine A2B receptor knock-out animals, as well as adenosine receptor agonists/antagonists and various enzymatic inhibitors, were used to study LIF expression and release in astrocytes. When needed, a one-way analysis of variance (ANOVA followed by Bonferroni post-hoc test was used for statistical analysis. Results We show here that glutamate-stressed cortical neurons induce LIF expression through activation of adenosine A2B receptor subtype in cultured astrocytes and require signaling of protein kinase C (PKC, mitogen-activated protein kinases (MAPKs: p38 and ERK1/2, and the nuclear transcription factor (NF-κB. Moreover, LIF concentration in the supernatant in response to 5′-N-ethylcarboxamide (NECA stimulation was directly correlated to de novo protein synthesis, suggesting that LIF release did not occur through a regulated release pathway. Immunocytochemistry experiments show that LIF-containing vesicles co-localize with clathrin and Rab11, but not with pHogrin, Chromogranin (CgA and CgB, suggesting that LIF might be secreted through recycling endosomes. We further show that pre-treatment with supernatants from NECA-treated astrocytes increased survival of cultured cortical neurons against glutamate, which was absent when the supernatants were pre-treated with an anti-LIF neutralizing antibody. Conclusions

  13. Expression of adenosine receptors in human retinal pigment epithelium cells in vitro

    Institute of Scientific and Technical Information of China (English)

    WAN Wen-juan; CUI Dong-mei; YANG Xiao; HU Jian-min; LI Chuan-xu; HU Shou-long; Klaus Trier; ZENG Jun-wen

    2011-01-01

    Background Adenosine receptors (ADORs) have been reported to play a role in experimental myopia. This study aimed to determine the distribution of ADORs in human retinal pigment epithelium (RPE) cells cultured in vitro.Methods Human RPE cells (cell line D407) were cultured in vitro. ADOR mRNA in RPE was detected by reverse transcription polymerase chain reaction. ADOR protein expression in RPE was confirmed by Western blotting analysis of cell lysates. Confocal fluorescence microscopy was used to study the subcellular distribution of ADORs.Results All four subtypes of ADORs mRNA and protein were expressed in human RPE. This was confirmed by Western blotting analysis. The ADOR subtypes were differently distributed within the cells. ADORA1 was expressed in nucleus, perinucleus and cytoplasm of RPE. ADORA2A was concentrated mainly in one side of the perinucleus and cytoplasm of RPE. ADORA2B was strongly expressed in the nucleus, perinucleus and the cytoplasm, and ADORA3 was expressed weakly in the cytoplasm of RPE.Conclusions ADORs are expressed in human RPE. The different distribution at the subcellular level suggests different functions of ADOR subtypes.

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

    Science.gov (United States)

    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.

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

    Science.gov (United States)

    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. PMID:24170241

  16. Increased desensitization of dopamine D₂ receptor-mediated response in the ventral tegmental area in the absence of adenosine A(2A) receptors.

    Science.gov (United States)

    Al-Hasani, R; Foster, J D; Metaxas, A; Ledent, C; Hourani, S M O; Kitchen, I; Chen, Y

    2011-09-01

    G-protein coupled receptors interact to provide additional regulatory mechanisms for neurotransmitter signaling. Adenosine A(2A) receptors are expressed at a high density in striatal neurons, where they closely interact with dopamine D₂ receptors and modulate effects of dopamine and responses to psychostimulants. A(2A) receptors are expressed at much lower densities in other forebrain neurons but play a more prominent yet opposing role to striatal receptors in response to psychostimulants in mice. It is, therefore, possible that A(2A) receptors expressed at low levels elsewhere in the brain may also regulate neurotransmitter systems and modulate neuronal functions. Dopamine D₂ receptors play an important role in autoinhibition of neuronal firing in dopamine neurons of the ventral tegmental area (VTA) and dopamine release in other brain areas. Here, we examined the effect of A(2A) receptor deletion on D₂ receptor-mediated inhibition of neuronal firing in dopamine neurons in the VTA. Spontaneous activity of dopamine neurons was recorded in midbrain slices, and concentration-dependent effects of the dopamine D₂ receptor agonist, quinpirole, was compared between wild-type and A(2A) knockout mice. The potency of quinpirole applied in single concentrations and the expression of D₂ receptors were not altered in the VTA of the knockout mice. However, quinpirole applied in stepwise escalating concentrations caused significantly reduced maximal inhibition in A(2A) knockout mice, indicating an enhanced agonist-induced desensitization of D₂ receptors in the absence of A(2A) receptors. The A(2A) receptor agonist, CGS21680, did not exert any effect on dopamine neuron firing or response to quinpirole, revealing a novel non-pharmacological interaction between adenosine A(2A) receptors and dopaminergic neurotransmission in midbrain dopamine neurons. Altered D₂ receptor desensitization may result in changes in dopamine neuron firing rate and pattern and dopamine

  17. Blockage of A2A and A3 adenosine receptors decreases the desensitization of human GABAA receptors microtransplanted to Xenopus oocytes

    Science.gov (United States)

    Roseti, Cristina; Palma, Eleonora; Martinello, Katiuscia; Fucile, Sergio; Morace, Roberta; Esposito, Vincenzo; Cantore, Gianpaolo; Arcella, Antonietta; Giangaspero, Felice; Aronica, Eleonora; Mascia, Addolorata; Di Gennaro, Giancarlo; Quarato, Pier Paolo; Manfredi, Mario; Cristalli, Gloria; Lambertucci, Catia; Marucci, Gabriella; Volpini, Rosaria; Limatola, Cristina; Eusebi, Fabrizio

    2009-01-01

    We previously found that the endogenous anticonvulsant adenosine, acting through A2A and A3 adenosine receptors (ARs), alters the stability of currents (IGABA) generated by GABAA receptors expressed in the epileptic human mesial temporal lobe (MTLE). Here we examined whether ARs alter the stability (desensitization) of IGABA expressed in focal cortical dysplasia (FCD) and in periglioma epileptic tissues. The experiments were performed with tissues from 23 patients, using voltage-clamp recordings in Xenopus oocytes microinjected with membranes isolated from human MTLE and FCD tissues or using patch-clamp recordings of pyramidal neurons in epileptic tissue slices. On repetitive activation, the epileptic GABAA receptors revealed instability, manifested by a large IGABA rundown, which in most of the oocytes (≈70%) was obviously impaired by the new A2A antagonists ANR82, ANR94, and ANR152. In most MTLE tissue-microtransplanted oocytes, a new A3 receptor antagonist (ANR235) significantly improved IGABA stability. Moreover, patch-clamped pyramidal neurons from human neocortical slices of periglioma epileptic tissues exhibited altered IGABA rundown on ANR94 treatment. Our findings indicate that antagonizing A2A and A3 receptors increases the IGABA stability in different epileptic tissues and suggest that adenosine derivatives may offer therapeutic opportunities in various forms of human epilepsy. PMID:19721003

  18. Adenosine Receptor Stimulation by Polydeoxyribonucleotide Improves Tissue Repair and Symptomology in Experimental Colitis.

    Science.gov (United States)

    Pallio, Giovanni; Bitto, Alessandra; Pizzino, Gabriele; Galfo, Federica; Irrera, Natasha; Squadrito, Francesco; Squadrito, Giovanni; Pallio, Socrate; Anastasi, Giuseppe P; Cutroneo, Giuseppina; Macrì, Antonio; Altavilla, Domenica

    2016-01-01

    Activation of the adenosine receptor pathway has been demonstrated to be effective in improving tissue remodeling and blunting the inflammatory response. Active colitis is characterized by an intense inflammatory reaction resulting in extensive tissue damage. Symptomatic improvement requires both control of the inflammatory process and repair and remodeling of damaged tissues. We investigated the ability of an A2A receptor agonist, polydeoxyribonucleotide (PDRN), to restore tissue structural integrity in two experimental colitis models using male Sprague-Dawley rats. In the first model, colitis was induced with a single intra-colonic instillation of dinitrobenzenesulfonic acid (DNBS), 25 mg diluted in 0.8 ml 50% ethanol. After 6 h, animals were randomized to receive either PDRN (8 mg/kg/i.p.), or PDRN + the A2A antagonist [3,7-dimethyl-1-propargylxanthine (DMPX); 10 mg/kg/i.p.], or vehicle (0.8 ml saline solution) daily. In the second model, dextran sulfate sodium (DSS) was dissolved in drinking water at a concentration of 8%. Control animals received standard drinking water. After 24 h animals were randomized to receive PDRN or PDRN+DMPX as described above. Rats were sacrificed 7 days after receiving DNBS or 5 days after DSS. In both experimental models of colitis, PDRN ameliorated the clinical symptoms and weight loss associated with disease as well as promoted the histological repair of damaged tissues. Moreover, PDRN reduced expression of inflammatory cytokines, myeloperoxidase activity, and malondialdehyde. All these effects were abolished by the concomitant administration of the A2A antagonist DMPX. Our study suggests that PDRN may represent a promising treatment for improving tissue repair during inflammatory bowel diseases. PMID:27601997

  19. Presynaptic facilitatory adenosine A2A receptors mediate fade induced by neuromuscular relaxants that exhibit anticholinesterase activity.

    Science.gov (United States)

    Bornia, Elaine Cs; Correia-de-Sá, Paulo; Alves-Do-Prado, Wilson

    2011-03-01

    1. Pancuronium, cisatracurium and vecuronium are antinicotinic agents that, in contrast with d-tubocurarine and hexamethonium, exhibit anticholinesterase activity. Pancuronium-, cisatracurium- and vecuronium-induced fade results from blockade of facilitatory nicotinic receptors on motor nerves, but fade produced by such agents also depends on the presynaptic activation of inhibitory muscarinic M2 receptors by acetylcholine released from motor nerve terminals and activation of inhibitory adenosine A1 receptors by adenosine released from motor nerves and muscles. The participation of presynaptic facilitatory A2A receptors in fade caused by pancuronium, cisatracurium and vecuronium has not yet been investigated. In the present study, we determined the effects of ZM241385, an antagonist of presynaptic facilitatory A2A receptors, on fade produced by these neuromuscular relaxants in the rat phrenic nerve-diaphragm (PND) preparation. 2. The muscles were stimulated indirectly at 75±3Hz to induce a sustained tetanizing muscular contraction. The lowest concentration at which each antinicotinic agent produced fade without modifying initial tetanic tension (presynaptic action) was determined. 3. d-Tubocurarine-induced fade occurred only at 55 nmol/L, a concentration that also reduced maximal tetanic tension (post-synaptic action). At 10 nmol/L, ZM 241385 alone did not produce fade, but it did attenuate pancuronium (0.32 μmol/L)-, cisatracurium (0.32 μmol/L)- and vecuronium (0.36 μmol/L)-induced fade. 4. The fade induced by the 'pure' antinicotinic agents d-tubocurarine (55 nmol/L) and hexamethonium (413 μmol/L) was not altered by 10 nmol/L ZM 241385, indicating that presynaptic adenosine A2A receptors play a significant role in the fade produced by antinicotinic agents when such agents have anticholinesterase activity.

  20. Adenosine A(2A) receptor modulation of hippocampal CA3-CA1 synapse plasticity during associative learning in behaving mice.

    Science.gov (United States)

    Fontinha, Bruno M; Delgado-García, José M; Madroñal, Noelia; Ribeiro, Joaquim A; Sebastião, Ana M; Gruart, Agnès

    2009-06-01

    Previous in vitro studies have characterized the electrophysiological and molecular signaling pathways of adenosine tonic modulation on long-lasting synaptic plasticity events, particularly for hippocampal long-term potentiation (LTP). However, it remains to be elucidated whether the long-term changes produced by endogenous adenosine in the efficiency of synapses are related to those required for learning and memory formation. Our goal was to understand how endogenous activation of adenosine excitatory A(2A) receptors modulates the associative learning evolution in conscious behaving mice. We have studied here the effects of the application of a highly selective A(2A) receptor antagonist, SCH58261, upon a well-known associative learning paradigm-classical eyeblink conditioning. We used a trace paradigm, with a tone as the conditioned stimulus (CS) and an electric shock presented to the supraorbital nerve as the unconditioned stimulus (US). A single electrical pulse was presented to the Schaffer collateral-commissural pathway to evoke field EPSPs (fEPSPs) in the pyramidal CA1 area during the CS-US interval. In vehicle-injected animals, there was a progressive increase in the percentage of conditioning responses (CRs) and in the slope of fEPSPs through conditioning sessions, an effect that was completely prevented (and lost) in SCH58261 (0.5 mg/kg, i.p.) -injected animals. Moreover, experimentally evoked LTP was impaired in SCH58261-injected mice. In conclusion, the endogenous activation of adenosine A(2A) receptors plays a pivotal effect on the associative learning process and its relevant hippocampal circuits, including activity-dependent changes at the CA3-CA1 synapse.

  1. Optimizing Ligand Efficiency of Selective Androgen Receptor Modulators (SARMs).

    Science.gov (United States)

    Handlon, Anthony L; Schaller, Lee T; Leesnitzer, Lisa M; Merrihew, Raymond V; Poole, Chuck; Ulrich, John C; Wilson, Joseph W; Cadilla, Rodolfo; Turnbull, Philip

    2016-01-14

    A series of selective androgen receptor modulators (SARMs) containing the 1-(trifluoromethyl)benzyl alcohol core have been optimized for androgen receptor (AR) potency and drug-like properties. We have taken advantage of the lipophilic ligand efficiency (LLE) parameter as a guide to interpret the effect of structural changes on AR activity. Over the course of optimization efforts the LLE increased over 3 log units leading to a SARM 43 with nanomolar potency, good aqueous kinetic solubility (>700 μM), and high oral bioavailability in rats (83%).

  2. Structural basis for AMPA receptor activation and ligand selectivity

    DEFF Research Database (Denmark)

    Hogner, A; Kastrup, Jette Sandholm Jensen; Jin, R;

    2002-01-01

    and binding experiments, has been used to increase our knowledge concerning the ionotropic glutamate receptor GluR2 at the molecular level. Five high-resolution X-ray structures of the ligand-binding domain of GluR2 (S1S2J) complexed with the three agonists (S)-2-amino-3-[3-hydroxy-5-(2-methyl-2H-tetrazol-5...

  3. Modulation of Opioid Receptor Ligand Affinity and Efficacy Using Active and Inactive State Receptor Models

    OpenAIRE

    Anand, Jessica P.; Purington, Lauren C.; Pogozheva, Irina D.; Traynor, John R.; Mosberg, Henry I.

    2012-01-01

    Mu opioid receptor (MOR) agonists are widely used for the treatment of pain; however chronic use results in the development of tolerance and dependence. It has been demonstrated that co-administration of a MOR agonist with a delta opioid receptor (DOR) antagonist maintains the analgesia associated with MOR agonists, but with reduced negative side effects. Using our newly refined opioid receptor models for structure-based ligand design, we have synthesized several pentapeptides with tailored a...

  4. Adenosine A{sub 1} receptors in contrast media-induced renal dysfunction in the normal rat

    Energy Technology Data Exchange (ETDEWEB)

    Liss, Per; Palm, Fredrik [Department of Diagnostic Radiology, University Hospital, 75185, Uppsala (Sweden); Department of Medical Cell Biology, University Hospital, 75185, Uppsala (Sweden); Carlsson, Per-Ola [Department of Medical Cell Biology, University Hospital, 75185, Uppsala (Sweden); Department of Medical Sciences, University Hospital, 75185, Uppsala (Sweden); Hansell, Peter [Department of Medical Cell Biology, University Hospital, 75185, Uppsala (Sweden)

    2004-07-01

    Renal vasoconstriction with resultant tissue hypoxia, especially in the renal medulla, has been suggested to play a role in contrast media (CM)-induced nephropathy. In this study we investigated the effects of injection of the non-ionic low-osmolar CM iopromide with and without pretreatment with the selective adenosine A{sub 1}-receptor antagonist DPCPX. The effects were evaluated on regional renal blood flow, outer medullary oxygen tension (PO{sub 2}) and urine output in normal anaesthetised rats. A laser-Doppler technique was used for recording haemodynamic changes while oxygen microelectrodes were used for oxygen measurements. The A{sub 1}-receptor antagonist per se elevated glomerular filtration rate (+44%), cortical blood flow (+15%) and urine output (threefold) while reducing outer medullary PO{sub 2} (-24%). Administration of CM reduced outer medullary blood flow (OMBF; -26%) and PO{sub 2} (-80%) but did not affect cortical blood flow. Urine output increased 28-fold by CM while arterial blood pressure was reduced. The CM-mediated effect on haemodynamics, PO{sub 2}, urine output and blood pressure was unaffected by the A{sub 1}-receptor antagonist. Adenosine A{sub 1}-receptors are not important mediators of the depression of outer medullary blood flow and PO{sub 2} caused by the CM iopromide in the normal rat; however, A{sub 1}-receptors are tonically active to regulate renal haemodynamics, PO{sub 2} and urine production during normal physiological conditions. (orig.)

  5. Genetic blockade of adenosine A2A receptors induces cognitive impairments and anatomical changes related to psychotic symptoms in mice.

    Science.gov (United States)

    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. PMID:27133030

  6. Steroid receptors and their ligands: Effects on male gamete functions

    Energy Technology Data Exchange (ETDEWEB)

    Aquila, Saveria; De Amicis, Francesca, E-mail: francesca.deamicis@unical.it

    2014-11-01

    In recent years a new picture of human sperm biology is emerging. It is now widely recognized that sperm contain nuclear encoded mRNA, mitochondrial encoded RNA and different transcription factors including steroid receptors, while in the past sperm were considered incapable of transcription and translation. One of the main targets of steroid hormones and their receptors is reproductive function. Expression studies on Progesterone Receptor, estrogen receptor, androgen receptor and their specific ligands, demonstrate the presence of these systems in mature spermatozoa as surface but also as nuclear conventional receptors, suggesting that both systemic and local steroid hormones, through sperm receptors, may influence male reproduction. However, the relationship between the signaling events modulated by steroid hormones and sperm fertilization potential as well as the possible involvement of the specific receptors are still controversial issues. The main line of this review highlights the current research in human sperm biology examining new molecular systems of response to the hormones as well as specific regulatory pathways controlling sperm cell fate and biological functions. Most significant studies regarding the identification of steroid receptors are reported and the mechanistic insights relative to signaling pathways, together with the change in sperm metabolism energy influenced by steroid hormones are discussed.The reviewed evidences suggest important effects of Progesterone, Estrogen and Testosterone and their receptors on spermatozoa and implicate the involvement of both systemic and local steroid action in the regulation of male fertility potential. - Highlights: • One of the main targets of steroid hormones and their receptors is reproductive function. • Pg/PR co-work to stimulate enzymatic activities to sustain a capacitation process. • E2/ERs regulate sperm motility, capacitation and acrosome reaction and act as survival factors. • Androgens

  7. Adenosine A1 receptor-mediated transactivation of the EGF receptor produces a neuroprotective effect on cortical neurons in vitro

    Institute of Scientific and Technical Information of China (English)

    Ke-qiang XIE; Li-min ZHANG; Yan CAO; Jun ZHU; Lin-yin FENG

    2009-01-01

    Aim:To understand the mechanism of the transactivation of the epidermal growth factor receptor (EGFR) mediated by the adenosine A1 receptor (A1R).Methods:Primary cultured rat cortical neurons subjected to oxygen-glucose deprivation (OGD) and HEK293/A1R cells were treated with the A1R-specific agonist N6-cyclopentyladenosine (CPA).Phospho-EGFR,Akt,and ERK1/2 were observed by Western blot.An interaction between EGFR and AIR was detected using immunoprecipitation and immunocytochemistry.Results:The A1R agonist CPA causes protein kinase B (Akt) activation and protects primary cortical neurons from oxygen-glucose deprivation (OGD) insult.A1R and EGFR co-localize in the membranes of neurons and form an immunocomplex.A1R stimulation induces significant EGFR phosphorylation via a P13K and Src kinase signaling pathway;this stimulation provides a neuroprotective effect in cortical neurons.CPA leads to sustained phosphorylation of extracellularly regulated kinases 1 and 2 (ERK1/2) in cortical neurons,but only to transient phosphorylation in HEK 293/A1R cells.The response to the AtR agonist is mediated primarily through EGFR trans-activation that is dependent on pertussis toxin (PTX)-sensitive G1 protein and metalloproteases in HEK 293/A1R.Conclusion:A1R-mediated EGFR transactivation confers a neuroprotective effect in primary cortical neurons.P13 kinase and Src kinase play pivotal roles in this response.

  8. Feedback, receptor clustering, and receptor restriction to single cells yield large Turing spaces for ligand-receptor-based Turing models

    Science.gov (United States)

    Kurics, Tamás; Menshykau, Denis; Iber, Dagmar

    2014-08-01

    Turing mechanisms can yield a large variety of patterns from noisy, homogenous initial conditions and have been proposed as patterning mechanism for many developmental processes. However, the molecular components that give rise to Turing patterns have remained elusive, and the small size of the parameter space that permits Turing patterns to emerge makes it difficult to explain how Turing patterns could evolve. We have recently shown that Turing patterns can be obtained with a single ligand if the ligand-receptor interaction is taken into account. Here we show that the general properties of ligand-receptor systems result in very large Turing spaces. Thus, the restriction of receptors to single cells, negative feedbacks, regulatory interactions among different ligand-receptor systems, and the clustering of receptors on the cell surface all greatly enlarge the Turing space. We further show that the feedbacks that occur in the FGF10-SHH network that controls lung branching morphogenesis are sufficient to result in large Turing spaces. We conclude that the cellular restriction of receptors provides a mechanism to sufficiently increase the size of the Turing space to make the evolution of Turing patterns likely. Additional feedbacks may then have further enlarged the Turing space. Given their robustness and flexibility, we propose that receptor-ligand-based Turing mechanisms present a general mechanism for patterning in biology.

  9. Targeting the inflammasome and adenosine type-3 receptors improves outcome of antibiotic therapy in murine anthrax

    Institute of Scientific and Technical Information of China (English)

    Serguei; G; Popov; Taissia; G; Popova; Fatah; Kashanchi; Charles; Bailey

    2011-01-01

    AIM:To establish whether activation of adenosine type-3 receptors(A3Rs)and inhibition of interleukin- 1β-induced inflammation is beneficial in combination with antibiotic therapy to increase survival of mice challenged with anthrax spores. METHODS:DBA/2 mice were challenged with Bacillus anthracis spores of the toxigenic Sterne strain 43F2. Survival of animals was monitored for 15 d.Ciprofloxacin treatment(50 mg/kg,once daily,intraperitoneally) was initiated at day+1 simultaneously with the ad- ministration of inhibitors,and continued for 10 d.Two doses(2.5 mg/kg and 12.5 mg/kg)of acetyl-tyrosylvalyl-alanyl-aspartyl-chloromethylketone(YVAD)and three doses(0.05,0.15 and 0.3 mg/kg)of 1-[2-Chloro- 6-[[(3-iodophenyl)methyl]amino]-9H-purin-9-yl]-1- deoxy-N-methyl-β-D-ribofuranuronamide(Cl-IB-MECA) were tested.Animals received YVAD on days 1-4,and Cl-IB-MECA on days 1-10 once daily,subcutaneously. Human lung epithelial cells in culture were challenged with spores or edema toxin and the effects of IB-MECAon phosphorylation of AKT and generation of cAMP were tested. RESULTS:We showed that the outcome of antibiotic treatment in a murine anthrax model could be substantially improved by co-administration of the caspase-1/4 inhibitor YVAD and the A3R agonist Cl-IB-MECA.Combination treatment with these substances and ciprofloxacin resulted in up to 90%synergistic protection.All untreated mice died,and antibiotic alone protected only 30% of animals.We conclude that both substances target the aberrant host signaling that underpins anthrax mortality. CONCLUSION:Our findings suggest new possibilities for combination therapy of anthrax with antibiotics,A3R agonists and caspase-1 inhibitors.

  10. Central or peripheral delivery of an adenosine A1 receptor agonist improves mechanical allodynia in a mouse model of painful diabetic neuropathy.

    Science.gov (United States)

    Katz, N K; Ryals, J M; Wright, D E

    2015-01-29

    Diabetic peripheral neuropathy is a common complication of diabetes mellitus, and a significant proportion of individuals suffer debilitating pain that significantly affects their quality of life. Unfortunately, symptomatic treatment options have limited efficacy, and often carry significant risk of systemic adverse effects. Activation of the adenosine A1 receptor (A1R) by the analgesic small molecule adenosine has been shown to have antinociceptive benefits in models of inflammatory and neuropathic pain. The current study used a mouse model of painful diabetic neuropathy to determine the effect of diabetes on endogenous adenosine production, and if central or peripheral delivery of adenosine receptor agonists could alleviate signs of mechanical allodynia in diabetic mice. Diabetes was induced using streptozocin in male A/J mice. Mechanical withdrawal thresholds were measured weekly to characterize neuropathy phenotype. Hydrolysis of AMP into adenosine by ectonucleotidases was determined in the dorsal root ganglia (DRG) and spinal cord at 8 weeks post-induction of diabetes. AMP, adenosine and the specific A1R agonist, N(6)-cyclopentyladenosine (CPA), were administered both centrally (intrathecal) and peripherally (intraplantar) to determine the effect of activation of adenosine receptors on mechanical allodynia in diabetic mice. Eight weeks post-induction, diabetic mice displayed significantly decreased hydrolysis of extracellular AMP in the DRG; at this same time, diabetic mice displayed significantly decreased mechanical withdrawal thresholds compared to nondiabetic controls. Central delivery AMP, adenosine and CPA significantly improved mechanical withdrawal thresholds in diabetic mice. Surprisingly, peripheral delivery of CPA also improved mechanical allodynia in diabetic mice. This study provides new evidence that diabetes significantly affects endogenous AMP hydrolysis, suggesting that altered adenosine production could contribute to the development of

  11. Halogenated benzamides as ligands for cerebral dopamine receptors

    International Nuclear Information System (INIS)

    In the past several years the authors' has synthesized a series of high affinity iodine-123 and fluorine-18 labeled substituted benzamide ligands for SPECT and PET visualization of the dopamine D-2 receptors in brain regions with low receptor density outside the striatum. Radioiodination and radiofluorination in high yield and high specific activity was achieved by using the tributyltin precursor and nucleophilic displacement of the saturation analysis revealed that the optimal striatum-to-cerebellum uptake ratio in the rat brain is highly correlated with the product of Kw and KD. The authors have used [125I] and [123I] epidepride to detect extra striatal dopamine D2 receptors in vitro by saturation analysis and in vivo with high resolution SPECT imaging

  12. Perinatal caffeine, acting on maternal adenosine A(1 receptors, causes long-lasting behavioral changes in mouse offspring.

    Directory of Open Access Journals (Sweden)

    Olga Björklund

    Full Text Available BACKGROUND: There are lingering concerns about caffeine consumption during pregnancy or the early postnatal period, partly because there may be long-lasting behavioral changes after caffeine exposure early in life. METHODOLOGY/PRINCIPAL FINDINGS: We show that pregnant wild type (WT mice given modest doses of caffeine (0.3 g/l in drinking water gave birth to offspring that as adults exhibited increased locomotor activity in an open field. The offspring also responded to cocaine challenge with greater locomotor activity than mice not perinatally exposed to caffeine. We performed the same behavioral experiments on mice heterozygous for adenosine A(1 receptor gene (A(1RHz. In these mice signaling via adenosine A(1 receptors is reduced to about the same degree as after modest consumption of caffeine. A(1RHz mice had a behavioral profile similar to WT mice perinatally exposed to caffeine. Furthermore, it appeared that the mother's genotype, not offspring's, was critical for behavioral changes in adult offspring. Thus, if the mother partially lacked A(1 receptors the offspring displayed more hyperactivity and responded more strongly to cocaine stimulation as adults than did mice of a WT mother, regardless of their genotype. This indicates that long-term behavioral alterations in the offspring result from the maternal effect of caffeine, and not a direct effect on fetus. WT offspring from WT mother but having a A(1R Hz grandmother preserved higher locomotor response to cocaine. CONCLUSIONS/SIGNIFICANCE: We suggest that perinatal caffeine, by acting on adenosine A(1 receptors in the mother, causes long-lasting behavioral changes in the offspring that even manifest themselves in the second generation.

  13. Acute hyperammonemia and systemic inflammation is associated with increased extracellular brain adenosine in rats

    DEFF Research Database (Denmark)

    Bjerring, Peter Nissen; Dale, Nicholas; Larsen, Fin Stolze

    2015-01-01

    Acute liver failure (ALF) can lead to brain edema, cerebral hyperperfusion and intracranial hypertension. These complications are thought to be mediated by hyperammonemia and inflammation leading to altered brain metabolism. As increased levels of adenosine degradation products have been found...... in brain tissue of patients with ALF we investigated whether hyperammonemia could induce adenosine release in brain tissue. Since adenosine is a potent vasodilator and modulator of cerebral metabolism we furthermore studied the effect of adenosine receptor ligands on intracranial pressure (ICP......) and cerebral blood flow (CBF). We measured the adenosine concentration with biosensors in rat brain slices exposed to ammonia and in a rat model with hyperammonemia and systemic inflammation. Exposure to ammonia in concentrations from 0.15-10 mM led to increases in the cortical adenosine concentration up to 18...

  14. Allosteric modulation and constitutive activity of fusion proteins between the adenosine A1 receptor and different 351Cys-mutated Gi α-subunits

    NARCIS (Netherlands)

    Klaasse, E.; Ligt, R.A.F.de; Roerink, S.F.; Lorenzen, A.; Milligan, G.; Leurs, R.; IJzerman, A.P.

    2004-01-01

    We studied fusion proteins between the human adenosine A1 receptor and different 351Cys-mutated Gi1 α-subunits (A1-Giα) with respect to two important concepts in receptor pharmacology, i.e. allosteric modulation and constitutive activity/inverse agonism. The aim of our study was twofold. We first an

  15. In vivo evaluation of [11C]preladenant positron emission tomography for quantification of adenosine A2A receptors in the rat brain

    NARCIS (Netherlands)

    Zhou, Xiaoyun; Khanapur, Shivashankar; de Jong, Johan R; Willemsen, Antoon T.M.; Dierckx, Rudi Ajo; Elsinga, Philip H; de Vries, Erik Fj

    2016-01-01

    [(11)C]Preladenant was developed as a novel adenosine A2A receptor positron emission tomography radioligand. The present study aims to evaluate the suitability of [(11)C]preladenant positron emission tomography for the quantification of striatal A2A receptor density and the assessment of striatal A2

  16. 1,2,4-Triazolo[1,5-a]quinoxaline derivatives: synthesis and biological evaluation as adenosine receptor antagonists.

    Science.gov (United States)

    Catarzi, Daniela; Colotta, Vittoria; Varano, Flavia; Filacchioni, Guido; Martini, Claudia; Trincavelli, Letizia; Lucacchini, Antonio

    2004-02-01

    Since most of the reported adenosine receptor antagonists are 2-(hetero)aryl-substituted tricyclic heteroaromatic derivatives, in the present study we report the synthesis and the biological evaluation of a new set of 4-amino-1,2,4-triazolo[1,5-a]quinoxalines containing at position-2 an ethyl carboxylate group or a hydrogen atom. The structure-activity relationships on these compounds were in accordance with those of a previously reported series of analogous size and shape, thus suggesting a similar A(1)-binding mode. In particular, the binding data indicate that alkylation of the 4-amino group of these derivatives lead to potent A(1)-receptor antagonists. Moreover, as new results, this study has pointed out that the ethyl 2-carboxylate group can advantageously replace the 2-(hetero)aryl ring of previously reported triazoloquinoxaline derivatives, affording an ameliorated interaction with the A(1)-receptor subtype.

  17. Presynaptic Adenosine Receptor-Mediated Regulation of Diverse Thalamocortical Short-Term Plasticity in the Mouse Whisker Pathway.

    Science.gov (United States)

    Ferrati, Giovanni; Martini, Francisco J; Maravall, Miguel

    2016-01-01

    Short-term synaptic plasticity (STP) sets the sensitivity of a synapse to incoming activity and determines the temporal patterns that it best transmits. In "driver" thalamocortical (TC) synaptic populations, STP is dominated by depression during stimulation from rest. However, during ongoing stimulation, lemniscal TC connections onto layer 4 neurons in mouse barrel cortex express variable STP. Each synapse responds to input trains with a distinct pattern of depression or facilitation around its mean steady-state response. As a result, in common with other synaptic populations, lemniscal TC synapses express diverse rather than uniform dynamics, allowing for a rich representation of temporally varying stimuli. Here, we show that this STP diversity is regulated presynaptically. Presynaptic adenosine receptors of the A1R type, but not kainate receptors (KARs), modulate STP behavior. Blocking the receptors does not eliminate diversity, indicating that diversity is related to heterogeneous expression of multiple mechanisms in the pathway from presynaptic calcium influx to neurotransmitter release.

  18. Presynaptic adenosine receptor-mediated regulation of diverse thalamocortical short-term plasticity in the mouse whisker pathway

    Directory of Open Access Journals (Sweden)

    Giovanni eFerrati

    2016-02-01

    Full Text Available Short-term synaptic plasticity (STP sets the sensitivity of a synapse to incoming activity and determines the temporal patterns that it best transmits. In driver thalamocortical (TC synaptic populations, STP is dominated by depression during stimulation from rest. However, during ongoing stimulation, lemniscal TC connections onto layer 4 neurons in mouse barrel cortex express variable STP. Each synapse responds to input trains with a distinct pattern of depression or facilitation around its mean steady-state response. As a result, in common with other synaptic populations, lemniscal TC synapses express diverse rather than uniform dynamics, allowing for a rich representation of temporally varying stimuli. Here we show that this STP diversity is regulated presynaptically. Presynaptic adenosine receptors of the A1R type, but not kainate receptors, modulate STP behavior. Blocking the receptors does not eliminate diversity, indicating that diversity is related to heterogeneous expression of multiple mechanisms in the pathway from presynaptic calcium influx to neurotransmitter release.

  19. Somatostatin receptor ligands and resistance to treatment in pituitary adenomas.

    Science.gov (United States)

    Cuevas-Ramos, Daniel; Fleseriu, Maria

    2014-06-01

    Somatostatin (SST), an inhibitory polypeptide with two biologically active forms SST14 and SST28, inhibits GH, prolactin (PRL), TSH, and ACTH secretion in the anterior pituitary gland. SST also has an antiproliferative effect inducing cell cycle arrest and apoptosis. Such actions are mediated through five G-protein-coupled somatostatin receptors (SSTR): SSTR1-SSTR5. In GH-secreting adenomas, SSTR2 expression predominates, and somatostatin receptor ligands (SRLs; octreotide and lanreotide) directed to SSTR2 are presently the mainstays of medical therapy. However, about half of patients show incomplete biochemical remission, but the definition of resistance per se remains controversial. We summarize here the determinants of SRL resistance in acromegaly patients, including clinical, imaging features as well as molecular (mutations, SSTR variants, and polymorphisms), and histopathological (granulation pattern, and proteins and receptor expression) predictors. The role of SSTR5 may explain the partial responsiveness to SRLs in patients with adequate SSTR2 density in the cell membrane. In patients with ACTH-secreting pituitary adenomas, i.e. Cushing's disease (CD), SSTR5 is the most abundant receptor expressed and tumors show low SSTR2 density due to hypercortisolism-induced SSTR2 down-regulation. Clinical studies with pasireotide, a multireceptor-targeted SRL with increased SSTR5 activity, lead to approval of pasireotide for treatment of patients with CD. Other SRL delivery modes (oral octreotide), multireceptor-targeted SRL (somatoprim) or chimeric compounds targeting dopamine D2 receptors and SSTR2 (dopastatin), are briefly discussed. PMID:24647046

  20. Identification of Putative Receptors for the Novel Adipokine CTRP3 Using Ligand-Receptor Capture Technology

    Science.gov (United States)

    Li, Ying; Ozment, Tammy; Wright, Gary L.

    2016-01-01

    C1q TNF Related Protein 3 (CTRP3) is a member of a family of secreted proteins that exert a multitude of biological effects. Our initial work identified CTRP3’s promise as an effective treatment for Nonalcoholic fatty liver disease (NAFLD). Specifically, we demonstrated that mice fed a high fat diet failed to develop NAFLD when treated with CTRP3. The purpose of this current project is to identify putative receptors which mediate the hepatic actions of CTRP3. Methods We used Ligand-receptor glycocapture technology with TriCEPS™-based ligand-receptor capture (LRC-TriCEPS; Dualsystems Biotech AG). The LRC-TriCEPS experiment with CTRP3-FLAG protein as ligand and insulin as a control ligand was performed on the H4IIE rat hepatoma cell line. Results Initial analysis demonstrated efficient coupling of TriCEPS to CTRP3. Further, flow cytometry analysis (FACS) demonstrated successful oxidation and crosslinking of CTRP3-TriCEPS and Insulin-TriCEPS complexes to cell surface glycans. Demonstrating the utility of TriCEPS under these conditions, the insulin receptor was identified in the control dataset. In the CTRP3 treated cells a total enrichment of 261 peptides was observed. From these experiments 5 putative receptors for CTRP3 were identified with two reaching statistically significance: Lysosomal-associated membrane protein 1 (LAMP-1) and Lysosome membrane protein 2 (LIMP II). Follow-up Co-immunoprecipitation analysis confirmed the association between LAMP1 and CTRP3 and further testing using a polyclonal antibody to block potential binding sites of LAMP1 prevented CTRP3 binding to the cells. Conclusion The LRC-TriCEPS methodology was successful in identifying potential novel receptors for CTRP3. Relevance The identification of the receptors for CTRP3 are important prerequisites for the development of small molecule drug candidates, of which none currently exist, for the treatment NAFLD. PMID:27727322

  1. Cherry-picked ligands at histamine receptor subtypes.

    Science.gov (United States)

    Sadek, Bassem; Stark, Holger

    2016-07-01

    Histamine, a biogenic amine, is considered as a principle mediator of multiple physiological effects through binding to its H1, H2, H3, and H4 receptors (H1-H4Rs). Currently, the HRs have gained attention as important targets for the treatment of several diseases and disorders ranging from allergy to Alzheimer's disease and immune deficiency. Accordingly, medicinal chemistry studies exploring histamine-like molecules and their physicochemical properties by binding and interacting with the four HRs has led to the development of a diversity of agonists and antagonists that display selectivity for each HR subtype. An overview on H1-R4Rs and developed ligands representing some key steps in development is provided here combined with a short description of structure-activity relationships for each class. Main chemical diversities, pharmacophores, and pharmacological profiles of most innovative H1-H4R agonists and antagonists are highlighted. Therefore, this overview should support the rational choice for the optimal ligand selection based on affinity, selectivity and efficacy data in biochemical and pharmacological studies. This article is part of the Special Issue entitled 'Histamine Receptors'. PMID:26581501

  2. Membrane omega-3 fatty acids modulate the oligomerisation kinetics of adenosine A2A and dopamine D2 receptors

    Science.gov (United States)

    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.

  3. The Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) Uses its C-Terminus to Regulate the A2B Adenosine Receptor.

    Science.gov (United States)

    Watson, Michael J; Lee, Shernita L; Marklew, Abigail J; Gilmore, Rodney C; Gentzsch, Martina; Sassano, Maria F; Gray, Michael A; Tarran, Robert

    2016-01-01

    CFTR is an apical membrane anion channel that regulates fluid homeostasis in many organs including the airways, colon, pancreas and sweat glands. In cystic fibrosis, CFTR dysfunction causes significant morbidity/mortality. Whilst CFTR's function as an ion channel has been well described, its ability to regulate other proteins is less understood. We have previously shown that plasma membrane CFTR increases the surface density of the adenosine 2B receptor (A2BR), but not of the β2 adrenergic receptor (β2AR), leading to an enhanced, adenosine-induced cAMP response in the presence of CFTR. In this study, we have found that the C-terminal PDZ-domain of both A2BR and CFTR were crucial for this interaction, and that replacing the C-terminus of A2BR with that of β2AR removed this CFTR-dependency. This observation extended to intact epithelia and disruption of the actin cytoskeleton prevented A2BR-induced but not β2AR-induced airway surface liquid (ASL) secretion. We also found that CFTR expression altered the organization of the actin cytoskeleton and PDZ-binding proteins in both HEK293T cells and in well-differentiated human bronchial epithelia. Furthermore, removal of CFTR's PDZ binding motif (ΔTRL) prevented actin rearrangement, suggesting that CFTR insertion in the plasma membrane results in local reorganization of actin, PDZ binding proteins and certain GPCRs. PMID:27278076

  4. The effects of methylmercury on motor activity are sex- and age-dependent, and modulated by genetic deletion of adenosine receptors and caffeine administration.

    Science.gov (United States)

    Björklund, Olga; Kahlström, Johan; Salmi, Peter; Ogren, Sven Ove; Vahter, Marie; Chen, Jiang-Fan; Fredholm, Bertil B; Daré, Elisabetta

    2007-11-30

    Adenosine and its receptors are, as part of the brain stress response, potential targets for neuroprotective drugs. We have investigated if the adenosine receptor system affects the developmental neurotoxicity caused by the fish pollutant methylmercury (MeHg). Behavioral outcomes of low dose perinatal MeHg exposure were studied in mice where the A(1) and A(2A) adenosine receptors were either partially blocked by caffeine treatment or eliminated by genetic modification (A(1)R and A(2A)R knock-out mice). From gestational day 7 to day 7 of lactation dams were administered doses that mimic human intake via normal diet, i.e. 1microM MeHg and/or 0.3g/l caffeine in the drinking water. This exposure to MeHg resulted in a doubling of brain Hg levels in wild type females and males at postnatal day 21 (PND21). Open field analysis was performed at PND21 and 2 months of age. MeHg caused time-dependent behavioral alterations preferentially in male mice. A decreased response to amphetamine in 2-month-old males pointed to disturbances in dopaminergic functions. Maternal caffeine intake induced long-lasting changes in the offspring evidenced by an increased motor activity and a modified response to psychostimulants in adult age, irrespectively of sex. Similar alterations were observed in A(1)R knock-out mice, suggesting that adenosine A(1) receptors are involved in the alterations triggered by caffeine exposure during development. Perinatal caffeine treatment and, to some extent, genetic elimination of adenosine A(1) receptors, attenuated the behavioral consequences of MeHg in males. Importantly, also deletion of the A(2A) adenosine receptor reduced the vulnerability to MeHg, consistent with the neuroprotective effects of adenosine A(2A) receptor inactivation observed in hypoxia and Parkinson's disease. Thus, the consequences of MeHg toxicity during gestation and lactation can be reduced by adenosine A(1) and A(2A) receptor inactivation, either via their genetic deletion or by

  5. Preparation and first evaluation of [{sup 18}F]FE-SUPPY: a new PET tracer for the adenosine A{sub 3} receptor

    Energy Technology Data Exchange (ETDEWEB)

    Wadsak, Wolfgang [Dept. of Nuclear Medicine, Medical Univ. of Vienna, A-1090 Vienna (Austria)]|[Dept. of Inorganic Chemistry, Univ. of Vienna, A-1090 Vienna (Austria); Mien, Leonhard-Key [Dept. of Nuclear Medicine, Medical Univ. of Vienna, A-1090 Vienna (Austria)]|[Dept. of Pharmaceutical Technology and Biopharmaceutics, Univ. of Vienna, A-1090 Vienna (Austria)]|[Dept. of Psychiatry and Psychotherapy, Medical Univ. of Vienna, A-1090 Vienna (Austria); Shanab, Karem [Dept. of Drug and Natural Product Synthesis, Faculty of Life Sciences, Univ. of Vienna, A-1090 Vienna (Austria); Ettlinger, Dagmar E. [Dept. of Nuclear Medicine, Medical Univ. of Vienna, A-1090 Vienna (Austria); Haeusler, Daniela [Dept. of Nuclear Medicine, Medical Univ. of Vienna, A-1090 Vienna (Austria)]|[Dept. of Pharmaceutical Technology and Biopharmaceutics, Univ. of Vienna, A-1090 Vienna (Austria); Sindelar, Karoline [Dept. of Nuclear Medicine, Medical Univ. of Vienna, A-1090 Vienna (Austria); Lanzenberger, Rupert R. [Dept. of Psychiatry and Psychotherapy, Medical Univ. of Vienna, A-1090 Vienna (Austria); Spreitzer, Helmut [Dept. of Drug and Natural Product Synthesis, Faculty of Life Sciences, Univ. of Vienna, A-1090 Vienna (Austria); Viernstein, Helmut [Dept. of Pharmaceutical Technology and Biopharmaceutics, Univ. of Vienna, A-1090 Vienna (Austria); Keppler, Bernhard K. [Dept. of Inorganic Chemistry, Univ. of Vienna, A-1090 Vienna (Austria); Dudczak, Robert; Kletter, Kurt [Dept. of Nuclear Medicine, Medical Univ. of Vienna, A-1090 Vienna (Austria); Mitterhauser, Markus [Dept. of Nuclear Medicine, Medical Univ. of Vienna, A-1090 Vienna (Austria)]|[Dept. of Pharmaceutical Technology and Biopharmaceutics, Univ. of Vienna, A-1090 Vienna (Austria)]|[Hospital Pharmacy of the General Hospital of Vienna, A-1090 Vienna (Austria)], E-mail: markus.mitterhouser@meduniwien.ac.at

    2008-01-15

    Introduction: Changes of the adenosine A{sub 3} receptor subtype (A3AR) expression have been shown in a variety of pathologies, especially neurological and affective disorders, cardiac diseases and oncological and inflammation processes. Recently, 5-(2-fluoroethyl) 2,4-diethyl-3-(ethylsulfanylcarbonyl)-6-phenylpyridine-5-carboxylate (FE-SUPPY) was presented as a high-affinity ligand for the A3AR with good selectivity. Our aims were the development of a suitable labeling precursor, the establishment of a reliable radiosynthesis for the fluorine-18-labeled analogue [{sup 18}F]FE-SUPPY and a first evaluation of [{sup 18}F]FE-SUPPY in rats. Methods: [{sup 18}F]FE-SUPPY was prepared in a feasible and reliable manner by radiofluorination of the corresponding tosylated precursor. Biodistribution was carried out in rats, and organs were removed and counted. Autoradiography was performed on rat brain slices in the presence or absence of 2-Cl-IB-MECA. Results: Overall yields and radiochemical purity were sufficient for further preclinical and clinical applications. The uptake pattern of [{sup 18}F]FE-SUPPY found in rats mainly followed the described mRNA distribution pattern of the A3AR. Specific uptake in brain was demonstrated by blocking with a selective A3AR agonist. Conclusion: We conclude that [{sup 18}F]FE-SUPPY has the potential to serve as the first positron emission tomography tracer for the A3AR.

  6. Biotinylated recombinant human erythropoietins: Bioactivity and utility as receptor ligand

    Energy Technology Data Exchange (ETDEWEB)

    Wojchowski, D.M.; Caslake, L. (Pennsylvania State Univ., University Park (USA))

    1989-08-15

    Recombinant human erythropoietin labeled covalently with biotin at sialic acid moieties has been prepared, and has been shown to possess high biological activity plus utility as a receptor ligand. Initially, the effects on biological activity of covalently attaching biotin to erythropoietin alternatively at carboxylate, amino, or sialic acid groups were compared. Biotinylation of erythropoietin at carboxylate groups using biotin-amidocaproyl hydrazide plus 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide led to substantial biological inactivation, although biotinylated molecules retained detectable activity when prepared at low stoichiometries. Biotinylation at amino groups using sulfosuccinimidyl 6-(biotinamido) hexanoate resulted in a high level of biological inactivation with little, if any, retention of biological activity, regardless of labeling stoichiometries. Biotinylation at sialic acid moieties using periodate and biotinamidocaproyl hydrazide proceeded efficiently (greater than 95% and 80% labeling efficiencies for human urinary and recombinant erythropoietin, respectively) and yielded stably biotinylated erythropoietin molecules possessing comparably high biological activity (ie, 45% of the activity of unmodified hormone). Utility of recombinant biotin-(sialyl)-erythropoietin (in combination with 125I-streptavidin) in the assay of cell surface receptors was demonstrated using two distinct murine erythroleukemia cell lines, Friend 745 and Rauscher Red 1. The densities and affinities of specific hormone binding sites were 116 +/- 4 sites, 3.3 +/- 0.4 nmol/L kd and 164 +/- 5 sites, 2.7 +/- 0.4 nmol/L kd, respectively. It is predicted that the present development of biotin-(sialyl)-erythropoietin as a chemically and biologically stable, bioactive ligand will assist in advancing an understanding of the regulated expression and physicochemistry of the human and murine erythropoietin receptors.

