Sample records for adenosine receptor agonists

  1. AMP is an adenosine A1 receptor agonist. (United States)

    Rittiner, Joseph E; Korboukh, Ilia; Hull-Ryde, Emily A; Jin, Jian; Janzen, William P; Frye, Stephen V; Zylka, Mark J


    Numerous receptors for ATP, ADP, and adenosine exist; however, it is currently unknown whether a receptor for the related nucleotide adenosine 5'-monophosphate (AMP) exists. Using a novel cell-based assay to visualize adenosine receptor activation in real time, we found that AMP and a non-hydrolyzable AMP analog (deoxyadenosine 5'-monophosphonate, ACP) directly activated the adenosine A(1) receptor (A(1)R). In contrast, AMP only activated the adenosine A(2B) receptor (A(2B)R) after hydrolysis to adenosine by ecto-5'-nucleotidase (NT5E, CD73) or prostatic acid phosphatase (PAP, ACPP). Adenosine and AMP were equipotent human A(1)R agonists in our real-time assay and in a cAMP accumulation assay. ACP also depressed cAMP levels in mouse cortical neurons through activation of endogenous A(1)R. Non-selective purinergic receptor antagonists (pyridoxalphosphate-6-azophenyl-2',4'-disulfonic acid and suramin) did not block adenosine- or AMP-evoked activation. Moreover, mutation of His-251 in the human A(1)R ligand binding pocket reduced AMP potency without affecting adenosine potency. In contrast, mutation of a different binding pocket residue (His-278) eliminated responses to AMP and to adenosine. Taken together, our study indicates that the physiologically relevant nucleotide AMP is a full agonist of A(1)R. In addition, our study suggests that some of the physiological effects of AMP may be direct, and not indirect through ectonucleotidases that hydrolyze this nucleotide to adenosine.

  2. 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


    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.

  3. Identification of A3 adenosine receptor agonists as novel non-narcotic analgesics. (United States)

    Janes, K; Symons-Liguori, A M; Jacobson, K A; Salvemini, D


    Chronic pain negatively impacts the quality of life in a variety of patient populations. The current therapeutic repertoire is inadequate in managing patient pain and warrants the development of new therapeutics. Adenosine and its four cognate receptors (A1 , A2A , A2B and A3 ) have important roles in physiological and pathophysiological states, including chronic pain. Preclinical and clinical studies have revealed that while adenosine and agonists of the A1 and A2A receptors have antinociceptive properties, their therapeutic utility is limited by adverse cardiovascular side effects. In contrast, our understanding of the A3 receptor is only in its infancy, but exciting preclinical observations of A3 receptor antinociception, which have been bolstered by clinical trials of A3 receptor agonists in other disease states, suggest pain relief without cardiovascular side effects and with sufficient tolerability. Our goal herein is to briefly discuss adenosine and its receptors in the context of pathological pain and to consider the current data regarding A3 receptor-mediated antinociception. We will highlight recent findings regarding the impact of the A3 receptor on pain pathways and examine the current state of selective A3 receptor agonists used for these studies. The adenosine-to-A3 receptor pathway represents an important endogenous system that can be targeted to provide safe, effective pain relief from chronic pain.

  4. 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


    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 (

  5. Adenosine-A1 receptor agonist induced hyperalgesic priming type II. (United States)

    Araldi, Dioneia; Ferrari, Luiz F; Levine, Jon D


    We have recently shown that repeated exposure of the peripheral terminal of the primary afferent nociceptor to the mu-opioid receptor (MOR) agonist DAMGO ([D-Ala, N-Me-Phe, Gly-ol]-enkephalin acetate salt) induces a model of transition to chronic pain that we have termed type II hyperalgesic priming. Similar to type I hyperalgesic priming, there is a markedly prolonged response to subsequent administration of proalgesic cytokines, prototypically prostaglandin E2 (PGE2). However, type II hyperalgesic priming differs from type I in being rapidly induced, protein kinase A (PKA), rather than PKCε dependent, not reversed by a protein translation inhibitor, occurring in female as well as in male rats, and isolectin B4-negative neuron dependent. We report that, as with the repeated injection of a MOR agonist, the repeated administration of an agonist at the A1-adenosine receptor, also a Gi-protein coupled receptor, N-cyclopentyladenosine (CPA), also produces priming similar to DAMGO-induced type II hyperalgesic priming. In this study, we demonstrate that priming induced by repeated exposure to this A1-adenosine receptor agonist shares the same mechanisms, as MOR-agonist induced priming. However, the prolongation of PGE2 hyperalgesia induced by repeated administration of CPA depends on G-protein αi subunit activation, differently from DAMGO-induced type II priming, in which it depends on the β/γ subunit. These data implicate a novel form of Gi-protein signaling pathway in the type II hyperalgesic priming induced by repeated administration of an agonist at A1-adenosine receptor to the peripheral terminal of the nociceptor.

  6. The synthesis of a series of adenosine A3 receptor agonists. (United States)

    Broadley, Kenneth J; Burnell, Erica; Davies, Robin H; Lee, Alan T L; Snee, Stephen; Thomas, Eric J


    A series of 1'-(6-aminopurin-9-yl)-1'-deoxy-N-methyl-β-d-ribofuranuronamides that were characterised by 2-dialkylamino-7-methyloxazolo[4,5-b]pyridin-5-ylmethyl substituents on N6 of interest for screening as selective adenosine A3 receptor agonists, have been synthesised. This work involved the synthesis of 2-dialkylamino-5-aminomethyl-7-methyloxazolo[4,5-b]pyridines and analogues that were coupled with the known 1'-(6-chloropurin-9-yl)-1'-deoxy-N-methyl-β-d-ribofuranuronamide. The oxazolo[4,5-b]pyridines were synthesized by regioselective functionalisation of 2,4-dimethylpyridine N-oxides. The regioselectivities of these reactions were found to depend upon the nature of the heterocycle with 2-dimethylamino-5,7-dimethyloxazolo[4,5-b]pyridine-N-oxide undergoing regioselective functionalisation at the 7-methyl group on reaction with trifluoroacetic anhydride in contrast to the reaction of 4,6-dimethyl-3-hydroxypyridine-N-oxide with acetic anhydride that resulted in functionalisation of the 6-methyl group. To optimise selectivity for the A3 receptor, 5-aminomethyl-7-bromo-2-dimethylamino-4-[(3-methylisoxazol-5-yl)methoxy]benzo[d]oxazole was synthesised and coupled with the 1'-(6-chloropurin-9-yl)-1'-deoxy-N-methyl-β-d-ribofuranuronamide. The products were active as selective adenosine A3 agonists.

  7. Structural and energetic effects of A2A adenosine receptor mutations on agonist and antagonist binding.

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    Henrik Keränen

    Full Text Available To predict structural and energetic effects of point mutations on ligand binding is of considerable interest in biochemistry and pharmacology. This is not only useful in connection with site-directed mutagenesis experiments, but could also allow interpretation and prediction of individual responses to drug treatment. For G-protein coupled receptors systematic mutagenesis has provided the major part of functional data as structural information until recently has been very limited. For the pharmacologically important A(2A adenosine receptor, extensive site-directed mutagenesis data on agonist and antagonist binding is available and crystal structures of both types of complexes have been determined. Here, we employ a computational strategy, based on molecular dynamics free energy simulations, to rationalize and interpret available alanine-scanning experiments for both agonist and antagonist binding to this receptor. These computer simulations show excellent agreement with the experimental data and, most importantly, reveal the molecular details behind the observed effects which are often not immediately evident from the crystal structures. The work further provides a distinct validation of the computational strategy used to assess effects of point-mutations on ligand binding. It also highlights the importance of considering not only protein-ligand interactions but also those mediated by solvent water molecules, in ligand design projects.

  8. The effects of the adenosine A3 receptor agonist IB-MECA on sodium taurocholate-induced experimental acute pancreatitis. (United States)

    Prozorow-Krol, Beata; Korolczuk, Agnieszka; Czechowska, Grazyna; Slomka, Maria; Madro, Agnieszka; Celinski, Krzysztof


    The role of adenosine A3 receptors and their distribution in the gastrointestinal tract have been widely investigated. Most of the reports discuss their role in intestinal inflammations. However, the role of adenosine A3 receptor agonist in pancreatitis has not been well established. The aim of this study is [corrected] to evaluate the effects of the adenosine A3 receptor agonist on the course of sodium taurocholate-induced experimental acute pancreatitis (EAP). The experiments were performed on 80 male Wistar rats, 58 of which survived, subdivided into 3 groups: C--control rats, I--EAP group, and II--EAP group treated with the adenosine A3 receptor agonist IB-MECA (1-deoxy-1-6[[(3-iodophenyl) methyl]amino]-9H-purin-9-yl)-N-methyl-B-D-ribofuronamide at a dose of 0.75 mg/kg b.w. i.p. at 48, 24, 12 and 1 h before and 1 h after the injection of 5% sodium taurocholate solution into the biliary-pancreatic duct. Serum for α-amylase and lipase determinations and tissue samples for morphological examinations were collected at 2, 6, and 24 h of the experiment. In the IB-MECA group, α-amylase activity was decreased with statistically high significance compared to group I. The activity of lipase was not significantly different among the experimental groups but higher than in the control group. The administration of IB-MECA attenuated the histological parameters of inflammation as compared to untreated animals. The use of A3 receptor agonist IB-MECA attenuates EAP. Our findings suggest that stimulation of adenosine A3 receptors plays a positive role in the sodium taurocholate-induced EAP in rats.

  9. Direct visualisation of internalization of the adenosine A3 receptor and localization with arrestin3 using a fluorescent agonist. (United States)

    Stoddart, Leigh A; Vernall, Andrea J; Briddon, Stephen J; Kellam, Barrie; Hill, Stephen J


    Fluorescence based probes provide a novel way to study the dynamic internalization process of G protein-coupled receptors (GPCRs). Recent advances in the rational design of fluorescent ligands for GPCRs have been used here to generate new fluorescent agonists containing tripeptide linkers for the adenosine A3 receptor. The fluorescent agonist BY630-X-(D)-A-(D)-A-G-ABEA was found to be a highly potent agonist at the adenosine A3 receptor in both reporter gene (pEC50 = 8.48 ± 0.09) and internalization assays (pEC50 = 7.47 ± 0.11). Confocal imaging studies showed that BY630-X-(D)-A-(D)-A-G-ABEA was internalized with A3 linked to yellow fluorescent protein, which was blocked by the competitive antagonist MRS1220. Internalization of untagged adenosine A3 could also be visualized with BY630-X-(D)-A-(D)-A-G-ABEA treatment. Further, BY630-X-(D)-A-(D)-A-G-ABEA stimulated the formation of receptor-arrestin3 complexes and was found to localize with these intracellular complexes. This highly potent agonist with excellent imaging properties should be a valuable tool to study receptor internalization. This article is part of the Special Issue entitled 'Fluorescent Tools in Neuropharmacology'.

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

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    Esposito Emanuela


    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

  11. Differential effects of the adenosine A1 receptor agonist adenosine amine congener on renal, femoral and carotid vascular conductance in preterm fetal sheep. (United States)

    Booth, Lindsea C; Tummers, Leonie; Jensen, Ellen C; Barrett, Carolyn J; Malpas, Simon C; Gunn, Alistair J; Bennet, Laura


    1. Adenosine A(1) receptor activation is critical for endogenous neuroprotection from hypoxia-ischaemia, raising the possibility that treatment with A(1) receptor agonists may be an effective physiological protection strategy for vulnerable preterm infants. However, the A(1) receptor can mediate unwanted systemic effects, including vasoconstriction of the afferent glomerular arteriole. There is limited information on whether this occurs at doses that improve cerebral perfusion in the immature brain. 2. Therefore, in the present study, we examined whether infusion of the selective A(1) receptor agonist adenosine amine congener (ADAC) is associated with reduced renal perfusion in chronically instrumented preterm (0.7 gestation) fetal sheep. In the present study, ADAC was given in successive doses of 2.5, 5.0 and 15.0 microg, 45 min apart. 3. Treatment with ADAC was associated with a marked reduction in renal vascular conductance (and blood flow), whereas carotid conductance was increased and there was no significant effect on femoral conductance. In contrast with the stable effects of increasing ADAC dose on vascular conductance, there was a significant dose-related fall in fetal heart rate and blood pressure. 4. In conclusion, these short-term data support the concern that A(1) receptor agonist infusion can selectively impair renal perfusion, even at low doses.

  12. Adenosine and adenosine receptors: Newer therapeutic perspective

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    Manjunath S


    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.

  13. Promotion of Wound Healing by an Agonist of Adenosine A2A Receptor Is Dependent on Tissue Plasminogen Activator. (United States)

    Montesinos, M Carmen; Desai-Merchant, Avani; Cronstein, Bruce N


    Impaired wound healing, as it occurs in diabetes mellitus or long-term corticoid treatment, is commonly associated with disability, diminished quality of life, and high economic costs. Selective agonists of the A2A receptor subtype of adenosine, an endogenous regulator of inflammation, promote tissue repair in animal models, both healthy and with impaired healing. Plasmin-mediated proteolysis of fibrin and other matrix proteins is essential for cell migration at sites of injury. Since adenosine A2A receptor activation increases plasminogen activator release from macrophages and mast cells, we studied the effect of a selective agonist, CGS-21680, on full-thickness excisional wound closure in wild-type, urokinase plasminogen activator (uPA)-deficient, and tissue plasminogen activator (tPA)-deficient mice. Wound closure was impaired in tPA- and uPA-deficient mice as compared with wild-type mice, and topical application of CGS-21680 significantly increased the rate at which wounds closed in wild-type mice and uPA-deficient mice, but not in tPA-deficient mice. Immunostaining of tissue sections showed that tPA was present in endothelial cells and histiocytes by day 3 post-wound and also by day 6. In contrast, uPA was more prominent in these cell types only by day 6 post-wound. Our results confirm that plasminogen activation contributes to wound repair and are consistent with the hypothesis that adenosine A2A receptor activation promotes wound closure by a mechanism that depends upon tPA, but not uPA. Moreover, our results suggest that topical adenosine A2A receptor agonists may be useful in promotion of wound closure in patients with impaired wound healing.

  14. Effects of synthetic A3 adenosine receptor agonists on cell proliferation and viability are receptor independent at micromolar concentrations. (United States)

    Mlejnek, Petr; Dolezel, Petr; Frydrych, Ivo


    The question as to whether A3 adenosine receptor (A3AR) agonists, N (6)-(3-iodobenzyl)-adenosine-5'-N- methyluronamide (IB-MECA) and 2-chloro-N (6)-(3-iodobenzyl)-adenosine-5'-N-methyluronamide (Cl-IB-MECA), could exert cytotoxic effects at high concentrations with or without the involvement of A3AR has been a controversial issue for a long time. The initial findings suggesting that A3AR plays a crucial role in the induction of cell death upon treatment with micromolar concentrations of IB-MECA or Cl-IB-MECA were revised, however, the direct and unequivocal evidence is still missing. Therefore, the sensitivity of Chinese hamster ovary (CHO) cells transfected with human recombinant A3AR (A3-CHO) and their counter partner wild-type CHO cells, which do not express any of adenosine receptors, to micromolar concentrations of IB-MECA and Cl-IB-MECA was studied. We observed that IB-MECA and Cl-IB-MECA exhibited a strong inhibitory effect on cell proliferation due to the blockage of cell cycle progression at G1/S and G2/M transitions in both A3-CHO and CHO cells. Further analysis revealed that IB-MECA and Cl-IB-MECA attenuated the Erk1/2 signalling irrespectively to A3AR expression. In addition, Cl-IB-MECA induced massive cell death mainly with hallmarks of a necrosis in both cell lines. In contrast, IB-MECA affected cell viability only slightly independently of A3AR expression. IB-MECA induced cell death that exhibited apoptotic hallmarks. In general, the sensitivity of A3-CHO cells to micromolar concentrations of IB-MECA and Cl-IB-MECA was somewhat, but not significantly, higher than that observed in the CHO cells. These results strongly suggest that IB-MECA and Cl-IB-MECA exert cytotoxic effects at micromolar concentrations independently of A3AR expression.

  15. Structure-kinetics relationships of Capadenoson derivatives as adenosine A1 receptor agonists. (United States)

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


    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.

  16. 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


    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...... and in combination with D-amphetamine and (-)-apomorphine. The monkeys were videotaped after drug administration and the tapes were rated for EPS and psychosis-like symptoms. CGS 21680 decreased apomorphine-induced behavioural unrest, arousal (0.01-0.05 mg/kg) and stereotypies (0.05 mg/kg) while amphetamine...... showed a functional anti-dopaminergic effect in Cebus apella monkeys without production of EPS. This further substantiates that adenosine A2A receptor agonists may have potential as antipsychotics with atypical profiles....

  17. 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;


    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...... and in combination with D-amphetamine and (-)-apomorphine. The monkeys were videotaped after drug administration and the tapes were rated for EPS and psychosis-like symptoms. CGS 21680 decreased apomorphine-induced behavioural unrest, arousal (0.01-0.05 mg/kg) and stereotypies (0.05 mg/kg) while amphetamine...

  18. Central or peripheral delivery of an adenosine A1 receptor agonist improves mechanical allodynia in a mouse model of painful diabetic neuropathy. (United States)

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


    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

  19. The role of adenosine receptor agonist and antagonist on Hippocampal MDMA detrimental effects; a structural and behavioral study. (United States)

    Kermanian, Fatemeh; Mehdizadeh, Mehdi; Soleimani, Mansureh; Ebrahimzadeh Bideskan, Ali Reza; Asadi-Shekaari, Majid; Kheradmand, Hamed; Haghir, Hossein


    There is abundant evidence showing that repeated use of MDMA (3, 4-Methylenedioxymethamphetamine, ecstasy) has been associated with depression, anxiety and deficits in learning and memory, suggesting detrimental effects on hippocampus. Adenosine is an endogenous purine nucleoside that has a neuromodulatory role in the central nervous system. In the present study, we investigated the role of A2a adenosine receptors agonist (CGS) and antagonist (SCH) on the body temperature, learning deficits, and hippocampal cell death induced by MDMA administration. In this study, 63 adult, male, Sprague - Dawley rats were subjected to MDMA (10 and 20 mg/kg) followed by intraperitoneal CGS (0.03 mg/kg) or SCH (0.03 mg/kg) injection. The animals were tested for spatial learning in the Morris water maze (MWM) task performance, accompanied by a recording of body temperature, electron microscopy and stereological study. Our results showed that MDMA treatment increased body temperature significantly, and impaired the ability of rats to locate the hidden platform(P mechanism of these interactions requires further studies.