  7. Characterization and ligand identification of a membrane progesterone receptor in fungi: existence of a novel PAQR in Sporothrix schenckii

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    Gonzalez-Velazquez Waleska

    2012-09-01

    Full Text Available Abstract Background Adaptive responses in fungi result from the interaction of membrane receptors and extracellular ligands. Many different classes of receptors have been described in eukaryotic cells. Recently a new family of receptors classified as belonging to the progesterone-adiponectin receptor (PAQR family has been identified. These receptors have the seven transmembrane domains characteristic of G-protein coupled receptors, but their activity has not been associated directly to G proteins. They share sequence similarity to the eubacterial hemolysin III proteins. Results A new receptor, SsPAQR1 (Sporothrixschenckiiprogesterone-adiponectinQ receptor1, was identified as interacting with Sporothrix schenckii G protein alpha subunit SSG-2 in a yeast two-hybrid assay. The receptor was identified as a member of the PAQR family. The cDNA sequence revealed a predicted ORF of 1542 bp encoding a 514 amino acids protein with a calculated molecular weight of 57.8 kDa. Protein domain analysis of SsPAQR1 showed the 7 transmembrane domains (TM characteristic of G protein coupled receptors and the presence of the distinctive motifs that characterize PAQRs. A yeast-based assay specific for PAQRs identified progesterone as the agonist. S. schenckii yeast cells exposed to progesterone (0.50 mM showed an increase in intracellular levels of 3′, 5′ cyclic adenosine monophosphate (cAMP within the first min of incubation with the hormone. Different progesterone concentrations were tested for their effect on the growth of the fungus. Cultures incubated at 35°C did not grow at concentrations of progesterone of 0.05 mM or higher. Cultures incubated at 25°C grew at all concentrations tested (0.01 mM-0.50 mM with growth decreasing gradually with the increase in progesterone concentration. Conclusion This work describes a receptor associated with a G protein alpha subunit in S. schenckii belonging to the PAQR family. Progesterone was identified as the ligand

  8. Activation profiles of opioid ligands in HEK cells expressing δ opioid receptors

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    Clark J

    2002-11-01

    Full Text Available Abstract Background The aim of the present study was to characterize the activation profiles of 15 opioid ligands in transfected human embryonic kidney cells expressing only δ opioid receptors. Activation profiles of most of these ligands at δ opioid receptors had not been previously characterized in vitro. Receptor activation was assessed by measuring the inhibition of forskolin-stimulated cAMP production. Results Naltrexone and nalorphine were classified as antagonists at δ opioid receptor. The other ligands studied were agonists at δ opioid receptors and demonstrated IC50 values of 0.1 nM to 2 μM, maximal inhibition of 39–77% and receptor binding affinities of 0.5 to 243 nM. The rank order of efficacy of the ligands tested was metazocine = xorphanol ≥ fentanyl = SKF 10047 = etorphine = hydromorphone = butorphanol = lofentanil > WIN 44,441 = Nalbuphine = cyclazocine ≥ met-enkephalin >> morphine = dezocine. For the first time these data describe and compare the function and relative efficacy of several ligands at δ opioid receptors. Conclusions The data produced from this study can lead to elucidation of the complete activation profiles of several opioid ligands, leading to clarification of the mechanisms involved in physiological effects of these ligands at δ opioid receptors. Furthermore, these data can be used as a basis for novel use of existing opioid ligands based on their pharmacology at δ opioid receptors.

  9. Adenosine in the tuberomammillary nucleus inhibits the histaminergic system via A1 receptors and promotes non-rapid eye movement sleep.

    Science.gov (United States)

    Oishi, Yo; Huang, Zhi-Li; Fredholm, Bertil B; Urade, Yoshihiro; Hayaishi, Osamu

    2008-12-16

    Adenosine has been proposed to promote sleep through A(1) receptors (A(1)R's) and/or A(2A) receptors in the brain. We previously reported that A(2A) receptors mediate the sleep-promoting effect of prostaglandin D(2), an endogenous sleep-inducing substance, and that activation of these receptors induces sleep and blockade of them by caffeine results in wakefulness. On the other hand, A(1)R has been suggested to increase sleep by inhibition of the cholinergic region of the basal forebrain. However, the role and target sites of A(1)R in sleep-wake regulation remained controversial. In this study, immunohistochemistry revealed that A(1)R was expressed in histaminergic neurons of the rat tuberomammillary nucleus (TMN). In vivo microdialysis showed that the histamine release in the frontal cortex was decreased by microinjection into the TMN of N(6)-cyclopentyladenosine (CPA), an A(1)R agonist, adenosine or coformycin, an inhibitor of adenosine deaminase, which catabolizes adenosine to inosine. Bilateral injection of CPA into the rat TMN significantly increased the amount and the delta power density of non-rapid eye movement (non-REM; NREM) sleep but did not affect REM sleep. CPA-promoted sleep was observed in WT mice but not in KO mice for A(1)R or histamine H(1) receptor, indicating that the NREM sleep promoted by A(1)R-specific agonist depended on the histaminergic system. Furthermore, the bilateral injection of adenosine or coformycin into the rat TMN increased NREM sleep, which was completely abolished by coadministration of 1,3-dimethyl-8-cyclopenthylxanthine, a selective A(1)R antagonist. These results indicate that endogenous adenosine in the TMN suppresses the histaminergic system via A(1)R to promote NREM sleep.

  10. Differential Expression of Adenosine P1 Receptor ADORA1 and ADORA2A Associated with Glioma Development and Tumor-Associated Epilepsy.

    Science.gov (United States)

    Huang, Jun; Chen, Ming-Na; Du, Juan; Liu, Hao; He, Yu-Jiao; Li, Guo-Liang; Li, Shu-Yu; Liu, Wei-Ping; Long, Xiao-Yan

    2016-07-01

    Level of adenosine, an endogenous astrocyte-based neuromodulator, is primarily regulated by adenosine P1 receptors. This study assessed expression of adenosine P1 receptors, ADORA1 (adenosine A1 receptor) and ADORA2A (adenosine A2a receptor) and their association with glioma development and epilepsy in glioma patients. Expression of ADORA1/ADORA2A was assessed immunohistochemically in 65 surgically removed glioma tissue and 21 peri-tumor tissues and 8 cases of normal brain tissues obtained from hematoma patients with cerebral trauma. Immunofluorescence, Western blot, and qRT-PCR were also used to verify immunohistochemical data. Adenosine P1 receptor ADORA1 and ADORA2A proteins were localized in the cell membrane and cytoplasm and ADORA1/ADORA2A immunoreactivity was significantly stronger in glioma and peri-tumor tissues that contained infiltrating tumor cells than in normal brain tissues (p < 0.05). The World Health Organization (WHO) grade III gliomas expressed even higher level of ADORA1 and ADORA2A. Western blot and qRT-PCR confirmed immunohistochemical data. Moreover, higher levels of ADORA1 and ADORA2A expression occurred in high-grade gliomas, in which incidence of epilepsy were lower (p < 0.05). In contrast, a lower level of ADORA1/ADORA2A expression was found in peri-tumor tissues with tumor cell presence from patients with epilepsy compared to patients without epilepsy (p < 0.05). The data from the current study indicates that dysregulation in ADORA1/ADORA2A expression was associated with glioma development, whereas low level of ADORA1/ADORA2A expression could increase susceptibility of tumor-associated epilepsy. PMID:27038930

  11. Ligands specify estrogen receptor alpha nuclear localization and degradation

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    Caze-Subra Stéphanie

    2010-12-01

    Full Text Available Abstract Background The estrogen receptor alpha (ERα is found predominately in the nucleus, both in hormone stimulated and untreated cells. Intracellular distribution of the ERα changes in the presence of agonists but the impact of different antiestrogens on the fate of ERα is a matter of debate. Results A MCF-7 cell line stably expressing GFP-tagged human ERα (SK19 cell line was created to examine the localization of ligand-bound GFP-ERα. We combined digitonin-based cell fractionation analyses with fluorescence and immuno-electron microscopy to determine the intracellular distribution of ligand-bound ERα and/or GFP-ERα. Using fluorescence- and electron microscopy we demonstrate that both endogenous ERα and GFP-ERα form numerous nuclear focal accumulations upon addition of agonist, 17β-estradiol (E2, and pure antagonists (selective estrogen regulator disruptor; SERD, ICI 182,780 or RU58,668, while in the presence of partial antagonists (selective estrogen regulator modulator; SERM, 4-hydroxytamoxifen (OHT or RU39,411, diffuse nuclear staining persisted. Digitonin based cell fractionation analyses confirmed that endogenous ERα and GFP-ERα predominantly reside in the nuclear fraction. Overall ERα protein levels were reduced after estradiol treatment. In the presence of SERMs ERα was stabilized in the nuclear soluble fraction, while in the presence of SERDs protein levels decreased drastically and the remaining ERα was largely found in a nuclear insoluble fraction. mRNA levels of ESR1 were reduced compared to untreated cells in the presence of all ligands tested, including E2. E2 and SERDs induced ERα degradation occurred in distinct nuclear foci composed of ERα and the proteasome providing a simple explanation for ERα sequestration in the nucleus. Conclusions Our results indicate that chemical structure of ligands directly affect the nuclear fate and protein turnover of the estrogen receptor alpha independently of their impact on

  12. Differential role of nitric oxide in regional sympathetic responses to stimulation of NTS A2a adenosine receptors.

    Science.gov (United States)

    Scislo, Tadeusz J; Tan, Nobusuke; O'Leary, Donal S

    2005-02-01

    Our previous studies showed that preganglionic adrenal (pre-ASNA), renal (RSNA), lumbar, and postganglionic adrenal sympathetic nerve activities (post-ASNA) are inhibited after stimulation of arterial baroreceptors, nucleus of the solitary tract (NTS), and glutamatergic and P2x receptors and are activated after stimulation of adenosine A1 receptors. However, stimulation of adenosine A2a receptors inhibited RSNA and post-ASNA, whereas it activated pre-ASNA. Because the effects evoked by NTS A2a receptors may be mediated via activation of nitric oxide (NO) mechanisms in NTS neurons, we tested the hypothesis that NO synthase (NOS) inhibitors would attenuate regional sympathetic responses to NTS A2a receptor stimulation, whereas NO donors would evoke contrasting responses from pre-ASNA versus RSNA and post-ASNA. Therefore, in chloralose/urethane-anesthetized rats, we compared hemodynamic and regional sympathetic responses to microinjections of selective A2a receptor agonist (CGS-21680, 20 pmol/50 nl) after pretreatment with NOS inhibitors Nomega-nitro-L-arginine methyl ester (10 nmol/100 nl) and 1-[2-(trifluoromethyl)phenyl]imidazole (100 pmol/100 nl) versus pretreatment with vehicle (100 nl). In addition, responses to microinjections into the NTS of different NO donors [40 and 400 pmol/50 nl sodium nitroprusside (SNP); 0.5 and 5 nmol/50 nl 3,3-bis(aminoethyl)-1-hydroxy-2-oxo-1-triazene (DETA NONOate, also known as NOC-18), and 2 nmol/50 nl 3-(2-hydroxy-2-nitroso-1-propylhydrazino)-1-propanamine (PAPA NONOate, also known as NOC-15)], the NO precursor L-arginine (10-50 nmol/50 nl), and sodium glutamate (500 pmol/50 nl) were evaluated. SNP, DETA NONOate, and PAPA NONOate activated pre-ASNA and inhibited RSNA and post-ASNA, whereas l-arginine and glutamate microinjected into the same site of the NTS inhibited all these sympathetic outputs. Decreases in heart rate and depressor or biphasic responses accompanied the neural responses. Pretreatment with NOS inhibitors

  13. Adenosine-to-inosine RNA editing affects trafficking of the gamma-aminobutyric acid type A (GABA(A)) receptor.

    Science.gov (United States)

    Daniel, Chammiran; Wahlstedt, Helene; Ohlson, Johan; Björk, Petra; Ohman, Marie

    2011-01-21

    Recoding by adenosine-to-inosine RNA editing plays an important role in diversifying proteins involved in neurotransmission. We have previously shown that the Gabra-3 transcript, coding for the α3 subunit of the GABA(A) receptor is edited in mouse, causing an isoleucine to methionine (I/M) change. Here we show that this editing event is evolutionarily conserved from human to chicken. Analyzing recombinant GABA(A) receptor subunits expressed in HEK293 cells, our results suggest that editing at the I/M site in α3 has functional consequences on receptor expression. We demonstrate that I/M editing reduces the cell surface and the total number of α3 subunits. The reduction in cell surface levels is independent of the subunit combination as it is observed for α3 in combination with either the β2 or the β3 subunit. Further, an amino acid substitution at the corresponding I/M site in the α1 subunit has a similar effect on cell surface presentation, indicating the importance of this site for receptor trafficking. We show that the I/M editing during brain development is inversely related to the α3 protein abundance. Our results suggest that editing controls trafficking of α3-containing receptors and may therefore facilitate the switch of subunit compositions during development as well as the subcellular distribution of α subunits in the adult brain. PMID:21030585

  14. Adenosine-to-Inosine RNA Editing Affects Trafficking of the γ-Aminobutyric Acid Type A (GABAA) Receptor*

    Science.gov (United States)

    Daniel, Chammiran; Wahlstedt, Helene; Ohlson, Johan; Björk, Petra; Öhman, Marie

    2011-01-01

    Recoding by adenosine-to-inosine RNA editing plays an important role in diversifying proteins involved in neurotransmission. We have previously shown that the Gabra-3 transcript, coding for the α3 subunit of the GABAA receptor is edited in mouse, causing an isoleucine to methionine (I/M) change. Here we show that this editing event is evolutionarily conserved from human to chicken. Analyzing recombinant GABAA receptor subunits expressed in HEK293 cells, our results suggest that editing at the I/M site in α3 has functional consequences on receptor expression. We demonstrate that I/M editing reduces the cell surface and the total number of α3 subunits. The reduction in cell surface levels is independent of the subunit combination as it is observed for α3 in combination with either the β2 or the β3 subunit. Further, an amino acid substitution at the corresponding I/M site in the α1 subunit has a similar effect on cell surface presentation, indicating the importance of this site for receptor trafficking. We show that the I/M editing during brain development is inversely related to the α3 protein abundance. Our results suggest that editing controls trafficking of α3-containing receptors and may therefore facilitate the switch of subunit compositions during development as well as the subcellular distribution of α subunits in the adult brain. PMID:21030585

  15. Adenosine receptors in the immature brain : with special reference to their role in hypoxic ischemia

    OpenAIRE

    Ådén, Ulrika

    2001-01-01

    Although the newborn brain tolerates a much longer period of oxygen deprivation and ischemia than does the adult brain, perinatal hypoxic ischemia probably is an important cause of neurological dysfunction, cerebral palsy and epilepsy later in life. Hence it is important to investigate the mechanisms that modulate the extent of perinatal ischernic brain damage. There is good evidence that endogenous adenosine acts as a neuroprotective agent in models of ischemia in the m...

  16. Nucleoside-derived antagonists to A3 adenosine receptors lower mouse intraocular pressure and act across species.

    Science.gov (United States)

    Wang, Zhao; Do, Chi Wai; Avila, Marcel Y; Peterson-Yantorno, Kim; Stone, Richard A; Gao, Zhan-Guo; Joshi, Bhalchandra; Besada, Pedro; Jeong, Lak Shin; Jacobson, Kenneth A; Civan, Mortimer M

    2010-01-01

    The purpose of the study was to determine whether novel, selective antagonists of human A3 adenosine receptors (ARs) derived from the A3-selective agonist Cl-IB-MECA lower intraocular pressure (IOP) and act across species. IOP was measured invasively with a micropipette by the Servo-Null Micropipette System (SNMS) and by non-invasive pneumotonometry during topical drug application. Antagonist efficacy was also assayed by measuring inhibition of adenosine-triggered shrinkage of native bovine nonpigmented ciliary epithelial (NPE) cells. Five agonist-based A3AR antagonists lowered mouse IOP measured with SNMS tonometry by 3-5 mm Hg within minutes of topical application. Of the five agonist derivatives, LJ 1251 was the only antagonist to lower IOP measured by pneumotonometry. No effect was detected pneumotonometrically over 30 min following application of the other four compounds, consonant with slower, smaller responses previously measured non-invasively following topical application of A3AR agonists and the dihydropyridine A3AR antagonist MRS 1191. Latanoprost similarly lowered SNMS-measured IOP, but not IOP measured non-invasively over 30 min. Like MRS 1191, agonist-based A3AR antagonists applied to native bovine NPE cells inhibited adenosine-triggered shrinkage. In summary, the results indicate that antagonists of human A3ARs derived from the potent, selective A3 agonist Cl-IB-MECA display efficacy in mouse and bovine cells, as well. When intraocular delivery was enhanced by measuring mouse IOP invasively, five derivatives of the A3AR agonist Cl-IB-MECA lowered IOP but only one rapidly reduced IOP measured non-invasively after topical application. We conclude that derivatives of the highly-selective A3AR agonist Cl-IB-MECA can reduce IOP upon reaching their intraocular target, and that nucleoside-based derivatives are promising A3 antagonists for study in multiple animal models. PMID:19878673

  17. Origin and evolution of the ligand-binding ability of nuclear receptors.

    Science.gov (United States)

    Markov, Gabriel V; Laudet, Vincent

    2011-03-01

    The origin of the ligand-binding ability of nuclear receptors is still a matter of discussion. Current opposing models are the early evolution of an ancestral receptor that would bind a specific ligand with high affinity and the early evolution of an ancestral orphan that was a constitutive transcription factor. Here we review the arguments in favour or against these two hypotheses, and we discuss an alternative possibility that the ancestor was a ligand sensor, which would be able to explain the apparently contradictory data generated in previous models for the evolution of ligand binding in nuclear receptors. PMID:21055443

  18. Identification of VDR Antagonists among Nuclear Receptor Ligands Using Virtual Screening

    Directory of Open Access Journals (Sweden)

    Kelly Teske

    2014-04-01

    Full Text Available Herein, we described the development of two virtual screens to identify new vitamin D receptor (VDR antagonists among nuclear receptor (NR ligands. Therefore, a database of 14330 nuclear receptor ligands and their NR affinities was assembled using the online available “Binding Database.” Two different virtual screens were carried out in conjunction with a reported VDR crystal structure applying a stringent and less stringent pharmacophore model to filter docked NR ligand conformations. The pharmacophore models were based on the spatial orientation of the hydroxyl functionalities of VDR's natural ligands 1,25(OH2D3 and 25(OH2D3. The first virtual screen identified 32 NR ligands with a calculated free energy of VDR binding of more than -6.0 kJ/mol. All but nordihydroguaiaretic acid (NDGA are VDR ligands, which inhibited the interaction between VDR and coactivator peptide SRC2-3 with an IC50 value of 15.8 μM. The second screen identified 162 NR ligands with a calculated free energy of VDR binding of more than -6.0 kJ/mol. More than half of these ligands were developed to bind VDR followed by ERα/β ligands (26%, TRα/β ligands (7%, and LxRα/β ligands (7%. The binding between VDR and ERα ligand H6036 as well as TRα/β ligand triiodothyronine and a homoserine analog thereof was confirmed by fluorescence polarization.

  19. Toll-Like Receptors, Their Ligands, and Atherosclerosis

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    Conrad P. Hodgkinson

    2011-01-01

    Full Text Available Atherosclerosis is a disease characterized by inflammation in the arterial wall. Atherogenesis is dependent on the innate immune response involving activation of Toll-like receptors (TLRs and the expression of inflammatory proteins. TLRs, which recognize various pathogen-associated molecular patterns, are expressed in various cell types within the atherosclerotic plaque. Microbial agents are associated with an increased risk of atherosclerosis and this is, in part, due to activation of TLRs. Recently considerable evidence has been provided suggesting that endogenous proteins promote atherosclerosis by binding to TLRs. In this review, we describe the role of TLRs in atherosclerosis with particular emphasis on those atherogenic endogenous proteins that have been implicated as TLR ligands.

  20. CF102 an A3 adenosine receptor agonist mediates anti-tumor and anti-inflammatory effects in the liver.

    Science.gov (United States)

    Cohen, S; Stemmer, S M; Zozulya, G; Ochaion, A; Patoka, R; Barer, F; Bar-Yehuda, S; Rath-Wolfson, L; Jacobson, K A; Fishman, P

    2011-09-01

    The Gi protein-associated A(3) adenosine receptor (A(3) AR) is a member of the adenosine receptor family. Selective agonists at the A(3) AR, such as CF101 and CF102 were found to induce anti-inflammatory and anti-cancer effects. In this study, we examined the differential effect of CF102 in pathological conditions of the liver. The anti-inflammatory protective effect of CF101 was tested in a model of liver inflammation induced by Concanavalin A (Con. A) and the anti-cancer effect of CF102 was examined in vitro and in a xenograft animal model utilizing Hep-3B hepatocellular carcinoma (HCC) cells. The mechanism of action was explored by following the expression levels of key signaling proteins in the inflamed and tumor liver tissues, utilizing Western blot (WB) analysis. In the liver inflammation model, CF102 (100 µg/kg) markedly reduced the secretion of serum glutamic oxaloacetic transaminase and serum glutamic pyruvic transaminase in comparison to the vehicle-treated group. Mechanistically, CF102 treatment decreased the expression level of phosphorylated glycogen synthase kinase-3β, NF-κB, and TNF-α and prevented apoptosis in the liver. This was demonstrated by decreased expression levels of Fas receptor (FasR) and of the pro-apoptotic proteins Bax and Bad in liver tissues. In addition, CF102-induced apoptosis of Hep-3B cells both in vitro and in vivo via de-regulation of the PI3K-NF-κB signaling pathway, resulting in up-regulation of pro-apoptotic proteins. Taken together, CF102 acts as a protective agent in liver inflammation and inhibits HCC tumor growth. These results suggest that CF102 through its differential effect is a potential drug candidate to treat various pathological liver conditions. PMID:21660967

  1. Imaging of a glioma using peripheral benzodiazepine receptor ligands

    Energy Technology Data Exchange (ETDEWEB)

    Starosta-Rubinstein, S.; Ciliax, B.J.; Penney, J.B.; McKeever, P.; Young, A.B.

    1987-02-01

    Two types of benzodiazepine receptors have been demonstrated in mammalian tissues, one which is localized on neuronal elements in brain and the other, on glial cells and in peripheral tissues such as kidney. In vivo administration of /sup 3/H-labeled PK 11195 (1-(2-chlorophenyl-N-methyl-N-(1-methylpropyl)-3-isoquinoline carboxamide) or (/sup 3/H)flunitrazepam with 5 mg of clonazepam per kg to rats with intracranial C6 gliomas resulted in high levels of tritiated-drug binding to the tumor as shown by quantitative autoradiography. Pharmacological studies indicated that the bound drugs labeled the peripheral benzodiazepine binding site. Binding to the peripheral benzodiazepine site was confirmed primarily to malignant cells with little binding to adjacent normal brain tissue or to necrotic tissue. Tumor cell binding was completely inhibited by preadministration of the peripheral benzodiazepine blocking agent PK 11195 at 5 mg/kg. The centrally selective benzodiazepine ligand clonazepam had no effect on PK 11195 binding to the tumor cells. When binding to other tumor cell lines grown in nude mice and nude athymic rats was evaluated, little or no peripheral benzodiazepine binding was detected on human pheochromocytoma (RN1) and neuroblastoma (SK-N-MC, SK-N-SH) tumor cells, respectively. However, high densities of peripheral benzodiazepine binding sites were observed on tumors derived from a human glioma cell line (ATCC HTB 14, U-87 MG). The presence of high concentrations of specific peripheral benzodiazepine receptors on glial tumors suggests that human primary central nervous system tumors could be imaged and diagnosed using peripheral benzodiazepine ligands labeled with positron- or gamma-emitting isotopes.

  2. Imaging of a glioma using peripheral benzodiazepine receptor ligands

    International Nuclear Information System (INIS)

    Two types of benzodiazepine receptors have been demonstrated in mammalian tissues, one which is localized on neuronal elements in brain and the other, on glial cells and in peripheral tissues such as kidney. In vivo administration of 3H-labeled PK 11195 [1-(2-chlorophenyl-N-methyl-N-(1-methylpropyl)-3-isoquinoline carboxamide] or [3H]flunitrazepam with 5 mg of clonazepam per kg to rats with intracranial C6 gliomas resulted in high levels of tritiated-drug binding to the tumor as shown by quantitative autoradiography. Pharmacological studies indicated that the bound drugs labeled the peripheral benzodiazepine binding site. Binding to the peripheral benzodiazepine site was confirmed primarily to malignant cells with little binding to adjacent normal brain tissue or to necrotic tissue. Tumor cell binding was completely inhibited by preadministration of the peripheral benzodiazepine blocking agent PK 11195 at 5 mg/kg. The centrally selective benzodiazepine ligand clonazepam had no effect on PK 11195 binding to the tumor cells. When binding to other tumor cell lines grown in nude mice and nude athymic rats was evaluated, little or no peripheral benzodiazepine binding was detected on human pheochromocytoma (RN1) and neuroblastoma (SK-N-MC, SK-N-SH) tumor cells, respectively. However, high densities of peripheral benzodiazepine binding sites were observed on tumors derived from a human glioma cell line (ATCC HTB 14, U-87 MG). The presence of high concentrations of specific peripheral benzodiazepine receptors on glial tumors suggests that human primary central nervous system tumors could be imaged and diagnosed using peripheral benzodiazepine ligands labeled with positron- or gamma-emitting isotopes

  3. Structural and functional characterization of a novel type of ligand-independent RXR-USP receptor

    OpenAIRE

    Iwema, Thomas; Billas, Isabelle ML; Beck, Yannick; Bonneton, François; Nierengarten, Hélène; Chaumot, Arnaud; Richards, Geoff; Laudet, Vincent; Moras, Dino

    2007-01-01

    Retinoid X receptor (RXR) and Ultraspiracle (USP) play a central role as ubiquitous heterodimerization partners of many nuclear receptors. While it has long been accepted that a wide range of ligands can activate vertebrate/mollusc RXRs, the existence and necessity of specific endogenous ligands activating RXR-USP in vivo is still matter of intense debate. Here we report the existence of a novel type of RXR-USP with a ligand-independent functional conformation. Our studies involved Tribolium ...

  4. Bioluminescent Ligand-Receptor Binding Assays for Protein or Peptide Hormones.

    Science.gov (United States)

    Liu, Ya-Li; Guo, Zhan-Yun

    2016-01-01

    Bioluminescence has been widely used in biomedical research due to its high sensitivity, low background, and broad linear range. In recent studies, we applied bioluminescence to ligand-receptor binding assays for some protein or peptide hormones based on a newly developed small monomeric Nanoluciferase (NanoLuc) reporter that has the so far brightest bioluminescence. The conventional ligand-receptor binding assays rely on radioligands that have drawbacks, such as radioactive hazards and short shelf lives. In contrast, the novel bioluminescent binding assays use the NanoLuc-based protein or peptide tracers that are safe, stable, and ultrasensitive. Thus, the novel bioluminescent ligand-receptor binding assay would be applied to more and more protein or peptide hormones for ligand-receptor interaction studies in future. In the present article, we provided detailed protocols for setting up the novel bioluminescent ligand-receptor binding assays using two representative protein hormones as examples. PMID:27424896

  5. Development of radioiodinated receptor ligands for cerebral single photon emission tomography

    International Nuclear Information System (INIS)

    In the last decade the use of radiolabeled ligands for the imaging of cerebral receptors by emission computed tomography (ECT) has seen rapid growth. The opportunity to routinely perform cerebral single photon emission tomography (SPET) with iodine-123-labeled ligands depends on the availability of receptor ligands into which iodine can be introduced without decreasing the required high target receptor specificity. The use of iodine-123-labeled receptor-specific ligands also depends on the availability of high purity iodine-123 at reasonable costs and the necessary imaging instrumentation. In this paper, the development and current stage of evaluation of various iodine-123-labeled ligands for SPET imaging of dopaminergic, serotonergic and muscarinic acetylcholinergic receptor classes are discussed

  6. Prevention of adenosine A2A receptor activation diminishes beat-to-beat alternation in human atrial myocytes.

    Science.gov (United States)

    Molina, Cristina E; Llach, Anna; Herraiz-Martínez, Adela; Tarifa, Carmen; Barriga, Montserrat; Wiegerinck, Rob F; Fernandes, Jacqueline; Cabello, Nuria; Vallmitjana, Alex; Benitéz, Raúl; Montiel, José; Cinca, Juan; Hove-Madsen, Leif

    2016-01-01

    Atrial fibrillation (AF) has been associated with increased spontaneous calcium release from the sarcoplasmic reticulum and linked to increased adenosine A2A receptor (A2AR) expression and activation. Here we tested whether this may favor atrial arrhythmogenesis by promoting beat-to-beat alternation and irregularity. Patch-clamp and confocal calcium imaging was used to measure the beat-to-beat response of the calcium current and transient in human atrial myocytes. Responses were classified as uniform, alternating or irregular and stimulation of Gs-protein coupled receptors decreased the frequency where a uniform response could be maintained from 1.0 ± 0.1 to 0.6 ± 0.1 Hz; p < 0.01 for beta-adrenergic receptors and from 1.4 ± 0.1 to 0.5 ± 0.1 Hz; p < 0.05 for A2ARs. The latter was linked to increased spontaneous calcium release and after-depolarizations. Moreover, A2AR activation increased the fraction of non-uniformly responding cells in HL-1 myocyte cultures from 19 ± 3 to 51 ± 9 %; p < 0.02, and electrical mapping in perfused porcine atria revealed that adenosine induced electrical alternans at longer cycle lengths, doubled the fraction of electrodes showing alternation, and increased the amplitude of alternations. Importantly, protein kinase A inhibition increased the highest frequency where uniform responses could be maintained from 0.84 ± 0.12 to 1.86 ± 0.11 Hz; p < 0.001 and prevention of A2AR-activation with exogenous adenosine deaminase selectively increased the threshold from 0.8 ± 0.1 to 1.2 ± 0.1 Hz; p = 0.001 in myocytes from patients with AF. In conclusion, A2AR-activation promotes beat-to-beat irregularities in the calcium transient in human atrial myocytes, and prevention of A2AR activation may be a novel means to maintain uniform beat-to-beat responses at higher beating frequencies in patients with atrial fibrillation.

  7. Computational approaches to modeling receptor flexibility upon ligand binding: Application to interfacially activated enzymes

    DEFF Research Database (Denmark)

    Wade, R.C.; Sobolev, V.; Ortiz, A.R. .;

    1998-01-01

    Receptors generally undergo conformational change upon ligand binding. We describe how fairly simple techniques may be used in docking and design studies to account for some of the changes in the conformations of proteins on ligand binding. Simulations of protein-ligand interactions that give...... a more complete description of the dynamics important for ligand binding are then discussed. These methods are illustrated for phospholipase A(2) and lipase, enzymes that both undergo interfacial activation....

  8. Greater adenosine A2A receptor densities in cardiac and skeletal muscle in endurance-trained men: a [11C]TMSX PET study

    International Nuclear Information System (INIS)

    We examined the densities of adenosine A2A receptors in cardiac and skeletal muscles between untrained and endurance-trained subjects using positron emission tomography (PET) and [7-methyl-11C]-(E)-8-(3,4,5-trimethoxystyryl)-1,3,7-trimethylxanthine ([11C]TMSX), a newly developed radioligand for mapping adenosine A2A receptors. Five untrained and five endurance-trained subjects participated in this study. The density of adenosine A2A receptors was evaluated as the distribution volume of [11C]TMSX in cardiac and triceps brachii muscles in the resting state using PET. The distribution volume of [11C]TMSX in the myocardium was significantly greater than in the triceps brachii muscle in both groups. Further, distribution volumes [11C]TMSX in the trained subjects were significantly grater than those in untrained subjects (myocardium, 3.6±0.3 vs. 3.1±0.4 ml g-1; triceps brachii muscle, 1.7±0.3 vs. 1.2±0.2 ml g-1, respectively). These results indicate that the densities of adenosine A2A receptors in the cardiac and skeletal muscles are greater in the endurance-trained men than in the untrained men

  9. SELECTIVITY AND SPECIFICITY OF SPHINGOSINE 1-PHOSPHATE RECEPTOR LIGANDS: ‘OFF-TARGETS’ OR COMPLEX PHARMACOLOGY?

    Directory of Open Access Journals (Sweden)

    Nigel John Pyne

    2011-05-01

    Full Text Available A recent perspective published in frontiers of Pharmacology by Salomone and Waeber (2011 discussed the selectivity and specificity of sphingosine 1-phosphate (S1P receptor ligands. This perspective surveyed the use of various S1P receptor ligands and attempted to reconcile a number of inconsistencies in the predicted biological outcomes: these were interpreted as ‘off-target’ effects. Therefore the perspective cautioned against the use of these S1P receptor ligands. Here we highlight the complex pharmacology of S1P receptors, which along with ‘inside-out’ signalling might provide an alternative explanation for ‘off-target’ effects.

  10. Progesterone in pregnancy; receptor-ligand interaction and signaling pathways.

    Science.gov (United States)

    Szekeres-Bartho, Julia; Halasz, Melinda; Palkovics, Tamas

    2009-12-01

    Progesterone is indispensable in creating a suitable endometrial environment for implantation, and also for the maintenance of pregnancy. Successful pregnancy depends on an appropriate maternal immune response to the fetus. Along with its endocrine effects, progesterone also acts as an "immunosteroid", by contributing to the establishment of a pregnancy protective immune milieu. Progesterone plays a role in uterine homing of NK cells and upregulates HLA-G gene expression, the ligand for NK inhibitory and activating receptors. At high concentrations, progesterone is a potent inducer of Th2-type cytokines as well as of LIF and M-CSF production by T cells. A protein called progesterone-induced blocking factor (PIBF), by inducing a Th2-dominant cytokine production mediates the immunological effects of progesterone. PIBF binds to a novel type of the IL-4 receptor and signals via the Jak/STAT pathway, to induce a number of genes, that not only affect the immune response, but might also play a role in trophoblast invasiveness. PMID:19880194

  11. Pharmacological profiles of the metabotropic glutamate receptor ligands.

    Science.gov (United States)

    Naples, M A; Hampson, D R

    2001-01-01

    Metabotropic glutamate receptors (mGluRs) are a family of G-protein coupled receptors that are expressed in the central and peripheral nervous systems. The purpose of this study was to compare the ligand binding selectivity profiles of the mGluR agonist [(3)H]L-AP4 and the novel radiolabeled phenylglycine antagonist [(3)H]CPPG at all eight rat mGluR subtypes expressed in transfected human embryonic kidney cells. At a concentration of 30 nM [(3)H]L-AP4, no specific binding was detected in membranes expressing the group I receptors mGluR1a or mGluR5a, or in membranes expressing the group II mGluRs, mGluR2 and mGluR3. Among the group III mGluRs, specific [(3)H]L-AP4 binding was detected in cells expressing mGluR4a and mGluR8a but not in cells expressing mGluR6 or mGluR7a. The binding of [(3)H]CPPG showed an exceptional pattern of selectivity amongst the mGluR subtypes; at a concentration of 20 nM [(3)H]CPPG, a high level of specific binding was seen in membranes containing mGluR8a but not in any of the other mGluR subtypes. The affinity constant (K(D)) calculated for [(3)H]CPPG binding to mGluR8a was 183 nM. In competition experiments, the phosphono-substituted phenylglycine congeners including MPPG, (RS)-PPG, and unlabeled CPPG were the most potent inhibitors of [(3)H]CPPG binding while non-phosphonated compounds such as L-glutamate and MCPG were substantially less potent. These results demonstrate that [(3)H]L-AP4 and [(3)H]CPPG can be used as probes to selectively label group III mGluRs and that CPPG and related phenylglycine derivatives are useful for studying differences in the ligand recognition sites of highly homologous mGluRs. PMID:11114395

  12. The adenosine A2A receptor agonist CGS 21680 exhibits antipsychotic-like activity in Cebus apella monkeys

    DEFF Research Database (Denmark)

    Andersen, M B; Fuxe, K; Werge, T;

    2002-01-01

    The adenosine A2A receptor agonist CGS 21680 has shown effects similar to dopamine antagonists in behavioural assays in rats predictive for antipsychotic activity, without induction of extrapyramidal side-effects (EPS). In the present study, we examined whether this functional dopamine antagonism...... and lack of EPS in rodents could also be observed in non-human primates. We investigated the effects of CGS 21680 on behaviours induced by D-amphetamine and (-)-apomorphine in EPS-sensitized Cebus apella monkeys. CGS 21680 was administered s.c. in doses of 0.01, 0.025 and 0.05 mg/kg, alone......-induced behaviours (unrest, stereotypies, arousal) were unaffected. EPS were not observed at any dose. At 0.05 mg/kg CGS 21680 produced vomiting. The two lower doses did not produce observable side-effects. Though the differential effect on amphetamine- and apomorphine-induced behaviours is intriguing, CGS 21680...

  13. ATP induced vasodilatation and purinergic receptors in the human leg: roles of nitric oxide, prostaglandins and adenosine

    DEFF Research Database (Denmark)

    Mortensen, Stefan P; Gonzalez-Alonso, Jose; Bune, Laurids;

    2009-01-01

    -arterial infusion of ATP (0.45-2.45 micromol/min; mean+/-SEM) in 19 healthy, male subjects with and without co-infusion of NG-mono-methyl-L-arginine (L-NMMA; NO formation inhibitor; 12.3+/-0.3 mg/min), indomethacin (INDO; prostaglandin formation blocker; 613+/-12 microg/min) and/or theophylline (adenosine receptor...... blocker; 400+/-26 mg). During control conditions, ATP infusion increased leg blood flow (LBF) from baseline conditions by 1.82+/-0.14 L/min. When ATP was co-infused with either L-NMMA, INDO or L-NMMA+INDO combined, the increase in LBF was reduced by 14+/-6, 15+/-9, and 39+/-8 %, respectively (P

  14. Progress in the discovery of selective, high affinity A2B adenosine receptor antagonists as clinical candidates

    OpenAIRE

    Kalla, Rao V.; Zablocki, Jeff

    2008-01-01

    The selective, high affinity A2B adenosine receptor (AdoR) antagonists that were synthesized by several research groups should aid in determining the role of the A2B AdoR in inflammatory diseases like asthma or rheumatoid arthritis (RA) and angiogenic diseases like diabetic retinopathy or cancer. CV Therapeutics scientists discovered the selective, high affinity A2B AdoR antagonist 10, a 8-(4-pyrazolyl)-xanthine derivative [CVT-6883, Ki(hA2B) = 22 nM; Ki(hA1) = 1,940 nM; Ki(hA2A) = 3,280; and...

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

    Institute of Scientific and Technical Information of China (English)

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

    2006-01-01

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

  16. Caffeine acts through neuronal adenosine A2A receptors to prevent mood and memory dysfunction triggered by chronic stress.

    Science.gov (United States)

    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. PMID:26056314

  17. Caffeine promotes anti-tumor immune response during tumor initiation: Involvement of the adenosine A2A receptor.

    Science.gov (United States)

    Eini, Hadar; Frishman, Valeria; Yulzari, Robert; Kachko, Leonid; Lewis, Eli C; Chaimovitz, Cidio; Douvdevani, Amos

    2015-11-01

    Epidemiologic studies depict a negative correlation between caffeine consumption and incidence of tumors in humans. The main pharmacological effects of caffeine are mediated by antagonism of the adenosine receptor, A2AR. Here, we examine whether the targeting of A2AR by caffeine plays a role in anti-tumor immunity. In particular, the effects of caffeine are studied in wild-type and A2AR knockout (A2AR(-/-)) mice. Tumor induction was achieved using the carcinogen 3-methylcholanthrene (3-MCA). Alternatively, tumor cells, comprised of 3-MCA-induced transformed cells or B16 melanoma cells, were inoculated into animal footpads. Cytokine release was determined in a mixed lymphocyte tumor reaction (MLTR). According to our findings, caffeine-consuming mice (0.1% in water) developed tumors at a lower rate compared to water-consuming mice (14% vs. 53%, respectively, p=0.0286, n=15/group). Within the caffeine-consuming mice, tumor-free mice displayed signs of autoimmune alopecia and pronounced leukocyte recruitment intocarcinogen injection sites. Similarly, A2AR(-/-) mice exhibited reduced rates of 3-MCA-induced tumors. In tumor inoculation studies, caffeine treatment resulted in inhibition of tumor growth and elevation in proinflammatory cytokine release over water-consuming mice, as depicted by MLTR. Addition of the adenosine receptor agonist, NECA, to MLTR resulted in a sharp decrease in IFNγ levels; this was reversed by the highly selective A2AR antagonist, ZM241385. Thus, immune response modulation through either caffeine or genetic deletion of A2AR leads to a Th1 immune profile and suppression of carcinogen-induced tumorigenesis. Taken together, our data suggest that the use of pharmacologic A2AR antagonists may hold therapeutic potential in diminishing the rate of cancer development.

  18. Adenosine A(2A receptor up-regulates retinal wave frequency via starburst amacrine cells in the developing rat retina.

    Directory of Open Access Journals (Sweden)

    Pin-Chien Huang

    Full Text Available BACKGROUND: Developing retinas display retinal waves, the patterned spontaneous activity essential for circuit refinement. During the first postnatal week in rodents, retinal waves are mediated by synaptic transmission between starburst amacrine cells (SACs and retinal ganglion cells (RGCs. The neuromodulator adenosine is essential for the generation of retinal waves. However, the cellular basis underlying adenosine's regulation of retinal waves remains elusive. Here, we investigated whether and how the adenosine A(2A receptor (A(2AR regulates retinal waves and whether A(2AR regulation of retinal waves acts via presynaptic SACs. METHODOLOGY/PRINCIPAL FINDINGS: We showed that A(2AR was expressed in the inner plexiform layer and ganglion cell layer of the developing rat retina. Knockdown of A(2AR decreased the frequency of spontaneous Ca²⁺ transients, suggesting that endogenous A(2AR may up-regulate wave frequency. To investigate whether A(2AR acts via presynaptic SACs, we targeted gene expression to SACs by the metabotropic glutamate receptor type II promoter. Ca²⁺ transient frequency was increased by expressing wild-type A(2AR (A2AR-WT in SACs, suggesting that A(2AR may up-regulate retinal waves via presynaptic SACs. Subsequent patch-clamp recordings on RGCs revealed that presynaptic A(2AR-WT increased the frequency of wave-associated postsynaptic currents (PSCs or depolarizations compared to the control, without changing the RGC's excitability, membrane potentials, or PSC charge. These findings suggest that presynaptic A(2AR may not affect the membrane properties of postsynaptic RGCs. In contrast, by expressing the C-terminal truncated A(2AR mutant (A(2AR-ΔC in SACs, the wave frequency was reduced compared to the A(2AR-WT, but was similar to the control, suggesting that the full-length A(2AR in SACs is required for A(2AR up-regulation of retinal waves. CONCLUSIONS/SIGNIFICANCE: A(2AR up-regulates the frequency of retinal waves via

  19. Presynaptic Adenosine Receptor-Mediated Regulation of Diverse Thalamocortical Short-Term Plasticity in the Mouse Whisker Pathway

    Science.gov (United States)

    Ferrati, Giovanni; Martini, Francisco J.; Maravall, Miguel

    2016-01-01

    Short-term synaptic plasticity (STP) sets the sensitivity of a synapse to incoming activity and determines the temporal patterns that it best transmits. In “driver” thalamocortical (TC) synaptic populations, STP is dominated by depression during stimulation from rest. However, during ongoing stimulation, lemniscal TC connections onto layer 4 neurons in mouse barrel cortex express variable STP. Each synapse responds to input trains with a distinct pattern of depression or facilitation around its mean steady-state response. As a result, in common with other synaptic populations, lemniscal TC synapses express diverse rather than uniform dynamics, allowing for a rich representation of temporally varying stimuli. Here, we show that this STP diversity is regulated presynaptically. Presynaptic adenosine receptors of the A1R type, but not kainate receptors (KARs), modulate STP behavior. Blocking the receptors does not eliminate diversity, indicating that diversity is related to heterogeneous expression of multiple mechanisms in the pathway from presynaptic calcium influx to neurotransmitter release. PMID:26941610

  20. The anti-inflammatory target A(3) adenosine receptor is over-expressed in rheumatoid arthritis, psoriasis and Crohn's disease.

    Science.gov (United States)

    Ochaion, A; Bar-Yehuda, S; Cohen, S; Barer, F; Patoka, R; Amital, H; Reitblat, T; Reitblat, A; Ophir, J; Konfino, I; Chowers, Y; Ben-Horin, S; Fishman, P

    2009-01-01

    The Gi protein associated A(3) adenosine receptor (A(3)AR) was recently defined as a novel anti-inflammatory target. The aim of this study was to look at A(3)AR expression levels in peripheral blood mononuclear cells (PBMCs) of patients with autoimmune inflammatory diseases and to explore transcription factors involved receptor expression. Over-expression of A(3)AR was found in PBMCs derived from patients with rheumatoid arthritis (RA), psoriasis and Crohn's disease compared with PBMCs from healthy subjects. Bioinformatics analysis demonstrated the presence of DNA binding sites for nuclear factor-kappaB (NF-kappaB) and cyclic AMP-responsive element binding protein (CREB) in the A(3)AR gene promoter. Up-regulation of NF-kappaB and CREB was found in the PBMCs from patients with RA, psoriasis and Crohn's disease. The PI3K-PKB/Akt signaling pathway, known to regulate both the NF-kappaB and CREB, was also up-regulated in the patients' PBMCs. Taken together, NF-kappaB and CREB are involved with the over-expression of A(3)AR in patients with autoimmune inflammatory diseases. The receptor may be considered as a specific target to combat inflammation. PMID:19426966

  1. Cell surface receptors for signal transduction and ligand transport: a design principles study.

    Directory of Open Access Journals (Sweden)

    Harish Shankaran

    2007-06-01

    Full Text Available Receptors constitute the interface of cells to their external environment. These molecules bind specific ligands involved in multiple processes, such as signal transduction and nutrient transport. Although a variety of cell surface receptors undergo endocytosis, the systems-level design principles that govern the evolution of receptor trafficking dynamics are far from fully understood. We have constructed a generalized mathematical model of receptor-ligand binding and internalization to understand how receptor internalization dynamics encodes receptor function and regulation. A given signaling or transport receptor system represents a particular implementation of this module with a specific set of kinetic parameters. Parametric analysis of the response of receptor systems to ligand inputs reveals that receptor systems can be characterized as being: i avidity-controlled where the response control depends primarily on the extracellular ligand capture efficiency, ii consumption-controlled where the ability to internalize surface-bound ligand is the primary control parameter, and iii dual-sensitivity where both the avidity and consumption parameters are important. We show that the transferrin and low-density lipoprotein receptors are avidity-controlled, the vitellogenin receptor is consumption-controlled, and the epidermal growth factor receptor is a dual-sensitivity receptor. Significantly, we show that ligand-induced endocytosis is a mechanism to enhance the accuracy of signaling receptors rather than merely serving to attenuate signaling. Our analysis reveals that the location of a receptor system in the avidity-consumption parameter space can be used to understand both its function and its regulation.

  2. An orally active adenosine A1 receptor antagonist, FK838, increases renal excretion and maintains glomerular filtration rate in furosemide-resistant rats

    Science.gov (United States)

    Schnackenberg, Christine G; Merz, Emily; Brooks, David P

    2003-01-01

    Loop and thiazide diuretics are common therapeutic agents for the treatment of sodium retention and oedema. However, resistance to diuretics and decreases in renal function can develop during diuretic therapy. Adenosine causes renal vasoconstriction, sodium reabsorption, and participates in the tubuloglomerular feedback mechanism for the regulation of glomerular filtration rate.We tested the hypothesis that the selective adenosine A1 receptor antagonist FK838 is orally active and causes diuresis and natriuresis, but maintains glomerular filtration rate in normal rats or in rats with furosemide resistance.In normal male Sprague – Dawley rats, FK838 dose-dependently increased urine flow and sodium and chloride excretion while sparing potassium. In combination with furosemide, FK838 enhanced the diuretic and natriuretic actions of furosemide to the same extent as hydrochlorothiazide and did not increase the potassium loss in normal rats. In furosemide-resistant rats, FK838 increased urine flow and electrolyte excretion to a greater extent than hydrochlorothiazide. In addition, hydrochlorothiazide significantly decreased glomerular filtration rate, whereas FK838 maintained glomerular filtration rate in furosemide-resistant rats.This study shows that the adenosine A1 receptor antagonist FK838 is orally active and causes potent diuresis and natriuresis and maintains glomerular filtration rate in normal or furosemide-resistant rats. Adenosine A1 receptor antagonists may be novel therapeutics for the treatment of oedema in normal or otherwise diuretic-resistant patients. PMID:12922924

  3. Caffeine prevents antihyperalgesic effect of gabapentin in an animal model of CRPS-I: evidence for the involvement of spinal adenosine A1 receptor.

    Science.gov (United States)

    Martins, Daniel F; Prado, Marcos R B; Daruge-Neto, Eduardo; Batisti, Ana P; Emer, Aline A; Mazzardo-Martins, Leidiane; Santos, Adair R S; Piovezan, Anna P

    2015-12-01

    This study was designed to determine whether 3 weeks of gabapentin treatment is effective in alleviating neuropathic pain-like behavior in animal models of complex regional pain syndrome type-I and partial sciatic nerve ligation (PSNL). We investigated the contribution of adenosine subtypes to the antihyperalgesic effect of gabapentin by examining the effect of caffeine, a non-selective adenosine A1 and A2 receptor antagonist or 1,3-dipropyl-8-cyclopentylxanthine (DPCPX), a selective adenosine A1 subtype receptor antagonist on this effect. Neuropathic pain was produced by unilateral prolonged hind paw ischemia and reperfusion (I/R) or PSNL procedures which resulted in stimulus-evoked mechanical hyperalgesia. After procedures, animals received gabapentin (10, 30, or 100 mg/kg intraperitoneal, respectively), caffeine (10 mg/kg intraperitoneal or 150 nmol intrathecally) or DPCPX (3 µg intrathecally) alone or in combination. Mice were tested for tactile mechanical hyperalgesia at 1, 2, and 3 weeks following procedures. Gabapentin produced dose-related inhibition of mechanical hyperalgesia over a 3-week period, and this effect was blocked by concomitant caffeine or DPCPX administration 1 week after injuries. The results of this study demonstrated that the mechanism through which gabapentin produces its effect may involve the activation of adenosine A1 subtype receptor.

  4. Glucagon-like peptide-1 receptor ligand interactions: structural cross talk between ligands and the extracellular domain.

    Directory of Open Access Journals (Sweden)

    Graham M West

    Full Text Available Activation of the glucagon-like peptide-1 receptor (GLP-1R in pancreatic β-cells potentiates insulin production and is a current therapeutic target for the treatment of type 2 diabetes mellitus (T2DM. Like other class B G protein-coupled receptors (GPCRs, the GLP-1R contains an N-terminal extracellular ligand binding domain. N-terminal truncations on the peptide agonist generate antagonists capable of binding to the extracellular domain, but not capable of activating full length receptor. The main objective of this study was to use Hydrogen/deuterium exchange (HDX to identify how the amide hydrogen bonding network of peptide ligands and the extracellular domain of GLP-1R (nGLP-1R were altered by binding interactions and to then use this platform to validate direct binding events for putative GLP-1R small molecule ligands. The HDX studies presented here for two glucagon-like peptide-1 receptor (GLP-1R peptide ligands indicates that the antagonist exendin-4[9-39] is significantly destabilized in the presence of nonionic detergents as compared to the agonist exendin-4. Furthermore, HDX can detect stabilization of exendin-4 and exendin-4[9-39] hydrogen bonding networks at the N-terminal helix [Val19 to Lys27] upon binding to the N-terminal extracellular domain of GLP-1R (nGLP-1R. In addition we show hydrogen bonding network stabilization on nGLP-1R in response to ligand binding, and validate direct binding events with the extracellular domain of the receptor for putative GLP-1R small molecule ligands.

  5. Synthesis of BODIPY derivatives substituted with various bioconjugatable linker groups: a construction kit for fluorescent labeling of receptor ligands.

    Science.gov (United States)

    Heisig, Fabian; Gollos, Sabrina; Freudenthal, Sven J; El-Tayeb, Ali; Iqbal, Jamshed; Müller, Christa E

    2014-01-01

    The goal of the present study was to design small, functionalized green-emitting BODIPY dyes, which can readily be coupled to target molecules such as receptor ligands, or even be integrated into their pharmacophores. A simple two-step one-pot procedure starting from 2,4-dimethylpyrrole and ω-bromoalkylcarboxylic acid chlorides was used to obtain new ω-bromoalkyl-substituted BODIPY fluorophores (1a-1f) connected via alkyl spacers of different length to the 8-position of the fluorescent dye. The addition of radical inhibitors reduced the amount of side products. The ω-bromoalkyl-substituted BODIPYs were further converted to introduce various functional groups: iodo-substituted dyes were obtained by Finkelstein reaction in excellent yields; microwave-assisted reaction with methanolic ammonia led to fast and clean conversion to the amino-substituted dyes; a hydroxyl-substituted derivative was prepared by reaction with sodium ethylate, and thiol-substituted BODIPYs were obtained by reaction of 1a-1f with potassium thioacetate followed by alkaline cleavage of the thioesters. Water-soluble derivatives were prepared by introducing sulfonate groups into the 2- and 6-position of the BODIPY core. The synthesized BODIPY derivatives showed high fluorescent yields and appeared to be stable under basic, reducing and oxidative conditions. As a proof of concept, 2-thioadenosine was alkylated with bromoethyl-BODIPY 1b. The resulting fluorescent 2-substituted adenosine derivative 15 displayed selectivity for the A3 adenosine receptor (ARs) over the other AR subtypes, showed agonistic activity, and may thus become a useful tool for studying A3ARs, or a lead structure for further optimization. The new functionalized dyes may be widely used for fluorescent labeling allowing the investigation of biological targets and processes. PMID:24052460

  6. Synthesis of BODIPY derivatives substituted with various bioconjugatable linker groups: a construction kit for fluorescent labeling of receptor ligands.

    Science.gov (United States)

    Heisig, Fabian; Gollos, Sabrina; Freudenthal, Sven J; El-Tayeb, Ali; Iqbal, Jamshed; Müller, Christa E

    2014-01-01

    The goal of the present study was to design small, functionalized green-emitting BODIPY dyes, which can readily be coupled to target molecules such as receptor ligands, or even be integrated into their pharmacophores. A simple two-step one-pot procedure starting from 2,4-dimethylpyrrole and ω-bromoalkylcarboxylic acid chlorides was used to obtain new ω-bromoalkyl-substituted BODIPY fluorophores (1a-1f) connected via alkyl spacers of different length to the 8-position of the fluorescent dye. The addition of radical inhibitors reduced the amount of side products. The ω-bromoalkyl-substituted BODIPYs were further converted to introduce various functional groups: iodo-substituted dyes were obtained by Finkelstein reaction in excellent yields; microwave-assisted reaction with methanolic ammonia led to fast and clean conversion to the amino-substituted dyes; a hydroxyl-substituted derivative was prepared by reaction with sodium ethylate, and thiol-substituted BODIPYs were obtained by reaction of 1a-1f with potassium thioacetate followed by alkaline cleavage of the thioesters. Water-soluble derivatives were prepared by introducing sulfonate groups into the 2- and 6-position of the BODIPY core. The synthesized BODIPY derivatives showed high fluorescent yields and appeared to be stable under basic, reducing and oxidative conditions. As a proof of concept, 2-thioadenosine was alkylated with bromoethyl-BODIPY 1b. The resulting fluorescent 2-substituted adenosine derivative 15 displayed selectivity for the A3 adenosine receptor (ARs) over the other AR subtypes, showed agonistic activity, and may thus become a useful tool for studying A3ARs, or a lead structure for further optimization. The new functionalized dyes may be widely used for fluorescent labeling allowing the investigation of biological targets and processes.

  7. Ion fluxes through KCa2 (SK) and Cav1 (L-type) channels contribute to chronoselectivity of adenosine A1 receptor-mediated actions in spontaneously beating rat atria

    OpenAIRE

    Paulo eCorreia-De-Sá

    2016-01-01

    Impulse generation in supraventricular tissue is inhibited by adenosine and acetylcholine via the activation of A1 and M2 receptors coupled to inwardly rectifying GIRK/KIR3.1/3.4 channels, respectively. Unlike M2 receptors, bradycardia produced by A1 receptors activation predominates over negative inotropy. Such difference suggests that other ion currents may contribute to adenosine chronoselectivity. In isolated spontaneously beating rat atria, blockade of KCa2/SK channels with apamin and Ca...

  8. Elimination of a ligand gating site generates a supersensitive olfactory receptor

    OpenAIRE

    Kanika Sharma; Gaurav Ahuja; Ashiq Hussain; Sabine Balfanz; Arnd Baumann; Korsching, Sigrun I.

    2016-01-01

    Olfaction poses one of the most complex ligand-receptor matching problems in biology due to the unparalleled multitude of odor molecules facing a large number of cognate olfactory receptors. We have recently deorphanized an olfactory receptor, TAAR13c, as a specific receptor for the death-associated odor cadaverine. Here we have modeled the cadaverine/TAAR13c interaction, exchanged predicted binding residues by site-directed mutagenesis, and measured the activity of the mutant receptors. Unex...

  9. Rescue of ligand binding of a mutant IGF-I receptor by complementation

    DEFF Research Database (Denmark)

    Chakravarty, Arjun Anders; Hinrichsen, Jane; Whittaker, Linda;

    2005-01-01

    from a wild-type receptor monomer and a mutant receptor monomer devoid of binding activity. Receptor hybrids were generated by transient co-transfection of cDNAs encoding wild-type and mutant receptors with unique epitope tags. Hybrid receptors were purified from transfected cells by sequential immuno......-affinity chromatography and their ligand-binding properties were determined. Complementation produced a hybrid with near wild-type affinity. Dissociation studies demonstrated that the hybrid did not exhibit negative cooperativity....

  10. CLE Peptides in Plants: Proteolytic Processing,Structure-Activity Relationship, and Ligand-Receptor Interaction

    Institute of Scientific and Technical Information of China (English)

    Xiaoming Gao; Yongfeng Guo

    2012-01-01

    Ligand-receptor signaling initiated by the CLAVATA3/ENDOSPERM SURROUNDING REGION (CLE) family peptides is critical in regulating cell division and differentiation in meristematic tissues in plants.Biologically active CLE peptides are released from precursor proteins via proteolytic processing.The mature form of CLE ligands consists of 12-13 amino acids with several post-translational modifications.This review summarizes recent progress toward understanding the proteolytic activities that cleave precursor proteins to release CLE peptides,the molecular structure and function of mature CLE ligands,and interactions between CLE ligands and corresponding leucine-rich repeat (LRR) receptor-like kinases (RLKs).

  11. Large-scale functional expression of WT and truncated human adenosine A2A receptor in Pichia pastoris bioreactor cultures

    Directory of Open Access Journals (Sweden)

    Strange Philip G

    2008-10-01

    Full Text Available Abstract Background The large-scale production of G-protein coupled receptors (GPCRs for functional and structural studies remains a challenge. Recent successes have been made in the expression of a range of GPCRs using Pichia pastoris as an expression host. P. pastoris has a number of advantages over other expression systems including ability to post-translationally modify expressed proteins, relative low cost for production and ability to grow to very high cell densities. Several previous studies have described the expression of GPCRs in P. pastoris using shaker flasks, which allow culturing of small volumes (500 ml with moderate cell densities (OD600 ~15. The use of bioreactors, which allow straightforward culturing of large volumes, together with optimal control of growth parameters including pH and dissolved oxygen to maximise cell densities and expression of the target receptors, are an attractive alternative. The aim of this study was to compare the levels of expression of the human Adenosine 2A receptor (A2AR in P. pastoris under control of a methanol-inducible promoter in both flask and bioreactor cultures. Results Bioreactor cultures yielded an approximately five times increase in cell density (OD600 ~75 compared to flask cultures prior to induction and a doubling in functional expression level per mg of membrane protein, representing a significant optimisation. Furthermore, analysis of a C-terminally truncated A2AR, terminating at residue V334 yielded the highest levels (200 pmol/mg so far reported for expression of this receptor in P. pastoris. This truncated form of the receptor was also revealed to be resistant to C-terminal degradation in contrast to the WT A2AR, and therefore more suitable for further functional and structural studies. Conclusion Large-scale expression of the A2AR in P. pastoris bioreactor cultures results in significant increases in functional expression compared to traditional flask cultures.

  12. The orthosteric GABAA receptor ligand Thio-4-PIOL displays distinctly different functional properties at synaptic and extrasynaptic receptors

    DEFF Research Database (Denmark)

    Hoestgaard-Jensen, K; O'Connor, R M; Dalby, Nils Ole;

    2013-01-01

    Explorations into the heterogeneous population of native GABA type A receptors (GABAA Rs) and the physiological functions governed by the multiple GABAA R subtypes have for decades been hampered by the lack of subtype-selective ligands....

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

    Directory of Open Access Journals (Sweden)

    Md. Kaimul eAhsan

    2014-04-01

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

  14. Cell surface receptors for signal transduction and ligand transport - a design principles study

    Energy Technology Data Exchange (ETDEWEB)

    Shankaran, Harish; Resat, Haluk; Wiley, H. S.

    2007-06-01

    Although many different receptors undergo endocytosis, the system-level design principles that govern the evolution of receptor dynamics are far from fully understood. We have constructed a generalized mathematical model to understand how receptor internalization dynamics encodes receptor function and regulation. Parametric analysis of the response of receptor systems to ligand inputs reveals that receptors can be categorized a being: i) avidity-controlled where the response control depends primarily on the extracelluar ligand capture efficiency, ii) consumption-controlled where the ability to internalize surface-bound ligand is the primary control parameter, and iii) dual-sensitivity where both the avidity and consumption parameters are important. We show that the transferrin and low-density lipoprotein receptors are avidity-controlled, the vitellogenin receptor is consumption-controlled and epidermal growth factor receptor is a dual-sensitivity receptor. Significantly, we show that ligand-induced endocytosis is a mechanism to anhance the accuracy of signaling receptors rather than serving to attenuate signaling. Our analysis reveals that the location of a receptor system in the avidity-consumption parameter space can be used to understand both its function and its regulations.

  15. Complex pharmacology of novel allosteric free fatty acid 3 receptor ligands

    DEFF Research Database (Denmark)

    Hudson, Brian D; Christiansen, Elisabeth; Murdoch, Hannah;

    2014-01-01

    Analysis of the roles of the short chain fatty acid receptor, free fatty acid 3 receptor (FFA3), has been severely limited by the low potency of its endogenous ligands, the crossover of function of these on the closely related free fatty acid 2 receptor, and a dearth of FFA3-selective synthetic l...

  16. 1,2,3-triazolyl amino acids as AMPA receptor ligands

    DEFF Research Database (Denmark)

    Stanley, Nathan J.; Pedersen, Daniel Sejer; Nielsen, Birgitte;

    2010-01-01

    The central nervous system glutamate receptors are an important target for drug discovery. Herein we report initial investigations into the synthesis and glutamate receptor activity of 1,2,3-triazolyl amino acids. Two compounds were found to be selective AMPA receptor ligands, which warrant further...

  17. Ligand Promiscuity of Aryl Hydrocarbon Receptor Agonists and Antagonists Revealed by Site-Directed Mutagenesis

    OpenAIRE

    Soshilov, Anatoly A; DENISON, Michael S.

    2014-01-01

    The aryl hydrocarbon receptor (AhR) is a ligand-dependent transcription factor that can be activated by structurally diverse chemicals. To examine the mechanisms responsible for the promiscuity in AhR ligand binding, we determined the effects of mutations within the AhR ligand-binding domain (LBD) on the activity of diverse AhR ligands. Site-directed mutagenesis identified Ile319 of the mouse AhR and, to a lesser extent, Phe318 as residues involved in ligand-selective modulation of AhR transf...

  18. Multiparameter flow cytometry of a pH sensitive ligand bound to receptors and inside cells

    Energy Technology Data Exchange (ETDEWEB)

    Fay, S.P.; Habbersett, R.; Posner, R.G.; Domalewski, M.D.; Freer, R.J.; Pierson, E.; Whittaker, J.; Haugland, R.P.; Sklar, L.A. (Univ. of New Mexico, Albuquerque (United States) Los Alamos National Lab., NM (United States))

    1993-01-01

    Because fluoresceinated ligands of the neutrophil formyl peptide receptor can be protonated either upon binding to the receptor on the cell surface or in acidified intracellular compartments, the authors synthesized a ligand conjugated to the pH sensitive fluorescent probe SNAFL (CHO-Met-Leu-Phe-Phe-Lys-SNAFL). In the three laser flow cytometer at LANL, protonated dye is excited at 488 nm and emits at 530 nm; unprotonated dye is excited at 568 nm and emits at 650 nm. Detection at the isobestic and isoemissive points at 528 and 600 nm is used to keep track of variations in ligand concentration from sample to sample. The SNAFL-ligand bound to HL-60 cells (which overexpress the formyl peptide receptor) was compared to the free ligand in solution over a pH range from 6.5 to 9.0. The results suggest that the ligand bound to cell surface receptors was protonated in the binding pocket, possibly by virtue of its proximity to His 90, based on sequence data. When the cells were raised from 4[degrees] to 37[degrees], they also observed a time-dependent acidification of the ligand, indicative of ligand-receptor processing beginning 3-4 minutes after internalization.

  19. Monitoring ligand-receptor interactions by photonic force microscopy

    Energy Technology Data Exchange (ETDEWEB)

    Jeney, Sylvia [M E Mueller Institute for Structural Biology, Biozentrum, University of Basel, Klingelbergstrasse 70, Basel, 4056 (Switzerland); Mor, Flavio; Forro, Laszlo [Laboratory of Complex Matter Physics (LPMC), Ecole Polytechnique Federale de Lausanne (EPFL), CH-1015 Lausanne (Switzerland); Koszali, Roland [Institute for Information and Communication Technologies (IICT), University of Applied Sciences of Western Switzerland (HEIG-VD), Rue Galilee 15, CH 1401 Yverdon-les-bains (Switzerland); Moy, Vincent T, E-mail: sylvia.jeney@unibas.ch, E-mail: vmoy@miami.edu [Department of Physiology and Biophysics, University of Miami Miller School of Medicine, 1600 NW 10th Avenue, Miami, FL 33136 (United States)

    2010-06-25

    We introduce a method for the acquisition of single molecule force measurements of ligand-receptor interactions using the photonic force microscope (PFM). Biotin-functionalized beads, manipulated with an optical trap, and a streptavidin-functionalized coverslip were used to measure the effect of different pulling forces on the lifetime of individual streptavidin-biotin complexes. By optimizing the design of the optical trap and selection of the appropriate bead size, pulling forces in excess of 50 pN were achieved. Based on the amplitude of three-dimensional (3D) thermal position fluctuations of the attached bead, we were able to select for a bead-coverslip interaction that was mediated by a single streptavidin-biotin complex. Moreover, the developed experimental system was greatly accelerated by automation of data acquisition and analysis. In force-dependent kinetic measurements carried out between streptavidin and biotin, we observed that the streptavidin-biotin complex exhibited properties of a catch bond, with the lifetime increasing tenfold when the pulling force increased from 10 to 20 pN. We also show that silica beads were more appropriate than polystyrene beads for the force measurements, as tethers, longer than 200 nm, could be extracted from polystyrene beads.

  20. Mutual inactivation of Notch receptors and ligands facilitates developmental patterning.

    Directory of Open Access Journals (Sweden)

    David Sprinzak

    2011-06-01

    Full Text Available Developmental patterning requires juxtacrine signaling in order to tightly coordinate the fates of neighboring cells. Recent work has shown that Notch and Delta, the canonical metazoan juxtacrine signaling receptor and ligand, mutually inactivate each other in the same cell. This cis-interaction generates mutually exclusive sending and receiving states in individual cells. It generally remains unclear, however, how this mutual inactivation and the resulting switching behavior can impact developmental patterning circuits. Here we address this question using mathematical modeling in the context of two canonical pattern formation processes: boundary formation and lateral inhibition. For boundary formation, in a model motivated by Drosophila wing vein patterning, we find that mutual inactivation allows sharp boundary formation across a broader range of parameters than models lacking mutual inactivation. This model with mutual inactivation also exhibits robustness to correlated gene expression perturbations. For lateral inhibition, we find that mutual inactivation speeds up patterning dynamics, relieves the need for cooperative regulatory interactions, and expands the range of parameter values that permit pattern formation, compared to canonical models. Furthermore, mutual inactivation enables a simple lateral inhibition circuit architecture which requires only a single downstream regulatory step. Both model systems show how mutual inactivation can facilitate robust fine-grained patterning processes that would be difficult to implement without it, by encoding a difference-promoting feedback within the signaling system itself. Together, these results provide a framework for analysis of more complex Notch-dependent developmental systems.

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

    Science.gov (United States)

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

    2015-02-01

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

  2. Renoprotective Effects of a Highly Selective A3 Adenosine Receptor Antagonist in a Mouse Model of Adriamycin-induced Nephropathy.

    Science.gov (United States)

    Min, Hye Sook; Cha, Jin Joo; Kim, Kitae; Kim, Jung Eun; Ghee, Jung Yeon; Kim, Hyunwook; Lee, Ji Eun; Han, Jee Young; Jeong, Lak Shin; Cha, Dae Ryong; Kang, Young Sun

    2016-09-01

    The concentration of adenosine in the normal kidney increases markedly during renal hypoxia, ischemia, and inflammation. A recent study reported that an A3 adenosine receptor (A3AR) antagonist attenuated the progression of renal fibrosis. The adriamycin (ADX)-induced nephropathy model induces podocyte injury, which results in severe proteinuria and progressive glomerulosclerosis. In this study, we investigated the preventive effect of a highly selective A3AR antagonist (LJ1888) in ADX-induced nephropathy. Three groups of six-week-old Balb/c mice were treated with ADX (11 mg/kg) for four weeks and LJ1888 (10 mg/kg) for two weeks as following: 1) control; 2) ADX; and 3) ADX + LJ1888. ADX treatment decreased body weight without a change in water and food intake, but this was ameliorated by LJ1888 treatment. Interestingly, LJ1888 lowered plasma creatinine level, proteinuria, and albuminuria, which had increased during ADX treatment. Furthermore, LJ1888 inhibited urinary nephrin excretion as a podocyte injury marker, and urine 8-isoprostane and kidney lipid peroxide concentration, which are markers of oxidative stress, increased after injection of ADX. ADX also induced the activation of proinflammatory and profibrotic molecules such as TGF-β1, MCP-1, PAI-1, type IV collagen, NF-κB, NOX4, TLR4, TNFα, IL-1β, and IFN-γ, but they were remarkably suppressed after LJ1888 treatment. In conclusion, our results suggest that LJ1888 has a renoprotective effect in ADX-induced nephropathy, which might be associated with podocyte injury through oxidative stress. Therefore, LJ1888, a selective A3AR antagonist, could be considered as a potential therapeutic agent in renal glomerular diseases which include podocyte injury and proteinuria. PMID:27510383

  3. Effect of size and conformation of the ligand on asialoglycoprotein receptor-mediated ligand internalization and degradation in rat hepatocytes

    International Nuclear Information System (INIS)

    The rates of internalization and degradation of 125-I-labeled desialylated cyanogen bromide fragment I of orosomucoid (AS-CNBr-I) and its reduced and carboxymethylated derivative (AS-RC-CNBr-I) were compared with those of 125I-labeled asialoorosomucoid (ASOR) in rat hepatocytes. At 30 nM the rates of internalization and degradation of 125I-AS-CNBr-I were greater than those of 125I-ASOR. 125I-AS-RC-CNBr-I also had a lower rate of internalization and degradation. In contrast to 125I-ASOR, when degradation was inhibited by 5 μM colchicine there was a significant intracellular accumulation of the smaller ligands. At 40C the hepatocytes were found to bind the fragmented ligands more than 125I-ASOR. Incubation of the cells with bound ligand at 370 indicated that diacytosis of 125I-ASOR was greater than the smaller ligands. Colchincine markedly enhanced diacytosis of 125I-ASOR. On the other hand, there were marked accumulation of the smaller ligands by colchicine. These results suggest that the rates of internalization, degradation and diacytosis of the ligand are affected by the size and conformation of the ligand through different rates of receptor binding and intracellular transport

  4. Challenges predicting ligand-receptor interactions of promiscuous proteins: the nuclear receptor PXR.

    Directory of Open Access Journals (Sweden)

    Sean Ekins

    2009-12-01

    Full Text Available Transcriptional regulation of some genes involved in xenobiotic detoxification and apoptosis is performed via the human pregnane X receptor (PXR which in turn is activated by structurally diverse agonists including steroid hormones. Activation of PXR has the potential to initiate adverse effects, altering drug pharmacokinetics or perturbing physiological processes. Reliable computational prediction of PXR agonists would be valuable for pharmaceutical and toxicological research. There has been limited success with structure-based modeling approaches to predict human PXR activators. Slightly better success has been achieved with ligand-based modeling methods including quantitative structure-activity relationship (QSAR analysis, pharmacophore modeling and machine learning. In this study, we present a comprehensive analysis focused on prediction of 115 steroids for ligand binding activity towards human PXR. Six crystal structures were used as templates for docking and ligand-based modeling approaches (two-, three-, four- and five-dimensional analyses. The best success at external prediction was achieved with 5D-QSAR. Bayesian models with FCFP_6 descriptors were validated after leaving a large percentage of the dataset out and using an external test set. Docking of ligands to the PXR structure co-crystallized with hyperforin had the best statistics for this method. Sulfated steroids (which are activators were consistently predicted as non-activators while, poorly predicted steroids were docked in a reverse mode compared to 5alpha-androstan-3beta-ol. Modeling of human PXR represents a complex challenge by virtue of the large, flexible ligand-binding cavity. This study emphasizes this aspect, illustrating modest success using the largest quantitative data set to date and multiple modeling approaches.

  5. The sigma receptor as a ligand-regulated auxiliary potassium channel subunit.

    Science.gov (United States)

    Aydar, Ebru; Palmer, Christopher P; Klyachko, Vitaly A; Jackson, Meyer B

    2002-04-25

    The sigma receptor is a novel protein that mediates the modulation of ion channels by psychotropic drugs through a unique transduction mechanism depending neither on G proteins nor protein phosphorylation. The present study investigated sigma receptor signal transduction by reconstituting responses in Xenopus oocytes. Sigma receptors modulated voltage-gated K+ channels (Kv1.4 or Kv1.5) in different ways in the presence and absence of ligands. Association between Kv1.4 channels and sigma receptors was demonstrated by coimmunoprecipitation. These results indicate a novel mechanism of signal transduction dependent on protein-protein interactions. Domain accessibility experiments suggested a structure for the sigma receptor with two cytoplasmic termini and two membrane-spanning segments. The ligand-independent effects on channels suggest that sigma receptors serve as auxiliary subunits to voltage-gated K+ channels with distinct functional interactions, depending on the presence or absence of ligand.

  6. The A3 adenosine receptor (A3AR): therapeutic target and predictive biological marker in rheumatoid arthritis.

    Science.gov (United States)

    Fishman, Pnina; Cohen, Shira

    2016-09-01

    The Gi protein-associated A3 adenosine receptor (A3AR) is over-expressed in inflammatory cells, and this high expression is also reflected in the peripheral blood mononuclear cells of patients with autoimmune inflammatory diseases such as rheumatoid arthritis, psoriasis, and Crohn's disease. CF101, a selective agonist with high affinity to the A3AR, is known to induce robust anti-inflammatory effect in experimental animal models of adjuvant-, collagen-, and tropomyosin-induced arthritis. The effect is mediated via a definitive molecular mechanism entailing deregulation of the nuclear factor-κB (NF-κB) and the Wnt signal transduction pathways resulting in apoptosis of inflammatory cells. CF101 was found to be safe and well tolerated in all preclinical, phase I, and phase II human clinical studies. In two phase II clinical studies where CF101 was administered to rheumatoid arthritis (RA) patients as a stand-alone drug, a significant anti-rheumatic effect and a direct significant correlation were found between receptor expression at baseline and patients' response to the drug, suggesting that A3AR may be utilized as a predictive biomarker. The A3AR is a promising therapeutic target in rheumatoid arthritis and can be used also as a biological marker to predict patients' response to CF101. This is a unique type of a personalized medicine approach which may pave the way for a safe and efficacious treatment for this patient population. PMID:26886128

  7. Effects of a Proprietary Standardized Orthosiphon stamineus Ethanolic Leaf Extract on Enhancing Memory in Sprague Dawley Rats Possibly via Blockade of Adenosine A2A Receptors

    Directory of Open Access Journals (Sweden)

    Annie George

    2015-01-01

    Full Text Available The aim of the study was to explore a propriety standardized ethanolic extract from leaves of Orthosiphon stamineus Benth in improving impairments in short-term social memory in vivo, possibly via blockade of adenosine A2A receptors (A2AR. The ethanolic extract of O. stamineus leaves showed significant in vitro binding activity of A2AR with 74% inhibition at 150 μg/ml and significant A2AR antagonist activity with 98% inhibition at 300 μg/mL. A significant adenosine A1 receptor (A1R antagonist activity with 100% inhibition was observed at 300 μg/mL. Its effect on learning and memory was assessed via social recognition task using Sprague Dawley rats whereby the ethanolic extract of O. stamineus showed significant (p<0.001 change in recognition index (RI at 300 mg/kg and 600 mg/kg p.o and 120 mg/kg i.p., respectively, compared to the vehicle control. In comparison, the ethanolic extract of Polygonum minus aerial parts showed small change in inflexion; however, it remained insignificant in RI at 200 mg/kg p.o. Our findings suggest that the ethanolic extract of O. stamineus leaves improves memory by reversing age-related deficits in short-term social memory and the possible involvement of adenosine A1 and adenosine A2A as a target bioactivity site in the restoration of memory.

  8. Vasopressin V1 receptors contribute to hemodynamic and sympathoinhibitory responses evoked by stimulation of adenosine A2a receptors in NTS.

    Science.gov (United States)

    Scislo, Tadeusz J; O'Leary, Donal S

    2006-05-01

    Activation of adenosine A2a receptors in the nucleus of the solitary tract (NTS) decreases mean arterial pressure (MAP), heart rate (HR), and renal sympathetic nerve activity (RSNA), whereas increases in preganglionic adrenal sympathetic nerve activity (pre-ASNA) occur, a pattern similar to that observed during hypotensive hemorrhage. Central vasopressin V1 receptors may contribute to posthemorrhagic hypotension and bradycardia. Both V1 and A2a receptors are densely expressed in the NTS, and both of these receptors are involved in cardiovascular control; thus they may interact. The responses elicited by NTS A2a receptors are mediated mostly via nonglutamatergic mechanisms, possibly via release of vasopressin. Therefore, we investigated whether blockade of NTS V1 receptors alters the autonomic response patterns evoked by stimulation of NTS A2a receptors (CGS-21680, 20 pmol/50 nl) in alpha-chloralose-urethane anesthetized male Sprague-Dawley rats. In addition, we compared the regional sympathetic responses to microinjections of vasopressin (0.1-100 ng/50 nl) into the NTS. Blockade of V1 receptors reversed the normal decreases in MAP into increases (-95.6 +/- 28.3 vs. 51.4 +/- 15.7 integralDelta%), virtually abolished the decreases in HR (-258.3 +/- 54.0 vs. 18.9 +/- 57.8 integralDeltabeats/min) and RSNA (-239.3 +/- 47.4 vs. 15.9 +/- 36.1 integralDelta%), and did not affect the increases in pre-ASNA (279.7 +/- 48.3 vs. 233.1 +/- 54.1 integralDelta%) evoked by A2a receptor stimulation. The responses partially returned toward normal values approximately 90 min after the blockade. Microinjections of vasopressin into the NTS evoked dose-dependent decreases in HR and RSNA and variable MAP and pre-ASNA responses with a tendency toward increases. We conclude that the decreases in MAP, HR, and RSNA in response to NTS A2a receptor stimulation may be mediated via release of vasopressin from neural terminals in the NTS. The differential effects of NTS V1 and A2a receptors on

  9. Potential applications for sigma receptor ligands in cancer diagnosis and therapy.

    Science.gov (United States)

    van Waarde, Aren; Rybczynska, Anna A; Ramakrishnan, Nisha K; Ishiwata, Kiichi; Elsinga, Philip H; Dierckx, Rudi A J O

    2015-10-01

    Sigma receptors (sigma-1 and sigma-2) represent two independent classes of proteins. Their endogenous ligands may include the hallucinogen N,N-dimethyltryptamine (DMT) and sphingolipid-derived amines which interact with sigma-1 receptors, besides steroid hormones (e.g., progesterone) which bind to both sigma receptor subpopulations. The sigma-1 receptor is a ligand-regulated molecular chaperone with various ion channels and G-protein-coupled membrane receptors as clients. The sigma-2 receptor was identified as the progesterone receptor membrane component 1 (PGRMC1). Although sigma receptors are over-expressed in tumors and up-regulated in rapidly dividing normal tissue, their ligands induce significant cell death only in tumor tissue. Sigma ligands may therefore be used to selectively eradicate tumors. Multiple mechanisms appear to underlie cell killing after administration of sigma ligands, and the signaling pathways are dependent both on the type of ligand and the type of tumor cell. Recent evidence suggests that the sigma-2 receptor is a potential tumor and serum biomarker for human lung cancer and an important target for inhibiting tumor invasion and cancer progression. Current radiochemical efforts are focused on the development of subtype-selective radioligands for positron emission tomography (PET) imaging. Right now, the mostpromising tracers are [18F]fluspidine and [18F]FTC-146 for sigma-1 receptors and [11C]RHM-1 and [18F]ISO-1 for the sigma-2 subtype. Nanoparticles coupled to sigma ligands have shown considerable potential for targeted delivery of antitumor drugs in animal models of cancer, but clinical studies exploring this strategy in cancer patients have not yet been reported. This article is part of a Special Issue entitled: Membrane channels and transporters in cancers. PMID:25173780

  10. Imaging G protein-coupled receptors while quantifying their ligand-binding free-energy landscape.

    Science.gov (United States)

    Alsteens, David; Pfreundschuh, Moritz; Zhang, Cheng; Spoerri, Patrizia M; Coughlin, Shaun R; Kobilka, Brian K; Müller, Daniel J

    2015-09-01

    Imaging native membrane receptors and testing how they interact with ligands is of fundamental interest in the life sciences but has proven remarkably difficult to accomplish. Here, we introduce an approach that uses force-distance curve-based atomic force microscopy to simultaneously image single native G protein-coupled receptors in membranes and quantify their dynamic binding strength to native and synthetic ligands. We measured kinetic and thermodynamic parameters for individual protease-activated receptor-1 (PAR1) molecules in the absence and presence of antagonists, and these measurements enabled us to describe PAR1's ligand-binding free-energy landscape with high accuracy. Our nanoscopic method opens an avenue to directly image and characterize ligand binding of native membrane receptors. PMID:26167642

  11. Ligands, cell-based models, and readouts required for Toll-like receptor action.

    LENUS (Irish Health Repository)

    Dellacasagrande, Jerome

    2012-02-01

    This chapter details the tools that are available to study Toll-like receptor (TLR) biology in vitro. This includes ligands, host cells, and readouts. The use of modified TLRs to circumvent some technical problems is also discussed.

  12. Computational Exploration of a Protein Receptor Binding Space with Student Proposed Peptide Ligands

    Science.gov (United States)

    King, Matthew D.; Phillips, Paul; Turner, Matthew W.; Katz, Michael; Lew, Sarah; Bradburn, Sarah; Andersen, Tim; McDougal, Owen M.

    2016-01-01

    Computational molecular docking is a fast and effective "in silico" method for the analysis of binding between a protein receptor model and a ligand. The visualization and manipulation of protein to ligand binding in three-dimensional space represents a powerful tool in the biochemistry curriculum to enhance student learning. The…

  13. Computer-aided design of a novel ligand for retinoic acid receptor in cancer chemotherapy

    Science.gov (United States)

    Silva, Carlos H. T. P.; Leopoldino, Andreia M.; Silva, Eloiza H. T.; Espinoza, V. A. A.; Taft, C. A.

    The isotypes of RAR and RXR are retinoic acid and retinoid X acid receptors, respectively, whose ligand-binding domain contains the ligand-dependent activation function, with distinct pharmacological targets for retinoids, involved in the treatment of various cancers and skin diseases. Due to the major challenge which cancer treatment and cure still imposes after many decades to the international scientific community, there is actually considerable interest in new ligands with increased bioactivity. We have focused on the retinoid acid receptor, which is considered an interesting target for drug design. In this work, we carried out density functional geometry optimizations and different docking procedures. We performed screening in a large database (hundreds of thousands of molecules which we optimized at the AM1 level) yielding a set of potential bioactive ligands. A new ligand was selected and optimized at the B3LYP/6-31G* level. A flexible docking program was used to investigate the interactions between the receptor and the new ligand. The result of this work is compared with several crystallographic ligands of RAR. Our theoretically more bioactive new ligand indicates stronger and more hydrogen bonds as well as hydrophobic interactions with the receptor.

  14. Expression and Purification of Functional Ligand-binding Domains of T1R3 Taste Receptors

    Energy Technology Data Exchange (ETDEWEB)

    Nie,Y.; Hobbs, J.; Vigues, S.; Olson, W.; Conn, G.; Munger, S.

    2006-01-01

    Chemosensory receptors, including odor, taste, and vomeronasal receptors, comprise the largest group of G protein-coupled receptors (GPCRs) in the mammalian genome. However, little is known about the molecular determinants that are critical for the detection and discrimination of ligands by most of these receptors. This dearth of understanding is due in part to difficulties in preparing functional receptors suitable for biochemical and biophysical analyses. Here we describe in detail two strategies for the expression and purification of the ligand-binding domain of T1R taste receptors, which are constituents of the sweet and umami taste receptors. These class C GPCRs contain a large extracellular N-terminal domain (NTD) that is the site of interaction with most ligands and that is amenable to expression as a separate polypeptide in heterologous cells. The NTD of mouse T1R3 was expressed as two distinct fusion proteins in Escherichia coli and purified by column chromatography. Spectroscopic analysis of the purified NTD proteins shows them to be properly folded and capable of binding ligands. This methodology should not only facilitate the characterization of T1R ligand interactions but may also be useful for dissecting the function of other class C GPCRs such as the large family of orphan V2R vomeronasal receptors.

  15. Impact of killer immunoglobulin-like receptor-human leukocyte antigens ligand incompatibility among renal transplantation.

    Science.gov (United States)

    Alam, S; Rangaswamy, D; Prakash, S; Sharma, R K; Khan, M I; Sonawane, A; Agrawal, S

    2015-01-01

    Killer immunoglobulin-like receptor (KIR) gene shows a high degree of polymorphism. Natural killer cell receptor gets activated once they bind to self-human leukocyte antigens (HLAs) with specific ligand. KIR gene and HLA ligand incompatibility due to the presence/absence of KIR in the recipient and the corresponding HLA ligand in the allograft may impact graft survival in solid organ transplantation. This study evaluates the effect of matches between KIR genes and known HLA ligands. KIR genotypes were determined using sequence specific primer polymerase chain reaction. Presence of certain KIR in a recipient, where the donor lacked the corresponding HLA ligand was considered a mismatch. The allograft was considered matched when both KIR receptor and HLA alloantigen reveald compatibility among recipient and donor. The data revealed better survival among individuals with matched inhibitory KIR receptors and their corresponding HLA ligands (KIR2DL2/DL3-HLAC2, KIR3DL1-HLABw4). On the contrary, no adverse effect was seen for matched activating KIR receptors and their corresponding HLA ligands. One of the activating gene KIR2DS4 showed risk (P = 0.0413, odds ratio = 1.91, 95% confidence interval = 1.02-3.57) association with renal allograft rejection. We conclude that the presence of inhibitory KIR gene leads to better survival; whereas activating motifs show no significant role in renal allograft survival.

  16. ASSESSMENT OF COCAINE-LIKE DISCRIMINATIVE STIMULUS EFFECTS OF DOPAMINE D-3 RECEPTOR LIGANDS

    NARCIS (Netherlands)

    ACRI, JB; CARTER, [No Value; ALLING, K; GETERDOUGLASS, B; DIJKSTRA, D; WIKSTROM, H; KATZ, JL; WITKIN, JM

    1995-01-01

    The highly selective dopamine D-3 receptor ligand, (+)-PD 128907 4aR10bR-(+)-trans-3,4,4a,10b-tetrahydro-4-n-propyl- 2H5H[4,3-b]-1,4-oxazin-9-ol), and other dopamine D-3 receptor ligands, (+/-)-7-hydroxy-2-(N,N-di-n-propylamino)tetralin and (+)-7-hydroxy-2-(N,N-di-n-propylamino)tetralin, substituted

  17. Ligand-gated chloride channels are receptors for biogenic amines in C. elegans

    OpenAIRE

    Ringstad, Niels; Abe, Namiko; Horvitz, H. Robert

    2009-01-01

    Biogenic amines such as serotonin and dopamine are intercellular signaling molecules that function widely as neurotransmitters and neuromodulators. We have identified in the nematode Caenorhabditis elegans three ligand-gated chloride channels that are receptors for biogenic amines: LGC-53 is a high-affinity dopamine receptor, LGC-55 is a high-affinity tyramine receptor, and LGC-40 is a low-affinity serotonin receptor that is also gated by choline and acetylcholine. lgc-55 mutants are defectiv...

  18. Modulation of glutamat AMPA receptors by adenosine, in physiological and hypoxic/ischemic conditions

    OpenAIRE

    Dias, Raquel Alice da Silva Baptista, 1983-

    2011-01-01

    Tese de doutoramento, Ciências Biomédicas (Neurociências), Universidade de Lisboa, Faculdade de Medicina, 2011 Most of the fast excitatory transmission in the brain is conveyed by ionotropic glutamate a-amino-3-hydroxy-5-methyl-4- isoxazolepropionic acid (AMPA) receptors, formed by tetrameric assemblies of different subunit (GluR1-GluR4) composition. Modulation of AMPA receptors enables profound changes in synaptic efficiency, underlying the maturation of neuronal networks t...

  19. A ligand channel through the G protein coupled receptor opsin.

    Directory of Open Access Journals (Sweden)

    Peter W Hildebrand

    Full Text Available The G protein coupled receptor rhodopsin contains a pocket within its seven-transmembrane helix (TM structure, which bears the inactivating 11-cis-retinal bound by a protonated Schiff-base to Lys296 in TM7. Light-induced 11-cis-/all-trans-isomerization leads to the Schiff-base deprotonated active Meta II intermediate. With Meta II decay, the Schiff-base bond is hydrolyzed, all-trans-retinal is released from the pocket, and the apoprotein opsin reloaded with new 11-cis-retinal. The crystal structure of opsin in its active Ops* conformation provides the basis for computational modeling of retinal release and uptake. The ligand-free 7TM bundle of opsin opens into the hydrophobic membrane layer through openings A (between TM1 and 7, and B (between TM5 and 6, respectively. Using skeleton search and molecular docking, we find a continuous channel through the protein that connects these two openings and comprises in its central part the retinal binding pocket. The channel traverses the receptor over a distance of ca. 70 A and is between 11.6 and 3.2 A wide. Both openings are lined with aromatic residues, while the central part is highly polar. Four constrictions within the channel are so narrow that they must stretch to allow passage of the retinal beta-ionone-ring. Constrictions are at openings A and B, respectively, and at Trp265 and Lys296 within the retinal pocket. The lysine enforces a 90 degrees elbow-like kink in the channel which limits retinal passage. With a favorable Lys side chain conformation, 11-cis-retinal can take the turn, whereas passage of the all-trans isomer would require more global conformational changes. We discuss possible scenarios for the uptake of 11-cis- and release of all-trans-retinal. If the uptake gate of 11-cis-retinal is assigned to opening B, all-trans is likely to leave through the same gate. The unidirectional passage proposed previously requires uptake of 11-cis-retinal through A and release of photolyzed all

  20. Internalization mechanisms of the epidermal growth factor receptor after activation with different ligands.

    Directory of Open Access Journals (Sweden)

    Lasse Henriksen

    Full Text Available The epidermal growth factor receptor (EGFR regulates normal growth and differentiation, but dysregulation of the receptor or one of the EGFR ligands is involved in the pathogenesis of many cancers. There are eight ligands for EGFR, however most of the research into trafficking of the receptor after ligand activation focuses on the effect of epidermal growth factor (EGF and transforming growth factor-α (TGF-α. For a long time it was believed that clathrin-mediated endocytosis was the major pathway for internalization of the receptor, but recent work suggests that different pathways exist. Here we show that clathrin ablation completely inhibits internalization of EGF- and TGF-α-stimulated receptor, however the inhibition of receptor internalization in cells treated with heparin-binding EGF-like growth factor (HB-EGF or betacellulin (BTC was only partial. In contrast, clathrin knockdown fully inhibits EGFR degradation after all ligands tested. Furthermore, inhibition of dynamin function blocked EGFR internalization after stimulation with all ligands. Knocking out a number of clathrin-independent dynamin-dependent pathways of internalization had no effect on the ligand-induced endocytosis of the EGFR. We suggest that EGF and TGF-α lead to EGFR endocytosis mainly via the clathrin-mediated pathway. Furthermore, we suggest that HB-EGF and BTC also lead to EGFR endocytosis via a clathrin-mediated pathway, but can additionally use an unidentified internalization pathway or better recruit the small amount of clathrin remaining after clathrin knockdown.

  1. Directed evolution of estrogen receptor proteins with altered ligand-binding specificities.

    Science.gov (United States)

    Islam, Kazi Mohammed Didarul; Dilcher, Meik; Thurow, Corinna; Vock, Carsten; Krimmelbein, Ilga Kristine; Tietze, Lutz Friedjan; Gonzalez, Victor; Zhao, Huimin; Gatz, Christiane

    2009-01-01

    Transcriptional activators that respond to ligands with no cellular targets are powerful tools that can confer regulated expression of a transgene in almost all biological systems. In this study, we altered the ligand-binding specificity of the human estrogen receptor alpha (hER alpha) so that it would recognize a non-steroidal synthetic compound with structural similarities to the phytoestrogen resveratrol. For this purpose, we performed iterative rounds of site-specific saturation mutagenesis of a fixed set of ligand-contacting residues and subsequent random mutagenesis of the entire ligand-binding domain. Selection of the receptor mutants and quantification of the interaction were carried out by exploiting a yeast two-hybrid system that reports the ligand-dependent interaction between hER alpha and steroid receptor coactivator-1 (SRC-1). The screen was performed with a synthetic ligand (CV3320) that promoted growth of the reporter yeast strain to half maximal levels at a concentration of 3.7 microM. The optimized receptor mutant (L384F/L387M/Y537S) showed a 67-fold increased activity to the synthetic ligand CV3320 (half maximal yeast growth at 0.055 microM) and a 10-fold decreased activity to 17beta-estradiol (E2; half maximal yeast growth at 4 nM). The novel receptor-ligand pair partially fulfills the requirements for a specific 'gene switch' as it responds to concentrations of the synthetic ligand which do not activate the wildtype receptor. Due to its residual responsiveness to E2 at concentrations (4 nM) that might occur in vivo, further improvements have to be performed to render the system applicable in organisms with endogenous E2 synthesis.

  2. Histamine H4 receptor ligands: future applications and state of art.

    Science.gov (United States)

    Corrêa, Michelle Fidelis; dos Santos Fernandes, João Paulo

    2015-04-01

    Histamine is a chemical transmitter found practically in whole organism and exerts its effects through the interaction with H1 to H4 histaminergic receptors. Specifically, H4 receptors are found mainly in immune cells and blood-forming tissues, thus are involved in inflammatory and immune processes, as well as some actions in central nervous system. Therefore, H4 receptor ligands can have applications in the treatment of chronic inflammatory and immune diseases and may be novel therapeutic option in these conditions. Several H4 receptor ligands have been described from early 2000's until nowadays, being imidazole, indolecarboxamide, 2-aminopyrimidine, quinazoline, and quinoxaline scaffolds the most explored and discussed in this review. Moreover, several studies of molecular modeling using homology models of H4 receptor and QSAR data of the ligands are summarized. The increasing and promising therapeutic applications are leading these compounds to clinical trials, which probably will be part of the next generation of blockbuster drugs. PMID:25228262

  3. The type I interleukin-1 receptor mediates fever in the rat as shown by interleukin-1 receptor subtype selective ligands.