  20. Purine (N)-Methanocarba Nucleoside Derivatives Lacking an Exocyclic Amine as Selective A3 Adenosine Receptor Agonists. (United States)

    Tosh, Dilip K; Ciancetta, Antonella; Warnick, Eugene; O'Connor, Robert; Chen, Zhoumou; Gizewski, Elizabeth; Crane, Steven; Gao, Zhan-Guo; Auchampach, John A; Salvemini, Daniela; Jacobson, Kenneth A


    Purine (N)-methanocarba-5'-N-alkyluronamidoriboside A3 adenosine receptor (A3AR) agonists lacking an exocyclic amine resulted from an unexpected reaction during a Sonogashira coupling and subsequent aminolysis. Because the initial C6-Me and C6-styryl derivatives had unexpectedly high A3AR affinity, other rigid nucleoside analogues lacking an exocyclic amine were prepared. Of these, the C6-Me-(2-phenylethynyl) and C2-(5-chlorothienylethynyl) analogues were particularly potent, with human A3AR Ki values of 6 and 42 nM, respectively. Additionally, the C2-(5-chlorothienyl)-6-H analogue was potent and selective at A3AR (MRS7220, Ki 60 nM) and also completely reversed mouse sciatic nerve mechanoallodynia (in vivo, 3 μmol/kg, po). The lack of a C6 H-bond donor while maintaining A3AR affinity and efficacy could be rationalized by homology modeling and docking of these hypermodified nucleosides. The modeling suggests that a suitable combination of stabilizing features can partially compensate for the lack of an exocyclic amine, an otherwise important contributor to recognition in the A3AR binding site.

  1. Calcium modulates calmodulin/α-actinin 1 interaction with and agonist-dependent internalization of the adenosine A2A receptor. (United States)

    Piirainen, Henni; Taura, Jaume; Kursula, Petri; Ciruela, Francisco; Jaakola, Veli-Pekka


    Adenosine receptors are G protein-coupled receptors that sense extracellular adenosine to transmit intracellular signals. One of the four adenosine receptor subtypes, the adenosine A2A receptor (A2AR), has an exceptionally long intracellular C terminus (A2AR-ct) that mediates interactions with a large array of proteins, including calmodulin and α-actinin. Here, we aimed to ascertain the α-actinin 1/calmodulin interplay whilst binding to A2AR and the role of Ca(2+) in this process. First, we studied the A2AR-α-actinin 1 interaction by means of native polyacrylamide gel electrophoresis, isothermal titration calorimetry, and surface plasmon resonance, using purified recombinant proteins. α-Actinin 1 binds the A2AR-ct through its distal calmodulin-like domain in a Ca(2+)-independent manner with a dissociation constant of 5-12μM, thus showing an ~100 times lower affinity compared to the A2AR-calmodulin/Ca(2+) complex. Importantly, calmodulin displaced α-actinin 1 from the A2AR-ct in a Ca(2+)-dependent fashion, disrupting the A2AR-α-actinin 1 complex. Finally, we assessed the impact of Ca(2+) on A2AR internalization in living cells, a function operated by the A2AR-α-actinin 1 complex. Interestingly, while Ca(2+) influx did not affect constitutive A2AR endocytosis, it abolished agonist-dependent internalization. In addition, we demonstrated that the A2AR/α-actinin interaction plays a pivotal role in receptor internalization and function. Overall, our results suggest that the interplay of A2AR with calmodulin and α-actinin 1 is fine-tuned by Ca(2+), a fact that might power agonist-mediated receptor internalization and function.

  2. Administration of caffeine inhibited adenosine receptor agonist-induced decreases in motor performance, thermoregulation, and brain neurotransmitter release in exercising rats. (United States)

    Zheng, Xinyan; Hasegawa, Hiroshi


    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.

  3. Adenosine A2A receptor agonist prevents cardiac remodeling and dysfunction in spontaneously hypertensive male rats after myocardial infarction (United States)

    da Silva, Jaqueline S; Gabriel-Costa, Daniele; Sudo, Roberto T; Wang, Hao; Groban, Leanne; Ferraz, Emanuele B; Nascimento, José Hamilton M; Fraga, Carlos Alberto M; Barreiro, Eliezer J; Zapata-Sudo, Gisele


    Background This work evaluated the hypothesis that 3,4-methylenedioxybenzoyl-2-thienylhydrazone (LASSBio-294), an agonist of adenosine A2A receptor, could be beneficial for preventing cardiac dysfunction due to hypertension associated with myocardial infarction (MI). Methods Male spontaneously hypertensive rats (SHR) were randomly divided into four groups (six animals per group): sham-operation (SHR-Sham), and myocardial infarction rats (SHR-MI) were treated orally either with vehicle or LASSBio-294 (10 and 20−1.d−1) for 4 weeks. Echocardiography and in vivo hemodynamic parameters measured left ventricle (LV) structure and function. Exercise tolerance was evaluated using a treadmill test. Cardiac remodeling was accessed by LV collagen deposition and tumor necrosis factor α expression. Results Early mitral inflow velocity was significantly reduced in the SHR-MI group, and there was significant recovery in a dose-dependent manner after treatment with LASSBio-294. Exercise intolerance observed in the SHR-MI group was prevented by 10−1.d−1 of LASS-Bio-294, and exercise tolerance exceeded that of the SHR-Sham group at 20−1.d−1. LV end-diastolic pressure increased after MI, and this was prevented by 10 and 20−1.d−1 of LASSBio-294. Sarcoplasmic reticulum Ca2+ ATPase levels were restored in a dose-dependent manner after treatment with LASSBio-294. Fibrosis and inflammatory processes were also counteracted by LASSBio-294, with reductions in LV collagen deposition and tumor necrosis factor α expression. Conclusion In summary, oral administration of LASSBio-294 after MI in a dose-dependent manner prevented the development of cardiac dysfunction, demonstrating this compound’s potential as an alternative treatment for heart failure in the setting of ischemic heart disease with superimposed chronic hypertension.

  4. Adenosine receptor neurobiology: overview. (United States)

    Chen, Jiang-Fan; Lee, Chien-fei; Chern, Yijuang


    Adenosine is a naturally occurring nucleoside that is distributed ubiquitously throughout the body as a metabolic intermediary. In the brain, adenosine functions as an important upstream neuromodulator of a broad spectrum of neurotransmitters, receptors, and signaling pathways. By acting through four G-protein-coupled receptors, adenosine contributes critically to homeostasis and neuromodulatory control of a variety of normal and abnormal brain functions, ranging from synaptic plasticity, to cognition, to sleep, to motor activity to neuroinflammation, and cell death. This review begun with an overview of the gene and genome structure and the expression pattern of adenosine receptors (ARs). We feature several new developments over the past decade in our understanding of AR functions in the brain, with special focus on the identification and characterization of canonical and noncanonical signaling pathways of ARs. We provide an update on functional insights from complementary genetic-knockout and pharmacological studies on the AR control of various brain functions. We also highlight several novel and recent developments of AR neurobiology, including (i) recent breakthrough in high resolution of three-dimension structure of adenosine A2A receptors (A2ARs) in several functional status, (ii) receptor-receptor heterodimerization, (iii) AR function in glial cells, and (iv) the druggability of AR. We concluded the review with the contention that these new developments extend and strengthen the support for A1 and A2ARs in brain as therapeutic targets for neurologic and psychiatric diseases.

  5. GLP-1 Receptor Agonists (United States)

    ... in Balance › GLP-1 Receptor Agonists Fact Sheet GLP-1 Receptor Agonists May, 2012 Download PDFs English Espanol Editors Silvio ... are too high or too low. What are GLP-1 receptor agonist medicines? GLP-1 receptor agonist medicines, also called ...

  6. Effects of adenosine agonist R-phenylisopropyl-adenosine on halothane anesthesia and antinociception in rats

    Institute of Scientific and Technical Information of China (English)

    Hai-chun MA; Yan-fen WANG; Chun-sheng FENG; Hua ZHAO; Shuji DOHI


    Aim: To investigate the antinociceptive effect of adenosine agonist Rphenylisopropyl-adenosine (R-PIA) given to conscious rats by intracerebroventricular (ICV) and intrathecal (IT), and identify the effect of R-PIA on minimum alveolar concentration (MAC) of halothane with pretreatment of A1 receptor an tagonist 8-cyclopentyl-1,3-dipropylxanthine (DPCPX) or K+ channel blocker 4-aminopyridine (4-AP). Methods: Sprague-Dawley rats were implanted with 24 gauge stainless steel guide cannula using stereotaxic apparatus and ICV method, and an IT catheter (PE-10, 8.5 cm) was inserted into the lumbar subarachnoid space, while the rats were under pentobarbital anesthesia. After one week of recovery from surgery, rats were randomly assigned to one of the following protocols: MAC of halothane, or tail-flick latency. All measurements were performed after R-PIA (0.8-2.0 μg) microinjection into ICV and IT with or without pretreatment of DPCPX or 4-AP. Results: Microinjection of adenosine agonist R PIA in doses of 0.8-2.0 μg into ICV and IT produced a significant dose- and time dependent antinociceptive action as reflected by increasing latency times and ICV administration of adenosine agonist R-PIA (0.8 μg) reducing halothane anes thetic requirements (by 29%). The antinociception and reducing halothane requirements effected by adenosine agonist R-PIA was abolished by DPCPX and 4-AP. Conclusion: ICV and IT administration of adenosine agonist R-PIA produced an antinociceptive effect in a dose-dependent manner and decreased hal othane MAC with painful stimulation through activation of A1 receptor subtype, and the underlying mechanism involves K+ channel activation.

  7. An adenosine A3 receptor agonist inhibits DSS-induced colitis in mice through modulation of the NF-κB signaling pathway. (United States)

    Ren, Tianhua; Tian, Ting; Feng, Xiao; Ye, Shicai; Wang, Hao; Wu, Weiyun; Qiu, Yumei; Yu, Caiyuan; He, Yanting; Zeng, Juncheng; Cen, Junwei; Zhou, Yu


    The role of the adenosine A3 receptor (A3AR) in experimental colitis is controversial. The A3AR agonist N(6)-(3-iodobenzyl)adenosine-5'-N-methyluronamide (IB-MECA) has been shown to have a clinical benefit, although studies in A3AR-deficient mice suggest a pro-inflammatory role. However, there are no studies on the effect of 2-Cl-IB-MECA and the molecular mechanism of action of A3AR in murine colitis models in vivo. Is it the same as that observed in vitro? The interaction between 2-CL-IB-MECA and A3AR in a murine colitis model and the signaling pathways associated with this interaction remain unclear. Here we demonstrate a role for the NF-κB signaling pathway and its effect on modifying the activity of proinflammatory factors in A3AR-mediated biological processes. Our results demonstrated that A3AR activation possessed marked effects on experimental colitis through the NF-κB signaling pathway.

  8. Topical application of the adenosine A2A receptor agonist CGS-21680 prevents phorbol-induced epidermal hyperplasia and inflammation in mice. (United States)

    Arasa, Jorge; Martos, Patricio; Terencio, María Carmen; Valcuende-Cavero, Francisca; Montesinos, María Carmen


    The nucleoside adenosine is a known regulator of immunity and inflammation that mediates, at least in part, the anti-inflammatory effect of methotrexate, an immunosuppressive agent widely used to treat autoimmune inflammatory diseases. Adenosine A2A receptors play a key role in the inhibition of the inflammatory process besides promoting wound healing. Therefore, we aimed to determine the topical effect of a selective agonist, CGS-21680, on a murine model of skin hyperplasia with a marked inflammatory component. Pretreatment with either CGS-21680 (5 μg per site) or the reference agent dexamethasone (200 μg/site) prevented the epidermal hyperplasia and inflammatory response induced by topical application of 12-O-tetradecanoylphorbol-13-acetate (TPA, 2 nmol/site) for three consecutive days. The histological analysis showed that both CGS-21680 and dexamethasone produced a marked reduction of inflammatory cell infiltrate, which correlated with diminished myeloperoxidase (MPO) activity in skin homogenates. Both treatments reduced the levels of the chemotactic mediators LTB4 and CXCL-1, and the inflammatory cytokine TNF-α, through the suppression of NFκB phosphorylation. The immunohistochemical analysis of the hyperproliferative markers cytokeratin 6 (CK6) and Ki67 revealed that while both agents inhibit the number of proliferating cells in the epidermis, CGS-21680 treatment promoted dermal fibroblasts proliferation. Consistently, increased collagen deposition in dermis was observed in tissue sections from agonist-treated mice. Our results showed that CGS 21680 efficiently prevents phorbol-induced epidermal hyperplasia and inflammation in mice without the deleterious atrophic effect of topical corticosteroids.

  9. Adenosine A2A  receptor agonist prevents cardiac remodeling and dysfunction in spontaneously hypertensive male rats after myocardial infarction

    Directory of Open Access Journals (Sweden)

    da Silva JS


    Full Text Available Jaqueline S da Silva,1 Daniele Gabriel-Costa,1 Roberto T Sudo,1 Hao Wang,2 Leanne Groban,2 Emanuele B Ferraz,3 José Hamilton M Nascimento,3 Carlos Alberto M Fraga,1 Eliezer J Barreiro,1 Gisele Zapata-Sudo1 1Research Program Development of Drugs, Institute of Biomedical Sciences, Universidade Federal do Rio de Janeiro, Rio de Janeiro, Brazil; 2Department of Anesthesiology, Wake Forest School of Medicine, Winston-Salem, NC, USA; 3Institute of Biophysics Carlos Chagas Filho, Universidade Federal do Rio de Janeiro, Rio de Janeiro, Brazil Background: This work evaluated the hypothesis that 3,4-methylenedioxybenzoyl-2- thienylhydrazone (LASSBio-294, an agonist of adenosine A2A  receptor, could be beneficial for preventing cardiac dysfunction due to hypertension associated with myocardial infarction (MI. Methods: Male spontaneously hypertensive rats (SHR were randomly divided into four groups (six animals per group: sham-operation (SHR-Sham, and myocardial infarction rats (SHR-MI were treated orally either with vehicle or LASSBio-294 (10 and 20 for 4 weeks. Echocardiography and in vivo hemodynamic parameters measured left ventricle (LV structure and function. Exercise tolerance was evaluated using a treadmill test. Cardiac remodeling was accessed by LV collagen deposition and tumor necrosis factor α expression. Results: Early mitral inflow velocity was significantly reduced in the SHR-MI group, and there was significant recovery in a dose-dependent manner after treatment with LASSBio-294. Exercise intolerance observed in the SHR-MI group was prevented by 10 of LASSBio-294, and exercise tolerance exceeded that of the SHR-Sham group at 20 LV end-diastolic pressure increased after MI, and this was prevented by 10 and 20 of LASSBio-294. Sarcoplasmic reticulum Ca2+ ATPase levels were restored in a dose-dependent manner after treatment with LASSBio-294. Fibrosis and inflammatory processes were also

  10. Glutamate receptor agonists

    DEFF Research Database (Denmark)

    Vogensen, Stine Byskov; Greenwood, Jeremy R; Bunch, Lennart;


    The neurotransmitter (S)-glutamate [(S)-Glu] is responsible for most of the excitatory neurotransmission in the central nervous system. The effect of (S)-Glu is mediated by both ionotropic and metabotropic receptors. Glutamate receptor agonists are generally a-amino acids with one or more...... stereogenic centers due to strict requirements in the agonist binding pocket of the activated state of the receptor. By contrast, there are many examples of achiral competitive antagonists. The present review addresses how stereochemistry affects the activity of glutamate receptor ligands. The review focuses...... mainly on agonists and discusses stereochemical and conformational considerations as well as biostructural knowledge of the agonist binding pockets, which is useful in the design of glutamate receptor agonists. Examples are chosen to demonstrate how stereochemistry not only determines how the agonist...

  11. CF101, An Agonist to the A3 Adenosine Receptor, Enhances the Chemotherapeutic Effect of 5-Fluorouracil in a Colon Carcinoma Murine Model

    Directory of Open Access Journals (Sweden)

    Sara Bar-Yehuda


    Full Text Available NF-κB and the upstream kinase PKB/Akt are highly expressed in chemoresistance tumor cells and may hamper the apoptotic pathway. CF101, a specific agonist to the A3 adenosine receptor, inhibits the development of colon carcinoma growth in cell cultures and xenograft murine models. Because CF101 has been shown to downregulate PKB/Akt and NF-κB protein expression level, we presumed that its combination with chemotherapy will enhance the antitumor effect of the cytotoxic drug. In this study, we utilized 3-[4,5Dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT and colony formation assays and a colon carcinoma xenograft model. It has been shown that a combined treatment of CF101 and 5-fluorouracil (5-FU enhanced the cytotoxic effect of the latter on HCT-116 human colon carcinoma growth. Downregulation of PKB/Akt, NF-κB, and cyclin D1, and upregulation of caspase-3 protein expression level were observed in cells and tumor lesions on treatment with a combination of CF101 and 5-FU. Moreover, in mice treated with the combined therapy, myelotoxicity was prevented as was evidenced by normal white blood cell and neutrophil counts. These results show that CF101 potentiates the cytotoxic effect of 5-FU, thus preventing drug resistance. The myeloprotective effect of CF101 suggests its development as an add-on treatment to 5-FU.