    Science.gov (United States)

    Malinowsky, D; Chai, Z; Bristulf, J; Simoncsits, A; Bartfai, T

    1995-12-01

    The interleukin-1 (IL-1) system possesses two distinct receptors (type I and type II) which, together with the accessory protein, mediate a multitude of responses to IL-1 alpha and IL-1 beta, including fever. So far, no receptor subtype-specific ligands have been described. Since both types of IL-1 receptors occur in the thermoregulatory areas it was unclear which IL-1 receptor type mediates fever. We report here that for a series of deletion mutants of human recombinant IL-1 beta (hrIL-1 beta), the affinity of these ligands for the type I IL-1 receptor correlates with their efficacy to evoke the fever response (hrIL-1 beta > des-SND52-54 > des-QGE48-50 > des-I56). Thus, the results suggest that agonist occupancy of the type I IL-1 receptor is essential for IL-1 beta-mediated fever.

  4. The novel heteromeric bivalent ligand SB9 potently antagonizes P2Y(1) receptor-mediated responses.

    Science.gov (United States)

    Lambrecht, G; Ganso, M; Bäumert, H G; Spatz-Kümbel, G; Hildebrandt, C; Braun, K; Mutschler, E

    2000-07-01

    Effects of 6-[(4,6,8-trisulfo-1-naphthyl)iminocarbonyl-1, 3-(4-methylphenylene)iminocarbonyl-1, 3-phenylene-azo]-pyridoxal-5'-phosphate (SB9), a heterodimeric bivalent ligand consisting of pyridoxal-5'-phosphate and the suramin monomer, were studied on contractions of the rat vas deferens elicited by alpha beta-methylene ATP (alpha beta meATP; mediated by P2X(1)-like receptors), contractions of the guinea-pig ileal longitudinal smooth muscle elicited by adenosine 5'-O-(2-thiodiphosphate) (ADP beta S mediated by P2Y(1)-like receptors), and the degradation of ATP by ecto-nucleotidases in folliculated Xenopus laevis oocytes. SB9 (0.1-10 microM) antagonized contractile responses produced by alpha beta meATP or ADP beta S in a concentration-dependent manner. Schild analysis yielded linear regression lines of unit slope, indicating competitive antagonism. From the rightward shifts of the agonist concentration-response curves pA(2) values of 6.05+/-0.13 (vas deferens) and 6.98+/-0.07 (ileum) were derived. In both preparations, SB9 behaved as a slow onset, slow offset antagonist. Incubation of three oocytes in the presence of ATP produced an increase in inorganic phosphate (P(i)) over a 30-min period, which amounted to 35.1+/-1.9 microM P(i) from 100 microM ATP. SB9 (10-1000 microM) reduced this degradation (pIC(50)=4.33+/-0.10). The results illustrate that SB9 is a high-affinity P2Y(1) receptor antagonist with a remarkable selectivity for P2Y(1) vs. P2X(1) receptors (about 10-fold) and ecto-nucleotidases (447-fold). These properties make it unique among the pyridoxal-5'-phosphate and suramin derivatives reported to date.

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

    Science.gov (United States)

    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

  6. Vasopressin is a major vasoconstrictor involved in hindlimb vascular responses to stimulation of adenosine A1 receptors in the nucleus of the solitary tract

    OpenAIRE

    McClure, Joseph M.; Rossi, Noreen F.; Chen, Haiping; O'Leary, Donal S.; Scislo, Tadeusz J.

    2009-01-01

    Our previous study showed that stimulation of adenosine A1 receptors located in the nucleus of the solitary tract (NTS) exerts counteracting effects on the iliac vascular bed: activation of the adrenal medulla and β-adrenergic vasodilation versus vasoconstriction mediated by neural and unknown humoral factors. In the present study we investigated the relative contribution of three major potential humoral vasoconstrictors: vasopressin, angiotensin II, and norepinephrine in this response. In ur...

  7. Rescuing the Corticostriatal Synaptic Disconnection in the R6/2 Mouse Model of Huntington’s Disease: Exercise, Adenosine Receptors and Ampakines

    OpenAIRE

    Cepeda, C.; Cummings, D. M.; Hickey, M. A.; Kleiman-Weiner, M.; Chen, J. Y.; Watson, J B; Levine, M.S.

    2010-01-01

    In the R6/2 mouse model of Huntington's disease (HD) we examined the effects of a number of behavioral and pharmacological manipulations aimed at rescuing the progressive loss of synaptic communication between cerebral cortex and striatum. Two cohorts of transgenic mice with ~110 and 210 CAG repeats were utilized. Exercise prevented the reduction in striatal medium-sized spiny neuron membrane capacitance but did not reestablish synaptic communication. Activation of adenosine A2A type receptor...

  8. An ELISA Based Binding and Competition Method to Rapidly Determine Ligand-receptor Interactions.

    Science.gov (United States)

    Syedbasha, Mohameedyaseen; Linnik, Janina; Santer, Deanna; O'Shea, Daire; Barakat, Khaled; Joyce, Michael; Khanna, Nina; Tyrrell, D Lorne; Houghton, Michael; Egli, Adrian

    2016-01-01

    A comprehensive understanding of signaling pathways requires detailed knowledge regarding ligand-receptor interaction. This article describes two fast and reliable point-by-point protocols of enzyme-linked immunosorbent assays (ELISAs) for the investigation of ligand-receptor interactions: the direct ligand-receptor interaction assay (LRA) and the competition LRA. As a case study, the ELISA based analysis of the interaction between different lambda interferons (IFNLs) and the alpha subunit of their receptor (IL28RA) is presented: the direct LRA is used for the determination of dissociation constants (KD values) between receptor and IFN ligands, and the competition LRA for the determination of the inhibitory capacity of an oligopeptide, which was designed to compete with the IFNLs at their receptor binding site. Analytical steps to estimate KD and half maximal inhibitory concentration (IC50) values are described. Finally, the discussion highlights advantages and disadvantages of the presented method and how the results enable a better molecular understanding of ligand-receptor interactions.

  9. Following a TRAIL:Update on a ligand and its five receptors

    Institute of Scientific and Technical Information of China (English)

    Fiona C. KIMBERLEY; Gavin R. SCREATON

    2004-01-01

    Identification of tumour necrosis factor apoptosis inducing ligand (TRAIL), a TNF family ligand, sparked a torrent of research, following an initial observation that it could kill tumour cells, but spare normal cells. Almost a decade after its discovery, and with five known receptors, the true physiological role of TRAIL is still debated and its anti-tumorigenic properties limited by potential toxicity. This review takes a comprehensive look at the story of this enigmatic ligand,addressing its remaining potential as a therapeutic and providing an overview of the TRAIL receptors themselves.

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

    Directory of Open Access Journals (Sweden)

    A. Heidarianpour

    2008-10-01

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

  11. Domain interplay in the urokinase receptor. Requirement for the third domain in high affinity ligand binding and demonstration of ligand contact sites in distinct receptor domains

    DEFF Research Database (Denmark)

    Behrendt, N; Ronne, E; Dano, K

    1996-01-01

    . This result shows that in addition to D1, which has an established function in ligand binding (Behrendt, N., Ploug, M., Patthy, L., Houen, G., Blasi, F., and Dano, K. (1991) J. Biol. Chem. 266, 7842-7847), D3 has an important role in governing a high affinity in the intact receptor. Real-time biomolecular...

  12. Administration of caffeine inhibited adenosine receptor agonist-induced decreases in motor performance, thermoregulation, and brain neurotransmitter release in exercising rats.

    Science.gov (United States)

    Zheng, Xinyan; Hasegawa, Hiroshi

    2016-01-01

    We examined the effects of an adenosine receptor agonist on caffeine-induced changes in thermoregulation, neurotransmitter release in the preoptic area and anterior hypothalamus, and endurance exercise performance in rats. One hour before the start of exercise, rats were intraperitoneally injected with either saline alone (SAL), 10 mg kg(-1) caffeine and saline (CAF), a non-selective adenosine receptor agonist (5'-N-ethylcarboxamidoadenosine [NECA]: 0.5 mg kg(-1)) and saline (NECA), or the combination of caffeine and NECA (CAF+NECA). Rats ran until fatigue on the treadmill with a 5% grade at a speed of 18 m min(-1) at 23 °C. Compared to the SAL group, the run time to fatigue (RTTF) was significantly increased by 52% following caffeine administration and significantly decreased by 65% following NECA injection (SAL: 91 ± 14.1 min; CAF: 137 ± 25.8 min; NECA: 31 ± 13.7 min; CAF+NECA: 85 ± 11.8 min; pcaffeine injection inhibited the NECA-induced decreases in the RTTF, Tcore, heat production, heat loss, and extracellular DA release. Neither caffeine nor NECA affected extracellular noradrenaline or serotonin release. These results support the findings of previous studies showing improved endurance performance and overrides in body limitations after caffeine administration, and imply that the ergogenic effects of caffeine may be associated with the adenosine receptor blockade-induced increases in brain DA release.

  13. Intracellular adenosine 3',5'-phosphate formation is essential for down-regulation of surface adenosine 3',5'-phosphate receptors in Dictyostelium

    OpenAIRE

    Van Haastert, Peter J. M.

    1994-01-01

    Dictyostelium discoideum cells contain cell surface cyclic AMP (cAMP) receptors that bind cAMP as a first messenger and intracellular cAMP receptors that bind cAMP as a second messenger. Prolonged incubation of Dictyostelium cells with cAMP induces a sequential process of phosphorylation, sequestration and down-regulation of the surface receptors. The role of intracellular cAMP in down-regulation of surface receptors was investigated. Down-regulation of receptors does not occur under conditio...

  14. Role of the T cell receptor ligand affinity in T cell activation by bacterial superantigens

    DEFF Research Database (Denmark)

    Andersen, P S; Geisler, C; Buus, S;

    2001-01-01

    (SEC3) with up to a 150-fold increase in TCR affinity. By stimulating T cells with SEC3 molecules immobilized onto plastic surfaces, we demonstrate that increasing the affinity of the SEC3/TCR interaction caused a proportional increase in the ability of SEC3 to activate T cells. Thus, the potency......Similar to native peptide/MHC ligands, bacterial superantigens have been found to bind with low affinity to the T cell receptor (TCR). It has been hypothesized that low ligand affinity is required to allow optimal TCR signaling. To test this, we generated variants of Staphylococcus enterotoxin C3...... correlation between ligand affinity and ligand potency indicating that it is the density of receptor-ligand complexes in the T cell contact area that determines TCR signaling strength....

  15. Revealing a steroid receptor ligand as a unique PPARγagonist

    Institute of Scientific and Technical Information of China (English)

    Shengchen Lin; Ying Han; Yuzhe Shi; Hui Rong; Songyang Zheng; Shikan Jin; Shu-Yong Lin; Sheng-Cai Lin; Yong Li

    2012-01-01

    Peroxisome proliferator-activated receptor gamma (PPARγ) regulates metabolic homeostasis and is a molecular target for anti-diabetic drugs.We report here the identification of a steroid receptor ligand,RU-486,as an unexpected PPARγ agonist,thereby uncovering a novel signaling route for this steroid drug.Similar to rosiglitazone,RU486 modulates the expression of key PPARγ target genes and promotes adipocyte differentiation,but with a lower adipogenic activity.Structural and functional studies of receptor-ligand interactions reveal the molecular basis for a unique binding mode for RU-486 in the PPARγ ligand-binding pocket with distinctive properties and epitopes,providing the molecular mechanisms for the discrimination of RU-486 from thiazolidinediones (TZDs) drugs.Our findings together indicate that steroid compounds may represent an alternative approach for designing non-TZD PPARγligands in the treatment of insulin resistance.

  16. Revealing a steroid receptor ligand as a unique PPAR[gamma] agonist

    Energy Technology Data Exchange (ETDEWEB)

    Lin, Shengchen; Han, Ying; Shi, Yuzhe; Rong, Hui; Zheng, Songyang; Jin, Shikan; Lin, Shu-Yong; Lin, Sheng-Cai; Li, Yong (Pitt); (Xiamen)

    2012-06-28

    Peroxisome proliferator-activated receptor gamma (PPAR{gamma}) regulates metabolic homeostasis and is a molecular target for anti-diabetic drugs. We report here the identification of a steroid receptor ligand, RU-486, as an unexpected PPAR{gamma} agonist, thereby uncovering a novel signaling route for this steroid drug. Similar to rosiglitazone, RU-486 modulates the expression of key PPAR{gamma} target genes and promotes adipocyte differentiation, but with a lower adipogenic activity. Structural and functional studies of receptor-ligand interactions reveal the molecular basis for a unique binding mode for RU-486 in the PPAR{gamma} ligand-binding pocket with distinctive properties and epitopes, providing the molecular mechanisms for the discrimination of RU-486 from thiazolidinediones (TZDs) drugs. Our findings together indicate that steroid compounds may represent an alternative approach for designing non-TZD PPAR{gamma} ligands in the treatment of insulin resistance.

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

    Science.gov (United States)

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

    2016-08-25

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

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

    Science.gov (United States)

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

    2016-08-01

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

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

    Science.gov (United States)

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

    2016-01-01

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

  20. Adenosine A2A Receptor and IL-10 in Peripheral Blood Mononuclear Cells of Patients with Mild Cognitive Impairment

    Directory of Open Access Journals (Sweden)

    Beatrice Arosio

    2011-01-01

    Full Text Available Adenosine suppresses immune responses through the A2A receptor (A2AR. This study investigated the interleukin 10 (IL-10 genetic profile and the expression of A2AR in peripheral blood mononuclear cells (PBMCs of patients with mild cognitive impairment (MCI, Alzheimer disease (AD, and age-matched controls to verify, if they may help distinguish different forms of cognitive decline. We analyzed the IL-10 genotype and the expression of A2AR in 41 subjects with AD, 10 with amnestic MCI (a-MCI, 49 with multiple cognitive domain MCI (mcd-MCI, and 46 controls. There was a significant linear increase in A2AR mRNA levels and A2AR density from mcd-MCI to a-MCI, with intermediate levels being found in AD. The IL-10 AA genotype frequency was 67% in a-MCI, 46% in AD, 35% in mcd-MCI, and 20% in controls. These data suggest that the assessment of the IL-10 genotype and the expression of A2AR in PBMCs may be a valuable means of differentiating between a-MCI and mcd-MCI.

  1. Visualization and ligand-induced modulation of dopamine receptor dimerization at the single molecule level.

    Science.gov (United States)

    Tabor, Alina; Weisenburger, Siegfried; Banerjee, Ashutosh; Purkayastha, Nirupam; Kaindl, Jonas M; Hübner, Harald; Wei, Luxi; Grömer, Teja W; Kornhuber, Johannes; Tschammer, Nuska; Birdsall, Nigel J M; Mashanov, Gregory I; Sandoghdar, Vahid; Gmeiner, Peter

    2016-01-01

    G protein-coupled receptors (GPCRs), including dopamine receptors, represent a group of important pharmacological targets. An increased formation of dopamine receptor D2 homodimers has been suggested to be associated with the pathophysiology of schizophrenia. Selective labeling and ligand-induced modulation of dimerization may therefore allow the investigation of the pathophysiological role of these dimers. Using TIRF microscopy at the single molecule level, transient formation of homodimers of dopamine receptors in the membrane of stably transfected CHO cells has been observed. The equilibrium between dimers and monomers was modulated by the binding of ligands; whereas antagonists showed a ratio that was identical to that of unliganded receptors, agonist-bound D2 receptor-ligand complexes resulted in an increase in dimerization. Addition of bivalent D2 receptor ligands also resulted in a large increase in D2 receptor dimers. A physical interaction between the protomers was confirmed using high resolution cryogenic localization microscopy, with ca. 9 nm between the centers of mass. PMID:27615810

  2. Potential applications for sigma receptor ligands in cancer diagnosis and therapy

    NARCIS (Netherlands)

    van Waarde, Aren; Rybczynska, Anna A.; Kuzhuppilly Ramakrishnan, Nisha; Ishiwata, Kiichi; Elsinga, Philip H.; Dierckx, Rudi A. J. O.

    2015-01-01

    Sigma receptors (sigma-1 and sigma-2) represent two independent classes of proteins. Their endogenous ligands may include the hallucinogen N,N-dimethyltryptamine (DMT) and sphingolipid-derived amines which interact with sigma-1 receptors, besides steroid hormones (e.g., progesterone) which bind to b

  3. Increased accuracy of ligand sensing by receptor diffusion on cell surface

    Science.gov (United States)

    Aquino, Gerardo; Endres, Robert G.

    2010-10-01

    The physical limit with which a cell senses external ligand concentration corresponds to the perfect absorber, where all ligand particles are absorbed and overcounting of same ligand particles does not occur. Here, we analyze how the lateral diffusion of receptors on the cell membrane affects the accuracy of sensing ligand concentration. Specifically, we connect our modeling to neurotransmission in neural synapses where the diffusion of glutamate receptors is already known to refresh synaptic connections. We find that receptor diffusion indeed increases the accuracy of sensing for both the glutamate α -Amino-3-hydroxy-5-Methyl-4-isoxazolePropionic Acid (AMPA) and N -Methyl-D-aspartic Acid (NMDA) receptor, although the NMDA receptor is overall much noisier. We propose that the difference in accuracy of sensing of the two receptors can be linked to their different roles in neurotransmission. Specifically, the high accuracy in sensing glutamate is essential for the AMPA receptor to start membrane depolarization, while the NMDA receptor is believed to work in a second stage as a coincidence detector, involved in long-term potentiation and memory.

  4. Modeling multivalent ligand-receptor interactions with steric constraints on configurations of cell surface receptor aggregates

    Energy Technology Data Exchange (ETDEWEB)

    Monine, Michael [Los Alamos National Laboratory; Posner, Richard [TRANSLATION GENOMICS RESAEARCH INSTITUTE; Savage, Paul [BYU; Faeder, James [UNIV OF PITTSBURGH; Hlavacek, William S [UNM

    2008-01-01

    Signal transduction generally involves multivalent protein-protein interactions, which can produce various protein complexes and post-translational modifications. The reaction networks that characterize these interactions tend to be so large as to challenge conventional simulation procedures. To address this challenge, a kinetic Monte Carlo (KMC) method has been developed that can take advantage of a model specification in terms of reaction rules for molecular interactions. A set of rules implicitly defines the reactions that can occur as a result of the interactions represented by the rules. With the rule-based KMC method, explicit generation of the underlying chemical reaction network implied by rules is avoided. Here, we apply and extend this method to characterize the interactions of a trivalent ligand with a bivalent cell-surface receptor. This system is also studied experimentally. We consider the following kinetic models: an equivalent-site model, an extension of this model, which takes into account steric constraints on the configurations of receptor aggregates, and finally, a model that accounts for cyclic receptor aggregates. Simulation results for the equivalent-site model are consistent with an equilibrium continuum model. Using these models, we investigate the effects of steric constraints and the formation of cyclic aggregates on the kinetics and equilibria of small and large aggregate formation and the percolation phase transition that occurs in this system.

  5. Adenosine A1 receptor activation modulates N-methyl-d-aspartate (NMDA) preconditioning phenotype in the brain.

    Science.gov (United States)

    Constantino, Leandra C; Pamplona, Fabrício A; Matheus, Filipe C; Ludka, Fabiana K; Gomez-Soler, Maricel; Ciruela, Francisco; Boeck, Carina R; Prediger, Rui D; Tasca, Carla I

    2015-04-01

    N-methyl-d-aspartate (NMDA) preconditioning is induced by subtoxic doses of NMDA and it promotes a transient state of resistance against subsequent lethal insults. Interestingly, this mechanism of neuroprotection depends on adenosine A1 receptors (A1R), since blockade of A1R precludes this phenomenon. In this study we evaluated the consequences of NMDA preconditioning on the hippocampal A1R biology (i.e. expression, binding properties and functionality). Accordingly, we measured A1R expression in NMDA preconditioned mice (75mg/kg, i.p.; 24h) and showed that neither the total amount of receptor, nor the A1R levels in the synaptic fraction was altered. In addition, the A1R binding affinity to the antagonist [(3)H] DPCPX was slightly increased in total membrane extracts of hippocampus from preconditioned mice. Next, we evaluated the impact of NMDA preconditioning on A1R functioning by measuring the A1R-mediated regulation of glutamate uptake into hippocampal slices and on behavioral responses in the open field and hot plate tests. NMDA preconditioning increased glutamate uptake into hippocampal slices without altering the expression of glutamate transporter GLT-1. Interestingly, NMDA preconditioning also induced antinociception in the hot plate test and both effects were reversed by post-activation of A1R with the agonist CCPA (0.2mg/kg, i.p.). NMDA preconditioning or A1R modulation did not alter locomotor activity in the open field. Overall, the results described herein provide new evidence that post-activation of A1R modulates NMDA preconditioning-mediated responses, pointing to the importance of the cross-talk between glutamatergic and adenosinergic systems to neuroprotection.

  6. DMPD: Toll-like receptors, Notch ligands, and cytokines drive the chronicity of lunginflammation. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 18073395 Toll-like receptors, Notch ligands, and cytokines drive the chronicity of lunginflammation. Raymond...ors, Notch ligands, and cytokines drive the chronicity of lunginflammation. Authors Raymond T, Schaller M, H

  7. Adenosine A(2A receptors measured with [C]TMSX PET in the striata of Parkinson's disease patients.

    Directory of Open Access Journals (Sweden)

    Masahiro Mishina

    Full Text Available Adenosine A(2A receptors (A2ARs are thought to interact negatively with the dopamine D(2 receptor (D2R, so selective A2AR antagonists have attracted attention as novel treatments for Parkinson's disease (PD. However, no information about the receptor in living patients with PD is available. The purpose of this study was to investigate the relationship between A2ARs and the dopaminergic system in the striata of drug-naïve PD patients and PD patients with dyskinesia, and alteration of these receptors after antiparkinsonian therapy. We measured binding ability of striatal A2ARs using positron emission tomography (PET with [7-methyl-(11C]-(E-8-(3,4,5-trimethoxystyryl-1,3,7-trimethylxanthine ([(11C]TMSX in nine drug-naïve patients with PD, seven PD patients with mild dyskinesia and six elderly control subjects using PET. The patients and eight normal control subjects were also examined for binding ability of dopamine transporters and D2Rs. Seven of the drug-naïve patients underwent a second series of PET scans following therapy. We found that the distribution volume ratio of A2ARs in the putamen were larger in the dyskinesic patients than in the control subjects (p<0.05, Tukey-Kramer post hoc test. In the drug-naïve patients, the binding ability of the A2ARs in the putamen, but not in the head of caudate nucleus, was significantly lower on the more affected side than on the less affected side (p<0.05, paired t-test. In addition, the A2ARs were significantly increased after antiparkinsonian therapy in the bilateral putamen of the drug-naïve patients (p<0.05, paired t-test but not in the bilateral head of caudate nucleus. Our study demonstrated that the A2ARs in the putamen were increased in the PD patients with dyskinesia, and also suggest that the A2ARs in the putamen compensate for the asymmetrical decrease of dopamine in drug-naïve PD patients and that antiparkinsonian therapy increases the A2ARs in the putamen. The A2ARs may play an

  8. Up-regulation of striatal adenosine A(2A) receptors with iron deficiency in rats: effects on locomotion and cortico-striatal neurotransmission.

    Science.gov (United States)

    Quiroz, César; Pearson, Virginia; Gulyani, Seema; Allen, Richard; Earley, Christopher; Ferré, Sergi

    2010-07-01

    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 A(2A) receptors are localized in glutamatergic terminals contacting GABAergic dynorphinergic neurons and their function can be analyzed by the ability of A(2A) receptor antagonists to block the motor output induced by cortical electrical stimulation. Postsynaptic A(2A) receptors are localized in the dendritic field of GABAergic enkephalinergic neurons and their function can be analyzed by studying the ability of A(2A) receptor antagonists to produce locomotor activity and to counteract striatal ERK1/2 phosphorylation induced by cortical electrical stimulation. Increased density of striatal A(2A) receptors was found in rats fed during 3 weeks with an iron-deficient diet during the post-weaning period. In iron-deficient rats, the selective A(2A) receptor antagonist MSX-3, at doses of 1 and 3 mg/kg, was more effective at blocking motor output induced by cortical electrical stimulation (presynaptic A(2A) receptor-mediated effect) and at enhancing locomotor activation and blocking striatal ERK phosphorylation induced by cortical electrical stimulation (postsynaptic A(2A) receptor-mediated effects). These results indicate that brain iron deficiency induces a functional up-regulation of both striatal pre- and postsynaptic A(2A) receptor, which could be involved in sensory-motor disorders associated with iron deficiency such as RLS.

  9. Changing the insulin receptor to possess insulin-like growth factor I ligand specificity

    International Nuclear Information System (INIS)

    To examine the role of the N-terminal part of the insulin-like growth factor I (IGF-I) receptor and insulin receptor in determining ligand specificity, the authors prepared an expression vector encoding a hybrid receptor where exon 1 (encoding the signal peptide and seven amino acids of the α-subunit), exon 2, and exon 3 of the insulin receptor were replaced with the corresponding IGF-I receptor cDNA (938 nucleotides). To allow direct quantitative comparison of the binding capabilities of this hybrid receptor with those of the human IGF-I receptor and the insulin receptor, all three receptors were expressed in baby hamster kidney (BHK) cells as soluble molecules and partially purified before characterization. The hybrid IGF-I/insulin receptor bound IGF-I with an affinity comparable to that of the wild-type IGF-I receptor. In contrast, the hybrid receptor no longer displayed high-affinity binding of insulin. These results directly demonstrate that it is possible to change the specificity of the insulin receptor to that of the IGF-I receptor and, furthermore, that the binding specificity for IGF-I is encoded within the nucleotide sequence from 135 to 938 of the IGF-I receptor cDNA. Since the hybrid receptor only bound insulin with low affinity, the insulin binding region is likely to be located within exons 2 and 3 of the insulin receptor

  10. Changing the insulin receptor to possess insulin-like growth factor I ligand specificity

    Energy Technology Data Exchange (ETDEWEB)

    Andersen, A.S.; Kjeldsen, T.; Wiberg, F.C.; Christensen, P.M.; Rasmussen, J.S.; Norris, K.; Moeller, K.B.; Moeller, N.P.H. (Biopharmaceuticals Div., Bagsvaerd (Denmark))

    1990-08-14

    To examine the role of the N-terminal part of the insulin-like growth factor I (IGF-I) receptor and insulin receptor in determining ligand specificity, the authors prepared an expression vector encoding a hybrid receptor where exon 1 (encoding the signal peptide and seven amino acids of the {alpha}-subunit), exon 2, and exon 3 of the insulin receptor were replaced with the corresponding IGF-I receptor cDNA (938 nucleotides). To allow direct quantitative comparison of the binding capabilities of this hybrid receptor with those of the human IGF-I receptor and the insulin receptor, all three receptors were expressed in baby hamster kidney (BHK) cells as soluble molecules and partially purified before characterization. The hybrid IGF-I/insulin receptor bound IGF-I with an affinity comparable to that of the wild-type IGF-I receptor. In contrast, the hybrid receptor no longer displayed high-affinity binding of insulin. These results directly demonstrate that it is possible to change the specificity of the insulin receptor to that of the IGF-I receptor and, furthermore, that the binding specificity for IGF-I is encoded within the nucleotide sequence from 135 to 938 of the IGF-I receptor cDNA. Since the hybrid receptor only bound insulin with low affinity, the insulin binding region is likely to be located within exons 2 and 3 of the insulin receptor.

  11. The Different Ligand-Binding Modes of Relaxin Family Peptide Receptors RXFP1 and RXFP2

    OpenAIRE

    Scott, Daniel J.; Rosengren, K. Johan; Bathgate, Ross A. D.

    2012-01-01

    Relaxin and insulin-like peptide 3 (INSL3) are peptide hormones with a number of important physiological roles in reproduction, regulation of extracellular matrix turnover, and cardiovascular function. Relaxin and INSL3 mediate their actions through the closely related G-protein coupled receptors, relaxin family peptide receptors 1 and 2 (RXFP1 and RXFP2), respectively. These receptors have large extracellular domains (ECD) that contain high-affinity ligand-binding sites within their 10 leuci...

  12. Characterization of the ligand-dependent transactivation domain of thyroid hormone receptor.

    OpenAIRE

    Barettino, D; Vivanco Ruiz, M M; Stunnenberg, H.G.

    1994-01-01

    Transcriptional activation by nuclear receptors is achieved through autonomous activation functions (AFs), a constitutive N-terminal AF-1 and a C-terminal, ligand-dependent AF-2 that comprises a motif conserved between nuclear receptors. We have performed an extensive mutational analysis of the putative AF-2 domain of chicken thyroid hormone receptor alpha (cT3R alpha). We show that the AF-2 region mediates transactivation as well as transcriptional interference (squelching), not only between...

  13. Melanocortin-1 receptor-mediated signalling pathways activated by NDP-MSH and HBD3 ligands

    OpenAIRE

    Beaumont, Kimberley A.; Smit, Darren J.; Liu, Yan Yan; Chai, Eric; Patel, Mira P.; Millhauser, Glenn L.; Smith, Jennifer J.; Alewood, Paul F.; Sturm, Richard A.

    2012-01-01

    Binding of melanocortin peptide agonists to the melanocortin-1 receptor of melanocytes results in eumelanin production, whereas binding of the agouti signalling protein inverse agonist results in pheomelanin synthesis. Recently, a novel melanocortin-1 receptor ligand was reported. A β-defensin gene mutation was found to beresponsible for black coat colour in domestic dogs. Notably, the human equivalent, β-defensin 3, was found to bind with high affinity to the melanocortin-1 receptor; however...

  14. Bombesin family receptor and ligand gene expression in human colorectal cancer and normal mucosa

    OpenAIRE

    Chave, H S; Gough, A C; Palmer, K.; Preston, S. R.; Primrose, J N

    1999-01-01

    Bombesin-like peptides and their receptors are widely distributed throughout the gut and are potential mitogens for a number of gastrointestinal (GI) cancers. We have analysed the expression of bombesin-like peptides and their receptor subtypes in normal and neoplastic colorectal tissue. Expression was analysed by reverse transcription polymerase chain reaction (RT-PCR) using receptor and ligand subtype-specific primers and then expression localized by in situ hybridization (ISH) with ribopro...

  15. Synergistic antidepressant-like effect of the joint administration of caffeine and NMDA receptor ligands in the forced swim test in mice.

    Science.gov (United States)

    Serefko, Anna; Szopa, Aleksandra; Wlaź, Aleksandra; Wośko, Sylwia; Wlaź, Piotr; Poleszak, Ewa

    2016-04-01

    The optimal treatment of depressed patients remains one of the most important challenges concerning depression. The identification of the best treatment strategies and development of new, safer, and more effective agents are crucial. The glutamatergic system seems to be a promising drug target, and consequently the use of the NMDA receptor ligands, particularly in co-administration with other substances exerting the antidepressant activity, has emerged among the new ideas. The objective of this study was to examine the effect of caffeine on the performance of mice treated with various NMDA modulators in the forced swim test. We demonstrated a significant interaction between caffeine (5 mg/kg) and the following NMDA receptor ligands: MK-801 (an antagonist binding in the ion channel, 0.05 mg/kg), CGP 37849 (an antagonist of the glutamate site, 0.312 mg/kg), L-701,324 (an antagonist of the glycine site, 1 mg/kg), and D-cycloserine (a high-efficacy partial agonist of the glycine site, 2.5 mg/kg), while the interaction between caffeine and the inorganic modulators, i.e., Zn(2+) (2.5 mg/kg) and Mg(2+) (10 mg/kg), was not considered as significant. Based on the obtained results, the simultaneous blockage of the adenosine and NMDA receptors may be a promising target in the development of new antidepressants.

  16. Preclinical tools in PET-tracer development : automatisation and biopharmaceutical evaluation with special emphasis on the adenosine A3 receptor

    International Nuclear Information System (INIS)

    Positron Emission Tomography (PET) is the first choice technology for the visualization and quantification of receptors and transporters, enabling examination of e.g. neurological, psychiatric and oncological diseases on a molecular level. Therefore, new and innovative PET-radiopharmaceuticals need to be developed to get further insights into the biochemical mechanisms involved in pathological changes. PET-tracer development starts with the idea or modelling of the chemical structure of a (new) molecule with (hopefully) good binding characteristics to the desired target site. As next steps, the compound needs to be synthesized and radiolabelled with a suitable PET-nuclide. Then it has to be evaluated regarding its parameters in various preclinical experimental settings. Hence, two major tools are crucial in the development-process of new PET-tracers: 1) a fast and reliable production method, most desirable and optimal in an automated set-up, and 2) proof of tracer suitability (high affinity, high selectivity and specificity, beside low unspecific binding) through preclinical evaluation in an animal model, prior to human application. Both aspects, the radiochemical preparation and automatisation, as well as the biopharmaceutical evaluation are presented in the thesis in 5 different manuscripts. In detail, the development and preclinical evaluation of 4 different PET-tracers ([11C]DASB, [18F]FE SUPPY, [18F]FE SUPPY:2, and [18F]FE CIT) for 3 targets, the serotonin transporter (SERT), the adenosine A3 receptor (A3R) and the dopamine transporter (DAT), respectively, are covered in the present thesis. The first manuscript presents a method for a fast, reliable and fully-automated radiosynthesis of [11C]DASB (a tracer for the imaging of the SERT in human brain in e.g. depression patients) will facilitate further clinical investigations (e.g. for the department of psychiatry and psychotherapy of the medical university of Vienna) with this tracer. [18F]FE SUPPY was

  17. Adenosine A1 receptor-mediated inhibition of in vitro prolactin secretion from the rat anterior pituitary

    Directory of Open Access Journals (Sweden)

    D.L.W. Picanço-Diniz

    2006-11-01

    Full Text Available In previous studies, we demonstrated biphasic purinergic effects on prolactin (PRL secretion stimulated by an adenosine A2 agonist. In the present study, we investigated the role of the activation of adenosine A1 receptors by (R-N6-(2-phenylisopropyladenosine (R-PIA at the pituitary level in in vitro PRL secretion. Hemipituitaries (one per cuvette in five replicates from adult male rats were incubated. Administration of R-PIA (0.001, 0.01, 0.1, 1, and 10 µM induced a reduction of PRL secretion into the medium in a U-shaped dose-response curve. The maximal reduction was obtained with 0.1 µM R-PIA (mean ± SEM, 36.01 ± 5.53 ng/mg tissue weight (t.w. treatment compared to control (264.56 ± 15.46 ng/mg t.w.. R-PIA inhibition (0.01 µM = 141.97 ± 15.79 vs control = 244.77 ± 13.79 ng/mg t.w. of PRL release was blocked by 1 µM cyclopentyltheophylline, a specific A1 receptor antagonist (1 µM = 212.360 ± 26.560 ng/mg t.w., whereas cyclopentyltheophylline alone (0.01, 0.1, 1 µM had no effect. R-PIA (0.001, 0.01, 0.1, 1 µM produced inhibition of PRL secretion stimulated by both phospholipase C (0.5 IU/mL; 977.44 ± 76.17 ng/mg t.w. and dibutyryl cAMP (1 mM; 415.93 ± 37.66 ng/mg t.w. with nadir established at the dose of 0.1 µM (225.55 ± 71.42 and 201.9 ± 19.08 ng/mg t.w., respectively. Similarly, R-PIA (0.01 µM decreased (242.00 ± 24.00 ng/mg t.w. the PRL secretion stimulated by cholera toxin (0.5 mg/mL; 1050.00 ± 70.00 ng/mg t.w.. In contrast, R-PIA had no effect (468.00 ± 34.00 ng/mg t.w. on PRL secretion stimulation by pertussis toxin (0.5 mg/mL; 430.00 ± 26.00 ng/mg t.w.. These results suggest that inhibition of PRL secretion after A1 receptor activation by R-PIA is mediated by a Gi protein-dependent mechanism.

  18. Engineering and optimization of an allosteric biosensor protein for peroxisome proliferator-activated receptor γ ligands.

    Science.gov (United States)

    Li, Jingjing; Gierach, Izabela; Gillies, Alison R; Warden, Charles D; Wood, David W

    2011-11-15

    The peroxisome proliferator-activated receptor gamma (PPARγ or PPARG) belongs to the nuclear receptor superfamily, and is a potential drug target for a variety of diseases. In this work, we constructed a series of bacterial biosensors for the identification of functional PPARγ ligands. These sensors entail modified Escherichia coli cells carrying a four-domain fusion protein, comprised of the PPARγ ligand binding domain (LBD), an engineered mini-intein domain, the E. coli maltose binding protein (MBD), and a thymidylate synthase (TS) reporter enzyme. E. coli cells expressing this protein exhibit hormone ligand-dependent growth phenotypes. Unlike our published estrogen (ER) and thyroid receptor (TR) biosensors, the canonical PPARγ biosensor cells displayed pronounced growth in the absence of ligand. They were able to distinguish agonists and antagonists, however, even in the absence of agonist. To improve ligand sensitivity of this sensor, we attempted to engineer and optimize linker peptides flanking the PPARγ LBD insertion point. Truncation of the original linkers led to decreased basal growth and significantly enhanced ligand sensitivity of the PPARγ sensor, while substitution of the native linkers with optimized G(4)S (Gly-Gly-Gly-Gly-Ser) linkers further increased the sensitivity. Our studies demonstrate that the properties of linkers, especially the C-terminal linker, greatly influence the efficiency and fidelity of the allosteric signal induced by ligand binding. Our work also suggests an approach to increase allosteric behavior in this multidomain sensor protein, without modification of the functional LBD. PMID:21893405

  19. Theoretical investigation of interaction between the set of ligands and α7 nicotinic acetylcholine receptor

    Science.gov (United States)

    Glukhova, O. E.; Prytkova, T. R.; Shmygin, D. S.

    2016-03-01

    Nicotinic acetylcholine receptors (nAChRs) are neuron receptor proteins that provide a transmission of nerve impulse through the synapses. They are composed of a pentametric assembly of five homologous subunits (5 α7 subunits for α7nAChR, for example), oriented around the central pore. These receptors might be found in the chemical synapses of central and peripheral nervous system, and also in the neuromuscular synapses. Transmembrane domain of the one of such receptors constitutes ion channel. The conductive properties of ion channel strongly depend on the receptor conformation changes in the response of binding with some molecule, f.e. acetylcholine. Investigation of interaction between ligands and acetylcholine receptor is important for drug design. In this work we investigate theoretically the interaction between the set of different ligands (such as vanillin, thymoquinone, etc.) and the nicotinic acetylcholine receptor (primarily with subunit of the α7nAChR) by different methods and packages (AutodockVina, GROMACS, KVAZAR, HARLEM, VMD). We calculate interaction energy between different ligands in the subunit using molecular dynamics. On the base of obtained calculation results and using molecular docking we found an optimal location of different ligands in the subunit.

  20. Why do receptor-ligand bonds in cell adhesion cluster into discrete focal-adhesion sites?

    Science.gov (United States)

    Gao, Zhiwen; Gao, Yanfei

    2016-10-01

    Cell adhesion often exhibits the clustering of the receptor-ligand bonds into discrete focal-adhesion sites near the contact edge, thus resembling a rosette shape or a contracting membrane anchored by a small number of peripheral forces. The ligands on the extracellular matrix are immobile, and the receptors in the cell plasma membrane consist of two types: high-affinity integrins (that bond to the substrate ligands and are immobile) and low-affinity integrins (that are mobile and not bonded to the ligands). Thus the adhesion energy density is proportional to the high-affinity integrin density. This paper provides a mechanistic explanation for the clustering/assembling of the receptor-ligand bonds from two main points: (1) the cellular contractile force leads to the density evolution of these two types of integrins, and results into a large high-affinity integrin density near the contact edge and (2) the front of a propagating crack into a decreasing toughness field will be unstable and wavy. From this fracture mechanics perspective, the chemomechanical equilibrium is reached when a small number of patches with large receptor-ligand bond density are anticipated to form at the cell periphery, as opposed to a uniform distribution of bonds on the entire interface. Cohesive fracture simulations show that the de-adhesion force can be significantly enhanced by this nonuniform bond density field, but the de-adhesion force anisotropy due to the substrate elastic anisotropy is significantly reduced.

  1. 3,5-T2 is an alternative ligand for the thyroid hormone receptor β1.

    Science.gov (United States)

    Mendoza, A; Navarrete-Ramírez, P; Hernández-Puga, G; Villalobos, P; Holzer, G; Renaud, J P; Laudet, V; Orozco, A

    2013-08-01

    Several liganded nuclear receptors have alternative ligands acting in a tissue-specific fashion and playing important biological roles. We present evidence that 3,5-diiodothyronine (T(2)), a naturally occurring iodothyronine that results from T(3) outer-ring deiodination, is an alternative ligand for thyroid hormone receptor β1 (TRβ1). In tilapia, 2 TRβ isoforms differing by 9 amino acids in the ligand-binding domain were cloned. Binding and transactivation studies showed that T(2) activates the human and the long tilapia TRβ1 isoform, but not the short one. A chimeric human TRβ1 (hTRβ1) that contained the 9-amino-acid insert showed no response to T(2), suggesting that the conformation of the hTRβ1 naturally allows T(2) binding and that other regions of the receptor are implicated in TR activation by T(2). Indeed, further analysis showed that the N terminus is essential for T(2)-mediated transactivation but not for that by T(3) in the long and hTRβ1, suggesting a functional interaction between the N-terminal domain and the insertion in the ligand-binding domain. To establish the functional relevance of T(2)-mediated TRβ1 binding and activation, mRNA expression and its regulation by T(2) and T(3) was evaluated for both isoforms. Our data show that long TRβ1expression is 10(6)-fold higher than that of the short isoform, and T(3) and T(2) differentially regulate the expression of these 2 TRβ1 isoforms in vivo. Taken together, our results prompted a reevaluation of the role and mechanism of action of thyroid hormone metabolites previously believed to be inactive. More generally, we propose that classical liganded receptors are only partially locked to very specific ligands and that alternative ligands may play a role in the tissue-specific action of receptors. PMID:23736295

  2. Discriminating agonist and antagonist ligands of the nuclear receptors using 3D-pharmacophores.

    Science.gov (United States)

    Lagarde, Nathalie; Delahaye, Solenne; Zagury, Jean-François; Montes, Matthieu

    2016-01-01

    Nuclear receptors (NRs) constitute an important class of therapeutic targets. We evaluated the performance of 3D structure-based and ligand-based pharmacophore models in predicting the pharmacological profile of NRs ligands using the NRLiSt BDB database. We could generate selective pharmacophores for agonist and antagonist ligands and we found that the best performances were obtained by combining the structure-based and the ligand-based approaches. The combination of pharmacophores that were generated allowed to cover most of the chemical space of the NRLiSt BDB datasets. By screening the whole NRLiSt BDB on our 3D pharmacophores, we demonstrated their selectivity towards their dedicated NRs ligands. The 3D pharmacophores herein presented can thus be used as a predictor of the pharmacological activity of NRs ligands.Graphical AbstractUsing a combination of structure-based and ligand-based pharmacophores, agonist and antagonist ligands of the Nuclear Receptors included in the NRLiSt BDB database could be separated.

  3. Cell receptor and surface ligand density effects on dynamic states of adhering circulating tumor cells.

    Science.gov (United States)

    Zheng, Xiangjun; Cheung, Luthur Siu-Lun; Schroeder, Joyce A; Jiang, Linan; Zohar, Yitshak

    2011-10-21

    Dynamic states of cancer cells moving under shear flow in an antibody-functionalized microchannel are investigated experimentally and theoretically. The cell motion is analyzed with the aid of a simplified physical model featuring a receptor-coated rigid sphere moving above a solid surface with immobilized ligands. The motion of the sphere is described by the Langevin equation accounting for the hydrodynamic loadings, gravitational force, receptor-ligand bindings, and thermal fluctuations; the receptor-ligand bonds are modeled as linear springs. Depending on the applied shear flow rate, three dynamic states of cell motion have been identified: (i) free motion, (ii) rolling adhesion, and (iii) firm adhesion. Of particular interest is the fraction of captured circulating tumor cells, defined as the capture ratio, via specific receptor-ligand bonds. The cell capture ratio decreases with increasing shear flow rate with a characteristic rate. Based on both experimental and theoretical results, the characteristic flow rate increases monotonically with increasing either cell-receptor or surface-ligand density within certain ranges. Utilizing it as a scaling parameter, flow-rate dependent capture ratios for various cell-surface combinations collapse onto a single curve described by an exponential formula.

  4. A mollusk retinoic acid receptor (RAR) ortholog sheds light on the evolution of ligand binding.

    Science.gov (United States)

    Gutierrez-Mazariegos, Juliana; Nadendla, Eswar Kumar; Lima, Daniela; Pierzchalski, Keely; Jones, Jace W; Kane, Maureen; Nishikawa, Jun-Ichi; Hiromori, Youhei; Nakanishi, Tsuyoshi; Santos, Miguel M; Castro, L Filipe C; Bourguet, William; Schubert, Michael; Laudet, Vincent

    2014-11-01

    Nuclear receptors are transcription factors that regulate networks of target genes in response to small molecules. There is a strong bias in our knowledge of these receptors because they were mainly characterized in classical model organisms, mostly vertebrates. Therefore, the evolutionary origins of specific ligand-receptor couples still remain elusive. Here we present the identification and characterization of a retinoic acid receptor (RAR) from the mollusk Nucella lapillus (NlRAR). We show that this receptor specifically binds to DNA response elements organized in direct repeats as a heterodimer with retinoid X receptor. Surprisingly, we also find that NlRAR does not bind all-trans retinoic acid or any other retinoid we tested. Furthermore, NlRAR is unable to activate the transcription of reporter genes in response to stimulation by retinoids and to recruit coactivators in the presence of these compounds. Three-dimensional modeling of the ligand-binding domain of NlRAR reveals an overall structure that is similar to vertebrate RARs. However, in the ligand-binding pocket (LBP) of the mollusk receptor, the alteration of several residues interacting with the ligand has apparently led to an overall decrease in the strength of the interaction with the ligand. Accordingly, mutations of NlRAR at key positions within the LBP generate receptors that are responsive to retinoids. Altogether our data suggest that, in mollusks, RAR has lost its affinity for all-trans retinoic acid, highlighting the evolutionary plasticity of its LBP. When put in an evolutionary context, our results reveal new structural and functional features of nuclear receptors validated by millions of years of evolution that were impossible to reveal in model organisms. PMID:25116705

  5. Adenosine A{sub 2A} receptor imaging with [{sup 11}C]KF18446 PET in the rat brain after quinolinic acid lesion. Comparison with the dopamine receptor imaging

    Energy Technology Data Exchange (ETDEWEB)

    Ishiwata, Kiichi; Ogi, Nobuo; Hayakawa, Nobutaka [Tokyo Metropolitan Inst. of Gerontology, Tokyo (Japan). Positron Medical Center] [and others

    2002-11-01

    We proposed [{sup 11}C]KF18446 as a selective radioligand for mapping the adenosine A{sub 2A} receptors being highly enriched in the striatum by positron emission tomography (PET). In the present study, we investigated whether [{sup 11}C]KF18446 PET can detect the change in the striatal adenosine A{sub 2A} receptors in the rat after unilateral injection of an excitotoxin quinolinic acid into the striatum, a Huntington's disease model, to demonstrate the usefulness of [{sup 11}C]KF18446. The extent of the striatal lesion was identified based on MRI, to which the PET was co-registered. The binding potential of [{sup 11}C]KF18446 significantly decreased in the quinolinic acid-lesioned striatum. The decrease was comparable to the decrease in the potential of [{sup 11}C] raclopride binding to dopamine D{sub 2} receptors in the lesioned striatum, but seemed to be larger than the decrease in the potential of [{sup 11}C]SCH23390 binding to dopamine D{sub 1} receptors. Ex vivo and in vitro autoradiography validated the PET signals. We concluded that [{sup 11}C]KF18446 PET can detect change in the adenosine A{sub 2A} receptors in the rat model, and will provide a new diagnostic tool for characterizing post-synaptic striatopallidal neurons in the stratum. (author)

  6. Metabotropic glutamate receptor ligands as potential therapeutics for addiction

    OpenAIRE

    Olive, M.F.

    2009-01-01

    There is now compelling evidence that the excitatory amino acid neurotransmitter glutamate plays a pivotal role in drug addiction and alcoholism. As a result, there has been increasing interest in developing glutamate-based therapies for the treatment of addictive disorders. Receptors for glutamate are primarily divided into two classes: ionotropic glutamate receptors (iGluRs) that mediate fast excitatory glutamate transmission, and metabotropic glutamate receptors (mGluRs), which are G-prote...