  12. Adenosine Receptors: Expression, Function and Regulation

    Directory of Open Access Journals (Sweden)

    Sandeep Sheth


    Full Text Available Adenosine receptors (ARs comprise a group of G protein-coupled receptors (GPCR which mediate the physiological actions of adenosine. To date, four AR subtypes have been cloned and identified in different tissues. These receptors have distinct localization, signal transduction pathways and different means of regulation upon exposure to agonists. This review will describe the biochemical characteristics and signaling cascade associated with each receptor and provide insight into how these receptors are regulated in response to agonists. A key property of some of these receptors is their ability to serve as sensors of cellular oxidative stress, which is transmitted by transcription factors, such as nuclear factor (NF-κB, to regulate the expression of ARs. Recent observations of oligomerization of these receptors into homo- and heterodimers will be discussed. In addition, the importance of these receptors in the regulation of normal and pathological processes such as sleep, the development of cancers and in protection against hearing loss will be examined.

  13. Adenosine modulation of [Ca2+]i in cerebellar granular cells: multiple adenosine receptors involved. (United States)

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


    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.

  14. Different efficacy of adenosine and NECA derivatives at the human A3 adenosine receptor: insight into the receptor activation switch. (United States)

    Dal Ben, Diego; Buccioni, Michela; Lambertucci, Catia; Kachler, Sonja; Falgner, Nico; Marucci, Gabriella; Thomas, Ajiroghene; Cristalli, Gloria; Volpini, Rosaria; Klotz, Karl-Norbert


    A3 Adenosine receptors are promising drug targets for a number of diseases and intense efforts are dedicated to develop selective agonists and antagonists of these receptors. A series of adenosine derivatives with 2-(ar)-alkynyl chains, with high affinity and different degrees of selectivity for human A3 adenosine receptors was tested for the ability to inhibit forskolin-stimulated adenylyl cyclase. All these derivatives are partial agonists at A3 adenosine receptors; their efficacy is not significantly modified by the introduction of small alkyl substituents in the N(6)-position. In contrast, the adenosine-5'-N-ethyluronamide (NECA) analogs of 2-(ar)-alkynyladenosine derivatives are full A3 agonists. Molecular modeling analyses were performed considering both the conformational behavior of the ligands and the impact of 2- and 5'-substituents on ligand-target interaction. The results suggest an explanation for the different agonistic behavior of adenosine and NECA derivatives, respectively. A sub-pocket of the binding site was analyzed as a crucial interaction domain for receptor activation.

  15. [Melatonin receptor agonist]. (United States)

    Uchiyama, Makoto


    Melatonin is a hormone secreted by the pineal gland and is involved in the regulation of human sleep-wake cycle and circadian rhythms. The melatonin MT1 and MT2 receptors located in the suprachiasmatic nucleus in the hypothalamus play a pivotal role in the sleep-wake regulation. Based on the fact that MT1 receptors are involved in human sleep onset process, melatonin receptor agonists have been developed to treat insomnia. In this article, we first reviewed functions of melatonin receptors with special reference to MT1 and MT2, and properties and clinical application of melatonin receptor agonists as hypnotics.

  16. The A3 adenosine receptor: history and perspectives. (United States)

    Borea, Pier Andrea; Varani, Katia; Vincenzi, Fabrizio; Baraldi, Pier Giovanni; Tabrizi, Mojgan Aghazadeh; Merighi, Stefania; Gessi, Stefania


    By general consensus, the omnipresent purine nucleoside adenosine is considered a major regulator of local tissue function, especially when energy supply fails to meet cellular energy demand. Adenosine mediation involves activation of a family of four G protein-coupled adenosine receptors (ARs): A(1), A(2)A, A(2)B, and A(3). The A(3) adenosine receptor (A(3)AR) is the only adenosine subtype to be overexpressed in inflammatory and cancer cells, thus making it a potential target for therapy. Originally isolated as an orphan receptor, A(3)AR presented a twofold nature under different pathophysiologic conditions: it appeared to be protective/harmful under ischemic conditions, pro/anti-inflammatory, and pro/antitumoral depending on the systems investigated. Until recently, the greatest and most intriguing challenge has been to understand whether, and in which cases, selective A(3) agonists or antagonists would be the best choice. Today, the choice has been made and A(3)AR agonists are now under clinical development for some disorders including rheumatoid arthritis, psoriasis, glaucoma, and hepatocellular carcinoma. More specifically, the interest and relevance of these new agents derives from clinical data demonstrating that A(3)AR agonists are both effective and safe. Thus, it will become apparent in the present review that purine scientists do seem to be getting closer to their goal: the incorporation of adenosine ligands into drugs with the ability to save lives and improve human health.

  17. Transport of A1 adenosine receptor agonist tecadenoson by human and mouse nucleoside transporters: evidence for blood-brain barrier transport by murine equilibrative nucleoside transporter 1 mENT1. (United States)

    Lepist, Eve-Irene; Damaraju, Vijaya L; Zhang, Jing; Gati, Wendy P; Yao, Sylvia Y M; Smith, Kyla M; Karpinski, Edward; Young, James D; Leung, Kwan H; Cass, Carol E


    The high density of A1 adenosine receptors in the brain results in significant potential for central nervous system (CNS)-related adverse effects with A1 agonists. Tecadenoson is a selective A1 adenosine receptor agonist with close similarity to adenosine. We studied the binding and transmembrane transport of tecadenoson by recombinant human equilibrative nucleoside transporters (hENTs) hENT1 and hENT2, and human concentrative nucleoside transporters (hCNTs) hCNT1, hCNT2, and hCNT3 in vitro and by mouse mENT1 in vivo. Binding affinities of the five recombinant human nucleoside transporters for tecadenoson differed (hENT1 > hCNT1 > hCNT3 > hENT2 > hCNT2), and tecadenoson was transported largely by hENT1. Pretreatment of mice with a phosphorylated prodrug of nitrobenzylmercaptopurine riboside, an inhibitor of mENT1, significantly decreased brain exposure to tecadenoson compared with that of the untreated (control) group, suggesting involvement of mENT1 in transport of tecadenoson across the blood-brain barrier (BBB). In summary, ENT1 was shown to mediate the transport of tecadenoson in vitro with recombinant and native human protein and in vivo with mice. The micromolar apparent Km value of tecadenoson for transport by native hENT1 in cultured cells suggests that hENT1 will not be saturated at clinically relevant (i.e., nanomolar) concentrations of tecadenoson, and that hENT1-mediated passage across the BBB may contribute to the adverse CNS effects observed in clinical trials. In contrast, in cases in which a CNS effect is desired, the present results illustrate that synthetic A1 agonists that are transported by hENT1 could be used to target CNS disorders because of enhanced delivery to the brain.

  18. Pharmacology of the Adenosine A3 Receptor in the Vasculature and Essential Hypertension (United States)

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


    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

  19. Presynaptic inhibition by kainate receptors converges mechanistically with presynaptic inhibition by adenosine and GABAB receptors. (United States)

    Partovi, Dara; Frerking, Matthew


    Kainate receptors are widely reported to regulate the release of neurotransmitter in the CNS, but the mechanisms involved remain controversial. Previous studies have found that the kainate receptor agonist ATPA, which selectively activates Glu(K5)-containing kainate receptors, depresses glutamate release at Schaffer-collateral synapses in the hippocampus. In the present study, we provide pharmacological evidence that this depressant effect is mediated by Glu(K5)-containing heteromers, but is distinct from a similar depressant effect engaged by the kainate receptor agonist domoate. The depressant effect of ATPA is insensitive to antagonists for GABA(A), GABA(B), and adenosine receptors, and is also unaffected by lowering the release probability by reducing extracellular calcium. However, the effect of ATPA is partly occluded by prior activation of GABA(B) receptors and completely occluded by prior activation of adenosine receptors, suggesting a mechanistic convergence of heteromeric Glu(K5) kainate receptor signaling with GABA(B) receptors and adenosine receptors. The effects of domoate are partially occluded by both adenosine and GABA(B) receptor agonists, indicating at least a partial convergence of Glu(K5)-lacking kainate receptor signaling with these other pathways. The depressant effect of ATPA is not blocked by inhibition of serine/threonine protein kinases. These results suggest that ATPA and domoate inhibit glutamate release through mechanisms that converge with those of classical metabotropic receptor agonists, although they do so through different receptors.

  20. 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


    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.

  1. Role of A3 adenosine receptor in diabetic neuropathy. (United States)

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


    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.

  2. Striatal adenosine-cannabinoid receptor interactions in rats over-expressing adenosine A2A receptors. (United States)

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


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

  3. Emerging GLP-1 receptor agonists

    DEFF Research Database (Denmark)

    Lund, Asger; Knop, Filip K; Vilsbøll, Tina


    Introduction: Recently, glucagon-like peptide-1 receptor (GLP-1R) agonists have become available for the treatment of type 2 diabetes. These agents exploit the physiological effects of GLP-1, which is able to address several of the pathophysiological features of type 2 diabetes. GLP-1R agonists...