  7. Perspectives on cognitive domains, H3 receptor ligands and neurological disease.

    Science.gov (United States)

    Hancock, Arthur A; Fox, Gerard B

    2004-10-01

    Histamine H(3) receptor agonists and antagonists have been evaluated in numerous in vitro and in vivo animal models to better understand how H(3) receptors modulate neurotransmitter function in the central nervous system. Likewise, behavioural models have explored the hypothesis that changes in neurotransmitter release could enhance cognitive function in human diseases. This review examines the reported effects of H(3) receptor ligands and how they influence cognitive behaviour. These data are interpreted on the basis of different cognitive domains that are relevant to neuropsychiatric diseases. Because of the diversity of H(3) receptors, their function and their influence on neurotransmitter systems, considerable promise exists for H(3) ligands to treat diseases in which aspects of learning and memory are impaired. However, because of the complexities of the histaminergic system and H(3) receptors and the lack of clinical data so far, proof of principle for use in human disease remains to be established.

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

    OpenAIRE

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

    2004-01-01

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

  9. Modeling of cell adhesion and deformation mediated by receptor-ligand interactions.

    Science.gov (United States)

    Golestaneh, Amirreza F; Nadler, Ben

    2016-04-01

    The current work is devoted to studying adhesion and deformation of biological cells mediated by receptors and ligands in order to enhance the existing models. Due to the sufficient in-plane continuity and fluidity of the phospholipid molecules, an isotropic continuum fluid membrane is proposed for modeling the cell membrane. The developed constitutive model accounts for the influence of the presence of receptors on the deformation and adhesion of the cell membrane through the introduction of spontaneous area dilation. Motivated by physics, a nonlinear receptor-ligand binding force is introduced based on charge-induced dipole interaction. Diffusion of the receptors on the membrane is governed by the receptor-ligand interaction via Fick's Law and receptor-ligand interaction. The developed model is then applied to study the deformation and adhesion of a biological cell. The proposed model is used to study the role of the material, binding, spontaneous area dilation and environmental properties on the deformation and adhesion of the cell. PMID:26093646

  10. Adenosine A2A receptor deficiency up-regulates cystatin F expression in white matter lesions induced by chronic cerebral hypoperfusion.

    Science.gov (United States)

    Duan, Wei; Ran, Hong; Zhou, Zhujuan; He, Qifen; Zheng, Jian

    2012-01-01

    In previous studies, we have shown that the inactivation of the adenosine A2A receptor exacerbates chronic cerebral hypoperfusion-induced white matter lesions (WMLs) by enhancing neuroinflammatory responses. However, the molecular mechanism underlying the effect of the adenosine A2A receptor remains unknown. Recent studies have demonstrated that cystatin F, a potent endogenous cysteine protease inhibitor, is selectively expressed in immune cells in association with inflammatory demyelination in central nervous system diseases. To understand the expression of cystatin F and its potential role in the effect of A2A receptor on WMLs induced through chronic cerebral hypoperfusion, we investigated cystatin F expression in the WMLs of A2A receptor gene knockout mice, the littermate wild-type mice and wild-type mice treated daily with the A2A receptor agonist CGS21680 or both CGS21680 and A2A receptor antagonist SCH58261 after chronic cerebral hypoperfusion. The results of quantitative-PCR and western blot analysis revealed that cystatin F mRNA and protein expression were significantly up-regulated in the WMLs after chronic cerebral hypoperfusion. In addition, cystatin F expression in the corpus callosum was significantly increased in A2A receptor gene knockout mice and markedly decreased in mice treated with CGS21680 on both the mRNA and protein levels. Additionally, SCH58261 counteracted the attenuation of cystatin F expression produced by CGS21680 after chronic cerebral hypoperfusion. Moreover, double immunofluorescence staining revealed that cystatin F was co-localized with the activated microglia marker CD11b. In conclusion, the cystatin F expression in the activated microglia is closely associated with the effect of the A2A receptors, which may be related to the neuroinflammatory responses occurring during the pathological process.

  11. Adenosine A2A receptor deficiency up-regulates cystatin F expression in white matter lesions induced by chronic cerebral hypoperfusion.

    Directory of Open Access Journals (Sweden)

    Wei Duan

    Full Text Available In previous studies, we have shown that the inactivation of the adenosine A2A receptor exacerbates chronic cerebral hypoperfusion-induced white matter lesions (WMLs by enhancing neuroinflammatory responses. However, the molecular mechanism underlying the effect of the adenosine A2A receptor remains unknown. Recent studies have demonstrated that cystatin F, a potent endogenous cysteine protease inhibitor, is selectively expressed in immune cells in association with inflammatory demyelination in central nervous system diseases. To understand the expression of cystatin F and its potential role in the effect of A2A receptor on WMLs induced through chronic cerebral hypoperfusion, we investigated cystatin F expression in the WMLs of A2A receptor gene knockout mice, the littermate wild-type mice and wild-type mice treated daily with the A2A receptor agonist CGS21680 or both CGS21680 and A2A receptor antagonist SCH58261 after chronic cerebral hypoperfusion. The results of quantitative-PCR and western blot analysis revealed that cystatin F mRNA and protein expression were significantly up-regulated in the WMLs after chronic cerebral hypoperfusion. In addition, cystatin F expression in the corpus callosum was significantly increased in A2A receptor gene knockout mice and markedly decreased in mice treated with CGS21680 on both the mRNA and protein levels. Additionally, SCH58261 counteracted the attenuation of cystatin F expression produced by CGS21680 after chronic cerebral hypoperfusion. Moreover, double immunofluorescence staining revealed that cystatin F was co-localized with the activated microglia marker CD11b. In conclusion, the cystatin F expression in the activated microglia is closely associated with the effect of the A2A receptors, which may be related to the neuroinflammatory responses occurring during the pathological process.

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

    Directory of Open Access Journals (Sweden)

    Li Yuesheng

    2012-12-01

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

  13. Embryonic caffeine exposure acts via A1 adenosine receptors to alter adult cardiac function and DNA methylation in mice.

    Directory of Open Access Journals (Sweden)

    Daniela L Buscariollo

    Full Text Available Evidence indicates that disruption of normal prenatal development influences an individual's risk of developing obesity and cardiovascular disease as an adult. Thus, understanding how in utero exposure to chemical agents leads to increased susceptibility to adult diseases is a critical health related issue. Our aim was to determine whether adenosine A1 receptors (A1ARs mediate the long-term effects of in utero caffeine exposure on cardiac function and whether these long-term effects are the result of changes in DNA methylation patterns in adult hearts. Pregnant A1AR knockout mice were treated with caffeine (20 mg/kg or vehicle (0.09% NaCl i.p. at embryonic day 8.5. This caffeine treatment results in serum levels equivalent to the consumption of 2-4 cups of coffee in humans. After dams gave birth, offspring were examined at 8-10 weeks of age. A1AR+/+ offspring treated in utero with caffeine were 10% heavier than vehicle controls. Using echocardiography, we observed altered cardiac function and morphology in adult mice exposed to caffeine in utero. Caffeine treatment decreased cardiac output by 11% and increased left ventricular wall thickness by 29% during diastole. Using DNA methylation arrays, we identified altered DNA methylation patterns in A1AR+/+ caffeine treated hearts, including 7719 differentially methylated regions (DMRs within the genome and an overall decrease in DNA methylation of 26%. Analysis of genes associated with DMRs revealed that many are associated with cardiac hypertrophy. These data demonstrate that A1ARs mediate in utero caffeine effects on cardiac function and growth and that caffeine exposure leads to changes in DNA methylation.

  14. Regulation of epithelial sodium channel a-subunit expression by adenosine receptor A2a in alveolar epithelial cells

    Institute of Scientific and Technical Information of China (English)

    DENG Wang; WANG Dao-xin; ZHANG Wei; LI Chang-yi

    2011-01-01

    Background The amiloride-sensitive epithelial sodium channel a-subunit (a-ENaC) is an important factor for alveolar fluid clearance during acute lung injury. The relationship between adenosine receptor A2a (A2aAR) expressed in alveolar epithelial cells and aα-ENaC is poorly understood. We targeted the A2aAR in this study to investigate its role in the expression of αa-ENaC and in acute lung injury.Methods A549 cells were incubated with different concentrations of A2aAR agonist CGS-21680 and with 100 μmol/L CGS-21680 for various times. Rats were treated with lipopolysaccharide (LPS) after CGS-21680 was injected. Animals were sacrificed and tissue was harvested for evaluation of lung injury by analysis of the lung wet-to-dry weight ratio, lung permeability and myeloperoxidase activity. RT-PCR and Western blotting were used to determine the mRNA and protein expression levels of α-ENaC in A549 cells and alveolar type II epithelial cells.Results Both mRNA and protein levels of α-ENaC were markedly higher from 4 hours to 24 hours after exposure to 100μmol/L CGS-21680. There were significant changes from 0.1 umol/L to 100 μmol/L CGS-21680, with a positive correlation between increased concentrations of CGS-21680 and expression of α-ENaC. Treatment with CGS-21680during LPS induced lung injury protected the lung and promoted α-ENaC expression in the alveolar epithelial cells.Conclusion Activation of A2aAR has a protective effect during the lung injury, which may be beneficial to the prognosis of acute lung injury.

  15. Ligands of estrogen receptors α and β, method of their preparation, and pharmaceuticals comprising them

    OpenAIRE

    Novák, P.; Sedlák, D. (David); Bartůněk, P. (Petr); Kotora, M. (Martin)

    2012-01-01

    The invention relates to novel ligands of the estrogen receptors α and β of general formula II, which are useful as an active substance of pharmaceuticals, for example pharmaceutical compositions useful for hormone replacement therapy, as well as for the treatment of tumors and inflammatory diseases. The invention also relates to a novel preparation method of these ligands comprising cyclotrimerization of ethynylestradiol with the appropriate diyne in an organic solvent. Further, th...

  16. The glucocorticoid receptor hormone binding domain mediates transcriptional activation in vitro in the absence of ligand.

    OpenAIRE

    Schmitt, J.; Stunnenberg, H G

    1993-01-01

    We show that recombinant rat glucocorticoid receptor (vvGR) expressed using vaccinia virus is indistinguishable from authentic GR with respect to DNA and hormone binding. In the absence of hormone, vvGR is mainly found in the cytoplasm in a complex with heat shock protein 90. Upon incubation with ligand, vvGR is released from this complex and translocated to the nucleus. Thus, the ligand binding domain displays the known biochemical properties. However, in vitro, transcription from a syntheti...

  17. Regulation of dendritic cell differentiation and function by estrogen receptor ligands

    OpenAIRE

    Kovats, Susan; Carreras, Esther

    2008-01-01

    Estrogen receptor (ER) ligands can modulate innate and adaptive immunity and hematopoiesis, which may explain the clear sex differences in immune responses during autoimmunity, infection or trauma. Dendritic cells (DC) are antigen-presenting cells important for initiation of innate and adaptive immunity, as well as immune tolerance. DC progenitors and terminally differentiated DC express ER, indicating the ER ligands may regulate DC at multiple developmental and functional stages. Although th...

  18. Biophysical characterization of G-protein coupled receptor-peptide ligand binding

    OpenAIRE

    Langelaan, David N.; Ngweniform, Pascaline; Rainey, Jan K.

    2011-01-01

    G-protein coupled receptors (GPCRs) are ubiquitous membrane proteins allowing intracellular response to extracellular factors that range from photons of light to small molecules to proteins. Despite extensive exploitation of GRCRs as therapeutic targets, biophysical characterization of GPCR-ligand interactions remains challenging. In this minireview, we focus on techniques which have been successfully employed for structural and biophysical characterization of peptide ligands binding to their...

  19. Signal processing in the TGF-beta superfamily ligand-receptor network.

    Directory of Open Access Journals (Sweden)

    Jose M G Vilar

    2006-01-01

    Full Text Available The TGF-beta pathway plays a central role in tissue homeostasis and morphogenesis. It transduces a variety of extracellular signals into intracellular transcriptional responses that control a plethora of cellular processes, including cell growth, apoptosis, and differentiation. We use computational modeling to show that coupling of signaling with receptor trafficking results in a highly versatile signal-processing unit, able to sense by itself absolute levels of ligand, temporal changes in ligand concentration, and ratios of multiple ligands. This coupling controls whether the response of the receptor module is transient or permanent and whether or not different signaling channels behave independently of each other. Our computational approach unifies seemingly disparate experimental observations and suggests specific changes in receptor trafficking patterns that can lead to phenotypes that favor tumor progression.

  20. 3-Substituted phenylalanines as selective AMPA- and kainate receptor ligands

    DEFF Research Database (Denmark)

    Szymanska, Ewa; Pickering, Darryl S; Nielsen, Birgitte;

    2009-01-01

    On the basis of X-ray structures of ionotropic glutamate receptor constructs in complex with amino acid-based AMPA and kainate receptor antagonists, a series of rigid as well as flexible biaromatic alanine derivatives carrying selected hydrogen bond acceptors and donors have been synthesized in o...

  1. Photoaffinity ligand for dopamine D2 receptors: azidoclebopride

    Energy Technology Data Exchange (ETDEWEB)

    Niznik, H.B.; Guan, J.H.; Neumeyer, J.L.; Seeman, P.

    1985-02-01

    In order to label D2 dopamine receptors selectively and covalently by means of a photosensitive compound, azidoclebopride was synthesized directly from clebopride. The dissociation constant (KD) of clebopride for the D2 dopamine receptor (canine brain striatum) was 1.5 nM, while that for azidoclebopride was 21 nM. The affinities of both clebopride and azidoclebopride were markedly reduced in the absence of sodium chloride. In the presence of ultraviolet light, azidoclebopride inactivated D2 dopamine receptors irreversibly, as indicated by the inability of the receptors to bind (/sup 3/H)spiperone. Maximal photoinactivation of about 60% of the D2 dopamine receptors occurred at 1 microM azidoclebopride; 30% of the receptors were inactivated at 80 nM azidoclebopride (pseudo-IC50). Dopamine agonists selectively protected the D2 receptors from being inactivated by azidoclebopride, the order of potency being (-)-N-n-propylnorapomorphine greater than apomorphine greater than (+/-)-6,7-dihydroxy-2-aminotetralin greater than (+)-N-n-propylnorapomorphine greater than dopamine greater than noradrenaline greater than serotonin. Similarly, dopaminergic antagonists prevented the photoinactivation of D2 receptors by azidoclebopride with the following order of potency: spiperone greater than (+)-butaclamol greater than haloperidol greater than clebopride greater than (-)-sulpiride greater than (-)-butaclamol.

  2. Genomic organization, annotation, and ligand-receptor inferences of chicken chemokines and chemokine receptor genes based on comparative genomics

    Directory of Open Access Journals (Sweden)

    Sze Sing-Hoi

    2005-03-01

    Full Text Available Abstract Background Chemokines and their receptors play important roles in host defense, organogenesis, hematopoiesis, and neuronal communication. Forty-two chemokines and 19 cognate receptors have been found in the human genome. Prior to this report, only 11 chicken chemokines and 7 receptors had been reported. The objectives of this study were to systematically identify chicken chemokines and their cognate receptor genes in the chicken genome and to annotate these genes and ligand-receptor binding by a comparative genomics approach. Results Twenty-three chemokine and 14 chemokine receptor genes were identified in the chicken genome. All of the chicken chemokines contained a conserved CC, CXC, CX3C, or XC motif, whereas all the chemokine receptors had seven conserved transmembrane helices, four extracellular domains with a conserved cysteine, and a conserved DRYLAIV sequence in the second intracellular domain. The number of coding exons in these genes and the syntenies are highly conserved between human, mouse, and chicken although the amino acid sequence homologies are generally low between mammalian and chicken chemokines. Chicken genes were named with the systematic nomenclature used in humans and mice based on phylogeny, synteny, and sequence homology. Conclusion The independent nomenclature of chicken chemokines and chemokine receptors suggests that the chicken may have ligand-receptor pairings similar to mammals. All identified chicken chemokines and their cognate receptors were identified in the chicken genome except CCR9, whose ligand was not identified in this study. The organization of these genes suggests that there were a substantial number of these genes present before divergence between aves and mammals and more gene duplications of CC, CXC, CCR, and CXCR subfamilies in mammals than in aves after the divergence.

  3. Determinants governing ligand specificity of the Vibrio harveyi LuxN quorum-sensing receptor.

    Science.gov (United States)

    Ke, Xiaobo; Miller, Laura C; Bassler, Bonnie L

    2015-01-01

    Quorum sensing is a process of bacterial cell-cell communication that relies on the production, release and receptor-driven detection of extracellular signal molecules called autoinducers. The quorum-sensing bacterium Vibrio harveyi exclusively detects the autoinducer N-((R)-3-hydroxybutanoyl)-L-homoserine lactone (3OH-C4 HSL) via the two-component receptor LuxN. To discover the principles underlying the exquisite selectivity LuxN has for its ligand, we identified LuxN mutants with altered specificity. LuxN uses three mechanisms to verify that the bound molecule is the correct ligand: in the context of the overall ligand-binding site, His210 validates the C3 modification, Leu166 surveys the chain-length and a strong steady-state kinase bias imposes an energetic hurdle for inappropriate ligands to elicit signal transduction. Affinities for the LuxN kinase on and kinase off states underpin whether a ligand will act as an antagonist or an agonist. Mutations that bias LuxN to the agonized, kinase off, state are clustered in a region adjacent to the ligand-binding site, suggesting that this region acts as the switch that triggers signal transduction. Together, our analyses illuminate how a histidine sensor kinase differentiates between ligands and exploits those differences to regulate its signaling activity.

  4. Lipoteichoic acid induces unique inflammatory responses when compared to other toll-like receptor 2 ligands.

    Directory of Open Access Journals (Sweden)

    Elizabeth M Long

    Full Text Available Toll-like receptors (TLRs recognize evolutionarily-conserved molecular patterns originating from invading microbes. In this study, we were interested in determining if microbial ligands, which use distinct TLR2-containing receptor complexes, represent unique signals to the cell and can thereby stimulate unique cellular responses. Using the TLR2 ligands, R-FSL1, S-FSL1, Pam2CSK4, Pam3CSK4, and lipoteichoic acid (LTA, we demonstrate that these ligands activate NF-kappaB and MAP Kinase pathways with ligand-specific differential kinetics in murine macrophages. Most strikingly, LTA stimulation of these pathways was substantially delayed when compared with the other TLR2 ligands. These kinetics differences were associated with a delay in the LTA-induced expression of a subset of genes as compared with another TLR2 ligand, R-FSL1. However, this did not translate to overall differences in gene expression patterns four hours following stimulation with different TLR2 ligands. We extended this study to evaluate the in vivo responses to distinct TLR2 ligands using a murine model of acute inflammation, which employs intravital microscopy to monitor leukocyte recruitment into the cremaster muscle. We found that, although R-FSL1, S-FSL1, Pam2CSK4, and Pam3CSK4 were all able to stimulate robust leukocyte recruitment in vivo, LTA remained functionally inert in this in vivo model. Therefore distinct TLR2 ligands elicit unique cellular responses, as evidenced by differences in the kinetic profiles of signaling and gene expression responses in vitro, as well as the physiologically relevant differences in the in vivo responses to these ligands.

  5. DEPENDENCE OF PPAR LIGAND-INDUCED MAPK SIGNALING ON EPIDERMAL GROWTH FACTOR RECEPTOR TRANSACTIVATION HEPARIN-BINDING EGF CLEAVAGE MEDIATES ZINC-INDUCED EGF RECEPTOR PHOSPHORYLATION

    Science.gov (United States)

    Peroxisome proliferator-activated receptors (PPARs) are nuclear hormone receptors that function as ligand-activated transcription factors regulating lipid metabolism and homeostasis. In addition to their ability to regulate PPAR-mediated gene transcription, PPARalpha and gamma li...

  6. Adenosine A₂A receptors in striatal glutamatergic terminals and GABAergic neurons oppositely modulate psychostimulant action and DARPP-32 phosphorylation.

    Directory of Open Access Journals (Sweden)

    Hai-Ying Shen

    Full Text Available Adenosine A2A receptors (A2AR are located postsynaptically in striatopallidal GABAergic neurons, antagonizing dopamine D2 receptor functions, and are also located presynaptically at corticostriatal terminals, facilitating glutamate release. To address the hypothesis that these two A2AR populations differently control the action of psychostimulants, we characterized A2AR modulation of cocaine-induced effects at the level of DARPP-32 phosphorylation at Thr-34 and Thr-75, c-Fos expression, and psychomotor activity using two lines of cell-type selective A2AR knockout (KO mice with selective A2AR deletion in GABAergic neurons (striatum-A2AR-KO mice, or with A2AR deletion in both striatal GABAergic neurons and projecting cortical glutamatergic neurons (forebrain-A2AR-KO mice. We demonstrated that striatum-A2AR KO mice lacked A2ARs exclusively in striatal GABAergic terminals whereas forebrain-A2AR KO mice lacked A2ARs in both striatal GABAergic and glutamatergic terminals leading to a blunted A2AR-mediated facilitation of synaptosomal glutamate release. The inactivation of A2ARs in GABAergic neurons reduced striatal DARPP-32 phosphorylation at Thr-34 and increased its phosphorylation at Thr-75. Conversely, the additional deletion of corticostriatal glutamatergic A2ARs produced opposite effects on DARPP-32 phosphorylation at Thr-34 and Thr-75. This distinct modulation of DARPP-32 phosphorylation was associated with opposite responses to cocaine-induced striatal c-Fos expression and psychomotor activity in striatum-A2AR KO (enhanced and forebrain-A2AR KO mice (reduced. Thus, A2ARs in glutamatergic corticostriatal terminals and in GABAergic striatal neurons modulate the action of psychostimulants and DARPP-32 phosphorylation in opposite ways. We conclude that A2ARs in glutamatergic terminals prominently control the action of psychostimulants and define a novel mechanism by which A2ARs fine-tune striatal activity by integrating GABAergic, dopaminergic and

  7. New halogenated tris-(phenylalkyl)amines as h5-HT2B receptor ligands.

    Science.gov (United States)

    Kapadia, Nirav; Ahmed, Shahrear; Harding, Wayne W

    2016-07-15

    A series of compounds in which various halogen substituents were incorporated into a phenyl ring of a tris-(phenylalkyl)amine scaffold, was synthesized and evaluated for affinity to h5-HT2 receptors. In general, all compounds were found to have good affinity for the 5-HT2B receptor and were selective over 5-HT2A and 5-HT2C receptors. Compound 9i was the most selective compound in this study and is the highest affinity 5-HT2B receptor ligand bearing a tris-(phenylalkyl)amine scaffold to date. PMID:27261181

  8. Therapeutic Potential of 5-HT2C Receptor Ligands

    Directory of Open Access Journals (Sweden)

    Nanna H. Jensen

    2010-01-01

    Full Text Available Serotonin 2C receptors are G protein-coupled receptors expressed by GABAergic, glutamatergic, and dopaminergic neurons. Anatomically, they are present in various brain regions, including cortical areas, hippocampus, ventral midbrain, striatum, nucleus accumbens, hypothalamus, and amygdala. A large body of evidence supports a critical role of serotonin 2C receptors in mediating the interaction between serotonergic and dopaminergic systems, which is at the basis of their proposed involvement in the regulation of mood, affective behavior, and memory. In addition, their expression in specific neuronal populations in the hypothalamus would be critical for their role in the regulation of feeding behavior. Modulation of these receptors has therefore been proposed to be of interest in the search for novel pharmacological strategies for the treatment of various pathological conditions, including schizophrenia and mood disorders, as well as obesity. More precisely, blockade of serotonin 2C receptors has been suggested to provide antidepressant and anxiolytic benefit, while stimulation of these receptors may offer therapeutic benefit for the treatment of psychotic symptoms in schizophrenia and obesity. In addition, modulation of serotonin 2C receptors may offer cognitive-enhancing potential, albeit still a matter of debate. In the present review, the most compelling evidence from the literature is presented and tentative hypotheses with respect to existing controversies are outlined.

  9. The caffeine-binding adenosine A2A receptor induces age-like HPA-axis dysfunction by targeting glucocorticoid receptor function

    Science.gov (United States)

    Batalha, Vânia L.; Ferreira, Diana G.; Coelho, Joana E.; Valadas, Jorge S.; Gomes, Rui; Temido-Ferreira, Mariana; Shmidt, Tatiana; Baqi, Younis; Buée, Luc; Müller, Christa E.; Hamdane, Malika; Outeiro, Tiago F.; Bader, Michael; Meijsing, Sebastiaan H.; Sadri-Vakili, Ghazaleh; Blum, David; Lopes, Luísa V.

    2016-01-01

    Caffeine is associated with procognitive effects in humans by counteracting overactivation of the adenosine A2A receptor (A2AR), which is upregulated in the human forebrain of aged and Alzheimer’s disease (AD) patients. We have previously shown that an anti-A2AR therapy reverts age-like memory deficits, by reestablishment of the hypothalamic-pituitary-adrenal (HPA) axis feedback and corticosterone circadian levels. These observations suggest that A2AR over-activation and glucocorticoid dysfunction are key events in age-related hippocampal deficits; but their direct connection has never been explored. We now show that inducing A2AR overexpression in an aging-like profile is sufficient to trigger HPA-axis dysfunction, namely loss of plasmatic corticosterone circadian oscillation, and promotes reduction of GR hippocampal levels. The synaptic plasticity and memory deficits triggered by GR in the hippocampus are amplified by A2AR over-activation and were rescued by anti-A2AR therapy; finally, we demonstrate that A2AR act on GR nuclear translocation and GR-dependent transcriptional regulation. We provide the first demonstration that A2AR is a major regulator of GR function and that this functional interconnection may be a trigger to age-related memory deficits. This supports the idea that the procognitive effects of A2AR antagonists, namely caffeine, on Alzheimer’s and age-related cognitive impairments may rely on its ability to modulate GR actions. PMID:27510168

  10. The caffeine-binding adenosine A2A receptor induces age-like HPA-axis dysfunction by targeting glucocorticoid receptor function.

    Science.gov (United States)

    Batalha, Vânia L; Ferreira, Diana G; Coelho, Joana E; Valadas, Jorge S; Gomes, Rui; Temido-Ferreira, Mariana; Shmidt, Tatiana; Baqi, Younis; Buée, Luc; Müller, Christa E; Hamdane, Malika; Outeiro, Tiago F; Bader, Michael; Meijsing, Sebastiaan H; Sadri-Vakili, Ghazaleh; Blum, David; Lopes, Luísa V

    2016-01-01

    Caffeine is associated with procognitive effects in humans by counteracting overactivation of the adenosine A2A receptor (A2AR), which is upregulated in the human forebrain of aged and Alzheimer's disease (AD) patients. We have previously shown that an anti-A2AR therapy reverts age-like memory deficits, by reestablishment of the hypothalamic-pituitary-adrenal (HPA) axis feedback and corticosterone circadian levels. These observations suggest that A2AR over-activation and glucocorticoid dysfunction are key events in age-related hippocampal deficits; but their direct connection has never been explored. We now show that inducing A2AR overexpression in an aging-like profile is sufficient to trigger HPA-axis dysfunction, namely loss of plasmatic corticosterone circadian oscillation, and promotes reduction of GR hippocampal levels. The synaptic plasticity and memory deficits triggered by GR in the hippocampus are amplified by A2AR over-activation and were rescued by anti-A2AR therapy; finally, we demonstrate that A2AR act on GR nuclear translocation and GR-dependent transcriptional regulation. We provide the first demonstration that A2AR is a major regulator of GR function and that this functional interconnection may be a trigger to age-related memory deficits. This supports the idea that the procognitive effects of A2AR antagonists, namely caffeine, on Alzheimer's and age-related cognitive impairments may rely on its ability to modulate GR actions. PMID:27510168

  11. The caffeine-binding adenosine A2A receptor induces age-like HPA-axis dysfunction by targeting glucocorticoid receptor function.

    Science.gov (United States)

    Batalha, Vânia L; Ferreira, Diana G; Coelho, Joana E; Valadas, Jorge S; Gomes, Rui; Temido-Ferreira, Mariana; Shmidt, Tatiana; Baqi, Younis; Buée, Luc; Müller, Christa E; Hamdane, Malika; Outeiro, Tiago F; Bader, Michael; Meijsing, Sebastiaan H; Sadri-Vakili, Ghazaleh; Blum, David; Lopes, Luísa V

    2016-01-01

    Caffeine is associated with procognitive effects in humans by counteracting overactivation of the adenosine A2A receptor (A2AR), which is upregulated in the human forebrain of aged and Alzheimer's disease (AD) patients. We have previously shown that an anti-A2AR therapy reverts age-like memory deficits, by reestablishment of the hypothalamic-pituitary-adrenal (HPA) axis feedback and corticosterone circadian levels. These observations suggest that A2AR over-activation and glucocorticoid dysfunction are key events in age-related hippocampal deficits; but their direct connection has never been explored. We now show that inducing A2AR overexpression in an aging-like profile is sufficient to trigger HPA-axis dysfunction, namely loss of plasmatic corticosterone circadian oscillation, and promotes reduction of GR hippocampal levels. The synaptic plasticity and memory deficits triggered by GR in the hippocampus are amplified by A2AR over-activation and were rescued by anti-A2AR therapy; finally, we demonstrate that A2AR act on GR nuclear translocation and GR-dependent transcriptional regulation. We provide the first demonstration that A2AR is a major regulator of GR function and that this functional interconnection may be a trigger to age-related memory deficits. This supports the idea that the procognitive effects of A2AR antagonists, namely caffeine, on Alzheimer's and age-related cognitive impairments may rely on its ability to modulate GR actions.

  12. The Role of the Enterohepatic Circulation of Bile Salts and Nuclear Hormone Receptors in the Regulation of Cholesterol Homeostasis: Bile Salts as Ligands for Nuclear Hormone Receptors

    OpenAIRE

    Redinger, Richard N.

    2003-01-01

    The coordinated effect of lipid activated nuclear hormone receptors; liver X receptor (LXR), bound by oxysterol ligands and farnesoid X receptor (FXR), bound by bile acid ligands, act as genetic transcription factors to cause feed-forward cholesterol catabolism to bile acids and feedback repression of bile acid synthesis, respectively. It is the coordinated action of LXR and FXR, each dimerized to retinoid X receptor, that signal nuclear DNA response elements to encode proteins that prevent e...

  13. Heart Failure Therapeutics on the Basis of a Biased Ligand of the Angiotensin-2 Type 1 Receptor Rationale and Design of the BLAST-AHF Study (Biased Ligand of the Angiotensin Receptor Study in Acute Heart Failure)

    NARCIS (Netherlands)

    Felker, G. Michael; Butler, Javed; Collins, Sean P.; Cotter, Gad; Davison, Beth A.; Ezekowitz, Justin A.; Filippatos, Gerasimos; Levy, Phillip D.; Metra, Marco; Ponikowski, Piotr; Soergel, David G.; Teerlink, John R.; Violin, Jonathan D.; Voors, Adriaan A.; Pang, Peter S.

    2015-01-01

    The BLAST-AHF (Biased Ligand of the Angiotensin Receptor Study in Acute Heart Failure) study is designed to test the efficacy and safety of TRV027, a novel biased ligand of the angiotensin-2 type 1 receptor, in patients with acute heart failure (AHF). AHF remains a major public health problem, and n

  14. Dissecting the chemistry of nicotinic receptor-ligand interactions with infrared difference spectroscopy.

    Science.gov (United States)

    Ryan, Stephen E; Hill, Danny G; Baenziger, John E

    2002-03-22

    The physical interactions that occur between the nicotinic acetylcholine receptor from Torpedo and the agonists carbamylcholine and tetramethylamine have been studied using both conventional infrared difference spectroscopy and a novel double-ligand difference technique. The latter was developed to isolate vibrational bands from residues in a membrane receptor that interact with individual functional groups on a small molecule ligand. The binding of either agonist leads to an increase in vibrational intensity at frequencies centered near 1663, 1655, 1547, 1430, and 1059 cm(-1) indicating that both induce a conformational change from the resting to the desensitized state. Vibrational shifts near 1580, 1516, 1455, 1334, and between 1300 and 1400 cm(-1) are assigned to structural perturbations of tyrosine and possibly both tryptophan and charged carboxylic acid residues upon the formation of receptor-quaternary amine interactions, with the relatively intense feature near 1516 cm(-1) indicating a key role for tyrosine. Other vibrational bands suggest the involvement of additional side chains in agonist binding. Two side-chain vibrational shifts from 1668 and 1605 cm(-1) to 1690 and 1620 cm(-1), respectively, could reflect the formation of a hydrogen bond between the ester carbonyl of carbamylcholine and an arginine residue. The results demonstrate the potential of the double-ligand difference technique for dissecting the chemistry of membrane receptor-ligand interactions and provide new insight into the nature of nicotinic receptor-agonist interactions. PMID:11782459

  15. Divergent Label-free Cell Phenotypic Pharmacology of Ligands at the Overexpressed β2-Adrenergic Receptors

    Science.gov (United States)

    Ferrie, Ann M.; Sun, Haiyan; Zaytseva, Natalya; Fang, Ye

    2014-01-01

    We present subclone sensitive cell phenotypic pharmacology of ligands at the β2-adrenergic receptor (β2-AR) stably expressed in HEK-293 cells. The parental cell line was transfected with green fluorescent protein (GFP)-tagged β2-AR. Four stable subclones were established and used to profile a library of sixty-nine AR ligands. Dynamic mass redistribution (DMR) profiling resulted in a pharmacological activity map suggesting that HEK293 endogenously expresses functional Gi-coupled α2-AR and Gs-coupled β2-AR, and the label-free cell phenotypic activity of AR ligands are subclone dependent. Pathway deconvolution revealed that the DMR of epinephrine is originated mostly from the remodeling of actin microfilaments and adhesion complexes, to less extent from the microtubule networks and receptor trafficking, and certain agonists displayed different efficacy towards the cAMP-Epac pathway. We demonstrate that receptor signaling and ligand pharmacology is sensitive to the receptor expression level, and the organization of the receptor and its signaling circuitry.

  16. Identifying and quantifying two ligand-binding sites while imaging native human membrane receptors by AFM

    Science.gov (United States)

    Pfreundschuh, Moritz; Alsteens, David; Wieneke, Ralph; Zhang, Cheng; Coughlin, Shaun R.; Tampé, Robert; Kobilka, Brian K.; Müller, Daniel J.

    2015-11-01

    A current challenge in life sciences is to image cell membrane receptors while characterizing their specific interactions with various ligands. Addressing this issue has been hampered by the lack of suitable nanoscopic methods. Here we address this challenge and introduce multifunctional high-resolution atomic force microscopy (AFM) to image human protease-activated receptors (PAR1) in the functionally important lipid membrane and to simultaneously localize and quantify their binding to two different ligands. Therefore, we introduce the surface chemistry to bifunctionalize AFM tips with the native receptor-activating peptide and a tris-N-nitrilotriacetic acid (tris-NTA) group binding to a His10-tag engineered to PAR1. We further introduce ways to discern between the binding of both ligands to different receptor sites while imaging native PAR1s. Surface chemistry and nanoscopic method are applicable to a range of biological systems in vitro and in vivo and to concurrently detect and localize multiple ligand-binding sites at single receptor resolution.

  17. Development of novel mixed ligand technetium complexes for imaging 5-HT1A neural system receptors

    International Nuclear Information System (INIS)

    The development of 99mTc complexes for imaging 5-HT1A neural system receptors using the 3 + 1 mixed ligand approach is described. Six novel complexes (I-VI) were designed using two different strategies. In complexes I-IV the pharmacophore 1-(2-methoxyphenyl)piperazine was attached to a monodentate thiol used as co-ligand and combined with tridentate dianionic aminothiols (SNS and NNS). On the other hand, complexes V and VI were obtained using thiophenol and 4-methoxy-thiophenol as co-ligand and a tridentate ligand (SNS) with the pharmacophore bound to the nitrogen through an alkyl chain. All complexes were prepared at tracer level using 99mTc-glucoheptonate as precursor. Ligand and co-ligand concentration, reaction time and temperature were optimized to achieve high substitution yield and radiochemical purity. Structure was studied at carrier level through the corresponding rhenium complexes. Complexes I and II presented the expected ReOLK structure and a distorted trigonal bipyramidal geometry. The structure of the other four complexes has not been completely elucidated yet. Biodistribution studies of all the complexes demonstrated selective brain uptake and retention. Uptake of complex I in receptor-rich hippocampus was significantly higher than that of the cerebellum (P = 0.05) 1 h post-injection. Oxorhenium complexes I and II showed affinity for the 5-HT1A receptor binding sites, with IC50 values in the nanomolar range. The results demonstrate the potential of the mixed ligand approach for the design of 99mTc complexes with the ability to bind neuroreceptors. However, the goal of imaging 5-HT1A receptors with technetium requires further development of complexes with improved biological profiles. (author)

  18. Adenosine and sleep

    International Nuclear Information System (INIS)

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

  19. Adenosine and sleep

    Energy Technology Data Exchange (ETDEWEB)

    Yanik, G.M. Jr.

    1987-01-01

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

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

    Science.gov (United States)

    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.

  1. The A3 Adenosine Receptor Agonist CF502 Inhibits the PI3K, PKB/Akt and NF-κB Signaling Pathway in Synoviocytes from Rheumatoid Arthritis Patients and in Adjuvant Induced Arthritis Rats

    OpenAIRE

    Ochaion, A.; BAR-YEHUDA, S.; Cohen, S.; Amital, H; Jacobson, K. A.; Joshi, B.V.; Gao, Z. G.; Barer, F.; PATOKA, R.; Del Valle, L; Perez-Liz, G.; Fishman, P

    2008-01-01

    The A3 adenosine receptor (A3AR) is over-expressed in inflammatory cells and was defined as a target to combat inflammation. Synthetic agonists to this receptor, such as IB-MECA and Cl-IB-MECA, exert an anti-inflammatory effect in experimental animal models of adjuvant and collagen induced arthritis.

  2. Preliminary Molecular Dynamic Simulations of the Estrogen Receptor Alpha Ligand Binding Domain from Antagonist to Apo

    Directory of Open Access Journals (Sweden)

    Adrian E. Roitberg

    2008-06-01

    Full Text Available Estrogen receptors (ER are known as nuclear receptors. They exist in the cytoplasm of human cells and serves as a DNA binding transcription factor that regulates gene expression. However the estrogen receptor also has additional functions independent of DNA binding. The human estrogen receptor comes in two forms, alpha and beta. This work focuses on the alpha form of the estrogen receptor. The ERα is found in breast cancer cells, ovarian stroma cells, endometrium, and the hypothalamus. It has been suggested that exposure to DDE, a metabolite of DDT, and other pesticides causes conformational changes in the estrogen receptor. Before examining these factors, this work examines the protein unfolding from the antagonist form found in the 3ERT PDB crystal structure. The 3ERT PDB crystal structure has the estrogen receptor bound to the cancer drug 4-hydroxytamoxifen. The 4-hydroxytamoxifen ligand was extracted before the simulation, resulting in new conformational freedom due to absence of van der Waals contacts between the ligand and the receptor. The conformational changes that result expose the binding clef of the co peptide beside Helix 12 of the receptor forming an apo conformation. Two key conformations in the loops at either end of the H12 are produced resulting in the antagonist to apo conformation transformation. The results were produced over a 42ns Molecular Dynamics simulation using the AMBER FF99SB force field.

  3. Doubling the Size of the Glucocorticoid Receptor Ligand Binding Pocket by Deacylcortivazol

    Energy Technology Data Exchange (ETDEWEB)

    Suino-Powell, Kelly; Xu, Yong; Zhang, Chenghai; Tao, Yong-guang; Tolbert, W. David; Simons, Jr., S. Stoney; Xu, H. Eric (NIH)

    2010-03-08

    A common feature of nuclear receptor ligand binding domains (LBD) is a helical sandwich fold that nests a ligand binding pocket within the bottom half of the domain. Here we report that the ligand pocket of glucocorticoid receptor (GR) can be continuously extended into the top half of the LBD by binding to deacylcortivazol (DAC), an extremely potent glucocorticoid. It has been puzzling for decades why DAC, which contains a phenylpyrazole replacement at the conserved 3-ketone of steroid hormones that are normally required for activation of their cognate receptors, is a potent GR activator. The crystal structure of the GR LBD bound to DAC and the fourth LXXLL motif of steroid receptor coactivator 1 reveals that the GR ligand binding pocket is expanded to a size of 1,070 {angstrom}{sup 3}, effectively doubling the size of the GR dexamethasone-binding pocket of 540 {angstrom}{sup 3} and yet leaving the structure of the coactivator binding site intact. DAC occupies only {approx}50% of the space of the pocket but makes intricate interactions with the receptor around the phenylpyrazole group that accounts for the high-affinity binding of DAC. The dramatic expansion of the DAC-binding pocket thus highlights the conformational adaptability of GR to ligand binding. The new structure also allows docking of various nonsteroidal ligands that cannot be fitted into the previous structures, thus providing a new rational template for drug discovery of steroidal and nonsteroidal glucocorticoids that can be specifically designed to reach the unoccupied space of the expanded pocket.

  4. Non-Ligand-Induced Dimerization is Sufficient to Initiate the Signalling and Endocytosis of EGF Receptor

    OpenAIRE

    Kourouniotis, George; Wang, Yi; Pennock, Steven; Chen, Xinmei; Wang, Zhixiang

    2016-01-01

    The binding of epidermal growth factor (EGF) to EGF receptor (EGFR) stimulates cell mitogenesis and survival through various signalling cascades. EGF also stimulates rapid EGFR endocytosis and its eventual degradation in lysosomes. The immediate events induced by ligand binding include receptor dimerization, activation of intrinsic tyrosine kinase and autophosphorylation. However, in spite of intensified efforts, the results regarding the roles of these events in EGFR signalling and internali...

  5. Receptor activator of nuclear factor kappa B ligand and osteoprotegerin levels in gingival crevicular fluid

    OpenAIRE

    Fatemeh Sarlati; Mandana Sattari; Shilan Razzaghi; Malihe Nasiri

    2012-01-01

    Background: Osteoclastogenesis is coordinated by the interaction of three members of the tumor necrosis factor (TNF) superfamily: Osteoprotegerin (OPG)/receptor activator of nuclear factor kappa B ligand (RANKL)/receptor activator of nuclear factor kappa B (RANK). The aim of this study was to investigate RANKL and OPG levels, and their relative ratio in gingival crevicular fluid (GCF) of patients with chronic and aggressive periodontitis, as well as healthy controls. Materials and Methods:...

  6. The Role of Adenosine A1 and A2A Receptors in the Caffeine Effect on MDMA-Induced DA and 5-HT Release in the Mouse Striatum

    OpenAIRE

    Górska, A. M.; Gołembiowska, K.

    2014-01-01

    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 ro...

  7. DMPD: Nucleic acid-sensing Toll-like receptors: beyond ligand search. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 18321608 Nucleic acid-sensing Toll-like receptors: beyond ligand search. Miyake K. ...Adv Drug Deliv Rev. 2008 Apr 29;60(7):782-5. Epub 2008 Feb 15. (.png) (.svg) (.html) (.csml) Show Nucleic ac...id-sensing Toll-like receptors: beyond ligand search. PubmedID 18321608 Title Nucleic acid-sensing Toll-like

  8. NFkappaB Selectivity of Estrogen Receptor Ligands Revealed By Comparative Crystallographic Analyses

    Energy Technology Data Exchange (ETDEWEB)

    Nettles, K.W.; Bruning, J.B.; Gil, G.; Nowak, J.; Sharma, S.K.; Hahm, J.B.; Kulp, K.; Hochberg, R.B.; Zhou, H.; Katzenellenbogen, J.A.; Katzenllenbogen, B.S.; Kim, Y.; Joachmiak, A.; Greene, G.L.

    2009-05-22

    Our understanding of how steroid hormones regulate physiological functions has been significantly advanced by structural biology approaches. However, progress has been hampered by misfolding of the ligand binding domains in heterologous expression systems and by conformational flexibility that interferes with crystallization. Here, we show that protein folding problems that are common to steroid hormone receptors are circumvented by mutations that stabilize well-characterized conformations of the receptor. We use this approach to present the structure of an apo steroid receptor that reveals a ligand-accessible channel allowing soaking of preformed crystals. Furthermore, crystallization of different pharmacological classes of compounds allowed us to define the structural basis of NF{kappa}B-selective signaling through the estrogen receptor, thus revealing a unique conformation of the receptor that allows selective suppression of inflammatory gene expression. The ability to crystallize many receptor-ligand complexes with distinct pharmacophores allows one to define structural features of signaling specificity that would not be apparent in a single structure.

  9. Ligand-selective activation of heterologously-expressed mammalian olfactory receptor.

    Science.gov (United States)

    Ukhanov, K; Bobkov, Y; Corey, E A; Ache, B W

    2014-10-01

    Mammalian olfactory receptors (ORs) appear to have the capacity to couple to multiple G protein-coupled signaling pathways in a ligand-dependent selective manner. To better understand the mechanisms and molecular range of such ligand selectivity, we expressed the mouse eugenol OR (mOR-EG) in HEK293T cells together with Gα15 to monitor activation of the phospholipase-C (PLC) signaling pathway and/or Gαolf to monitor activation of the adenylate cyclase (AC) signaling pathway, resulting in intracellular Ca(2+) release and/or Ca(2+) influx through a cyclic nucleotide-gated channel, respectively. PLC-dependent responses differed dynamically from AC-dependent responses, allowing them to be distinguished when Gα15 and Gαolf were co-expressed. The dynamic difference in readout was independent of the receptor, the heterologous expression system, and the ligand concentration. Of 17 reported mOR-EG ligands tested, including eugenol, its analogs, and structurally dissimilar compounds (mousse cristal, nootkatone, orivone), some equally activated both signaling pathways, some differentially activated both signaling pathways, and some had no noticeable effect even at 1-5mM. Our findings argue that mOR-EG, when heterologously expressed, can couple to two different signaling pathways in a ligand selective manner. The challenge now is to determine the potential of mOR-EG, and perhaps other ORs, to activate multiple signaling pathways in a ligand selective manner in native ORNs. PMID:25149566

  10. Peroxisome proliferator-activated receptor γ ligands regulate neural stem cell proliferation and differentiation in vitro and in vivo

    OpenAIRE

    Morales-García, José A.; Luna Medina, Rosario de; Alfaro-Cervello, Clara; Cortés-Canteli, Marta; Santos, Ángel; García-Verdugo, J. M.; Pérez Castillo, Ana

    2011-01-01

    Peroxisome proliferator-activated receptor gamma (PPARγ) belongs to a family of ligand-activated nuclear receptors and its ligands are known to control many physiological and pathological situations. Its role in the central nervous system has been under intense analysis during the last years. Here we show a novel function for PPARγ in controlling stem cell expansion in the adult mammalian brain. Adult rats treated with pioglitazone, a specific ligand of PPARγ, had elevated numbers of prolifer...

  11. Imaging GABAc Receptors with Ligand-Conjugated Quantum Dots

    Directory of Open Access Journals (Sweden)

    Ian D. Tomlinson

    2007-01-01

    Full Text Available We report a methodology for labeling the GABAc receptor on the surface membrane of intact cells. This work builds upon our earlier work with serotonin-conjugated quantum dots and our studies with PEGylated quantum dots to reduce nonspecific binding. In the current approach, a PEGylated derivative of muscimol was synthesized and attached via an amide linkage to quantum dots coated in an amphiphilic polymer derivative of a modified polyacrylamide. These conjugates were used to image GABAC receptors heterologously expressed in Xenopus laevis oocytes.

  12. Progress in the discovery of selective, high affinity A(2B) adenosine receptor antagonists as clinical candidates.

    Science.gov (United States)

    Kalla, Rao V; Zablocki, Jeff

    2009-03-01

    The selective, high affinity A(2B) adenosine receptor (AdoR) antagonists that were synthesized by several research groups should aid in determining the role of the A(2B) AdoR in inflammatory diseases like asthma or rheumatoid arthritis (RA) and angiogenic diseases like diabetic retinopathy or cancer. CV Therapeutics scientists discovered the selective, high affinity A(2B) AdoR antagonist 10, a 8-(4-pyrazolyl)-xanthine derivative [CVT-6883, K(i)(hA(2B)) = 22 nM; K(i)(hA(1)) = 1,940 nM; K(i)(hA(2A)) = 3,280; and K(i)(hA(3)) = 1,070 nM] that has favorable pharmacokinetic (PK) properties (t (1/2) = 4 h and F > 35% rat). Compound 10 demonstrated functional antagonism at the A(2B) AdoR (K(B) = 6 nM) and efficacy in a mouse model of asthma. In two phase 1 clinical trials, CVT-6883 was found to be safe, well tolerated, and suitable for once daily dosing. A second compound 20, 8-(5-pyrazolyl)-xanthine, has been nominated for development from Baraldi's group in conjunction with King Pharmaceuticals that has favorable A(2B) AdoR affinity and selectivity [K(i)(hA(2B)) = 5.5 nM; K(i)(hA(1)) > 1,000 nM; K(i)(hA(2A)) > 1,000; and K(i)(hA(3)) > 1,000 nM], and it has been demonstrated to be a functional antagonist. A third compound 32, a 2-aminopyrimidine, from the Almirall group has high A(2B) AdoR affinity and selectivity [K(i)(hA(2B)) = 17 nM; K(i)(hA(1)) > 1,000 nM; K(i)(hA(2A)) > 2,500; and K(i)(hA(3)) > 1,000 nM], and 32 has been moved into preclinical safety testing. Since three highly selective, high affinity A(2B) AdoR antagonists have been nominated for development with 10 (CVT-6883) being the furthest along in the development process, the role of the A(2B) AdoR in various disease states will soon be established. PMID:18568423

  13. Stochastic description of the ligand-receptor interaction of biologically active substances at extremely low doses.

    Science.gov (United States)

    Gurevich, Konstantin G; Agutter, Paul S; Wheatley, Denys N

    2003-04-01

    Signalling molecules can be effective at extraordinarily low concentrations (down to attomolar levels). To handle such cases, probabilistic methods have been used to describe the formal kinetics of action of biologically active substances in these low doses, although it has been necessary to review what is meant by such a term. The mean numbers of transformed/degraded molecules and their dispersions were calculated for the possible range of ligand-receptor binding schemes. We used both analytical equations and numerical simulations to calculate the coefficients of variation (ratio of standard deviation to mean) and demonstrated that the distribution of the coefficient is highly dependent on the reaction scheme. It may, therefore, be used as an additional factor for discriminating between cooperative and noncooperative models of ligand-receptor interaction over extreme ranges of ligand dilution. The relevance to signalling behaviour is discussed.

  14. Major advances in the development of histamine H4 receptor ligands.

    Science.gov (United States)

    Smits, Rogier A; Leurs, Rob; de Esch, Iwan J P

    2009-08-01

    The search for new and potent histamine H4 receptor ligands is leading to a steadily increasing number of scientific publications and patent applications. Several interesting and structurally diverse compounds have been found, but fierce IP competition for a preferred 2-aminopyrimidine scaffold is becoming apparent. Recent investigations into the role of the histamine H(4)R in (patho)physiology and the use of H4R ligands in in vivo disease models reveal enormous potential in the field of inflammation and allergy, among others. The development of ligands that display activity at two or more histamine receptor (HR) subtypes is another clinical opportunity that is currently being explored. Taken together, the histamine H4R field is gearing up for clinical studies and has the potential to deliver another generation of blockbuster drugs. PMID:19477292

  15. Triazoloquinazolinediones as novel high affinity ligands for the benzodiazepine site of GABA(A) receptors

    DEFF Research Database (Denmark)

    Nilsson, Jakob; Gidlöf, Ritha; Nielsen, Elsebet Østergaard;

    2011-01-01

    Based on a pharmacophore model of the benzodiazepine-binding site of GABA(A) receptors, a series of 2-aryl-2,6-dihydro[1,2,4]triazolo[4,3-c]quinazoline-3,5-diones (structure type I) were designed, synthesized, and identified as high-affinity ligands of the binding site. For several compounds, K(i...

  16. Ligand binding to G protein-coupled receptors in tethered cell membranes

    DEFF Research Database (Denmark)

    Martinez, Karen L.; Meyer, Bruno H.; Hovius, Ruud;

    2003-01-01

    G protein-coupled receptors (GPCRs) constitute a large class of seven transmembrane proteins, which bind selectively agonists or antagonists with important consequences for cellular signaling and function. Comprehension of the molecular details of ligand binding is important for the understanding...

  17. Oestrogen receptor β ligand: a novel treatment to enhance endogenous functional remyelination

    OpenAIRE

    Crawford, Daniel K.; Mangiardi, Mario; Song, Bingbing; Patel, Rhusheet; Du, Sienmi; Michael V Sofroniew; Voskuhl, Rhonda R; Tiwari-Woodruff, Seema K.

    2010-01-01

    Demyelinating diseases, such as multiple sclerosis, are characterized by inflammatory demyelination and neurodegeneration of the central nervous system. Therapeutic strategies that induce effective neuroprotection and enhance intrinsic repair mechanisms are central goals for future therapy of multiple sclerosis. Oestrogens and oestrogen receptor ligands are promising treatments to prevent multiple sclerosis-induced neurodegeneration. In the present study we investigated the capacity of oestro...

  18. REACTIVITY PROFILE OF LIGANDS OF MAMMALIAN RETINOIC ACID RECEPTORS: A PRELIMINARY COREPA ANALYSIS

    Science.gov (United States)

    Retinoic acid and associated derivatives comprise a class of endogenous hormones that bind to and activate different families of retinoic acid receptors (RARs, RXRs), and control many aspects of vertebrate development. Identification of potential RAR and RXR ligands is of interes...

  19. Tension-compression asymmetry in the binding affinity of membrane-anchored receptors and ligands

    Science.gov (United States)

    Xu, Guang-Kui; Liu, Zishun; Feng, Xi-Qiao; Gao, Huajian

    2016-03-01

    Cell adhesion plays a crucial role in many biological processes of cells, e.g., immune responses, tissue morphogenesis, and stem cell differentiation. An essential problem in the molecular mechanism of cell adhesion is to characterize the binding affinity of membrane-anchored receptors and ligands under different physiological conditions. In this paper, a theoretical model is presented to study the binding affinity between a large number of anchored receptors and ligands under both tensile and compressive stresses, and corroborated by demonstrating excellent agreement with Monte Carlo simulations. It is shown that the binding affinity becomes lower as the magnitude of the applied stress increases, and drops to zero at a critical tensile or compressive stress. Interestingly, the critical compressive stress is found to be substantially smaller than the critical tensile stress for relatively long and flexible receptor-ligand complexes. This counterintuitive finding is explained by using the Euler instability theory of slender columns under compression. The tension-compression asymmetry in the binding affinity of anchored receptors and ligands depends subtly on the competition between the breaking and instability of their complexes. This study helps in understanding the role of mechanical forces in cell adhesion mediated by specific binding molecules.

  20. The molecular basis of ligand interaction at free fatty acid receptor 4 (FFA4/GPR120)

    DEFF Research Database (Denmark)

    Hudson, Brian D; Shimpukade, Bharat; Milligan, Graeme;

    2014-01-01

    The long-chain fatty acid receptor FFA4 (previously GPR120) is receiving substantial interest as a novel target for the treatment of metabolic and inflammatory disease. This study examines for the first time the detailed mode of binding of both long-chain fatty acid and synthetic agonist ligands ...

  1. Unnatural amino acids as probes of ligand-receptor interactions and their conformational consequences

    DEFF Research Database (Denmark)

    Pless, Stephan Alexander; Ahern, Christopher A

    2013-01-01

    -edge synthetic and chemical biological approaches. Here we summarize recent advances in the use of site-directed incorporation of unnatural amino acids and chemical probes to study ligand-receptor interactions, determine the location of binding sites, and examine the downstream conformational consequences...

  2. Elimination of a ligand gating site generates a supersensitive olfactory receptor

    Science.gov (United States)

    Sharma, Kanika; Ahuja, Gaurav; Hussain, Ashiq; Balfanz, Sabine; Baumann, Arnd; Korsching, Sigrun I.

    2016-01-01

    Olfaction poses one of the most complex ligand-receptor matching problems in biology due to the unparalleled multitude of odor molecules facing a large number of cognate olfactory receptors. We have recently deorphanized an olfactory receptor, TAAR13c, as a specific receptor for the death-associated odor cadaverine. Here we have modeled the cadaverine/TAAR13c interaction, exchanged predicted binding residues by site-directed mutagenesis, and measured the activity of the mutant receptors. Unexpectedly we observed a binding site for cadaverine at the external surface of the receptor, in addition to an internal binding site, whose mutation resulted in complete loss of activity. In stark contrast, elimination of the external binding site generated supersensitive receptors. Modeling suggests this site to act as a gate, limiting access of the ligand to the internal binding site and thereby downregulating the affinity of the native receptor. This constitutes a novel mechanism to fine-tune physiological sensitivity to socially relevant odors. PMID:27323929

  3. In Silico Docking of HNF-1a Receptor Ligands

    Directory of Open Access Journals (Sweden)

    Gumpeny Ramachandra Sridhar

    2012-01-01

    Full Text Available Background. HNF-1a is a transcription factor that regulates glucose metabolism by expression in various tissues. Aim. To dock potential ligands of HNF-1a using docking software in silico. Methods. We performed in silico studies using HNF-1a protein 2GYP·pdb and the following softwares: ISIS/Draw 2.5SP4, ARGUSLAB 4.0.1, and HEX5.1. Observations. The docking distances (in angstrom units: 1 angstrom unit (Å = 0.1 nanometer or  metres with ligands in decreasing order are as follows: resveratrol (3.8 Å, aspirin (4.5 Å, stearic acid (4.9 Å, retinol (6.0 Å, nitrazepam (6.8 Å, ibuprofen (7.9 Å, azulfidine (9.0 Å, simvastatin (9.0 Å, elaidic acid (10.1 Å, and oleic acid (11.6 Å. Conclusion. HNF-1a domain interacted most closely with resveratrol and aspirin

  4. Ligand-based receptor tyrosine kinase partial agonists: New paradigm for cancer drug discovery?

    Science.gov (United States)

    Riese, David J.

    2010-01-01

    Introduction Receptor tyrosine kinases (RTKs) are validated targets for oncology drug discovery and several RTK antagonists have been approved for the treatment of human malignancies. Nonetheless, the discovery and development of RTK antagonists has lagged behind the discovery and development of agents that target G-protein coupled receptors. In part, this is because it has been difficult to discover analogs of naturally-occurring RTK agonists that function as antagonists. Areas covered Here we describe ligands of ErbB receptors that function as partial agonists for these receptors, thereby enabling these ligands to antagonize the activity of full agonists for these receptors. We provide insights into the mechanisms by which these ligands function as antagonists. We discuss how information concerning these mechanisms can be translated into screens for novel small molecule- and antibody-based antagonists of ErbB receptors and how such antagonists hold great potential as targeted cancer chemotherapeutics. Expert opinion While there have been a number of important key findings into this field, the identification of the structural basis of ligand functional specificity is still of the greatest importance. While it is true that, with some notable exceptions, peptide hormones and growth factors have not proven to be good platforms for oncology drug discovery; addressing the fundamental issues of antagonistic partial agonists for receptor tyrosine kinases has the potential to steer oncology drug discovery in new directions. Mechanism based approaches are now emerging to enable the discovery of RTK partial agonists that may antagonize both agonist-dependent and –independent RTK signaling and may hold tremendous promise as targeted cancer chemotherapeutics. PMID:21532939

  5. Design and synthesis of carborane-containing estrogen receptor-beta (ERβ)-selective ligands.

    Science.gov (United States)

    Ohta, Kiminori; Ogawa, Takumi; Oda, Akifumi; Kaise, Asako; Endo, Yasuyuki

    2015-10-01

    Candidates for highly selective estrogen receptor-beta (ERβ) ligands (6a-c, 7a-c, 8a and 8b) were designed and synthesized based on carborane-containing ER ligands 1 and 2 as lead compounds. Among them, p-carboranylcyclohexanol derivatives 8a and 8b exhibited high ERβ selectivity in competitive binding assay: for example, 8a showed 56-fold selectivity for ERβ over ERα. Docking studies of 8a and 8b with the ERα and ERβ ligand-binding domains (LBDs) suggested that the p-carborane cage of the ligands is located close to key amino acid residues that influence ER-subtype selectivity, that is, Leu384 in the ERα LBD and Met336 in the ERβ LBD. The p-carborane cage in 8a and 8b appears to play a crucial role in the increased ERβ selectivity. PMID:26298498

  6. Role of adenosine A2B receptor signaling in contribution of cardiac mesenchymal stem-like cells to myocardial scar formation.

    Science.gov (United States)

    Ryzhov, Sergey; Sung, Bong Hwan; Zhang, Qinkun; Weaver, Alissa; Gumina, Richard J; Biaggioni, Italo; Feoktistov, Igor

    2014-09-01

    Adenosine levels increase in ischemic hearts and contribute to the modulation of that pathological environment. We previously showed that A2B adenosine receptors on mouse cardiac Sca1(+)CD31(-) mesenchymal stromal cells upregulate secretion of paracrine factors that may contribute to the improvement in cardiac recovery seen when these cells are transplanted in infarcted hearts. In this study, we tested the hypothesis that A2B receptor signaling regulates the transition of Sca1(+)CD31(-) cells, which occurs after myocardial injury, into a myofibroblast phenotype that promotes myocardial repair and remodeling. In vitro, TGFβ1 induced the expression of the myofibroblast marker α-smooth muscle actin (αSMA) and increased collagen I generation in Sca1(+)CD31(-) cells. Stimulation of A2B receptors attenuated TGFβ1-induced collagen I secretion but had no effect on αSMA expression. In vivo, myocardial infarction resulted in a rapid increase in the numbers of αSMA-positive cardiac stromal cells by day 5 followed by a gradual decline. Genetic deletion of A2B receptors had no effect on the initial accumulation of αSMA-expressing stromal cells but hastened their subsequent decline; the numbers of αSMA-positive cells including Sca1(+)CD31(-) cells remained significantly higher in wild type compared with A2B knockout hearts. Thus, our study revealed a significant contribution of cardiac Sca1(+)CD31(-) cells to the accumulation of αSMA-expressing cells after infarction and implicated A2B receptor signaling in regulation of myocardial repair and remodeling by delaying deactivation of these cells. It is plausible that this phenomenon may contribute to the beneficial effects of transplantation of these cells to the injured heart.

  7. A python-based docking program utilizing a receptor bound ligand shape: PythDock.

    Science.gov (United States)

    Chung, Jae Yoon; Cho, Seung Joo; Hah, Jung-Mi

    2011-09-01

    PythDock is a heuristic docking program that uses Python programming language with a simple scoring function and a population based search engine. The scoring function considers electrostatic and dispersion/repulsion terms. The search engine utilizes a particle swarm optimization algorithm. A grid potential map is generated using the shape information of a bound ligand within the active site. Therefore, the searching area is more relevant to the ligand binding. To evaluate the docking performance of PythDock, two well-known docking programs (AutoDock and DOCK) were also used with the same data. The accuracy of docked results were measured by the difference of the ligand structure between x-ray structure, and docked pose, i.e., average root mean squared deviation values of the bound ligand were compared for fourteen protein-ligand complexes. Since the number of ligands' rotational flexibility is an important factor affecting the accuracy of a docking, the data set was chosen to have various degrees of flexibility. Although PythDock has a scoring function simpler than those of other programs (AutoDock and DOCK), our results showed that PythDock predicted more accurate poses than both AutoDock4.2 and DOCK6.2. This indicates that PythDock could be a useful tool to study ligand-receptor interactions and could also be beneficial in structure based drug design.

  8. Tools for investigating functional interactions between ligands and G-protein-coupled receptors.

    Science.gov (United States)

    Lerner, M R

    1994-04-01

    A general assay for evaluating functional interactions between ligands and G-protein-coupled receptors within minutes has been developed. The system uses the principles employed by animals such as reptiles, amphibians and fish to control their colors. In nature, activation of G-protein-coupled receptors expressed by skin cells called chromatophores effects pigment redistribution within the cells to change an animal's coloration. The in vitro 'chameleon in a dish' equivalent can use essentially any cloned G-protein-coupled receptor. PMID:7517590

  9. Labelling of central neural system receptor ligands with the fac-[Tc(CO)3]+ moiety

    International Nuclear Information System (INIS)

    During the period of the IAEA Co-ordinated Research Project on Development of Agents for the Imaging of CNS Receptors based on 99mTc, many efforts were made to find an improved system or alternative methods for the labelling of various central nervous system (CNS) receptor binding agents based on the fac-[Tc(CO)3]+ fragment. Within the same period the chemistry of the fac-[Tc(CO)3]+ fragment has been developed as a useful label more and more not only for the labelling of CNS receptor ligands but also for peptides, antibodies and other biologically active molecules such as B12. Especially the latter molecule is known to be taken up as well through the blood-brain barrier but is obviously not an CNS receptor ligand. One of the most important achievements over the whole period of the project has been the final formulation of a kit useful for the preparation of [99mTc(OH2)3(CO)3]+ without the requirement for using free CO. Much time was invested in that particular topic, since it will allow this relevant moiety to be applied not only on a routine basis but also for research into CNS ligands. A major achievement has thus been the commercial availability of these kits by the beginning of 2002. During the period of the project, a number of new systems were introduced, some of which were specially designed not only for CNS receptor ligands but also for other biomolecules. Among these is that for the syntheses of highly lipophilic ligands, the complex formation of which is based on classical co-ordination chemistry. In addition, the feasibility of the mixed ligand concept from a chemical point of view has been proved in principle. A number of complexes have been prepared where the CNS receptor ligand is attached to the monodentate ligand system. In principle it can also be attached to the bidentate moiety, allowing a screening of the biological behaviour as a function of the co-ligand. A major breakthrough could be achieved with the aqueous synthesis of cymantren

  10. Derivatives of serotonergic receptors ligands labeled with SPECT radionuclide for neutronal imaging

    International Nuclear Information System (INIS)

    Full text: Introduction: Serotonergic receptors are associated with a variety of pathophysiology of neuropsychiatric disorders. Serotonergic ligands have remained a very active area in the development of CNS drugs. In search of the ligands that recognize serotonergic receptor we have synthesized derivatives of methoxyphenylpiperazine. Long chain alkylation of methoxyphenylpiperazine was successfully carried out and a series of MPP based precursors were obtained which comprised of hydrocarbon chain of varied length. These derivatives were then conjugated to acyclic chelating system and efficiently labeled with SPECT radionuclide. Materials and Methods: Labeling was performed with high yield (>95%) and radiochemical purity (>98%) using very low ligand concentration. In vivo studies were done on Hela cell lines which overexpress serotonergic receptors. Further studies done includes in vivo distribution and gamma scintigraphy performed in rat and rabbit. Results: All the intermediates and final compounds were characterized by 1H, 13C NMR and Mass Spectroscopy. In vitro binding assays in rat hippocampal cultures demonstrated the high affinity of complexes for serotonergic receptors. Conclusion: We have optimized the synthesis of 2-methoxyphenylpiperazine based chelating agents. This series of imaging agents holds a promising future in imaging 5-HT receptors for the effective treatment of neuropathological disorders

  11. Structure of complement receptor 2 in complex with its C3d ligand.

    Science.gov (United States)

    Szakonyi, G; Guthridge, J M; Li, D; Young, K; Holers, V M; Chen, X S

    2001-06-01

    Complement receptor 2 (CR2/CD21) is an important receptor that amplifies B lymphocyte activation by bridging the innate and adaptive immune systems. CR2 ligands include complement C3d and Epstein-Barr virus glycoprotein 350/220. We describe the x-ray structure of this CR2 domain in complex with C3d at 2.0 angstroms. The structure reveals extensive main chain interactions between C3d and only one short consensus repeat (SCR) of CR2 and substantial SCR side-side packing. These results provide a detailed understanding of receptor-ligand interactions in this protein family and reveal potential target sites for molecular drug design. PMID:11387479

  12. Synthesis and biological activity of novel small peptides with aminophosphonates moiety as NOP receptor ligands.

    Science.gov (United States)

    Naydenova, Emilia D; Todorov, Petar T; Mateeva, Polina I; Zamfirova, Rositza N; Pavlov, Nikola D; Todorov, Simeon B

    2010-11-01

    The aim of the present study was the synthesis and the biological screening of new analogs of Ac-RYYRWK-NH2, modified at the N-terminal with 1-[(methoxyphosphono)methylamino]cycloalkanecarboxylic acids. The four newly synthesized ligands for the nociceptin/orphanin FQ (N/OFQ) receptor (NOP) have been prepared by solid-phase peptide synthesis--Fmoc-strategy. These compounds were tested for agonistic activity in vitro on electrically stimulated smooth-muscle preparations isolated from vas deferens of Wistar rats. Our data showed that substitution of Arg at position 1 with aminophosphonates moiety decreased significantly the affinity of ligands to the NOP receptor. Furthermore, the enlargement of the cycle (with 5-8 carbon atoms) additionally diminished both the activity and the selectivity for NOP-receptor.

  13. The Length and Flexibility of the 2-Substituent of 9-Ethyladenine Derivatives Modulate Affinity and Selectivity for the Human A2A Adenosine Receptor.

    Science.gov (United States)

    Thomas, Ajiroghene; Buccioni, Michela; Dal Ben, Diego; Lambertucci, Catia; Marucci, Gabriella; Santinelli, Claudia; Spinaci, Andrea; Kachler, Sonja; Klotz, Karl-Norbert; Volpini, Rosaria

    2016-08-19

    The A2A adenosine receptor (A2A AR) is a key target for the development of pharmacological tools for the treatment of central nervous system disorders. Previous works have demonstrated that the insertion of substituents at various positions on adenine leads to A2A AR antagonists with affinity in the micromolar to nanomolar range. In this work, a series of 9-ethyladenine derivatives bearing phenylalkylamino, phenylakyloxy or phenylakylthio groups of different lengths at the 2-position were synthesised and tested against the human adenosine receptors. The derivatives showed sub-micromolar affinity for these membrane proteins. The further introduction of a bromine atom at the 8-position has the effect of improving the affinity and selectivity for all ARs and led to compounds that are able bind to the A2A AR subtype at low nanomolar levels. Functional studies confirmed that the new adenine derivatives behave as A2A AR antagonists with half-maximal inhibitory concentration values in the nanomolar range. Molecular modelling studies provide a description of the possible binding mode of these compounds at the A2A AR and an interpretation of the affinity data at this AR subtype. PMID:27037522

  14. Neuroprotection of Persea major extract against oxygen and glucose deprivation in hippocampal slices involves increased glutamate uptake and modulation of A1 and A2A adenosine receptors

    Directory of Open Access Journals (Sweden)

    Marielli Letícia Fedalto

    2013-10-01

    Full Text Available Ischemic stroke is characterised by a lack of oxygen and glucose in the brain, leading to excessive glutamate release and neuronal cell death. Adenosine is produced in response to ATP depletion and acts as an endogenous neuromodulator that reduces excitotoxicity. Persea major (Meins. L.E. Kopp (Lauraceae is a medical plant that is indigenous to South Brazil, and the rural population has used it medicinally due to its anti-inflammatory properties. The aim of this study was to evaluate the neuroprotective effect of Persea major methanolic extract against oxygen and glucose deprivation and re-oxygenation as well as to determine its underlying mechanism of action in hippocampal brain slices. Persea major methanolic extract (0.5 mg/ml has a neuroprotective effect on hippocampal slices when added before or during 15 min of oxygen and glucose deprivation or 2 h of re-oxygenation. Hippocampal slices subjected to oxygen and glucose deprivation and re-oxygenation showed significantly reduced glutamate uptake, and the addition of Persea major methanolic extract in the re-oxygenation period counteracted the reduction of glutamate uptake. The presence of A1 or A2A, but not A2B or A3 receptor antagonists, abolished the neuroprotective effect of Persea major methanolic extract. In conclusion, the neuroprotective effect of Persea majormethanolic extract involves augmentation of glutamate uptake and modulation of A1 and A2B adenosine receptors.

  15. GABA Acts as a Ligand Chaperone in the Early Secretory Pathway to Promote Cell Surface Expression of GABAA Receptors

    OpenAIRE

    Eshaq, Randa S.; Stahl, Letha D.; Stone, Randolph; Smith, Sheryl S.; Robinson, Lucy C.; Leidenheimer, Nancy J.

    2010-01-01

    GABA (γ-aminobutyric acid) is the primary inhibitory neurotransmitter in brain. The fast inhibitory effect of GABA is mediated through the GABAA receptor, a postsynaptic ligand-gated chloride channel. We propose that GABA can act as a ligand chaperone in the early secretory pathway to facilitate GABAA receptor cell surface expression. Forty-two hrs of GABA treatment increased the surface expression of recombinant receptors expressed in HEK 293 cells, an effect accompanied by an increase in GA...

  16. A strategy using NMR peptide structures of thromboxane A2 receptor as templates to construct ligand-recognition pocket of prostacyclin receptor

    Directory of Open Access Journals (Sweden)

    Ruan Ke-He

    2005-11-01

    Full Text Available Abstract Background: Prostacyclin receptor (IP and thromboxane A2 receptor (TP belong to rhodopsin-type G protein-coupling receptors and respectively bind to prostacyclin and thromboxane A2 derived from arachidonic acid. Recently, we have determined the extracellular loop (eLP structures of the human TP receptor by 2-D 1H NMR spectroscopy using constrained peptides mimicking the individual eLP segments. The studies have identified the segment along with several residues in the eLP domains important to ligand recognition, as well as proposed a ligand recognition pocket for the TP receptor. Results: The IP receptor shares a similar primary structure in the eLPs with those of the TP receptor. Forty percent residues in the second eLPs of the receptors are identical, which is the major region involved in forming the ligand recognition pocket in the TP receptor. Based on the high homology score, the eLP domains of the IP receptor were constructed by the homology modeling approach using the NMR structures of the TP eLPs as templates, and then configured to the seven transmembrane (TM domains model constructed using the crystal structure of the bovine rhodopsin as a template. A NMR structure of iloprost was docked into the modeled IP ligand recognition pocket. After dynamic studies, the segments and residues involved in the IP ligand recognition were proposed. A key residue, Arg173 involved in the ligand recognition for the IP receptor, as predicted from the modeling, was confirmed by site-directed mutagenesis. Conclusion: A 3-D model of the human IP receptor was constructed by homology modeling using the crystal structure of bovine rhodopsin TM domains and the NMR structures of the synthetic constrained peptides of the eLP domains of the TP receptor as templates. This strategy can be applied to molecular modeling and the prediction of ligand recognition pockets for other prostanoid receptors.

  17. In search of selective P2 receptor ligands: interaction of dihydropyridine derivatives at recombinant rat P2X(2) receptors.

    Science.gov (United States)

    Jacobson, K A; Kim, Y C; King, B F

    2000-07-01

    1,4-Dihydropyridines are regarded as privileged structures for drug design, i.e. they tend to bind to a wide variety of receptor sites. We have shown that upon appropriate manipulation of the substituent groups on a 1,4-dihydropyridine template, high affinity and selectivity for the A(3) subtype of adenosine receptors ('P1 receptors') may be attained. In the present study we have begun to extend this approach to P2 receptors which are activated by ATP and other nucleotides. Nicardipine, a representative dihydropyridine, used otherwise as an L-type calcium channel blocker, was shown to be an antagonist at recombinant rat P2X(2) (IC(50)=25 microM) and P2X(4) (IC(50) approximately 220 microM) receptors expressed in Xenopus oocytes. Thus, this class of compounds represents a suitable lead for enhancement of affinity through chemical synthesis. In an attempt to modify the 1,4-dihydropyridine structure with a predicted P2 receptor recognition moiety, we have replaced one of the ester groups with a negatively charged phosphonate group. Several 4-phenyl-5-phosphonato-1,4-dihydropyridine derivatives, MRS 2154 (2, 6-dimethyl), MRS 2155 (6-methyl-2-phenyl), and MRS 2156 (2-methyl-6-phenyl), were synthesized through three component condensation reactions. These derivatives were not pure antagonists of the effects of ATP at P2X(2) receptors, rather were either inactive (MRS 2156) or potentiated the effects of ATP in a concentration-dependent manner (MRS 2154 in the 0.3-10 microM range and MRS 2155 at >1 microM). Antagonism of the effects of ATP at P2X(2) receptor superimposed on the potentiation was also observed at >10 microM (MRS 2154) or 0.3-1 microM (MRS 2155). Thus, while a conventional dihydropyridine, nicardipine, was found to antagonize rat P2X(2) receptors ninefold more potently than P2X(4) receptors, the effects of novel, anionic 5-phosphonate analogues at the receptor were more complex.

  18. Presynaptic muscarinic and adenosine receptors are involved in 2 Hz-induced train-of-four fade caused by antinicotinic neuromuscular relaxants in the rat.

    Science.gov (United States)

    Pereira, Mw; Bornia, Ecs; Correia-de-Sá, P; Alves-Do-Prado, W

    2011-11-01

    1. Train-of-four fade (TOF(fade) ) is a clinically useful parameter to monitor the degree of block of neuromuscular transmission in curarized patients. Experimentally, TOF(fade) has been attributed to the blockade of facilitatory nicotinic receptors on motor nerve terminals. There is less information regarding the involvement of coexistent presynaptic receptors (e.g. muscarinic M(1) and M(2) , adenosine A(1) and A(2A) ) in the TOF(fade) produced by antinicotinic agents. 2. In the present study, we evaluated the TOF(fade) caused by antinicotinic neuromuscular relaxants (hexamethonium, d-tubocurarine, vecuronium and rocuronium) as the ratio of the muscle tension produced in the rat diaphragm by the fourth to the first stimulus (T(4) /T(1) ) of a train-of-four stimuli delivered to the phrenic nerve trunk at a frequency of 2 Hz. 3. All antinicotinic agents, except hexamethonium, decreased the amplitude of muscle tension during the first stimulus. Hexamethonium, (5.47 mmol/L), d-tubocurarine- (1.1 μmol/L), vecuronium (4.7 μmol/L)- and rocuronium (9.8 μmol/L)-induced TOF(fade) was attenuated by 10 nmol/L pirenzepine (an M(1) receptor antagonist), 1 μmol/L methoctramine (an M(2) receptor antagonist) and 2.5 nmol/L 1,3-dipropyl-8-cyclopentylxanthine (an A(1) receptor antagonist). Blockade of the A(2A) receptor with 10 nmol/L ZM241385 partially reversed the TOF(fade) induced by d-tubocurarine, vecuronium and rocuronium, but not that caused by the 'pure' neuronal nicotinic receptor antagonist hexamethonium, unless one increased the concentration of ZM241385 to 50 nmol/L. 4. The data indicate that presynaptic M(1) , M(2) , A(1) and A(2A) receptors play a role in neuromuscular TOF(fade) caused by antinicotinic neuromuscular relaxants. Such interplay depends on adenosine tonus and on the affinity of neuromuscular blocking agents for neuronal versus muscular nicotinic receptors.

  19. Homology modeling of adenosine A2A receptor and molecular docking for exploration of appropriate potent antagonists for treatment of Parkinson's disease.

    Science.gov (United States)

    Singh, Vijai; Somvanshi, Pallavi

    2009-07-01

    Parkinson's disease (PD) is a neurodegenerative disorder of central nervous system (CNS) that impaired the patient motor skills, speech and other functions. Adenosine A2A receptors have a unique cellular distribution in the neuron, which is used as a potential target for PD. Homology modeling was used to construct the 3-D structure of A2A using the known template (PDB: 2VT4), and the stereochemical quality was validated. Several effective antagonist drugs were selected and active amino acid residues in A2A were targeted on the basis of robust binding affinity between protein-drug interactions in molecular docking. Six antagonists, Bromocriptine, Cabergoline, Etilevodopa, Lysuride, Melevodopa and Pramipexole, were found more potent for binding and the active amino acids residues were identified (http://www.rcsb.org/pdb/) in A2A receptor. It could be used as the basis for rationale designing of novel antagonist drugs against Parkinson's disease. PMID:20021407

  20. Acetylation of pregnane X receptor protein determines selective function independent of ligand activation

    International Nuclear Information System (INIS)

    Research highlights: → Pregnane X receptor (PXR), a major regulatory protein, is modified by acetylation. → PXR undergoes dynamic deacetylation upon ligand-mediated activation. → SIRT1 partially mediates PXR deacetylation. → PXR deacetylation per se induces lipogenesis mimicking ligand-mediated activation. -- Abstract: Pregnane X receptor (PXR), like other members of its class of nuclear receptors, undergoes post-translational modification [PTM] (e.g., phosphorylation). However, it is unknown if acetylation (a major and common form of protein PTM) is observed on PXR and, if it is, whether it is of functional consequence. PXR has recently emerged as an important regulatory protein with multiple ligand-dependent functions. In the present work we show that PXR is indeed acetylated in vivo. SIRT1 (Sirtuin 1), a NAD-dependent class III histone deacetylase and a member of the sirtuin family of proteins, partially mediates deacetylation of PXR. Most importantly, the acetylation status of PXR regulates its selective function independent of ligand activation.

  1. Ligand-stimulated downregulation of the alpha interferon receptor: role of protein kinase D2.

    Science.gov (United States)

    Zheng, Hui; Qian, Juan; Varghese, Bentley; Baker, Darren P; Fuchs, Serge

    2011-02-01

    Alpha interferon (IFN-α) controls homeostasis of hematopoietic stem cells, regulates antiviral resistance, inhibits angiogenesis, and suppresses tumor growth. This cytokine is often used to treat cancers and chronic viral infections. The extent of cellular responses to IFN-α is limited by the IFN-induced ubiquitination and degradation of the IFN-α/β receptor chain 1 (IFNAR1) chain of the cognate receptor. IFNAR1 ubiquitination is facilitated by the βTrcp E3 ubiquitin ligase that is recruited to IFNAR1 upon its degron phosphorylation, which is induced by the ligand. Here we report identification of protein kinase D2 (PKD2) as a kinase that mediates the ligand-inducible phosphorylation of IFNAR1 degron and enables binding of βTrcp to the receptor. Treatment of cells with IFN-α induces catalytic activity of PKD2 and stimulates its interaction with IFNAR1. Expression and kinase activity of PKD2 are required for the ligand-inducible stimulation of IFNAR1 ubiquitination and endocytosis and for accelerated proteolytic turnover of IFNAR1. Furthermore, inhibition or knockdown of PKD2 robustly augments intracellular signaling induced by IFN-α and increases the efficacy of its antiviral effects. The mechanisms of the ligand-inducible elimination of IFNAR1 are discussed, along with the potential medical significance of this regulation. PMID:21173164

  2. Active regions' setting of the extracellular ligand-binding domain of human interleukin-6 receptor

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

    The reliable three dimensional (3-D) structure of the extracellular ligand-binding domain (V106-P322) of human interleukin-6 receptor (hIL-6R) has been constructed by means of computer-guided homology modeling techniques using the crystal structure of the extracellular ligand-binding region (K52-L251) of human growth hormone receptor (hGHR) as templet. The space location of some key residues which influence the combination ability between the receptor and the ligand has been observed and the effects of point mutagenesis of the four conservative cysteine residues on the space conformation are analyzed. The results show that the space conformation of the side-chain carboxyl of E305 plays a key role in the ligand-binding ability. Furthermore, the space conformation of the side-chain carboxyl of E305 is very important for the electrostatic potential complementarity between hIL-6R and hIL-6 according to the docking method.

  3. Bile acid derivatives as ligands of the farnesoid x receptor: molecular determinants for bile acid binding and receptor modulation.

    Science.gov (United States)

    Gioiello, Antimo; Cerra, Bruno; Mostarda, Serena; Guercini, Chiara; Pellicciari, Roberto; Macchiarulo, Antonio

    2014-01-01

    Bile acids are a peculiar class of steroidal compounds that never cease to amaze. From being simple detergents with a primary role in aiding the absorption of fats and fat-soluble vitamins, bile acids are now widely considered as crucial hormones endowed with genomic and non-genomic functions that are mediated by their interaction with several proteins including the nuclear receptor Farnesoid X Receptor (FXR). Taking advantages of the peculiar properties of bile acids in interacting with the FXR receptor, several biliary derivatives have been synthesized and tested as FXR ligands. The availability of these compounds has contributed to characterize the receptor from a structural, patho-physiological and therapeutic standpoint. Among these, obeticholic acid is a first-in-class FXR agonist that is demonstrating hepatoprotective effects upon FXR activation in patients with liver diseases such as primary biliary cirrhosis and nonalcoholic steatohepatitis. This review provides an historical overview of the rationale behind the discovery of obeticholic acid and chemical tools generated to depict the molecular features and bio-pharmacological relevance of the FXR receptor, as well as to summarize structure-activity relationships of bile acid-based FXR ligands so far reported. PMID:25388535

  4. Evolutionary diversification of retinoic acid receptor ligand-binding pocket structure by molecular tinkering.

    Science.gov (United States)

    Gutierrez-Mazariegos, Juliana; Nadendla, Eswar Kumar; Studer, Romain A; Alvarez, Susana; de Lera, Angel R; Kuraku, Shigehiro; Bourguet, William; Schubert, Michael; Laudet, Vincent

    2016-03-01

    Whole genome duplications (WGDs) have been classically associated with the origin of evolutionary novelties and the so-called duplication-degeneration-complementation model describes the possible fates of genes after duplication. However, how sequence divergence effectively allows functional changes between gene duplicates is still unclear. In the vertebrate lineage, two rounds of WGDs took place, giving rise to paralogous gene copies observed for many gene families. For the retinoic acid receptors (RARs), for example, which are members of the nuclear hormone receptor (NR) superfamily, a unique ancestral gene has been duplicated resulting in three vertebrate paralogues: RARα, RARβ and RARγ. It has previously been shown that this single ancestral RAR was neofunctionalized to give rise to a larger substrate specificity range in the RARs of extant jawed vertebrates (also called gnathostomes). To understand RAR diversification, the members of the cyclostomes (lamprey and hagfish), jawless vertebrates representing the extant sister group of gnathostomes, provide an intermediate situation and thus allow the characterization of the evolutionary steps that shaped RAR ligand-binding properties following the WGDs. In this study, we assessed the ligand-binding specificity of cyclostome RARs and found that their ligand-binding pockets resemble those of gnathostome RARα and RARβ. In contrast, none of the cyclostome receptors studied showed any RARγ-like specificity. Together, our results suggest that cyclostome RARs cover only a portion of the specificity repertoire of the ancestral gnathostome RARs and indicate that the establishment of ligand-binding specificity was a stepwise event. This iterative process thus provides a rare example for the diversification of receptor-ligand interactions of NRs following WGDs. PMID:27069642

  5. In search of selective P2 receptor ligands: interaction of dihydropyridine derivatives at recombinant rat P2X2 receptors

    Science.gov (United States)

    Jacobson, Kenneth A.; Kim, Yong-Chul; King, Brian F.

    2012-01-01

    1,4-Dihydropyridines are regarded as privileged structures for drug design, i.e. they tend to bind to a wide variety of receptor sites. We have shown that upon appropriate manipulation of the substituent groups on a 1,4-dihydropyridine template, high affinity and selectivity for the A3 subtype of adenosine receptors (‘P1 receptors’) may be attained. In the present study we have begun to extend this approach to P2 receptors which are activated by ATP and other nucleotides. Nicardipine, a representative dihydropyridine, used otherwise as an L-type calcium channel blocker, was shown to be an antagonist at recombinant rat P2X2 (IC50 = 25 μM) and P2X4 (IC50 ~ 220 μM) receptors expressed in Xenopus oocytes. Thus, this class of compounds represents a suitable lead for enhancement of affinity through chemical synthesis. In an attempt to modify the 1,4-dihydropyridine structure with a predicted P2 receptor recognition moiety, we have replaced one of the ester groups with a negatively charged phosphonate group. Several 4-phenyl-5-phosphonato-1,4-dihydropyridine derivatives, MRS 2154 (2,6-dimethyl), MRS 2155 (6-methyl-2-phenyl), and MRS 2156 (2-methyl-6-phenyl), were synthesized through three component condensation reactions. These derivatives were not pure antagonists of the effects of ATP at P2X2 receptors, rather were either inactive (MRS 2156) or potentiated the effects of ATP in a concentration-dependent manner (MRS 2154 in the 0.3–10 μM range and MRS 2155 at >1 μM). Antagonism of the effects of ATP at P2X2 receptor superimposed on the potentiation was also observed at >10 μM (MRS 2154) or 0.3–1 μM (MRS 2155). Thus, while a conventional dihydropyridine, nicardipine, was found to antagonize rat P2X2 receptors ninefold more potently than P2X4 receptors, the effects of novel, anionic 5-phosphonate analogues at the receptor were more complex. PMID:10869714

  6. Fluorinated azabicycloesters as muscarinic receptor ligands for application with PET

    International Nuclear Information System (INIS)

    Human muscarinic acetylcholine receptors (MAR) play an important role in a number of physiological and behavioral responses. A correlation has been established between changes in the MAR density and human memory as well as to other specific neurodegenerative disorders such as Huntington's chorea or Alzheimer's dementia. MAR density has been observed, also, to decrease under the effect of several chemical agents such as organophosphorus compounds, barbiturates, ethanol or antidepressants. Most of the studies on human MAR were done on post-mortem samples obtained at autopsy and stored for variable times which may not reflect the actual in vivo status of such receptors. To carry out preliminary in vivo studies, the choice will be directed primarily to experimental animals. However, animal models for many of the neurodegenerative disorders may be inadequate. Several studies showed a dramatically increasing number of dementia cases which is leading to decreased survival among this group. Such a dramatic increase in Alzheimer's dementia cases and the inability to determine the density and distribution of MAR in vivo have stimulated the interest of many researchers to investigate MAR mapping

  7. Structure-Based Evolution of Subtype-Selective Neurotensin Receptor Ligands

    OpenAIRE

    Schaab, Carolin; Kling, Ralf Christian; Einsiedel, Jürgen; Hübner, Harald; Clark, Tim; Seebach, Dieter; Gmeiner, Peter

    2014-01-01

    Subtype-selective agonists of the neurotensin receptor NTS2 represent a promising option for the treatment of neuropathic pain, as NTS2 is involved in the mediation of μ-opioid-independent anti-nociceptive effects. Based on the crystal structure of the subtype NTS1 and previous structure–activity relationships (SARs) indicating a potential role for the sub-pocket around Tyr11 of NT(8–13) in subtype-specific ligand recognition, we have developed new NTS2-selective ligands. Starting from NT(8–1...

  8. 腺苷及其受体参与外周痛觉信息调控的机制%Mechanisms of adenosine and its receptors in pain modulation in the peripheral system

    Institute of Scientific and Technical Information of China (English)

    赵静; 米文丽; 毛应启梁

    2011-01-01

    Adenosine is an endogenous nucleoside that widely exists in the human body cells. Through activating different subgroups of adenosine receptors (A1, A2A, A2B, and A3 receptors), adenosine produces various effects in a broad spectrum of tissues, especially in the central nervous systems, which includes modulating physiological and pathological processes such as sleep, learning and memory, depression as well as anxiety. With the research development in the agonists and antagonists of adenosine receptors, the roles of adenosine and its receptors in the peripheral nervous system have been widely revealed. The researches reported that adenosine and its receptors are closely related to transmission and modulation of nociception in peripheral signals.%腺苷是一种遍布人体细胞的内源性核苷,通过其不同类型的受体(A1,A2A,A2B和A3受体)对机体的许多系统(特别是中枢神经系统)及组织发挥着重要的作用,参与调控睡眠、学习记忆、抑郁和焦虑等多种生理和病理过程.随着腺苷对受体亚型选择性激动剂和拮抗剂的开发,人们对腺苷及其受体在外周神经系统中的作用研究越来越深入,并逐步认识到腺苷及其受体与外周痛党信息的传递和调控密切相关.

  9. Differential ligand-dependent protein–protein interactions between nuclear receptors and a neuronal-specific cofactor

    OpenAIRE

    Greiner, Erich F.; Kirfel, Jutta; Greschik, Holger; Huang, DongYa; Becker, Peter; Kapfhammer, Josef P.; Schüle, Roland

    2000-01-01

    Nuclear receptors are transcription factors that require multiple protein–protein interactions to regulate target gene expression. We have cloned a 27-kDa protein, termed NIX1 (neuronal interacting factor X 1), that directly binds nuclear receptors in vitro and in vivo. Protein–protein interaction between NIX1 and ligand-activated or constitutive active nuclear receptors, including retinoid-related orphan receptor β (RORβ) (NR1F2), strictly depends on the conserved receptor C-terminal activat...

  10. Genetic deletion of the adenosine A(2A) receptor prevents nicotine-induced upregulation of α7, but not α4β2* nicotinic acetylcholine receptor binding in the brain.

    Science.gov (United States)

    Metaxas, Athanasios; Al-Hasani, Ream; Farshim, Pamela; Tubby, Kristina; Berwick, Amy; Ledent, Catherine; Hourani, Susanna; Kitchen, Ian; Bailey, Alexis

    2013-08-01

    Considerable evidence indicates that adenosine A(2A) receptors (A(2A)Rs) modulate cholinergic neurotransmission, nicotinic acetylcholine receptor (nAChR) function, and nicotine-induced behavioural effects. To explore the interaction between A(2A) and nAChRs, we examined if the complete genetic deletion of adenosine A(2A)Rs in mice induces compensatory alterations in the binding of different nAChR subtypes, and whether the long-term effects of nicotine on nAChR regulation are altered in the absence of the A(2A)R gene. Quantitative autoradiography was used to measure cytisine-sensitive [¹²⁵I]epibatidine and [¹²⁵I]α-bungarotoxin binding to α4β2* and α7 nAChRs, respectively, in brain sections of drug-naïve (n = 6) or nicotine treated (n = 5-7), wild-type and adenosine A(2A)R knockout mice. Saline or nicotine (7.8 mg/kg/day; free-base weight) were administered to male CD1 mice via subcutaneous osmotic minipumps for a period of 14 days. Blood plasma levels of nicotine and cotinine were measured at the end of treatment. There were no compensatory developmental alterations in nAChR subtype distribution or density in drug-naïve A(2A)R knockout mice. In nicotine treated wild-type mice, both α4β2* and α7 nAChR binding sites were increased compared with saline treated controls. The genetic ablation of adenosine A(2A)Rs prevented nicotine-induced upregulation of α7 nAChRs, without affecting α4β2* receptor upregulation. This selective effect was observed at plasma levels of nicotine that were within the range reported for smokers (10-50 ng ml⁻¹). Our data highlight the involvement of adenosine A(2A)Rs in the mechanisms of nicotine-induced α7 nAChR upregulation, and identify A(2A)Rs as novel pharmacological targets for modulating the long-term effects of nicotine on α7 receptors. PMID:23583933

  11. Ligands for SPECT and PET imaging of muscarinic-cholinergic receptors of the heart and brain

    Energy Technology Data Exchange (ETDEWEB)

    Knapp, F.F. Jr.; McPherson, D.W.; Luo, H. [and others

    1995-06-01

    Interest in the potential use of cerebral SPECT and PET imaging for determination of the density and activity of muscarinic-cholinergic receptors (mAChR) has been stimulated by the changes in these receptors which occur in many neurological diseases. In addition, the important involvement of mAChR in modulating negative inotropic cardiac activity suggests that such receptor ligands may have important applications in evaluation of changes which may occur in cardiac disease. In this paper, the properties of several key muscarinic receptor ligands being developed or which have been used for clinical SPECT and PET are discussed. In addition, the ORNL development of the new iodinated IQNP ligand based on QNB and the results of in vivo biodistribution studies in rats, in vitro competitive binding studies and ex vivo autoradiographic experiments are described. The use of radioiodinated IQNP may offer several advantages in comparison to IQNB because of its easy and high yield preparation and high brain uptake and the potential usefulness of the {open_quotes}partial{close_quotes} subtype selective IONP isomers. We also describe the development of new IQNP-type analogues which offer the opportunity for radiolabeling with positron-emitting radioisotopes (carbon-11, fluorine-18 and bromine-76) for potential use with PET.