  4. Emerging GLP-1 receptor agonists

    DEFF Research Database (Denmark)

    Lund, Asger; Knop, Filip K; Vilsbøll, Tina


    Introduction: Recently, glucagon-like peptide-1 receptor (GLP-1R) agonists have become available for the treatment of type 2 diabetes. These agents exploit the physiological effects of GLP-1, which is able to address several of the pathophysiological features of type 2 diabetes. GLP-1R agonists...... presently available are administered once or twice daily, but several once-weekly GLP-1R agonists are in late clinical development. Areas covered: The present review aims to give an overview of the clinical data on the currently available GLP-1R agonists used for treatment of type 2 diabetes, exenatide...

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

    Directory of Open Access Journals (Sweden)

    Noam eRudich


    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.

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


    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

  7. Severe hemorrhage attenuates cardiopulmonary chemoreflex control of regional sympathetic outputs via NTS adenosine receptors. (United States)

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


    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.

  8. Hydrogen sulfide inhibits A2A adenosine receptor agonist induced β-amyloid production in SH-SY5Y neuroblastoma cells via a cAMP dependent pathway.

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    Bhushan Vijay Nagpure

    Full Text Available Alzheimer's disease (AD is the leading cause of senile dementia in today's society. Its debilitating symptoms are manifested by disturbances in many important brain functions, which are influenced by adenosine. Hence, adenosinergic system is considered as a potential therapeutic target in AD treatment. In the present study, we found that sodium hydrosulfide (NaHS, an H2S donor, 100 µM attenuated HENECA (a selective A2A receptor agonist, 10-200 nM induced β-amyloid (1-42 (Aβ42 production in SH-SY5Y cells. NaHS also interfered with HENECA-stimulated production and post-translational modification of amyloid precursor protein (APP by inhibiting its maturation. Measurement of the C-terminal APP fragments generated from its enzymatic cleavage by β-site amyloid precursor protein cleaving enzyme 1 (BACE1 showed that NaHS did not have any significant effect on β-secretase activity. However, the direct measurements of HENECA-elevated γ-secretase activity and mRNA expressions of presenilins suggested that the suppression of Aβ42 production in NaHS pretreated cells was mediated by inhibiting γ-secretase. NaHS induced reductions were accompanied by similar decreases in intracellular cAMP levels and phosphorylation of cAMP responsive element binding protein (CREB. NaHS significantly reduced the elevated cAMP and Aβ42 production caused by forskolin (an adenylyl cyclase, AC agonist alone or forskolin in combination with IBMX (a phosphodiesterase inhibitor, but had no effect on those caused by IBMX alone. Moreover, pretreatment with NaHS significantly attenuated HENECA-elevated AC activity and mRNA expressions of various AC isoforms. These data suggest that NaHS may preferentially suppress AC activity when it was stimulated. In conclusion, H2S attenuated HENECA induced Aβ42 production in SH-SY5Y neuroblastoma cells through inhibiting γ-secretase via a cAMP dependent pathway.

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

    DEFF Research Database (Denmark)

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


    hamster or rat. However, the role of adenosine in human BAT is unknown. Here we show that adenosine activates human and murine brown adipocytes at low nanomolar concentrations. Adenosine is released in BAT during stimulation of sympathetic nerves as well as from brown adipocytes. The adenosine A2A...... of A2A receptors or injection of lentiviral vectors expressing the A2A receptor into white fat induces brown-like cells-so-called beige adipocytes. Importantly, mice fed a high-fat diet and treated with an A2A agonist are leaner with improved glucose tolerance. Taken together, our results demonstrate...... 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....

  10. Role of adenosine A1 and A2A receptors in the alcohol withdrawal syndrome. (United States)

    Kaplan, G B; Bharmal, N H; Leite-Morris, K A; Adams, W R


    The role of adenosine receptor-mediated signaling was examined in the alcohol withdrawal syndrome. CD-1 mice received a liquid diet containing ethanol (6.7%, v/v) or a control liquid diet that were abruptly discontinued after 14 days of treatment. Mice consuming ethanol showed a progressive increase in signs of intoxication throughout the drinking period. Following abrupt discontinuation of ethanol diet, mice demonstrated reversible signs of handling-induced hyperexcitability that were maximal between 5-8 h. Withdrawing mice received treatment with adenosine receptor agonists at the onset of peak withdrawal (5.5 h) and withdrawal signs were blindly rated (during withdrawal hours 6 and 7). Adenosine A1-receptor agonist R-N6(phenylisopropyl)adenosine (0.15 and 0.3 mg/ kg) reduced withdrawal signs 0.5 and 1.5 h after drug administration in a dose-dependent fashion. Adenosine A2A-selective agonist 2-p-(2-carboxyethyl)phenylethyl-amino-5'-N-ethylcarboxamidoadenosine (0.3 mg/kg) reduced withdrawal signs at both time points. In ethanol-withdrawing mice, there were significant decreases in adenosine transporter sites in striatum without changes in cortex or cerebellum. In ethanol-withdrawing mice, there were no changes in adenosine A1 and A2A receptor concentrations in cortex, striatum, or cerebellum. There appears to be a role for adenosine A1 and A2A receptors in the treatment of the ethanol withdrawal syndrome. Published by Elsevier Science Inc.

  11. Identification and function of adenosine A3 receptor in afferent arterioles. (United States)

    Lu, Yan; Zhang, Rui; Ge, Ying; Carlstrom, Mattias; Wang, Shaohui; Fu, Yiling; Cheng, Liang; Wei, Jin; Roman, Richard J; Wang, Lei; Gao, Xichun; Liu, Ruisheng


    Adenosine plays an important role in regulation of renal microcirculation. All receptors of adenosine, A1, A2A, A2B, and A3, have been found in the kidney. However, little is known about the location and function of the A3 receptor in the kidney. The present study determined the expression and role of A3 receptors in mediating the afferent arteriole (Af-Art) response and studied the interaction of A3 receptors with angiotensin II (ANG II), A1 and A2 receptors on the Af-Art. We found that the A3 receptor expressed in microdissected isolated Af-Art and the mRNA levels of A3 receptor were 59% of A1. In the isolated microperfused Af-Art, A3 receptor agonist IB-MECA did not have a constrictive effect. Activation of A3 receptor dilated the preconstricted Af-Art by norepinephrine and blunted the vasoconstrictive effect of both adenosine A1 receptor activation and ANG II on the Af-Art, respectively. Selective A2 receptor antagonist (both A2A and A2B) had no effect on A3 receptor agonist-induced vasodilation, indicating that the dilatory effect of A3 receptor activation is not mediated by activation of A2 receptor. We conclude that the A3 receptor is expressed in the Af-Art, and activation of the A3 receptor dilates the Af-Art.

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

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


    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.

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

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    Lei Sun

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

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

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


    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. The A2B adenosine receptor impairs the maturation and immunogenicity of dendritic cells. (United States)

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


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

  16. Muscarinic Receptor Agonists and Antagonists

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    David R. Kelly


    Full Text Available A comprehensive review of pharmacological and medical aspects of the muscarinic class of acetylcholine agonists and antagonists is presented. The therapeutic benefits of achieving receptor subtype selectivity are outlined and applications in the treatment of Alzheimer’s disease are discussed. A selection of chemical routes are described, which illustrate contemporary methodology for the synthesis of chiral medicinal compounds (asymmetric synthesis, chiral pool, enzymes. Routes to bicyclic intrannular amines and intramolecular Diels-Alder reactions are highlighted.

  17. The A3 adenosine receptor attenuates the calcium rise triggered by NMDA receptors in retinal ganglion cells. (United States)

    Zhang, Mei; Hu, Huiling; Zhang, Xiulan; Lu, Wennan; Lim, Jason; Eysteinsson, Thor; Jacobson, Kenneth A; Laties, Alan M; Mitchell, Claire H


    The A(3) adenosine receptor is emerging as an important regulator of neuronal signaling, and in some situations receptor stimulation can limit excitability. As the NMDA receptor frequently contributes to neuronal excitability, this study examined whether A(3) receptor activation could alter the calcium rise accompanying NMDA receptor stimulation. Calcium levels were determined from fura-2 imaging of isolated rat retinal ganglion cells as these neurons possess both receptor types. Brief application of glutamate or NMDA led to repeatable and reversible elevations of intracellular calcium. The A(3) agonist Cl-IB-MECA reduced the response to both glutamate and NMDA. While adenosine mimicked the effect of Cl-IB-MECA, the A(3) receptor antagonist MRS 1191 impeded the block by adenosine, implicating a role for the A(3) receptor in response to the natural agonist. The A(1) receptor antagonist DPCPX provided additional inhibition, implying a contribution from both A(1) and A(3) adenosine receptors. The novel A(3) agonist MRS 3558 (1'S,2'R,3'S,4'R,5'S)-4-(2-chloro-6-(3-chlorobenzylamino)-9H-purin-9-yl)-2,3-dihydroxy-N-methylbicyclo [3.1.0] hexane-1-carboxamide and mixed A(1)/A(3) agonist MRS 3630 (1'S,2'R,3'S,4'R,5'S)-4-(2-chloro-6-(cyclopentylamino)-9H-purin-9-yl)-2,3-dihydroxy-N-methylbicyclo [3.1.0] hexane-1-carboxamide also inhibited the calcium rise induced by NMDA. Low levels of MRS 3558 were particularly effective, with an IC(50) of 400 pM. In all cases, A(3) receptor stimulation inhibited only 30-50% of the calcium rise. In summary, stimulation of the A(3) adenosine receptor by either endogenous or synthesized agonists can limit the calcium rise accompanying NMDA receptor activation. It remains to be determined if partial block of the calcium rise by A(3) agonists can modify downstream responses to NMDA receptor stimulation.