  12. Molecular mechanism of ligand recognition by NR3 subtype glutamate receptors

    Energy Technology Data Exchange (ETDEWEB)

    Yao, Yongneng; Harrison, Chris B.; Freddolino, Peter L.; Schulten, Klaus; Mayer, Mark L. (UIUC); (NIH)

    2008-10-27

    NR3 subtype glutamate receptors have a unique developmental expression profile, but are the least well-characterized members of the NMDA receptor gene family, which have key roles in synaptic plasticity and brain development. Using ligand binding assays, crystallographic analysis, and all atom MD simulations, we investigate mechanisms underlying the binding by NR3A and NR3B of glycine and D-serine, which are candidate neurotransmitters for NMDA receptors containing NR3 subunits. The ligand binding domains of both NR3 subunits adopt a similar extent of domain closure as found in the corresponding NR1 complexes, but have a unique loop 1 structure distinct from that in all other glutamate receptor ion channels. Within their ligand binding pockets, NR3A and NR3B have strikingly different hydrogen bonding networks and solvent structures from those found in NR1, and fail to undergo a conformational rearrangement observed in NR1 upon binding the partial agonist ACPC. MD simulations revealed numerous interdomain contacts, which stabilize the agonist-bound closed-cleft conformation, and a novel twisting motion for the loop 1 helix that is unique in NR3 subunits.

  13. Domain architecture of a calcium-permeable AMPA receptor in a ligand-free conformation

    Directory of Open Access Journals (Sweden)

    Charles R. Midgett

    2012-01-01

    Full Text Available Ligand-gated ion channels couple the free energy of agonist binding to the gating of selective transmembrane ion pores, permitting cells to regulate ion flux in response to external chemical stimuli. However, the stereochemical mechanisms responsible for this coupling remain obscure. In the case of the ionotropic glutamate receptors (iGluRs, the modular nature of receptor subunits has facilitated structural analysis of the N-terminal domain (NTD, and of multiple conformations of the ligand-binding domain (LBD. Recently, the crystallographic structure of an antagonist-bound form of the receptor was determined. However, disulfide trapping of this conformation blocks channel opening, suggesting that channel activation involves additional quaternary packing arrangements. To explore the conformational space available to iGluR channels, we report here a second, clearly distinct domain architecture of homotetrameric, calcium-permeable AMPARs, determined by single-particle electron microscopy of untagged and fluorescently tagged constructs in a ligand-free state. It reveals a novel packing of NTD dimers, and a separation of LBD dimers across a central vestibule. In this arrangement, which reconciles diverse functional observations, agonist-induced cleft closure across LBD dimers can be converted into a twisting motion that provides a basis for receptor activation.

  14. International Union of Basic and Clinical Pharmacology. XC. multisite pharmacology: recommendations for the nomenclature of receptor allosterism and allosteric ligands.

    Science.gov (United States)

    Christopoulos, Arthur; Changeux, Jean-Pierre; Catterall, William A; Fabbro, Doriano; Burris, Thomas P; Cidlowski, John A; Olsen, Richard W; Peters, John A; Neubig, Richard R; Pin, Jean-Philippe; Sexton, Patrick M; Kenakin, Terry P; Ehlert, Frederick J; Spedding, Michael; Langmead, Christopher J

    2014-10-01

    Allosteric interactions play vital roles in metabolic processes and signal transduction and, more recently, have become the focus of numerous pharmacological studies because of the potential for discovering more target-selective chemical probes and therapeutic agents. In addition to classic early studies on enzymes, there are now examples of small molecule allosteric modulators for all superfamilies of receptors encoded by the genome, including ligand- and voltage-gated ion channels, G protein-coupled receptors, nuclear hormone receptors, and receptor tyrosine kinases. As a consequence, a vast array of pharmacologic behaviors has been ascribed to allosteric ligands that can vary in a target-, ligand-, and cell-/tissue-dependent manner. The current article presents an overview of allostery as applied to receptor families and approaches for detecting and validating allosteric interactions and gives recommendations for the nomenclature of allosteric ligands and their properties.

  15. Effect of adenosine receptors on 3, 4 methylene dioxy methamphetamine induced hyperthermic, neuroinflammatory and neurotoxic effects in mouse brain

    OpenAIRE

    Khairnar, Amit S.

    2010-01-01

    Previous studies of ours and other groups in mice have shown that 3, 4 Methylenedioxymethamphetamine (MDMA, ecstasy) produces neurotoxic damage to dopaminergic neurons and neuroinflammation and caffeine, an adenosine A1/A2A antagonist enhances glial activation induced by MDMA, suggesting potential facilitation of neurodegenerative processes. In the present study we want to investigate effect of caffeine on MDMA induced dopaminergic neurotoxicity in adult mice, whereas selective A1 ( DPCPX ) a...

  16. Regulation of adenosine levels during cerebral ischemia

    Institute of Scientific and Technical Information of China (English)

    Stephanie CHU; Wei XIONG; Dali ZHANG; Hanifi SOYLU; Chao SUN; Benedict C ALBENSI; Fiona E PARKINSON

    2013-01-01

    Adenosine is a neuromodulator with its level increasing up to 100-fold during ischemic events,and attenuates the excitotoxic neuronal injury.Adenosine is produced both intracellularly and extracellularly,and nucleoside transport proteins transfer adenosine across plasma membranes.Adenosine levels and receptor-mediated effects of adenosine are regulated by intracellular ATP consumption,cellular release of ATP,metabolism of extracellular ATP (and other adenine nucleotides),adenosine influx,adenosine efflux and adenosine metabolism.Recent studies have used genetically modified mice to investigate the relative contributions of intra-and extracellular pathways for adenosine formation.The importance of cortical or hippocampal neurons as a source or a sink of adenosine under basal and hypoxic/ischemic conditions was addressed through the use of transgenic mice expressing human equilibrative nucleoside transporter 1 (hENT1) under the control of a promoter for neuron-specific enolase.From these studies,we conclude that ATP consumption within neurons is the primary source of adenosine in neuronal cultures,but not in hippocampal slices or in vivo mice exposed to ischemic conditions.

  17. ONRLDB--manually curated database of experimentally validated ligands for orphan nuclear receptors: insights into new drug discovery.

    Science.gov (United States)

    Nanduri, Ravikanth; Bhutani, Isha; Somavarapu, Arun Kumar; Mahajan, Sahil; Parkesh, Raman; Gupta, Pawan

    2015-01-01

    Orphan nuclear receptors are potential therapeutic targets. The Orphan Nuclear Receptor Ligand Binding Database (ONRLDB) is an interactive, comprehensive and manually curated database of small molecule ligands targeting orphan nuclear receptors. Currently, ONRLDB consists of ∼11,000 ligands, of which ∼6500 are unique. All entries include information for the ligand, such as EC50 and IC50, number of aromatic rings and rotatable bonds, XlogP, hydrogen donor and acceptor count, molecular weight (MW) and structure. ONRLDB is a cross-platform database, where either the cognate small molecule modulators of a receptor or the cognate receptors to a ligand can be searched. The database can be searched using three methods: text search, advanced search or similarity search. Substructure search, cataloguing tools, and clustering tools can be used to perform advanced analysis of the ligand based on chemical similarity fingerprints, hierarchical clustering, binning partition and multidimensional scaling. These tools, together with the Tree function provided, deliver an interactive platform and a comprehensive resource for identification of common and unique scaffolds. As demonstrated, ONRLDB is designed to allow selection of ligands based on various properties and for designing novel ligands or to improve the existing ones. Database URL: http://www.onrldb.org/.

  18. Ligand-receptor binding kinetics in surface plasmon resonance cells: A Monte Carlo analysis

    CERN Document Server

    Carroll, Jacob; Forsten-Williams, Kimberly; Täuber, Uwe C

    2016-01-01

    Surface plasmon resonance (SPR) chips are widely used to measure association and dissociation rates for the binding kinetics between two species of chemicals, e.g., cell receptors and ligands. It is commonly assumed that ligands are spatially well mixed in the SPR region, and hence a mean-field rate equation description is appropriate. This approximation however ignores the spatial fluctuations as well as temporal correlations induced by multiple local rebinding events, which become prominent for slow diffusion rates and high binding affinities. We report detailed Monte Carlo simulations of ligand binding kinetics in an SPR cell subject to laminar flow. We extract the binding and dissociation rates by means of the techniques frequently employed in experimental analysis that are motivated by the mean-field approximation. We find major discrepancies in a wide parameter regime between the thus extracted rates and the known input simulation values. These results underscore the crucial quantitative importance of s...

  19. The phosphatase domains of CD45 are required for ligand induced T-cell receptor downregulation

    DEFF Research Database (Denmark)

    Kastrup, J; Lauritsen, Jens Peter Holst; Menné, C;

    2000-01-01

    Down-regulation of the T-cell receptor (TCR) plays an important role in modulating T-cell responses, both during T-cell development and in mature T cells. At least two distinct pathways exist for TCR down-regulation: down-regulation following TCR ligation; and down-regulation following activation...... of protein kinase C (PKC). Ligand-induced TCR down-regulation is dependent on protein tyrosine kinase (PTK) activity and seems to be closely related to T-cell activation. In addition, previous studies have indicated that ligand-induced TCR down-regulation is dependent on the expression of CD45, a...... transmembrane protein tyrosine phosphatase. The role of the different domains of CD45 in TCR down-regulation was investigated in this study. We found that the phosphatase domains of CD45 are required for efficient ligand-induced TCR down-regulation. In contrast, the extracellular domain of CD45 is dispensable...

  20. Neuroprotective and anti-inflammatory effects of the adenosine A(2A) receptor antagonist ST1535 in a MPTP mouse model of Parkinson's disease.

    Science.gov (United States)

    Frau, Lucia; Borsini, Franco; Wardas, Jadwiga; Khairnar, Amit S; Schintu, Nicoletta; Morelli, Micaela

    2011-03-01

    Adenosine A(2A) receptor antagonists are one of the most attractive classes of drug for the treatment of Parkinson's disease (PD) as they are effective in counteracting motor dysfunctions and display neuroprotective and anti-inflammatory effects in animal models of PD. In this study, we evaluated the neuroprotective and anti-inflammatory properties of the adenosine A(2A) receptor antagonist ST1535 in a subchronic 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) mouse model of PD. C57BL/6J mice were repeatedly administered with vehicle, MPTP (20 mg/kg), or MPTP + ST1535 (2 mg/kg). Mice were sacrificed three days after the last administration of MPTP. Immunohistochemistry for tyrosine hydroxylase (TH) and cresyl violet staining were employed to evaluate dopaminergic neuron degeneration in the substantia nigra pars compacta (SNc) and caudate-putamen (CPu). CD11b and glial fibrillary acidic protein (GFAP) immunoreactivity were, respectively, evaluated as markers of microglial and astroglial response in the SNc and CPu. Stereological analysis for TH revealed a 32% loss of dopaminergic neurons in the SNc after repeated MPTP administration, which was completely prevented by ST1535 coadministration. Similarly, CPu decrease in TH (25%) was prevented by ST1535. MPTP treatment induced an intense gliosis in both the SNc and CPu. ST1535 totally prevented CD11b immunoreactivity in both analyzed areas, but only partially blocked GFAP increase in the SNc and CPu. A(2A) receptor antagonism is a new opportunity for improving symptomatic PD treatment. With its neuroprotective effect on dopaminergic neuron toxicity induced by MPTP and its antagonism on glial activation, ST1535 represents a new prospect for a disease-modifying drug. PMID:20665698

  1. Labeling of receptor ligands with bromine radionuclides. Progress report, March 1, 1981-February 28, 1982

    International Nuclear Information System (INIS)

    In recent years there has been an interest in the use of various radioisotopes of bromine as labels for radiopharmaceuticals. Although radioisotopes of iodine have been used extensively as radiopharmaceutical labels, there are several advantages associated with the use of radiobromine as a label, due primarily to increased stability of bonds to the radiohalide and smaller steric perturbation resulting from substitution of the radiohalide. Methods of attaching radiobromine to receptor ligands with the potential of mapping estrogen receptors in mammary tumors and uteri were studied. Two ligands were studied extensively in vitro and in animal models; preliminary studies were also carried out in humans. To date, the only radioisotope of bromine used was bromine-77. In addition, a series of model compounds were labeled with bromine-77 using a recently described method for rapid bromination; the scope and limitations of this new rapid radiobromination technique were evaluated

  2. Structural Basis for Hydroxycholesterols as Natural Ligands of Orphan Nuclear Receptor ROR[gamma

    Energy Technology Data Exchange (ETDEWEB)

    Jin, Lihua; Martynowski, Dariusz; Zheng, Songyang; Wada, Taira; Xie, Wen; Li, Yong (Pitt); (Xiamen)

    2010-09-03

    The retinoic acid-related orphan receptor {gamma} (ROR{gamma}) has important roles in development and metabolic homeostasis. Although the biological functions of ROR{gamma} have been studied extensively, no ligands for ROR{gamma} have been identified, and no structure of ROR{gamma} has been reported. In this study, we showed that hydroxycholesterols promote the recruitment of coactivators by ROR{gamma} using biochemical assays. We also report the crystal structures of the ROR{gamma} ligand-binding domain bound with hydroxycholesterols. The structures reveal the binding modes of various hydroxycholesterols in the ROR{gamma} pocket, with the receptors all adopting the canonical active conformation. Mutations that disrupt the binding of hydroxycholesterols abolish the constitutive activity of ROR{gamma}. Our observations suggest an important role for the endogenous hydroxycholesterols in modulating ROR{gamma}-dependent biological processes.

  3. Communication: Free energy of ligand-receptor systems forming multimeric complexes

    Science.gov (United States)

    Di Michele, Lorenzo; Bachmann, Stephan J.; Parolini, Lucia; Mognetti, Bortolo M.

    2016-04-01

    Ligand-receptor interactions are ubiquitous in biology and have become popular in materials in view of their applications to programmable self-assembly. Although complex functionalities often emerge from the simultaneous interaction of more than just two linker molecules, state of the art theoretical frameworks enable the calculation of the free energy only in systems featuring one-to-one ligand/receptor binding. In this Communication, we derive a general formula to calculate the free energy of systems featuring simultaneous direct interaction between an arbitrary number of linkers. To exemplify the potential and generality of our approach, we apply it to the systems recently introduced by Parolini et al. [ACS Nano 10, 2392 (2016)] and Halverson and Tkachenko [J. Chem. Phys. 144, 094903 (2016)], both featuring functionalized Brownian particles interacting via three-linker complexes.

  4. The Oligomeric States of the Purified Sigma-1 Receptor Are Stabilized by Ligands*

    Science.gov (United States)

    Gromek, Katarzyna A.; Suchy, Fabian P.; Meddaugh, Hannah R.; Wrobel, Russell L.; LaPointe, Loren M.; Chu, Uyen B.; Primm, John G.; Ruoho, Arnold E.; Senes, Alessandro; Fox, Brian G.

    2014-01-01

    Sigma-1 receptor (S1R) is a mammalian member of the ERG2 and sigma-1 receptor-like protein family (pfam04622). It has been implicated in drug addiction and many human neurological disorders, including Alzheimer and Parkinson diseases and amyotrophic lateral sclerosis. A broad range of synthetic small molecules, including cocaine, (+)-pentazocine, haloperidol, and small endogenous molecules such as N,N-dimethyltryptamine, sphingosine, and steroids, have been identified as regulators of S1R. However, the mechanism of activation of S1R remains obscure. Here, we provide evidence in vitro that S1R has ligand binding activity only in an oligomeric state. The oligomeric state is prone to decay into an apparent monomeric form when exposed to elevated temperature, with loss of ligand binding activity. This decay is suppressed in the presence of the known S1R ligands such as haloperidol, BD-1047, and sphingosine. S1R has a GXXXG motif in its second transmembrane region, and these motifs are often involved in oligomerization of membrane proteins. Disrupting mutations within the GXXXG motif shifted the fraction of the higher oligomeric states toward smaller states and resulted in a significant decrease in specific (+)-[3H]pentazocine binding. Results presented here support the proposal that S1R function may be regulated by its oligomeric state. Possible mechanisms of molecular regulation of interacting protein partners by S1R in the presence of small molecule ligands are discussed. PMID:24847081

  5. The oligomeric states of the purified sigma-1 receptor are stabilized by ligands.

    Science.gov (United States)

    Gromek, Katarzyna A; Suchy, Fabian P; Meddaugh, Hannah R; Wrobel, Russell L; LaPointe, Loren M; Chu, Uyen B; Primm, John G; Ruoho, Arnold E; Senes, Alessandro; Fox, Brian G

    2014-07-18

    Sigma-1 receptor (S1R) is a mammalian member of the ERG2 and sigma-1 receptor-like protein family (pfam04622). It has been implicated in drug addiction and many human neurological disorders, including Alzheimer and Parkinson diseases and amyotrophic lateral sclerosis. A broad range of synthetic small molecules, including cocaine, (+)-pentazocine, haloperidol, and small endogenous molecules such as N,N-dimethyltryptamine, sphingosine, and steroids, have been identified as regulators of S1R. However, the mechanism of activation of S1R remains obscure. Here, we provide evidence in vitro that S1R has ligand binding activity only in an oligomeric state. The oligomeric state is prone to decay into an apparent monomeric form when exposed to elevated temperature, with loss of ligand binding activity. This decay is suppressed in the presence of the known S1R ligands such as haloperidol, BD-1047, and sphingosine. S1R has a GXXXG motif in its second transmembrane region, and these motifs are often involved in oligomerization of membrane proteins. Disrupting mutations within the GXXXG motif shifted the fraction of the higher oligomeric states toward smaller states and resulted in a significant decrease in specific (+)-[(3)H]pentazocine binding. Results presented here support the proposal that S1R function may be regulated by its oligomeric state. Possible mechanisms of molecular regulation of interacting protein partners by S1R in the presence of small molecule ligands are discussed. PMID:24847081

  6. Peroxisome proliferator-activated receptor ligands as antiatherogenic agents: panacea or another Pandora's box?

    Science.gov (United States)

    Molavi, Behzad; Rasouli, Neda; Mehta, Jawahar L

    2002-01-01

    Peroxisome proliferator activated receptors (PPARs) are members of the nuclear receptor super family that modulate gene expression upon ligand activation. They are 3 major subtypes of PPARs: alpha, delta (also called beta), and gamma. PPAR-gamma is widely expressed in the cardiovascular system and is involved in the regulation of tissue inflammation and smooth muscle cell growth pathways as well as in lipoprotein metabolism and coagulation cascades. PPAR-gamma ligands of (e.g., rosigitazone and pioglitazone) have been shown to exert antiatherogenic effects both in vitro and in vivo. PPAR-alpha ligands (e.g., clofibrate and benzofibrate) modulate lipoprotein metabolism, and affect inflammation and coagulation cascade. These effects may be helpful in resolving the dilemma arising from studies that showed significant mortality and morbidity benefits of fibrates in the face of minimal changes in HDL-cholesterol levels. The role of PPAR-delta in atherogenesis remains largely unknown, although it appears that PPAR-delta activation affects lipoprotein metabolism. PPAR ligands appear to be promising agents in limiting atherosclerosis; however, large-scale clinical trials are required to assess their safety and efficacy before they can be added to the clinicians' arsenal of antiatherosclerotic agents. PMID:12000972

  7. Effects of different ligands on epidermal growth factor receptor (EGFR) nuclear translocation.

    Science.gov (United States)

    Faria, Jerusa A Q A; de Andrade, Carolina; Goes, Alfredo M; Rodrigues, Michele A; Gomes, Dawidson A

    2016-09-01

    The epidermal growth factor receptor (EGFR) is activated through binding to specific ligands and generates signals for proliferation, differentiation, migration, and cell survival. Recent data show the role of nuclear EGFR in tumors. Although many EGFR ligands are upregulated in cancers, little is known about their effects on EGFR nuclear translocation. We have compared the effects of six EGFR ligands (EGF, HB-EGF, TGF-α, β-Cellulin, amphiregulin, and epiregulin) on nuclear translocation of EGFR, receptor phosphorylation, migration, and proliferation. Cell fractionation and confocal immunofluorescence detected EGFR in the nucleus after EGF, HB-EGF, TGF-α and β-Cellulin stimulation in a dose-dependent manner. In contrast, amphiregulin and epiregulin did not generate nuclear translocation of EGFR. EGF, HB-EGF, TGF-α and β-Cellulin showed correlations between a higher rate of wound closure and increased phosphorylation of residues in the carboxy-terminus of EGFR, compared to amphiregulin and epiregulin. The data indicate that EGFR is translocated to the nucleus after stimulation with EGF, HB-EGF, TGF-α and β-Cellulin, and that these ligands are related to increased phosphorylation of EGFR tyrosine residues, inducing migration of SkHep-1 cells. PMID:27462018

  8. Effects of coumestrol on lipid and glucose metabolism as a farnesoid X receptor ligand

    International Nuclear Information System (INIS)

    In the course of an effort to identify novel agonists of the farnesoid X receptor (FXR), coumestrol was determined to be one such ligand. Reporter and in vitro coactivator interaction assays revealed that coumestrol bound and activated FXR. Treatment of Hep G2 cells with coumestrol stimulated the expression of FXR target genes, thereby regulating the expression of target genes of the liver X receptor and hepatocyte nuclear factor-4α. Through these actions, coumestrol is expected to exert beneficial effects on lipid and glucose metabolism

  9. Antianxiety and antidepressant-like effects of AC-5216, a novel mitochondrial benzodiazepine receptor ligand

    OpenAIRE

    Kita, Atsuko; Kohayakawa, Hitoshi; Kinoshita, Tomoko; Ochi, Yoshiaki; Nakamichi, Keiko; Kurumiya, Satoshi; Furukawa, Kiyoshi; Oka, Makoto

    2004-01-01

    We investigated the ability of N-benzyl-N-ethyl-2-(7,8-dihydro-7-methyl-8-oxo-2-phenyl-9H-purin-9-yl)acetamide (AC-5216), a novel mitochondrial benzodiazepine receptor (MBR) ligand, to produce anti-anxiety and antidepressant-like effects in various animal models.AC-5216 showed high affinity for MBRs prepared from rat whole brain (Ki 0.297 nM), rat glioma cells (IC50 3.04 nM) and human glioma cells (IC50 2.73 nM), but only negligible affinity for the other main receptors including central benz...

  10. Differential effects of EGFR ligands on endocytic sorting of the receptor

    DEFF Research Database (Denmark)

    Roepstorff, Kirstine; Grandal, Michael Vibo; Henriksen, Lasse;

    2009-01-01

    Endocytic downregulation is a pivotal mechanism turning off signalling from the EGF receptor (EGFR). It is well established that whereas EGF binding leads to lysosomal degradation of EGFR, transforming growth factor (TGF)-alpha causes receptor recycling. TGF-alpha therefore leads to continuous...... recycling. EGF leads to lysosomal degradation of the majority but not all EGFRs. Amphiregulin does not target EGFR for lysosomal degradation but causes fast as well as slow EGFR recycling. The Cbl ubiquitin ligases, especially c-Cbl, are responsible for EGFR ubiquitination after stimulation with all ligands...

  11. Ligand-induced Coupling versus Receptor Pre-association: Cellular automaton simulations of FGF-2 binding

    OpenAIRE

    Gopalakrishnan, Manoj; Forsten-Williams, Kimberly; Tauber, Uwe C.

    2003-01-01

    The binding of basic fibroblast growth factor (FGF-2) to its cell surface receptor (CSR) and subsequent signal transduction is known to be enhanced by Heparan Sulfate Proteoglycans (HSPGs). HSPGs bind FGF-2 with low affinity and likely impact CSR-mediated signaling via stabilization of FGF-2-CSR complexes via association with both the ligand and the receptor. What is unknown is whether HSPG associates with CSR in the absence of FGF-2. In this paper, we determine conditions by which pre-associ...

  12. Structures of pattern recognition receptors reveal molecular mechanisms of autoinhibition, ligand recognition and oligomerization.

    Science.gov (United States)

    Chuenchor, Watchalee; Jin, Tengchuan; Ravilious, Geoffrey; Xiao, T Sam

    2014-02-01

    Pattern recognition receptors (PRRs) are essential sentinels for pathogens or tissue damage and integral components of the innate immune system. Recent structural studies have provided unprecedented insights into the molecular mechanisms of ligand recognition and signal transduction by several PRR families at distinct subcellular compartments. Here we highlight some of the recent discoveries and summarize the common themes that are emerging from these exciting studies. Better mechanistic understanding of the structure and function of the PRRs will improve future prospects of therapeutic targeting of these important innate immune receptors.

  13. High-throughput screening assay for new ligands at human melatonin receptors

    Institute of Scientific and Technical Information of China (English)

    Jian-hua YAN; Hao-ran SU; Jean A BOUTIN; M Pierre RENARD; Ming-wei WANG

    2008-01-01

    Aim: Melatonin (MT) is a neurohormone produced and secreted primarily by the pineal gland in a circadian manner, and mainly acta through 2 receptor subtypes: MT1 and MT2 in humans. The diversity in their tissue distribution is in favor of different functions for each receptor subtype. Selective modulators are therefore required to determine the physiological roles of these melatonin receptor sub-types and their implications in pathological processes. Methods: A homogenous MT1/MT2 receptor binding assay was established for high-throughput screening of new ligands at the hMT1 and/or hMT2 receptors. The functional properties (agonists or antagonists) were assessed by a conventional guanosine-5'[γ-35S] triphosphate (GTP-γS) assay. Results: Three hMT, receptor-selective small mol-ecule antagonists and 1 hMT2 receptor-selective small molecule antagonist with novel structural features were identified following a high-throughput screening campaign of 48 240 synthetic and natural compounds. Conclusion: The findings may assist in the expansion of chemical probes to these 2 receptor subtypes.

  14. The Prelude on Novel Receptor and Ligand Targets Involved in the Treatment of Diabetes Mellitus

    Directory of Open Access Journals (Sweden)

    Venu Gopal Jonnalagadda

    2014-05-01

    Full Text Available Metabolic disorders are a group of disorders, due to the disruption of the normal metabolic process at a cellular level. Diabetes Mellitus and Tyrosinaemia are the majorly reported metabolic disorders. Among them, Diabetes Mellitus is a one of the leading metabolic syndrome, affecting 5 to 7 % of the population worldwide and mainly characterised by elevated levels of glucose and is associated with two types of physiological event disturbances such as impaired insulin secretion and insulin resistance. Up to now, various treatment strategies are like insulin, alphaglucosidase inhibitors, biguanides, incretins were being followed. Concurrently, various novel therapeutic strategies are required to advance the therapy of Diabetes mellitus. For the last few decades, there has been an extensive research in understanding the metabolic pathways involved in Diabetes Mellitus at the cellular level and having the profound knowledge on cell-growth, cell-cycle, and apoptosis at a molecular level provides new targets for the treatment of Diabetes Mellitus. Receptor signalling has been involved in these mechanisms, to translate the information coming from outside. To understand the various receptors involved in these pathways, we must have a sound knowledge on receptors and ligands involved in it. This review mainly summarises the receptors and ligands which are involved the Diabetes Mellitus. Finally, researchers have to develop the alternative chemical moieties that retain their affinity to receptors and efficacy. Diabetes Mellitus being a metabolic disorder due to the glucose surfeit, demands the need for regular exercise along with dietary changes.

  15. HER2-mediated anticancer drug delivery: strategies to prepare targeting ligands highly specific for the receptor.

    Science.gov (United States)

    Calce, Enrica; Monfregola, Luca; Saviano, Michele; De Luca, Stefania

    2015-01-01

    HER2 receptor, for its involvement in tumorigenesis, has been largely studied as topic in cancer research. In particular, the employment of trastuzumab (Herceptin), a humanized anti-HER2 antibody, showed several clinical benefits in the therapy against the breast cancer. Moreover, for its accessible extracellular domain, this receptor is considered an ideal target to deliver anticancer drugs for the receptormediated anticancer therapy. By now, monoclonal antibody and its fragments, affibody, and some peptides have been employed as targeting agents in order to deliver various drugs to HER2 positive tumor cells. In particular, the ability to perform a fast and reliable screening of a large number of peptide molecules would make possible the selection of highly specific compounds to the receptor target. In this regard, the availability of preparing a simplified synthetic model which is a good mimetic of the receptor target and can be used in a reliable screening method of ligands would be of a strategic importance for the development of selective HER2-targeting peptide molecules. Herein, we illustrate the importance of HER2-targeted anticancer therapies. We also report on a synthetic and effective mimetic of the receptor, which revealed to be a useful tool for the selection of specific HER2 ligands. PMID:25994863

  16. Estrogen receptor determination in endometrial carcinoma: ligand binding assay versus enzyme immunoassay

    DEFF Research Database (Denmark)

    Nyholm, H C; Nielsen, Anette Lynge; Lyndrup, J;

    1995-01-01

    We compared concentrations of cytosolic estrogen receptors (ERc) measured in 35 postmenopausal endometrial carcinomas by ligand binding method (LBA) (dextran-coated charcoal assay) and enzyme immunoassay (EIA). Correlations between ERc, nuclear estrogen receptors (ERn) determined by EIA, and cyto......We compared concentrations of cytosolic estrogen receptors (ERc) measured in 35 postmenopausal endometrial carcinomas by ligand binding method (LBA) (dextran-coated charcoal assay) and enzyme immunoassay (EIA). Correlations between ERc, nuclear estrogen receptors (ERn) determined by EIA......, and cytosolic progesterone receptors (PR) measured by LBA were also studied. While ERc concentrations determined by LBA and EIA were highly correlated (r: 0.94), ERc values detected by LBA were approximately twice those found by EIA (median values of ERc: 155 vs. 64 fmol/mg cytosol protein, DCC vs. EIA......). The percentages of ERc positive tumors were 89% by LBA and 77% by EIA. The median fraction of total ER present as ERn was 63%. PR levels correlated positively with ERn concentrations (r: 0.73). We explore possible reasons why greater concentrations of ERc are determined by estradiol binding than by the ER-EIA kit...

  17. Synthesis and radiofluorination of putative NMDA receptor ligands

    International Nuclear Information System (INIS)

    In the course of this work on the synthesis of radioligands for the NMDA receptor the authentic standards and labeling precursors of four compounds with an amidine structure was performed. Synthesis of the precursors followed reaction conditions given in the literature and was successful. The imidoesters used for the synthesis were obtained from their nitriles in a Pinner synthesis, while 2-hydroxybenzylamine was synthesized in a reduction of 2-hydroxybenzonitrile using borane as a reducing agent. After a coupling reaction of the amine and the imidoester in DMF using triethylamine as base the precursors were obtained in good yields and purified by crystallization from methanol. The cyclic standard compound was synthesized directly from 2-(bromomethyl)- benzonitrile and 2-hydroxybenzylamine in a ring closing reaction. Similar to the other precursors, crystallization from methanol produced a pure compound. The authentic standards were synthesized starting from salicylaldehyde. In a four step synthesis the desired ortho-fluoroethoxybenzylamine was obtained in good yield. Coupling of the amine with the respective imidoester or in the case of the cyclic compound 2-(bromomethyl)-benzonitrile gave the desired product which was then purified by column chromatography or by crystallization from ethanol and water. For the labeling procedure 1-bromo-2-[18F]fluoroethane was synthesized following a previously published pathway starting from 1,2-dibromoethane. An alternative route of radiosynthesis for this prosthetic group was tested using ethyleneglycole- 1,2-ditosylate. The labeling reaction was performed on one of the precursors testing both DMF and DMSO as solvents and using NaOH as base. Yields of N-(2-fluoroethoxybenzyl)- cinnamamidine were about 78 % at 80 C after 30 minutes in DMSO. The desired product can now be synthesized in sufficient yields for in vitro and in vivo evaluation studies. Labeling on the cyclic precursor was attempted utilizing DMSO as solvent, but no

  18. Synthesis and radiofluorination of putative NMDA receptor ligands

    Energy Technology Data Exchange (ETDEWEB)

    Kronenberg, U.

    2011-01-15

    In the course of this work on the synthesis of radioligands for the NMDA receptor the authentic standards and labeling precursors of four compounds with an amidine structure was performed. Synthesis of the precursors followed reaction conditions given in the literature and was successful. The imidoesters used for the synthesis were obtained from their nitriles in a Pinner synthesis, while 2-hydroxybenzylamine was synthesized in a reduction of 2-hydroxybenzonitrile using borane as a reducing agent. After a coupling reaction of the amine and the imidoester in DMF using triethylamine as base the precursors were obtained in good yields and purified by crystallization from methanol. The cyclic standard compound was synthesized directly from 2-(bromomethyl)- benzonitrile and 2-hydroxybenzylamine in a ring closing reaction. Similar to the other precursors, crystallization from methanol produced a pure compound. The authentic standards were synthesized starting from salicylaldehyde. In a four step synthesis the desired ortho-fluoroethoxybenzylamine was obtained in good yield. Coupling of the amine with the respective imidoester or in the case of the cyclic compound 2-(bromomethyl)-benzonitrile gave the desired product which was then purified by column chromatography or by crystallization from ethanol and water. For the labeling procedure 1-bromo-2-[{sub 18}F]fluoroethane was synthesized following a previously published pathway starting from 1,2-dibromoethane. An alternative route of radiosynthesis for this prosthetic group was tested using ethyleneglycole- 1,2-ditosylate. The labeling reaction was performed on one of the precursors testing both DMF and DMSO as solvents and using NaOH as base. Yields of N-(2-fluoroethoxybenzyl)- cinnamamidine were about 78 % at 80 C after 30 minutes in DMSO. The desired product can now be synthesized in sufficient yields for in vitro and in vivo evaluation studies. Labeling on the cyclic precursor was attempted utilizing DMSO as solvent

  19. Increased Signaling via Adenosine A1 Receptors, Sleep Deprivation, Imipramine, and Ketamine Inhibit Depressive-like Behavior via Induction of Homer1a.

    Science.gov (United States)

    Serchov, Tsvetan; Clement, Hans-Willi; Schwarz, Martin K; Iasevoli, Felice; Tosh, Dilip K; Idzko, Marco; Jacobson, Kenneth A; de Bartolomeis, Andrea; Normann, Claus; Biber, Knut; van Calker, Dietrich

    2015-08-01

    Major depressive disorder is among the most commonly diagnosed disabling mental diseases. Several non-pharmacological treatments of depression upregulate adenosine concentration and/or adenosine A1 receptors (A1R) in the brain. To test whether enhanced A1R signaling mediates antidepressant effects, we generated a transgenic mouse with enhanced doxycycline-regulated A1R expression, specifically in forebrain neurons. Upregulating A1R led to pronounced acute and chronic resilience toward depressive-like behavior in various tests. Conversely, A1R knockout mice displayed an increased depressive-like behavior and were resistant to the antidepressant effects of sleep deprivation (SD). Various antidepressant treatments increase homer1a expression in medial prefrontal cortex (mPFC). Specific siRNA knockdown of homer1a in mPFC enhanced depressive-like behavior and prevented the antidepressant effects of A1R upregulation, SD, imipramine, and ketamine treatment. In contrast, viral overexpression of homer1a in the mPFC had antidepressant effects. Thus, increased expression of homer1a is a final common pathway mediating the antidepressant effects of different antidepressant treatments. PMID:26247862

  20. Natural ligands of nuclear receptors. Isolation, design, synthesis, biochemical decodification and potential therapeutic applications.

    OpenAIRE

    Ummarino, Raffaella

    2013-01-01

    Natural products have historically been a rich source of lead compounds in drug discovery. The biochemical investigation of marine organisms, through the deep collaboration between chemists and pharmacologists, focused on searching of new biologically active compounds, is a central issue of this kind of studies. My research work, described in this PhD thesis, has been developed in this research area and was addressed to the identification of new ligands of nuclear receptors, discovering ...

  1. Arrest functions of the MIF ligand/receptor axes in atherogenesis

    Directory of Open Access Journals (Sweden)

    Sabine eTillmann

    2013-05-01

    Full Text Available Macrophage migration inhibitory factor (MIF has been defined as an important chemokine-like function (CLF chemokine with an essential role in monocyte recruitment and arrest. Adhesion of monocytes to the vessel wall and their transendothelial migration are critical in atherogenesis and many other inflammatory diseases. Chemokines carefully control all steps of the monocyte recruitment process. Those chemokines specialized in controlling arrest are typically immobilized on the endothelial surface, mediating the arrest of rolling monocytes by chemokine receptor-triggered pathways. The chemokine receptor CXCR2 functions as an important arrest receptor on monocytes. An arrest function has been revealed for the bona fide CXCR2 ligands CXCL1 and CXCL8, but genetic studies also suggested that additional arrest chemokines are likely to be involved in atherogenic leukocyte recruitment. While CXCR2 is known to interact with numerous CXC chemokine ligands, the CLF-chemokine MIF, which structurally does not belong to the CXC chemokine sub-family, was surprisingly identified as a non-cognate ligand of CXCR2, responsible for critical arrest functions during the atherogenic process. MIF was originally identified as macrophage migration inhibitory factor, but is now known as a potent inflammatory cytokine with chemokine-like functions including chemotaxis and leukocyte arrest. This review will cover the mechanisms underlying these functions, including MIF’s effects on LFA1 integrin activity and signal transduction, and will discuss the structural similarities between MIF and the bona fide CXCR2 ligand CXCL8 while emphasizing the structural differences. As MIF also interacts with CXCR4, a chemokine receptor implicated in CXCL12-elicited lymphocyte arrest, the arrest potential of the MIF/CXCR4 axis will also be scrutinized as well as the recently identified role of pericyte MIF in attracting leukocytes exiting through venules as part of the pericyte 'motility

  2. Dopamine D3 receptor ligands for drug addiction treatment: update on recent findings.

    Science.gov (United States)

    Le Foll, Bernard; Collo, Ginetta; Rabiner, Eugenii A; Boileau, Isabelle; Merlo Pich, Emilio; Sokoloff, Pierre

    2014-01-01

    The dopamine D3 receptor is located in the limbic area and apparently mediates selective effects on motivation to take drugs and drug-seeking behaviors, so that there has been considerable interest on the possible use of D3 receptor ligands to treat drug addiction. However, only recently selective tools allowing studying this receptor have been developed. This chapter presents an overview of findings that were presented at a symposium on the conference Dopamine 2013 in Sardinia in May 2013. Novel neurobiological findings indicate that drugs of abuse can lead to significant structural plasticity in rodent brain and that this is dependent on the availability of functional dopamine D3 autoreceptor, whose activation increased phosphorylation in the ERK pathway and in the Akt/mTORC1 pathway indicating the parallel engagement of a series of intracellular signaling pathways all involved in cell growth and survival. Preclinical findings using animal models of drug-seeking behaviors confirm that D3 antagonists have a promising profile to treat drug addiction across drugs of abuse type. Imaging the D3 is now feasible in human subjects. Notably, the development of (+)-4-propyl-9-hydroxynaphthoxazine ligand used in positron emission tomography (PET) studies in humans allows to measure D3 and D2 receptors based on the area of the brain under study. This PET ligand has been used to confirm up-regulation of D3 sites in psychostimulant users and to reveal that tobacco smoking produces elevation of dopamine at the level of D3 sites. There are now novel antagonists being developed, but also old drugs such as buspirone, that are available to test the D3 hypothesis in humans. The first results of clinical investigations are now being provided. Overall, those recent findings support further exploration of D3 ligands to treat drug addiction. PMID:24968784

  3. Functional Selectivity and Antidepressant Activity of Serotonin 1A Receptor Ligands

    Directory of Open Access Journals (Sweden)

    Zdzisław Chilmonczyk

    2015-08-01

    Full Text Available Serotonin (5-HT is a monoamine neurotransmitter that plays an important role in physiological functions. 5-HT has been implicated in sleep, feeding, sexual behavior, temperature regulation, pain, and cognition as well as in pathological states including disorders connected to mood, anxiety, psychosis and pain. 5-HT1A receptors have for a long time been considered as an interesting target for the action of antidepressant drugs. It was postulated that postsynaptic 5-HT1A agonists could form a new class of antidepressant drugs, and mixed 5-HT1A receptor ligands/serotonin transporter (SERT inhibitors seem to possess an interesting pharmacological profile. It should, however, be noted that 5-HT1A receptors can activate several different biochemical pathways and signal through both G protein-dependent and G protein-independent pathways. The variables that affect the multiplicity of 5-HT1A receptor signaling pathways would thus result from the summation of effects specific to the host cell milieu. Moreover, receptor trafficking appears different at pre- and postsynaptic sites. It should also be noted that the 5-HT1A receptor cooperates with other signal transduction systems (like the 5-HT1B or 5-HT2A/2B/2C receptors, the GABAergic and the glutaminergic systems, which also contribute to its antidepressant and/or anxiolytic activity. Thus identifying brain specific molecular targets for 5-HT1A receptor ligands may result in a better targeting, raising a hope for more effective medicines for various pathologies.

  4. Adenosine A{sub 1} receptors in human sleep regulation studied by electroencephalography (EEG) and positron emission tomography (PET)[Dissertation 17227

    Energy Technology Data Exchange (ETDEWEB)

    Geissler, E

    2007-07-01

    Sleep is an essential physiological process. However, the functions of sleep and the endogenous mechanisms involved in sleep regulation are only partially understood. Convergent lines of evidence support the hypothesis that the build-up of sleep propensity during wakefulness and its decline during sleep are associated with alterations in brain adenosine levels and adenosine receptor concentrations. The non-selective A{sub 1} and A{sub 2A} adenosine receptor antagonist caffeine stimulates alertness and is known to attenuate changes in the waking and sleep electroencephalogram (EEG) typically observed after prolonged waking. Several findings point to an important function of the adenosine A{sub 1} receptor (A{sub 1}AR) in the modulation of vigilance states. The A{sub 1}AR is densely expressed in brain regions involved in sleep regulation, and pharmacological manipulations affecting the A{sub 1}AR were shown to influence sleep propensity and sleep depth. However, an involvement of the A{sub 2A} adenosine receptor (A{sub 2A}AR) is also assumed. The distinct functions of the A{sub 1} and A{sub 2A} receptor subtypes in sleep-wake regulation and in mediating the effects of caffeine have not been identified so far. The selective adenosine A{sub 1} receptor antagonist, 8-cyclopentyl-3-(3-{sup 18}Ffluoropropyl)- 1-propylxanthine ({sup 18}F-CPFPX), offers the opportunity to get further insights into adenosinergic mechanisms by in vivo imaging of the A{sub 1}AR subtype with positron emission tomography (PET). The aim of this thesis was to elucidate the role of adenosine A{sub 1} receptors in human sleep regulation, combining {sup 18}F-CPFPX PET brain imaging and EEG recordings, the gold standard in sleep research. It was hypothesized that sleep deprivation would induce adenosine accumulation and/or changes in A{sub 1}AR density. Thus, the question was addressed whether these effects of prolonged wakefulness can be visualized by altered {sup 18}F-CPFPX binding. Moreover, it was

  5. Expression of tumor necrosis factor related apoptosis inducing ligand receptor in glioblastoma

    Institute of Scientific and Technical Information of China (English)

    Dongling Gao; Zhongwei Zhao; Hongxin Zhang; Lan Zhang; Kuisheng Chen; Yunhan Zhang

    2008-01-01

    BACKGROUND: Receptors for tumor necrosis factor related apoptosis inducing ligand (TRAIL) include death receptor 4, death receptor 5, decoy receptor 1, and decoy receptor 2. Activation of death receptor 4 and 5 selectively kills tumor cells.OBJECTIVE: To detect TRAIL receptor expression in glioblastoma by immunohistochemistry and RT-PCR and to compare this expression to that in normal brain tissue.DESIGN: Observational analysis.SETTING: Department of Pathology, the First Affiliated Hospital of Zhengzhou University; Henan Tumor Pathology Key Laboratory.PARTICIPANTS: Twenty-five patients (17 males and 8 females) who received glioblastoma resection were selected from the Fifth Affiliated Hospital of Zhengzhou University, between September 2003 to June 2004. All glioblastoma samples were diagnosed pathologically. Twenty patients (12 males and 8 females) with craniocerebral injury who received normal brain tissue resection were selected in the same time period. There were no significant differences in sex and age between glioblastoma patients or between craniocerebral injury patients (P>0.05). All patients and appropriate relatives provided informed consent, and this study was approved by the local research ethics committee.METHODS: Polyclonal antibody against TRAIL receptors and an immunohistochemical kit (batch number: 200502) were purchased from Boster Company, Wuhan. Immunohistochemistry: Expression of death receptor 4, death receptor 5, decoy receptor 1, and decoy receptor 2 were observed in both glioblastoma and normal brain tissue. The experiment was performed according to the kit instructions, and positive staining was brown-yellow. Assessment: There were no positive signals (-); weakly positive signals, positive cells75% (++++). Evaluation: Expression levels of TRAIL receptors were estimated in both normal brain tissue and glioblastoma. Expression of decoy receptor 1 and decoy receptor 2 mRNA in glioblastoma were detected by reverse transcription polymerase

  6. Safety, tolerability, and initial efficacy of AZD6140, the first reversible oral adenosine diphosphate receptor antagonist, compared with clopidogrel, in patients with non-ST-segment elevation acute coronary syndrome: primary results of the DISPERSE-2 trial

    DEFF Research Database (Denmark)

    Cannon, Christopher P; Husted, Steen; Harrington, Robert A;

    2007-01-01

    OBJECTIVES: Our goal was to compare the safety and initial efficacy of AZD6140, the first reversible oral adenosine diphosphate receptor antagonist, with clopidogrel in patients with non-ST-segment elevation acute coronary syndromes (NSTE-ACS). BACKGROUND: AZD6140 achieves higher mean levels of p...

  7. Non-Ligand-Induced Dimerization is Sufficient to Initiate the Signalling and Endocytosis of EGF Receptor

    Science.gov (United States)

    Kourouniotis, George; Wang, Yi; Pennock, Steven; Chen, Xinmei; Wang, Zhixiang

    2016-01-01

    The binding of epidermal growth factor (EGF) to EGF receptor (EGFR) stimulates cell mitogenesis and survival through various signalling cascades. EGF also stimulates rapid EGFR endocytosis and its eventual degradation in lysosomes. The immediate events induced by ligand binding include receptor dimerization, activation of intrinsic tyrosine kinase and autophosphorylation. However, in spite of intensified efforts, the results regarding the roles of these events in EGFR signalling and internalization is still very controversial. In this study, we constructed a chimeric EGFR by replacing its extracellular domain with leucine zipper (LZ) and tagged a green fluorescent protein (GFP) at its C-terminus. We showed that the chimeric LZ-EGFR-GFP was constitutively dimerized. The LZ-EGFR-GFP dimer autophosphorylated each of its five well-defined C-terminal tyrosine residues as the ligand-induced EGFR dimer does. Phosphorylated LZ-EGFR-GFP was localized to both the plasma membrane and endosomes, suggesting it is capable of endocytosis. We also showed that LZ-EGFR-GFP activated major signalling proteins including Src homology collagen-like (Shc), extracellular signal-regulated kinase (ERK) and Akt. Moreover, LZ-EGFR-GFP was able to stimulate cell proliferation. These results indicate that non-ligand induced dimerization is sufficient to activate EGFR and initiate cell signalling and EGFR endocytosis. We conclude that receptor dimerization is a critical event in EGF-induced cell signalling and EGFR endocytosis. PMID:27463710

  8. Ligand-specific regulation of the extracellular surface of a G-protein-coupled receptor

    Energy Technology Data Exchange (ETDEWEB)

    Bokoch, Michael P.; Zou, Yaozhong; Rasmussen, Søren G.F.; Liu, Corey W.; Nygaard, Rie; Rosenbaum, Daniel M.; Fung, Juan José; Choi, Hee-Jung; Thian, Foon Sun; Kobilka, Tong Sun; Puglisi, Joseph D.; Weis, William I.; Pardo, Leonardo; Prosser, R. Scott; Mueller, Luciano; Kobilka, Brian K. (Stanford-MED); (Toronto); (BMS); (UAB, Spain)

    2010-01-14

    G-protein-coupled receptors (GPCRs) are seven-transmembrane proteins that mediate most cellular responses to hormones and neurotransmitters. They are the largest group of therapeutic targets for a broad spectrum of diseases. Recent crystal structures of GPCRs have revealed structural conservation extending from the orthosteric ligand-binding site in the transmembrane core to the cytoplasmic G-protein-coupling domains. In contrast, the extracellular surface (ECS) of GPCRs is remarkably diverse and is therefore an ideal target for the discovery of subtype-selective drugs. However, little is known about the functional role of the ECS in receptor activation, or about conformational coupling of this surface to the native ligand-binding pocket. Here we use NMR spectroscopy to investigate ligand-specific conformational changes around a central structural feature in the ECS of the {beta}{sub 2} adrenergic receptor: a salt bridge linking extracellular loops 2 and 3. Small-molecule drugs that bind within the transmembrane core and exhibit different efficacies towards G-protein activation (agonist, neutral antagonist and inverse agonist) also stabilize distinct conformations of the ECS. We thereby demonstrate conformational coupling between the ECS and the orthosteric binding site, showing that drugs targeting this diverse surface could function as allosteric modulators with high subtype selectivity. Moreover, these studies provide a new insight into the dynamic behaviour of GPCRs not addressable by static, inactive-state crystal structures.