  18. 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


    by CFTRinh-172, a cystic fibrosis transmembrane conductance regulator (CFTR) Cl− channel inhibitor. The adenosine A2B receptor agonist, BAY 60-6583, increased Isc and whole-cell Cl− currents through CFTR Cl− channels, whereas the A2A receptor agonist, CGS 21680, had negligible effects. The A2B receptor....... 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....

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

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    Felicita Pedata


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

  20. Adenosine Receptors Differentially Regulate the Expression of Regulators of G-Protein Signalling (RGS 2, 3 and 4 in Astrocyte-Like Cells.

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    Till Nicolas Eusemann

    Full Text Available The "regulators of g-protein signalling" (RGS comprise a large family of proteins that limit by virtue of their GTPase accelerating protein domain the signal transduction of G-protein coupled receptors. RGS proteins have been implicated in various neuropsychiatric diseases such as schizophrenia, drug abuse, depression and anxiety and aggressive behaviour. Since conditions associated with a large increase of adenosine in the brain such as seizures or ischemia were reported to modify the expression of some RGS proteins we hypothesized that adenosine might regulate RGS expression in neural cells. We measured the expression of RGS-2,-3, and -4 in both transformed glia cells (human U373 MG astrocytoma cells and in primary rat astrocyte cultures stimulated with adenosine agonists. Expression of RGS-2 mRNA as well as RGS2 protein was increased up to 30-fold by adenosine agonists in astrocytes. The order of potency of agonists and the blockade by the adenosine A2B-antagonist MRS1706 indicated that this effect was largely mediated by adenosine A2B receptors. However, a smaller effect was observed due to activation of adenosine A2A receptors. In astrocytoma cells adenosine agonists elicited an increase in RGS-2 expression solely mediated by A2B receptors. Expression of RGS-3 was inhibited by adenosine agonists in both astrocytoma cells and astrocytes. However while this effect was mediated by A2B receptors in astrocytoma cells it was mediated by A2A receptors in astrocytes as assessed by the order of potency of agonists and selective blockade by the specific antagonists MRS1706 and ZM241385 respectively. RGS-4 expression was inhibited in astrocytoma cells but enhanced in astrocytes by adenosine agonists.

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

    LENUS (Irish Health Repository)

    Wakai, A


    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.

  2. Modulatory effect of adenosine receptors on the ascending and descending neural reflex responses of rat ileum

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    Schusdziarra Volker


    Full Text Available Abstract Background Adenosine is known to act as a neuromodulator by suppressing synaptic transmission in the central and peripheral nervous system. Both the release of adenosine within the small intestine and the presence of adenosine receptors on enteric neurons have been demonstrated. The aim of the present study was to characterize a possible involvement of adenosine receptors in the modulation of the myenteric reflex. The experiments were carried out on ileum segments 10 cm in length incubated in an single chambered organ bath, and the reflex response was initiated by electrical stimulation (ES. Results ES caused an ascending contraction and a descending relaxation followed by a contraction. All motility responses to ES were completely blocked by tetrodotoxin, indicating that they are mediated by neural mechanisms. Atropine blocked the contractile effects, whereas the descending relaxation was significantly increased. The A1 receptor agonist N6-cyclopentyladenosine increased the ascending contraction, whereas the ascending contraction was reduced by the A1 receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine. Activation of the A1 receptor further reduced the descending relaxation and the latency of the peristaltic reflex. The A2B receptor antagonist alloxazine increased ascending contraction, whereas descending relaxation remained unchanged. For A2A and A3 receptors, we found contradictory effects of the agonists and antagonists, thus there is no clear physiological role for these receptors at this time. Conclusions This study suggests that the myenteric ascending and descending reflex response of the rat small intestine is modulated by release of endogenous adenosine via A1 receptors.

  3. Stabilizing effects of G protein on the active conformation of adenosine A1 receptor differ depending on G protein type. (United States)

    Tateyama, Michihiro; Kubo, Yoshihiro


    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.

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

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


    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.

  5. A3 Adenosine receptors mediate oligodendrocyte death and ischemic damage to optic nerve. (United States)

    González-Fernández, Estíbaliz; Sánchez-Gómez, María Victoria; Pérez-Samartín, Alberto; Arellano, Rogelio O; Matute, Carlos


    Adenosine receptor activation is involved in myelination and in apoptotic pathways linked to neurodegenerative diseases. In this study, we investigated the effects of adenosine receptor activation in the viability of oligodendrocytes of the rat optic nerve. Selective activation of A3 receptors in pure cultures of oligodendrocytes caused concentration-dependent apoptotic and necrotic death which was preceded by oxidative stress and mitochondrial membrane depolarization. Oligodendrocyte apoptosis induced by A3 receptor activation was caspase-dependent and caspase-independent. In addition to dissociated cultures, incubation of optic nerves ex vivo with adenosine and the A3 receptor agonist 2-CI-IB-MECA(1-[2-Chloro-6-[[(3-iodophenyl)methyl]amino]-9H-purin-9-yl]-1-deoxy-N-methyl-b-D-ribofuranuronamide)-induced caspase-3 activation, oligodendrocyte damage, and myelin loss, effects which were prevented by the presence of caffeine and the A3 receptor antagonist MRS 1220 (N-[9-Chloro-2-(2-furanyl)[1,2,4]-triazolo [1,5-c]quinazolin-5-yl]benzene acetamide). Finally, ischemia-induced injury and functional loss to the optic nerve was attenuated by blocking A3 receptors. Together, these results indicate that adenosine may trigger oligodendrocyte death via activation of A3 receptors and suggest that this mechanism contributes to optic nerve and white matter ischemic damage.

  6. Cloning of two adenosine receptor subtypes from mouse bone marrow-derived mast cells. (United States)

    Marquardt, D L; Walker, L L; Heinemann, S


    Adenosine potentiates the stimulated release of mast cell mediators. Pharmacologic studies suggest the presence of two adenosine receptors, one positively coupled to adenylate cyclase and the other coupled to phospholipase C activation. To identify mast cell adenosine receptor subtypes, cDNAs for the A1 and A2a adenosine receptors were obtained by screening a mouse brain cDNA library with the use of PCR-derived probes. Mouse bone marrow-derived mast cell cDNA libraries were constructed and screened with the use of A1 and A2a cDNA probes, which revealed the presence of A2a, but not A1, receptor clones. A putative A2b receptor was identified by using low stringency mast cell library screening. Northern blotting of mast cell poly(A)+ RNA with the use of receptor subtype probes labeled single mRNA bands of 2.4 kb and 1.8 kb for the A2a and A2b receptors, respectively. In situ cells. An A2a receptor-specific agonist failed to enhance mast cell mediator release, which suggests that the secretory process is modulated through the A2b and/or another receptor subtype. By using RNase protection assays, we found that mast cells that had been cultured in the presence of N-ethylcarboxamidoadenosine for 24 h exhibited a decrease in both A2a and A2b receptor RNA levels. Cells that had been cultured for 1 to 2 days in the presence of dexamethasone demonstrated increased amounts of A2a receptor mRNA, but no identifiable change in A2b receptor mRNA. Mast cells possess at least two adenosine receptor subtypes that may be differentially regulated.

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

    Directory of Open Access Journals (Sweden)

    Kubilay Oransay


    Full Text Available 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 prior to citalopram. Other rats were pretreated with erythro-9-(2-hydroxy-3-nonyl adenine (EHNA; inhibitor of adenosine deaminase and S-(4-Nitrobenzyl-6-thioinosine (NBTI; inhibitor of facilitated adenosine transport. After pretreatment, group 2 received 5% dextrose and group 3 received citalopram. Adenosine concentrations, mean arterial pressure (MAP, heart rate (HR,  QRS duration and QT interval were evaluated. Results: In the dextrose group, citalopram infusion caused a significant decrease in MAP and HR and caused a significant prolongation in QRS and QT. DPCPX infusion significantly prevented the prolongation of the QT interval when compared to control. In the second protocol, citalopram infusion did not cause a significant change in plasma adenosine concentrations, but a significant increase observed in EHNA/NBTI groups. In EHNA/NBTI groups, citalopram-induced MAP and HR reductions, QRS and QT prolongations were more significant than the dextrose group. Conclusions: Citalopram may lead to QT prolongation by stimulating adenosine A 1 receptors without affecting the release of adenosine.