  9. Non-Ligand-Induced Dimerization is Sufficient to Initiate the Signalling and Endocytosis of EGF Receptor

    Directory of Open Access Journals (Sweden)

    George Kourouniotis

    2016-07-01

    Full Text Available The binding of epidermal growth factor (EGF to EGF receptor (EGFR stimulates cell mitogenesis and survival through various signalling cascades. EGF also stimulates rapid EGFR endocytosis and its eventual degradation in lysosomes. The immediate events induced by ligand binding include receptor dimerization, activation of intrinsic tyrosine kinase and autophosphorylation. However, in spite of intensified efforts, the results regarding the roles of these events in EGFR signalling and internalization is still very controversial. In this study, we constructed a chimeric EGFR by replacing its extracellular domain with leucine zipper (LZ and tagged a green fluorescent protein (GFP at its C-terminus. We showed that the chimeric LZ-EGFR-GFP was constitutively dimerized. The LZ-EGFR-GFP dimer autophosphorylated each of its five well-defined C-terminal tyrosine residues as the ligand-induced EGFR dimer does. Phosphorylated LZ-EGFR-GFP was localized to both the plasma membrane and endosomes, suggesting it is capable of endocytosis. We also showed that LZ-EGFR-GFP activated major signalling proteins including Src homology collagen-like (Shc, extracellular signal-regulated kinase (ERK and Akt. Moreover, LZ-EGFR-GFP was able to stimulate cell proliferation. These results indicate that non-ligand induced dimerization is sufficient to activate EGFR and initiate cell signalling and EGFR endocytosis. We conclude that receptor dimerization is a critical event in EGF-induced cell signalling and EGFR endocytosis.

  10. Trafficking of epidermal growth factor receptor ligands in polarized epithelial cells.

    Science.gov (United States)

    Singh, Bhuminder; Coffey, Robert J

    2014-01-01

    A largely unilamellar epithelial layer lines body cavities and organ ducts such as the digestive tract and kidney tubules. This polarized epithelium is composed of biochemically and functionally separate apical and basolateral surfaces. The epidermal growth factor receptor (EGFR) signaling pathway is a critical regulator of epithelial homeostasis and is perturbed in a number of epithelial disorders. It is underappreciated that in vivo EGFR signaling is most often initiated by cell-surface delivery and processing of one of seven transmembrane ligands, resulting in release of the soluble form that binds EGFR. In polarized epithelial cells, EGFR is restricted largely to the basolateral surface, and apical or basolateral ligand delivery therefore has important biological consequences. In vitro approaches have been used to study the biosynthesis, cell-surface delivery, proteolytic processing, and release of soluble EGFR ligands in polarized epithelial cells. We review these results, discuss their relevance to normal physiology, and demonstrate the pathophysiological consequences of aberrant trafficking. These studies have uncovered a rich diversity of apico-basolateral trafficking mechanisms among the EGFR ligands, provided insights into the pathogenesis of an inherited magnesium-wasting disorder of the kidney (isolated renal hypomagnesemia), and identified a new mode of EGFR ligand signaling via exosomes. PMID:24215440

  11. The evolution of the ligand/receptor couple: a long road from comparative endocrinology to comparative genomics

    OpenAIRE

    Markov, Gabriel V.; Paris, Mathilde; Bertrand, Stephanie; Laudet, Vincent

    2008-01-01

    The evolution of the ligand/receptor couple: a long road from comparative endocrinology to comparative genomics FRANCE (Markov, Gabriel V.) FRANCE Received: 2008-02-11 Revised: 2008-05-14 Accepted: 2008-06-11

  12. Interleukin-6-type cytokines in neuroprotection and neuromodulation: Oncostatin M, but not leukemia inhibitory factor, requires neuronal Adenosine A1 receptor function

    NARCIS (Netherlands)

    Moidunny, S.; Dias, R.; Van Calker, D.; Boddeke, H.; Sebastiao, A.; Biber, K.

    2010-01-01

    Objective: Adenosine is a neuromodulator in the central nervous system exhibiting anticonvulsive, neuroprotective and sedating/sleep regulating properties. A pathophysiological importance of adenosine in various neuropsychiatric diseases (e.g. epilepsy, neurodegenerative disorders, apoplexia and moo

  13. Non-canonical kinase signaling by the death ligand TRAIL in cancer cells : discord in the death receptor family

    NARCIS (Netherlands)

    Azijli, K.; Weyhenmeyer, B.; Peters, G. J.; de Jong, S.; Kruyt, F. A. E.

    2013-01-01

    Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-based therapy is currently evaluated in clinical studies as a tumor cell selective pro-apoptotic approach. However, besides activating canonical caspase-dependent apoptosis by binding to TRAIL-specific death receptors, the TRAIL ligand

  14. Chemically engineering ligand selectivity at the free fatty acid receptor 2 based on pharmacological variation between species orthologs

    DEFF Research Database (Denmark)

    Hudson, Brian D; Christiansen, Elisabeth; Tikhonova, Irina G;

    2012-01-01

    When it is difficult to develop selective ligands within a family of related G-protein-coupled receptors (GPCRs), chemically engineered receptors activated solely by synthetic ligands (RASSLs) are useful alternatives for probing receptor function. In the present work, we explored whether a RASSL...... on this receptor and demonstrates that exploitation of pharmacological variation between species orthologs is a powerful method to generate novel chemically engineered GPCRs.-Hudson, B. D., Christiansen, E., Tikhonova, I. G., Grundmann, M., Kostenis, E., Adams, D. R., Ulven, T., Milligan, G. Chemically engineering...

  15. Potential clinical relevance of Eph receptors and ephrin ligands expressed in prostate carcinoma cell lines.

    Science.gov (United States)

    Fox, Brian P; Tabone, Christopher J; Kandpal, Raj P

    2006-04-21

    The family of Eph and ephrin receptors is involved in a variety of functions in normal cells, and the alterations in their expression profiles have been observed in several cancers. We have compared the transcripts for Eph receptors and ephrin ligands in cell lines established from normal prostate epithelium and several carcinoma cell lines isolated from prostate tumors of varying degree of metastasis. These cell lines included NPTX, CTPX, LNCaP, DU145, PC-3, and PC-3ML. The cell lines displayed characteristic pattern of expression for specific Eph receptors and ephrin ligands, thus allowing identification of Eph receptor signatures for a particular cell line. The sensitivity of these transcripts to genome methylation is also investigated by treating the cells with 5-aza-2'-deoxycytidine. The comparison of expression profiles revealed that normal prostate and primary prostate tumor cell lines differ in the expression of EphA3, EphB3, and ephrin A3 that are over-expressed in normal prostate. Furthermore, the transcript levels for EphA1 decrease progressively from normal prostate to primary prostate tumor cell line and metastatic tumor cells. A converse relationship was observed for ephrin B2. The treatment of cells with 5-aza-2'-deoxycytidine revealed the sensitivity of EphA3, EphA10, EphB3, and EphB6 to methylation status of genomic DNA. The utility of methylation specific PCR to identify prostate tumor cells and the importance of specific Eph receptors and ephrin ligands in initiation and progression of prostate tumor are discussed. PMID:16516143

  16. Data for amino acid alignment of Japanese stingray melanocortin receptors with other gnathostome melanocortin receptor sequences, and the ligand selectivity of Japanese stingray melanocortin receptors

    Directory of Open Access Journals (Sweden)

    Akiyoshi Takahashi

    2016-06-01

    Full Text Available This article contains structure and pharmacological characteristics of melanocortin receptors (MCRs related to research published in “Characterization of melanocortin receptors from stingray Dasyatis akajei, a cartilaginous fish” (Takahashi et al., 2016 [1]. The amino acid sequences of the stingray, D. akajei, MC1R, MC2R, MC3R, MC4R, and MC5R were aligned with the corresponding melanocortin receptor sequences from the elephant shark, Callorhinchus milii, the dogfish, Squalus acanthias, the goldfish, Carassius auratus, and the mouse, Mus musculus. These alignments provide the basis for phylogenetic analysis of these gnathostome melanocortin receptor sequences. In addition, the Japanese stingray melanocortin receptors were separately expressed in Chinese Hamster Ovary cells, and stimulated with stingray ACTH, α-MSH, β-MSH, γ-MSH, δ-MSH, and β-endorphin. The dose response curves reveal the order of ligand selectivity for each stingray MCR.

  17. Novel aza-analogous ergoline derived scaffolds as potent serotonin 5-HT6 and dopamine D2 receptor ligands

    DEFF Research Database (Denmark)

    Krogsgaard-Larsen, Niels; Jensen, Anders A.; Schrøder, T.J.;

    2014-01-01

    By introducing distal substituents on a tetracyclic scaffold resembling the ergoline structure, two series of analogues were achieved exhibiting subnanomolar receptor binding affinities for the dopamine D2 and serotonin 5-HT6 receptor subtype, respectively. While the 5-HT6 ligands were antagonist......, the D2 ligands displayed intrinsic activities ranging from full agonism to partial agonism with low intrinsic activity. These structures could potentially be interesting for treatment of neurological diseases such as schizophrenia, Parkinson’s disease, and cognitive deficits....

  18. Evaluation of 3-Ethyl-3-(phenylpiperazinylbutyl)oxindoles as PET Ligands for the Serotonin 5-HT7 Receptor

    DEFF Research Database (Denmark)

    Herth, Matthias M; Andersen, Valdemar L; Hansen, Hanne D;

    2015-01-01

    We have investigated several oxindole derivatives in the pursuit of a 5-HT7 receptor PET ligand. Herein the synthesis, chiral separation, and pharmacological profiling of two possible PET candidates toward a wide selection of CNS-targets are detailed. Subsequent (11)C-labeling and in vivo evaluat...... evaluation in Danish landrace pigs showed that both ligands displayed high brain uptake. However, neither of the radioligands could be displaced by the 5-HT7 receptor selective inverse agonist SB-269970....

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

    OpenAIRE

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

    2008-01-01

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

  20. Activation of epidermal growth factor receptor by metal-ligand complexes decreases levels of extracellular amyloid beta peptide.

    Science.gov (United States)

    Price, Katherine A; Filiz, Gulay; Caragounis, Aphrodite; Du, Tai; Laughton, Katrina M; Masters, Colin L; Sharples, Robyn A; Hill, Andrew F; Li, Qiao-Xin; Donnelly, Paul S; Barnham, Kevin J; Crouch, Peter J; White, Anthony R

    2008-01-01

    The epidermal growth factor receptor is a receptor tyrosine kinase expressed in a range of tissues and cell-types. Activation of the epidermal growth factor receptor by a number of ligands induces downstream signalling that modulates critical cell functions including growth, survival and differentiation. Abnormal epidermal growth factor receptor expression and activation is also involved in a number of cancers. In addition to its cognate ligands, the epidermal growth factor receptor can be activated by metals such as zinc (Zn) and copper (Cu). Due to the important role of these metals in a number of diseases including neurodegenerative disorders, therapeutic approaches are being developed based on the use of lipid permeable metal-complexing molecules. While these agents are showing promising results in animal models and clinical trials, little is known about the effects of metal-ligand complexes on cell signalling pathways. In this study, we investigated the effects of clioquinol (CQ)-metal complexes on activation of epidermal growth factor receptor. We show here that CQ-Cu complexes induced potent epidermal growth factor receptor phosphorylation resulting in downstream activation of extracellular signal-regulated kinase. Similar levels of epidermal growth factor receptor activation were observed with alternative lipid permeable metal-ligands including neocuproine and pyrrolidine dithiocarbamate. We found that CQ-Cu complexes induced a significant reduction in the level of extracellular Abeta1-40 in cell culture. Inhibition of epidermal growth factor receptor activation by PD153035 blocked extracellular signal-regulated kinase phosphorylation and restored Abeta1-40 levels. Activation of the epidermal growth factor receptor by CQ-Cu was mediated through up-regulation of src kinase activity by a cognate ligand-independent process involving membrane integrins. These findings provide the first evidence that metal-ligand complexes can activate the epidermal growth

  1. Comparison of SYBR Green and TaqMan methods in quantitative real-time polymerase chain reaction analysis of four adenosine receptor subtypes

    Directory of Open Access Journals (Sweden)

    Mohamadhasan Tajadini

    2014-01-01

    Full Text Available Background: Real-time polymerase chain reaction (PCR is based on the revolutionary method of PCR. This technique is the result of PCR enormous sensitivity and real-time monitoring combination. In quantitative gene expression analysis, two methods have more popularity, SYBR Green and TaqMan, SYBR Green is relatively cost benefit and easy to use and technically based on binding the fluorescent dye to double-stranded deoxyribonucleic acid (dsDNA where TaqMan method has more expensive and based on dual labeled oligonucleotide and exonuclease activity of Taq polymerase enzyme. Specificity is the most important concern with the usage of any non-specific dsDNA-binding Dyes such as SYBR Green whiles more specificity showed by labeled oligonucleotide method such as TaqMan. In this study, we compared two common RT PCR methods, TaqMan and SYBR Green in measurement gene expression profile of adenosine receptors. Materials and Methods: Gene expression profiles of A1, A2A, A2B and A3 Adenosine receptors were analyzed by optimized TaqMan and SYBR Green quantitative RT PCR in breast cancer tissues. Primary expression data was normalizing by B. actin reference gene. Results: Efficiencies were calculated more than 95% for TaqMan and SYBR Green methods in all genes. The correlations between means of normalized data of each gene in two methods were positive and significant (P < 0.05. Conclusion: Data analysis showed that with the use of high performance primer and by use proper protocols and material we can make precise data by SYBR Green as TaqMan method. In other word by optimization of SYBR Green method, its performance and quality could be comparable to TaqMan method.

  2. 腺苷A2a受体在炎症反应中的作用%Effects of A2a adenosine receptor on inflammatory reaction

    Institute of Scientific and Technical Information of China (English)

    郦铮铮; 郑荣远

    2009-01-01

    Adenosine has a wide distribution in organs and tissues of the human body. It plays diverse physio-logical roles including the inhibition of inflammatory reactions, the resistance of ischemia/hypoxia and the mod-ulation of immunological reactions through binding with G-protein-coupled receptors on cell surfaces. There are four kinds of adenosine receptors, A1R,A2aR,A2bR and A3R. A2aR is widely expressed in virtually all cell typos involving in inflammatory/immune responses. These cell types include glial cells, macrophages, dendritic cells, mast cells, NK cells, endothelial cells, as well as epithelial cells. A2aR serves to regulate conventional inflammatory reactions and immunological reactions.%腺苷广泛分布于全身各组织器官,可通过与细胞表面相应G-蛋白偶联受体结合发挥抑制炎症反应,对抗缺血/缺氧和免疫调节等生物学作用.目前发现腺苷受体有4种亚型,即A1R、A2aR、A2bR和A3R.其中A2a腺苷受体(A2aR)广泛分布于神经胶质细胞,巨噬细胞,树突状细胞,肥大细胞,自然杀伤细胞等免疫细胞及内皮细胞和上皮细胞中,在相应的腺苷配体作用下,可通过腺苷-腺苷受体系统介导炎症反应和发挥免疫调节作用.

  3. Cross sectional PET study of cerebral adenosine A{sub 1} receptors in premanifest and manifest Huntington's disease

    Energy Technology Data Exchange (ETDEWEB)

    Matusch, Andreas; Elmenhorst, David [Institute of Neuroscience and Medicine (INM-2), Juelich (Germany); Saft, Carsten; Kraus, Peter H.; Gold, Ralf [St. Josef Hospital, Ruhr University Bochum, Department of Neurology, Huntington Centre NRW, Bochum (Germany); Hartung, Hans-Peter [Heinrich Heine University Duesseldorf, Department of Neurology, Medical Faculty, Duesseldorf (Germany); Bauer, Andreas [Institute of Neuroscience and Medicine (INM-2), Juelich (Germany); Heinrich Heine University Duesseldorf, Department of Neurology, Medical Faculty, Duesseldorf (Germany)

    2014-06-15

    To study cerebral adenosine receptors (AR) in premanifest and manifest stages of Huntington's disease (HD). We quantified the cerebral binding potential (BP{sub ND}) of the A{sub 1}AR in carriers of the HD CAG trinucleotide repeat expansion using the radioligand [{sup 18} F]CPFPX and PET. Four groups were investigated: (i) premanifest individuals far (preHD-A; n = 7) or (ii) near (preHD-B; n = 6) to the predicted symptom onset, (iii) manifest HD patients (n = 8), and (iv) controls (n = 36). Cerebral A{sub 1}AR values of preHD-A subjects were generally higher than those of controls (by up to 31 %, p <.01, in the thalamus on average). Across stages a successive reduction of A{sub 1}AR BP{sub ND} was observed to the levels of controls in preHD-B and undercutting controls in manifest HD by down to 25 %, p <.01, in the caudatus and amygdala. There was a strong correlation between A{sub 1}AR BP{sub ND} and years to onset. Before onset of HD, the assumed annual rates of change of A{sub 1}AR density were -1.2 % in the caudatus, -1.7 % in the thalamus and -3.4 % in the amygdala, while the corresponding volume losses amounted to 0.6 %, 0.1 % and 0.2 %, respectively. Adenosine receptors switch from supra to subnormal levels during phenoconversion of HD. This differential regulation may play a role in the pathophysiology of altered energy metabolism. (orig.)

  4. The CXCL12/CXCR4 axis promotes ligand-independent activation of the androgen receptor.

    Science.gov (United States)

    Kasina, Sathish; Macoska, Jill A

    2012-04-01

    The molecular mechanisms responsible for the transition of some prostate cancers from androgen ligand-dependent to androgen ligand-independent are incompletely established. Molecules that are ligands for G protein coupled receptors (GPCRs) have been implicated in ligand-independent androgen receptor (AR) activation. The purpose of this study was to examine whether CXCL12, the ligand for the GPCR, CXCR4, might mediate prostate cancer cell proliferation through AR-dependent mechanisms involving functional transactivation of the AR in the absence of androgen. The results of these studies showed that activation of the CXCL12/CXCR4 axis promoted: The nuclear accumulation of both wild-type and mutant AR in several prostate epithelial cell lines; AR-dependent proliferative responses; nuclear accumulation of the AR co-regulator SRC-1 protein; SRC-1:AR protein:protein association; co-localization of AR and SRC-1 on the promoters of AR-regulated genes; AR- and SRC-1 dependent transcription of AR-regulated genes; AR-dependent secretion of the AR-regulated PSA protein; P13K-dependent phosphorylation of AR; MAPK-dependent phosphorylation of SRC-1, and both MAPK- and P13K-dependent secretion of the PSA protein, in the absence of androgen. Taken together, these studies identify CXCL12 as a novel, non-steroidal growth factor that promotes the growth of prostate epithelial cells through AR-dependent mechanisms in the absence of steroid hormones. These findings support the development of novel therapeutics targeting the CXCL12/CXCR4 axis as an ancillary to those targeting the androgen/AR axis to effectively treat castration resistant/recurrent prostate tumors.

  5. Structure-activity relationships of constrained phenylethylamine ligands for the serotonin 5-HT2 receptors.

    Directory of Open Access Journals (Sweden)

    Vignir Isberg

    Full Text Available Serotonergic ligands have proven effective drugs in the treatment of migraine, pain, obesity, and a wide range of psychiatric and neurological disorders. There is a clinical need for more highly 5-HT2 receptor subtype-selective ligands and the most attention has been given to the phenethylamine class. Conformationally constrained phenethylamine analogs have demonstrated that for optimal activity the free lone pair electrons of the 2-oxygen must be oriented syn and the 5-oxygen lone pairs anti relative to the ethylamine moiety. Also the ethyl linker has been constrained providing information about the bioactive conformation of the amine functionality. However, combined 1,2-constriction by cyclization has only been tested with one compound. Here, we present three new 1,2-cyclized phenylethylamines, 9-11, and describe their synthetic routes. Ligand docking in the 5-HT2B crystal structure showed that the 1,2-heterocyclized compounds can be accommodated in the binding site. Conformational analysis showed that 11 can only bind in a higher-energy conformation, which would explain its absent or low affinity. The amine and 2-oxygen interactions with D3.32 and S3.36, respectively, can form but shift the placement of the core scaffold. The constraints in 9-11 resulted in docking poses with the 4-bromine in closer vicinity to 5.46, which is polar only in the human 5-HT2A subtype, for which 9-11 have the lowest affinity. The new ligands, conformational analysis and docking expand the structure-activity relationships of constrained phenethylamines and contributes towards the development of 5-HT2 receptor subtype-selective ligands.

  6. Monitoring Solution Structures of Peroxisome Proliferator-Activated Receptor β/δ upon Ligand Binding.

    Science.gov (United States)

    Schwarz, Rico; Tänzler, Dirk; Ihling, Christian H; Sinz, Andrea

    2016-01-01

    Peroxisome proliferator-activated receptors (PPARs) have been intensively studied as drug targets to treat type 2 diabetes, lipid disorders, and metabolic syndrome. This study is part of our ongoing efforts to map conformational changes in PPARs in solution by a combination of chemical cross-linking and mass spectrometry (MS). To our best knowledge, we performed the first studies addressing solution structures of full-length PPAR-β/δ. We monitored the conformations of the ligand-binding domain (LBD) as well as full-length PPAR-β/δ upon binding of two agonists. (Photo-) cross-linking relied on (i) a variety of externally introduced amine- and carboxyl-reactive linkers and (ii) the incorporation of the photo-reactive amino acid p-benzoylphenylalanine (Bpa) into PPAR-β/δ by genetic engineering. The distances derived from cross-linking experiments allowed us to monitor conformational changes in PPAR-β/δ upon ligand binding. The cross-linking/MS approach proved highly advantageous to study nuclear receptors, such as PPARs, and revealed the interplay between DBD (DNA-binding domain) and LDB in PPAR-β/δ. Our results indicate the stabilization of a specific conformation through ligand binding in PPAR-β/δ LBD as well as full-length PPAR-β/δ. Moreover, our results suggest a close distance between the N- and C-terminal regions of full-length PPAR-β/δ in the presence of GW1516. Chemical cross-linking/MS allowed us gaining detailed insights into conformational changes that are induced in PPARs when activating ligands are present. Thus, cross-linking/MS should be added to the arsenal of structural methods available for studying nuclear receptors. PMID:26992147

  7. Epidermal growth factor receptor ligands as new extracellular targets for the metastasis-promoting S100A4 protein

    DEFF Research Database (Denmark)

    Klingelhöfer, Jörg; Møller, Henrik D.; Sumer, Eren U;

    2009-01-01

    that the most abundant peptide mimicked the F/YCC motif present in the epidermal growth factor domain of ErbB receptor ligands. S100A4 selectively interacted with a number of epidermal growth factor receptor (EGFR) ligands, demonstrating highest affinity for amphiregulin. Importantly, we found that S100A4...... stimulated EGFR/ErbB2 receptor signaling and enhanced the amphiregulin-mediated proliferation of mouse embryonic fibroblasts. S100A4-neutralizing antibodies, as well as EGFR- and ErbB2 receptor-specific tyrosine kinase inhibitors, blocked these effects. The present results suggest that extracellular S100A4...... regulates tumor progression by interacting with EGFR ligands, thereby enhancing EGFR/ErbB2 receptor signaling and cell proliferation. Structured digital abstract: * MINT-7256556: EGF (uniprotkb:P01133) binds (MI:0407) to S100A4 (uniprotkb:P26447) by far western blotting (MI:0047) * MINT-7256512: BC...

  8. Adenosine AA Receptor Antagonists Do Not Disrupt Rodent Prepulse Inhibition: An Improved Side Effect Profile in the Treatment of Parkinson's Disease

    Directory of Open Access Journals (Sweden)

    Carina J. Bleickardt

    2012-01-01

    Full Text Available Parkinson's disease (PD is characterized by loss of dopaminergic neurons in the substantia nigra. Current treatments for PD focus on dopaminergic therapies, including L-dopa and dopamine receptor agonists. However, these treatments induce neuropsychiatric side effects. Psychosis, characterized by delusions and hallucinations, is one of the most serious such side effects. Adenosine A2A receptor antagonism is a nondopaminergic treatment for PD with clinical and preclinical efficacy. The present studies assessed A2A antagonists SCH 412348 and istradefylline in rodent prepulse inhibition (PPI, a model of psychosis. Dopamine receptor agonists pramipexole (0.3–3 mg/kg, pergolide (0.3–3 mg/kg, and apomorphine (0.3–3 mg/kg significantly disrupted PPI; ropinirole (1–30 mg/kg had no effect; L-dopa (100–300 mg/kg disrupted rat but not mouse PPI. SCH 412348 (0.3–3 mg/kg did not disrupt rodent PPI; istradefylline (0.1–1 mg/kg marginally disrupted mouse but not rat PPI. These results suggest that A2A antagonists, unlike dopamine agonists, have an improved neuropsychiatric side effect profile.

  9. SPOT-Ligand: Fast and effective structure-based virtual screening by binding homology search according to ligand and receptor similarity.

    Science.gov (United States)

    Yang, Yuedong; Zhan, Jian; Zhou, Yaoqi

    2016-07-01

    Structure-based virtual screening usually involves docking of a library of chemical compounds onto the functional pocket of the target receptor so as to discover novel classes of ligands. However, the overall success rate remains low and screening a large library is computationally intensive. An alternative to this "ab initio" approach is virtual screening by binding homology search. In this approach, potential ligands are predicted based on similar interaction pairs (similarity in receptors and ligands). SPOT-Ligand is an approach that integrates ligand similarity by Tanimoto coefficient and receptor similarity by protein structure alignment program SPalign. The method was found to yield a consistent performance in DUD and DUD-E docking benchmarks even if model structures were employed. It improves over docking methods (DOCK6 and AUTODOCK Vina) and has a performance comparable to or better than other binding-homology methods (FINDsite and PoLi) with higher computational efficiency. The server is available at http://sparks-lab.org. © 2016 Wiley Periodicals, Inc. PMID:27074979

  10. Development of novel mixed ligand technetium complexes (3 + 1 combination) for imaging central neural system receptors

    International Nuclear Information System (INIS)

    A series of mixed ligand oxotechnetium-99m complexes carrying the 1-(2-methoxyphenyl) piperazine moiety has been synthesized. For structural characterization, and for in vitro binding assays, the analogous oxorhenium or oxotechnetium-99 complexes were prepared. As demonstrated by appropriate competition binding tests in rat hippocampal preparations, all oxorhenium analogues showed affinity for the 5-HT1A receptor binding sites with 50% inhibitory concentration values in the nanomolar range (IC50=6-106nM). All 99mTcO[SN(R)S]/[S] complexes showed a significant brain uptake in rats at 2 min post-injection (0.24-1.31 dose/organ). The regional distribution is inhomogeneous but the ratio between areas rich and poor in 5-HT1A receptor was not high. Structural modifications to this system may further improve the biological profile of these compounds and eventually provide efficient 99mTc receptor imaging agents. (author)

  11. The 2.1 A resolution structure of cyanopindolol-bound β1-adrenoceptor identifies an intramembrane Na+ ion that stabilises the ligand-free receptor.

    Directory of Open Access Journals (Sweden)

    Jennifer L Miller-Gallacher

    Full Text Available The β1-adrenoceptor (β1AR is a G protein-coupled receptor (GPCR that is activated by the endogenous agonists adrenaline and noradrenaline. We have determined the structure of an ultra-thermostable β1AR mutant bound to the weak partial agonist cyanopindolol to 2.1 Å resolution. High-quality crystals (100 μm plates were grown in lipidic cubic phase without the assistance of a T4 lysozyme or BRIL fusion in cytoplasmic loop 3, which is commonly employed for GPCR crystallisation. An intramembrane Na+ ion was identified co-ordinated to Asp872.50, Ser1283.39 and 3 water molecules, which is part of a more extensive network of water molecules in a cavity formed between transmembrane helices 1, 2, 3, 6 and 7. Remarkably, this water network and Na+ ion is highly conserved between β1AR and the adenosine A2A receptor (rmsd of 0.3 Å, despite an overall rmsd of 2.4 Å for all Cα atoms and only 23% amino acid identity in the transmembrane regions. The affinity of agonist binding and nanobody Nb80 binding to β1AR is unaffected by Na+ ions, but the stability of the receptor is decreased by 7.5°C in the absence of Na+. Mutation of amino acid side chains that are involved in the co-ordination of either Na+ or water molecules in the network decreases the stability of β1AR by 5-10°C. The data suggest that the intramembrane Na+ and associated water network stabilise the ligand-free state of β1AR, but still permits the receptor to form the activated state which involves the collapse of the Na+ binding pocket on agonist binding.

  12. ReFlexIn: a flexible receptor protein-ligand docking scheme evaluated on HIV-1 protease.

    Directory of Open Access Journals (Sweden)

    Simon Leis

    Full Text Available For many targets of pharmaceutical importance conformational changes of the receptor protein are relevant during the ligand binding process. A new docking approach, ReFlexIn (Receptor Flexibility by Interpolation, that combines receptor flexibility with the computationally efficient potential grid representation of receptor molecules has been evaluated on the retroviral HIV-1 (Human Immunodeficiency Virus 1 protease system. An approximate inclusion of receptor flexibility is achieved by using interpolation between grid representations of individual receptor conformations. For the retroviral protease the method was tested on an ensemble of protease structures crystallized in the presence of different ligands and on a set of structures obtained from morphing between the unbound and a ligand-bound protease structure. Docking was performed on ligands known to bind to the protease and several non-binders. For the binders the ReFlexIn method yielded in almost all cases ligand placements in similar or closer agreement with experiment than docking to any of the ensemble members without degrading the discrimination with respect to non-binders. The improved docking performance compared to docking to rigid receptors allows for systematic virtual screening applications at very small additional computational cost.

  13. The different ligand-binding modes of relaxin family peptide receptors RXFP1 and RXFP2.

    Science.gov (United States)

    Scott, Daniel J; Rosengren, K Johan; Bathgate, Ross A D

    2012-11-01

    Relaxin and insulin-like peptide 3 (INSL3) are peptide hormones with a number of important physiological roles in reproduction, regulation of extracellular matrix turnover, and cardiovascular function. Relaxin and INSL3 mediate their actions through the closely related G-protein coupled receptors, relaxin family peptide receptors 1 and 2 (RXFP1 and RXFP2), respectively. These receptors have large extracellular domains (ECD) that contain high-affinity ligand-binding sites within their 10 leucine-rich repeat (LRR)-containing modules. Although relaxin can bind and activate both RXFP1 and RXFP2, INSL3 can only bind and activate RXFP2. To investigate whether this difference is related to the nature of the high-affinity ECD binding site or to differences in secondary binding sites involving the receptor transmembrane (TM) domain, we created a suite of constructs with RXFP1/2 chimeric ECD attached to single TM helices. We show that by changing as little as one LRR, representing four amino acid substitutions, we were able to engineer a high-affinity INSL3-binding site into the ECD of RXFP1. Molecular modeling of the INSL3-RXFP2 interaction based on extensive experimental data highlights the differences in the binding mechanisms of relaxin and INSL3 to the ECD of their cognate receptors. Interestingly, when the engineered RXFP1/2 ECD were introduced into full-length RXFP1 constructs, INSL3 exhibited only low affinity and efficacy on these receptors. These results highlight critical differences both in the ECD binding and in the coordination of the ECD-binding site with the TM domain, and provide new mechanistic insights into the binding and activation events of RXFP1 and RXFP2 by their native hormone ligands. PMID:22973049

  14. Ion fluxes through KCa2 (SK and Cav1 (L-type channels contribute to chronoselectivity of adenosine A1 receptor-mediated actions in spontaneously beating rat atria

    Directory of Open Access Journals (Sweden)

    Paulo eCorreia-De-Sá

    2016-03-01

    Full Text Available Impulse generation in supraventricular tissue is inhibited by adenosine and acetylcholine via the activation of A1 and M2 receptors coupled to inwardly rectifying GIRK/KIR3.1/3.4 channels, respectively. Unlike M2 receptors, bradycardia produced by A1 receptors activation predominates over negative inotropy. Such difference suggests that other ion currents may contribute to adenosine chronoselectivity. In isolated spontaneously beating rat atria, blockade of KCa2/SK channels with apamin and Cav1 (L-type channels with nifedipine or verapamil, sensitized atria to the negative inotropic action of the A1 agonist, R-PIA, without affecting the nucleoside negative chronotropy. Patch-clamp experiments in the whole-cell configuration mode demonstrate that adenosine, via A1 receptors, activates the inwardly-rectifying GIRK/KIR3.1/KIR3.4 current resulting in hyperpolarization of atrial cardiomyocytes, which may slow down heart rate. Conversely, the nucleoside inactivates a small conductance Ca2+-activated KCa2/SK outward current, which eventually reduces the repolarizing force and thereby prolong action potentials duration Ca2+ influx into cardiomyocytes. Immunolocalization studies showed that differences in A1 receptors distribution between the sinoatrial node and surrounding cardiomyocytes do not afford a rationale for adenosine chronoselectivity. Immunolabelling of KIR3.1, KCa2.2, KCa2.3 and Cav1 was also observed throughout the right atrium. Functional data indicate that while both A1 and M2 receptors favor the opening of GIRK/KIR3.1/3.4 channels modulating atrial chronotropy, A1 receptors may additionally restrain KCa2/SK activation thereby compensating atrial inotropic depression by increasing the time available for Ca2+ influx through Cav1 (L-type channels.

  15. Ion Fluxes through KCa2 (SK) and Cav1 (L-type) Channels Contribute to Chronoselectivity of Adenosine A1 Receptor-Mediated Actions in Spontaneously Beating Rat Atria.

    Science.gov (United States)

    Bragança, Bruno; Oliveira-Monteiro, Nádia; Ferreirinha, Fátima; Lima, Pedro A; Faria, Miguel; Fontes-Sousa, Ana P; Correia-de-Sá, Paulo

    2016-01-01

    Impulse generation in supraventricular tissue is inhibited by adenosine and acetylcholine via the activation of A1 and M2 receptors coupled to inwardly rectifying GIRK/KIR3.1/3.4 channels, respectively. Unlike M2 receptors, bradycardia produced by A1 receptors activation predominates over negative inotropy. Such difference suggests that other ion currents may contribute to adenosine chronoselectivity. In isolated spontaneously beating rat atria, blockade of KCa2/SK channels with apamin and Cav1 (L-type) channels with nifedipine or verapamil, sensitized atria to the negative inotropic action of the A1 agonist, R-PIA, without affecting the nucleoside negative chronotropy. Patch-clamp experiments in the whole-cell configuration mode demonstrate that adenosine, via A1 receptors, activates the inwardly-rectifying GIRK/KIR3.1/KIR3.4 current resulting in hyperpolarization of atrial cardiomyocytes, which may slow down heart rate. Conversely, the nucleoside inactivates a small conductance Ca(2+)-activated KCa2/SK outward current, which eventually reduces the repolarizing force and thereby prolong action potentials duration and Ca(2+) influx into cardiomyocytes. Immunolocalization studies showed that differences in A1 receptors distribution between the sinoatrial node and surrounding cardiomyocytes do not afford a rationale for adenosine chronoselectivity. Immunolabelling of KIR3.1, KCa2.2, KCa2.3, and Cav1 was also observed throughout the right atrium. Functional data indicate that while both A1 and M2 receptors favor the opening of GIRK/KIR3.1/3.4 channels modulating atrial chronotropy, A1 receptors may additionally restrain KCa2/SK activation thereby compensating atrial inotropic depression by increasing the time available for Ca(2+) influx through Cav1 (L-type) channels. PMID:27014060

  16. Peroxisome Proliferator-Activated Receptor γ (PPARγ) and Ligand Choreography: Newcomers Take the Stage.

    Science.gov (United States)

    Garcia-Vallvé, Santiago; Guasch, Laura; Tomas-Hernández, Sarah; del Bas, Josep Maria; Ollendorff, Vincent; Arola, Lluís; Pujadas, Gerard; Mulero, Miquel

    2015-07-23

    Thiazolidinediones (TZDs), such as rosiglitazone and pioglitazone, are peroxisome proliferator-activated receptor γ (PPARγ) full agonists that have been widely used in the treatment of type 2 diabetes mellitus. Despite the demonstrated beneficial effect of reducing glucose levels in the plasma, TZDs also induce several adverse effects. Consequently, the search for new compounds with potent antidiabetic effects but fewer undesired effects is an active field of research. Interestingly, the novel proposed mechanisms for the antidiabetic activity of PPARγ agonists, consisting of PPARγ Ser273 phosphorylation inhibition, ligand and receptor mutual dynamics, and the presence of an alternate binding site, have recently changed the view regarding the optimal characteristics for the screening of novel PPARγ ligands. Furthermore, transcriptional genomics could bring essential information about the genome-wide effects of PPARγ ligands. Consequently, facing the new mechanistic scenario proposed for these compounds is essential for resolving the paradoxes among their agonistic function, antidiabetic activities, and side effects and should allow the rational development of better and safer PPARγ-mediated antidiabetic drugs. PMID:25734377

  17. Emerging importance of chemokine receptor CXCR3 and its ligands in cardiovascular diseases.

    Science.gov (United States)

    Altara, Raffaele; Manca, Marco; Brandão, Rita D; Zeidan, Asad; Booz, George W; Zouein, Fouad A

    2016-04-01

    The CXC chemokines, CXCL4, -9, -10, -11, CXCL4L1, and the CC chemokine CCL21, activate CXC chemokine receptor 3 (CXCR3), a cell-surface G protein-coupled receptor expressed mainly by Th1 cells, cytotoxic T (Tc) cells and NK cells that have a key role in immunity and inflammation. However, CXCR3 is also expressed by vascular smooth muscle and endothelial cells, and appears to be important in controlling physiological vascular function. In the last decade, evidence from pre-clinical and clinical studies has revealed the participation of CXCR3 and its ligands in multiple cardiovascular diseases (CVDs) of different aetiologies including atherosclerosis, hypertension, cardiac hypertrophy and heart failure, as well as in heart transplant rejection and transplant coronary artery disease (CAD). CXCR3 ligands have also proven to be valid biomarkers for the development of heart failure and left ventricular dysfunction, suggesting an underlining pathophysiological relation between levels of these chemokines and the development of adverse cardiac remodelling. The observation that several of the above-mentioned chemokines exert biological actions independent of CXCR3 provides both opportunities and challenges for developing effective drug strategies. In this review, we provide evidence to support our contention that CXCR3 and its ligands actively participate in the development and progression of CVDs, and may additionally have utility as diagnostic and prognostic biomarkers. PMID:26888559

  18. Allosteric ligands for the pharmacologically dark receptors GPR68 and GPR65.

    Science.gov (United States)

    Huang, Xi-Ping; Karpiak, Joel; Kroeze, Wesley K; Zhu, Hu; Chen, Xin; Moy, Sheryl S; Saddoris, Kara A; Nikolova, Viktoriya D; Farrell, Martilias S; Wang, Sheng; Mangano, Thomas J; Deshpande, Deepak A; Jiang, Alice; Penn, Raymond B; Jin, Jian; Koller, Beverly H; Kenakin, Terry; Shoichet, Brian K; Roth, Bryan L

    2015-11-26

    At least 120 non-olfactory G-protein-coupled receptors in the human genome are 'orphans' for which endogenous ligands are unknown, and many have no selective ligands, hindering the determination of their biological functions and clinical relevance. Among these is GPR68, a proton receptor that lacks small molecule modulators for probing its biology. Using yeast-based screens against GPR68, here we identify the benzodiazepine drug lorazepam as a non-selective GPR68 positive allosteric modulator. More than 3,000 GPR68 homology models were refined to recognize lorazepam in a putative allosteric site. Docking 3.1 million molecules predicted new GPR68 modulators, many of which were confirmed in functional assays. One potent GPR68 modulator, ogerin, suppressed recall in fear conditioning in wild-type but not in GPR68-knockout mice. The same approach led to the discovery of allosteric agonists and negative allosteric modulators for GPR65. Combining physical and structure-based screening may be broadly useful for ligand discovery for understudied and orphan GPCRs. PMID:26550826

  19. Peroxisome Proliferator-Activated Receptor γ (PPARγ) and Ligand Choreography: Newcomers Take the Stage.

    Science.gov (United States)

    Garcia-Vallvé, Santiago; Guasch, Laura; Tomas-Hernández, Sarah; del Bas, Josep Maria; Ollendorff, Vincent; Arola, Lluís; Pujadas, Gerard; Mulero, Miquel

    2015-07-23

    Thiazolidinediones (TZDs), such as rosiglitazone and pioglitazone, are peroxisome proliferator-activated receptor γ (PPARγ) full agonists that have been widely used in the treatment of type 2 diabetes mellitus. Despite the demonstrated beneficial effect of reducing glucose levels in the plasma, TZDs also induce several adverse effects. Consequently, the search for new compounds with potent antidiabetic effects but fewer undesired effects is an active field of research. Interestingly, the novel proposed mechanisms for the antidiabetic activity of PPARγ agonists, consisting of PPARγ Ser273 phosphorylation inhibition, ligand and receptor mutual dynamics, and the presence of an alternate binding site, have recently changed the view regarding the optimal characteristics for the screening of novel PPARγ ligands. Furthermore, transcriptional genomics could bring essential information about the genome-wide effects of PPARγ ligands. Consequently, facing the new mechanistic scenario proposed for these compounds is essential for resolving the paradoxes among their agonistic function, antidiabetic activities, and side effects and should allow the rational development of better and safer PPARγ-mediated antidiabetic drugs.

  20. The Combined Inhibitory Effect of the Adenosine A1 and Cannabinoid CB1 Receptors on cAMP Accumulation in the Hippocampus Is Additive and Independent of A1 Receptor Desensitization

    Directory of Open Access Journals (Sweden)

    André Serpa

    2015-01-01

    Full Text Available Adenosine A1 and cannabinoid CB1 receptors are highly expressed in hippocampus where they trigger similar transduction pathways. We investigated how the combined acute activation of A1 and CB1 receptors modulates cAMP accumulation in rat hippocampal slices. The CB1 agonist WIN55212-2 (0.3–30 μM decreased forskolin-stimulated cAMP accumulation with an EC50 of 6.6 ± 2.7 μM and an Emax⁡ of 31% ± 2%, whereas for the A1 agonist, N6-cyclopentyladenosine (CPA, 10–150 nM, an EC50 of 35 ± 19 nM, and an Emax⁡ of 29% ± 5 were obtained. The combined inhibitory effect of WIN55212-2 (30 μM and CPA (100 nM on cAMP accumulation was 41% ± 6% (n=4, which did not differ (P>0.7 from the sum of the individual effects of each agonist (43% ± 8% but was different (P<0.05 from the effects of CPA or WIN55212-2 alone. Preincubation with CPA (100 nM for 95 min caused desensitization of adenosine A1 activity, which did not modify the effect of WIN55212-2 (30 μM on cAMP accumulation. In conclusion, the combined effect of CB1 and A1 receptors on cAMP formation is additive and CB1 receptor activity is not affected by short-term A1 receptor desensitization.

  1. Crystallization and preliminary X-ray analysis of the human androgen receptor ligand-binding domain with a coactivator-like peptide and selective androgen receptor modulators

    International Nuclear Information System (INIS)

    The human androgen receptor ligand-binding domain has been crystallized as a ternary complex with a coactivator-like undecapeptide and two different synthetic ligands. The ligand-binding domain of the human androgen receptor has been cloned, overproduced and crystallized in the presence of a coactivator-like 11-mer peptide and two different nonsteroidal ligands. The crystals of the two ternary complexes were isomorphous and belonged to space group P212121, with one molecule in the asymmetric unit. They diffracted to 1.7 and 1.95 Å resolution, respectively. Structure determination of these two complexes will help in understanding the mode of binding of selective nonsteroidal androgens versus endogenous steroidal ligands and possibly the origin of their tissue selectivity

  2. GluVII:06--a highly conserved and selective anchor point for non-peptide ligands in chemokine receptors

    DEFF Research Database (Denmark)

    Rosenkilde, Mette M; Schwartz, Thue W

    2006-01-01

    to be crucially important for the binding and action of a number of non-peptide ligands in for example the CCR1, CCR2 and CCR5 receptors. It is proposed that in chemokine receptors in general GluVII:06 serves as a selective anchor point for the centrally located, positively charged nitrogen of the small molecule...

  3. The relaxin family peptide receptors and their ligands : new developments and paradigms in the evolution from jawless fish to mammals

    NARCIS (Netherlands)

    Yegorov, Sergey; Bogerd, Jan; Good, Sara V

    2014-01-01

    Relaxin family peptide receptors (Rxfps) and their ligands, relaxin (Rln) and insulin-like (Insl) peptides, are broadly implicated in the regulation of reproductive and neuroendocrine processes in mammals. Most placental mammals harbour genes for four receptors, namely rxfp1, rxfp2, rxfp3 and rxfp4.

  4. An endogenous aryl hydrocarbon receptor ligand inhibits proliferation and migration of human ovarian cancer cells.

    Science.gov (United States)

    Wang, Kai; Li, Yan; Jiang, Yi-Zhou; Dai, Cai-Feng; Patankar, Manish S; Song, Jia-Sheng; Zheng, Jing

    2013-10-28

    The aryl hydrocarbon receptor (AhR), a ligand-activated transcription factor mediates many biological processes. Herein, we investigated if 2-(1'H-indole-3'-carbonyl)-thiazole-4-carboxylic acid methyl ester (ITE, an endogenous AhR ligand) regulated proliferation and migration of human ovarian cancer cells via AhR. We found that AhR was widely present in many histotypes of ovarian cancer tissues. ITE suppressed OVCAR-3 cell proliferation and SKOV-3 cell migration in vitro, which were blocked by AhR knockdown. ITE also suppressed OVCAR-3 cell growth in mice. These data suggest that the ITE might potentially be used for therapeutic intervention for at least a subset of human ovarian cancer.

  5. Emerging Roles for CSF-1 Receptor and its Ligands in the Nervous System.

    Science.gov (United States)

    Chitu, Violeta; Gokhan, Şölen; Nandi, Sayan; Mehler, Mark F; Stanley, E Richard

    2016-06-01

    The colony-stimulating factor-1 receptor (CSF-1R) kinase regulates tissue macrophage homeostasis, osteoclastogenesis, and Paneth cell development. However, recent studies in mice have revealed that CSF-1R signaling directly controls the development and maintenance of microglia, and cell autonomously regulates neuronal differentiation and survival. While the CSF-1R-cognate ligands, CSF-1 and interleukin-34 (IL-34) compete for binding to the CSF-1R, they are expressed in a largely non-overlapping manner by mature neurons. The recent identification of a dominantly inherited, adult-onset, progressive dementia associated with inactivating mutations in the CSF-1R highlights the importance of CSF-1R signaling in the brain. We review the roles of the CSF-1R and its ligands in microglial and neural development and function, and their relevance to our understanding of neurodegenerative disease.

  6. Ligand binding affinities of arctigenin and its demethylated metabolites to estrogen receptor alpha.

    Science.gov (United States)

    Jin, Jong-Sik; Lee, Jong-Hyun; Hattori, Masao

    2013-01-01

    Phytoestrogens are defined as plant-derived compounds with estrogen-like activities according to their chemical structures and activities. Plant lignans are generally categorized as phytoestrogens. It was reported that (-)-arctigenin, the aglycone of arctiin, was demethylated to (-)-dihydroxyenterolactone (DHENL) by Eubacterium (E.) sp. ARC-2. Through stepwise demethylation, E. sp. ARC-2 produced six intermediates, three mono-desmethylarctigenins and three di-desmethylarctigenins. In the present study, ligand binding affinities of (-)-arctigenin and its seven metabolites, including DHENL, were investigated for an estrogen receptor alpha, and found that demethylated metabolites had stronger binding affinities than (-)-arctigenin using a ligand binding screen assay method. The IC(50) value of (2R,3R)-2-(4-hydroxy-3-methoxybenzyl)-3-(3,4-dihydroxybenzyl)-butyrolactone was 7.9 × 10⁻⁴ M. PMID:23325100

  7. Ligand-binding domains of nuclear receptors facilitate tight control of split CRISPR activity.

    Science.gov (United States)

    Nguyen, Duy P; Miyaoka, Yuichiro; Gilbert, Luke A; Mayerl, Steven J; Lee, Brian H; Weissman, Jonathan S; Conklin, Bruce R; Wells, James A

    2016-01-01

    Cas9-based RNA-guided nuclease (RGN) has emerged to be a versatile method for genome editing due to the ease of construction of RGN reagents to target specific genom