  8. Synthesis and anti-renal fibrosis activity of conformationally locked truncated 2-hexynyl-N(6)-substituted-(N)-methanocarba-nucleosides as A3 adenosine receptor antagonists and partial agonists. (United States)

    Nayak, Akshata; Chandra, Girish; Hwang, Inah; Kim, Kyunglim; Hou, Xiyan; Kim, Hea Ok; Sahu, Pramod K; Roy, Kuldeep K; Yoo, Jakyung; Lee, Yoonji; Cui, Minghua; Choi, Sun; Moss, Steven M; Phan, Khai; Gao, Zhan-Guo; Ha, Hunjoo; Jacobson, Kenneth A; Jeong, Lak Shin


    Truncated N(6)-substituted-(N)-methanocarba-adenosine derivatives with 2-hexynyl substitution were synthesized to examine parallels with corresponding 4'-thioadenosines. Hydrophobic N(6) and/or C2 substituents were tolerated in A3AR binding, but only an unsubstituted 6-amino group with a C2-hexynyl group promoted high hA2AAR affinity. A small hydrophobic alkyl (4b and 4c) or N(6)-cycloalkyl group (4d) showed excellent binding affinity at the hA3AR and was better than an unsubstituted free amino group (4a). A3AR affinities of 3-halobenzylamine derivatives 4f-4i did not differ significantly, with Ki values of 7.8-16.0 nM. N(6)-Methyl derivative 4b (Ki = 4.9 nM) was a highly selective, low efficacy partial A3AR agonist. All compounds were screened for renoprotective effects in human TGF-β1-stimulated mProx tubular cells, a kidney fibrosis model. Most compounds strongly inhibited TGF-β1-induced collagen I upregulation, and their A3AR binding affinities were proportional to antifibrotic effects; 4b was most potent (IC50 = 0.83 μM), indicating its potential as a good therapeutic candidate for treating renal fibrosis.

  9. Endogenous Receptor Agonists: Resolving Inflammation

    Directory of Open Access Journals (Sweden)

    Gerhard Bannenberg


    Full Text Available Controlled resolution or the physiologic resolution of a well-orchestrated inflammatory response at the tissue level is essential to return to homeostasis. A comprehensive understanding of the cellular and molecular events that control the termination of acute inflammation is needed in molecular terms given the widely held view that aberrant inflammation underlies many common diseases. This review focuses on recent advances in the understanding of the role of arachidonic acid and ω-3 polyunsaturated fatty acids (PUFA–derived lipid mediators in regulating the resolution of inflammation. Using a functional lipidomic approach employing LC-MS-MS–based informatics, recent studies, reviewed herein, uncovered new families of local-acting chemical mediators actively biosynthesized during the resolution phase from the essential fatty acids eicosapentaenoic acid (EPA and docosahexaenoic acid (DHA. These new families of local chemical mediators are generated endogenously in exudates collected during the resolution phase, and were coined resolvins and protectins because specific members of these novel chemical families control both the duration and magnitude of inflammation in animal models of complex diseases. Recent advances on the biosynthesis, receptors, and actions of these novel anti-inflammatory and proresolving lipid mediators are reviewed with the aim to bring to attention the important role of specific lipid mediators as endogenous agonists in inflammation resolution.

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

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


    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

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

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

    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.

  12. Subtype selective kainic acid receptor agonists

    DEFF Research Database (Denmark)

    Bunch, Lennart; Krogsgaard-Larsen, Povl


    (S)-Glutamic acid (Glu) is the major excitatory neurotransmitter in the mammalian central nervous system, activating the plethora of glutamate receptors (GluRs). In broad lines, the GluRs are divided into two major classes: the ionotropic Glu receptors (iGluRs) and the metabotropic Glu receptors (m......GluRs). Within the iGluRs, five subtypes (KA1, KA2, iGluR5-7) show high affinity and express full agonist activity upon binding of the naturally occurring amino acid kainic acid (KA). Thus these receptors have been named the KA receptors. This review describes all-to our knowledge-published KA receptor agonists...

  13. Muscimol as an ionotropic GABA receptor agonist. (United States)

    Johnston, Graham A R


    Muscimol, a psychoactive isoxazole from Amanita muscaria and related mushrooms, has proved to be a remarkably selective agonist at ionotropic receptors for the inhibitory neurotransmitter GABA. This historic overview highlights the discovery and development of muscimol and related compounds as a GABA agonist by Danish and Australian neurochemists. Muscimol is widely used as a ligand to probe GABA receptors and was the lead compound in the development of a range of GABAergic agents including nipecotic acid, tiagabine, 4,5,6,7-tetrahydroisoxazolo(5,4-c)pyridin-3-ol, (Gaboxadol(®)) and 4-PIOL.

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

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


    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

  15. Activation of NTS A2a adenosine receptors differentially resets baroreflex control of renal vs. adrenal sympathetic nerve activity. (United States)

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


    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.

  16. The role of glial adenosine receptors in neural resilience and the neurobiology of mood disorders

    NARCIS (Netherlands)

    Calker, D; Biber, K


    Adenosine receptors were classified into A(1)- and A(2)-receptors in the laboratory of Bernd Hamprecht more than 25 years ago. Adenosine receptors are instrumental to the neurotrophic effects of glia cells. Both microglia and astrocytes release after stimulation via adenosine receptors factors that

  17. Adenosine A1 Receptor Suppresses Tonic GABAA Receptor Currents in Hippocampal Pyramidal Cells and in a Defined Subpopulation of Interneurons. (United States)

    Rombo, Diogo M; Dias, Raquel B; Duarte, Sofia T; Ribeiro, Joaquim A; Lamsa, Karri P; Sebastião, Ana M


    Adenosine is an endogenous neuromodulator that decreases excitability of hippocampal circuits activating membrane-bound metabotropic A1 receptor (A1R). The presynaptic inhibitory action of adenosine A1R in glutamatergic synapses is well documented, but its influence on inhibitory GABAergic transmission is poorly known. We report that GABAA receptor (GABAAR)-mediated tonic, but not phasic, transmission is suppressed by A1R in hippocampal neurons. Adenosine A1R activation strongly inhibits GABAAR agonist (muscimol)-evoked currents in Cornu Ammonis 1 (CA1) pyramidal neurons and in a specific subpopulation of interneurons expressing axonal cannabinoid receptor type 1. In addition, A1R suppresses tonic GABAAR currents measured in the presence of elevated ambient GABA as well as in naïve slices. The inhibition of GABAergic currents involves both protein kinase A (PKA) and protein kinase C (PKC) signaling pathways and decreases GABAAR δ-subunit expression. On the contrary, no A1R-mediated modulation was detected in phasic inhibitory postsynaptic currents evoked either by afferent electrical stimulation or by spontaneous quantal release. The results show that A1R modulates extrasynaptic rather than synaptic GABAAR-mediated signaling, and that this modulation selectively occurs in hippocampal pyramidal neurons and in a specific subpopulation of inhibitory interneurons. We conclude that modulation of tonic GABAAR signaling by adenosine A1R in specific neuron types may regulate neuronal gain and excitability in the hippocampus.

  18. Ethologically based resolution of D2-like dopamine receptor agonist-versus antagonist-induced behavioral topography in dopamine- and adenosine 3',5'-monophosphate-regulated phosphoprotein of 32 kDa "knockout" mutants congenic on the C57BL/6 genetic background. (United States)

    Nally, Rachel E; Kinsella, Anthony; Tighe, Orna; Croke, David T; Fienberg, Allen A; Greengard, Paul; Waddington, John L


    Given the critical role of dopamine- and adenosine 3',5'-monophosphate-regulated phosphoprotein of 32 kDa (DARPP-32) in the regulation of dopaminergic function, DARPP-32-null mutant mice congenic on the inbred C57BL/6 strain for 10 generations were examined phenotypically for their ethogram of responsivity to the selective D2-like receptor agonist RU 24213 (N-n-propyl-N-phenylethyl-p-3-hydroxyphenylethylamine) and the selective D2-like receptor antagonist YM 09151-2 (cis-N-[1-benzyl-2-methyl-pyrrolidin-3-yl]-5-chloro-2-methoxy-4-methylaminobenzamide), using procedures that resolve all topographies of behavior in the natural repertoire. After vehicle challenge, levels of sniffing and rearing seated were reduced in DARPP-32 mutants; the injection procedure seems to constitute a "stressor" that reveals phenotypic effects of DARPP-32 deletion not apparent under natural conditions. Topographical effects of 0.3 to 10.0 mg/kg RU 24213, primarily induction of sniffing and ponderous locomotion with accompanying reductions in rearing, grooming, sifting and chewing, were not altered to any material extent in DARPP-32-null mice. However, topographical effects of 0.005 to 0.625 mg/kg YM 09151-2, namely, reduction in sniffing, locomotion, rearing, grooming, and chewing but not sifting, were essentially absent in DARPP-32 mutants. Thus, the D2-like receptor agonist-mediated ethogram was essentially conserved, whereas major elements of the corresponding D2-like receptor antagonist-mediated ethogram were essentially absent in DARPP-32-null mice. This suggests some relationship between 1) extent of tonic dopaminergic activation of DARPP-32 mechanisms and 2) compensatory mechanisms consequent to the developmental absence of DARPP-32, which may emerge to act differentially on individual elements of the DARPP-32 system. Critically, the present data indicate that phenotypic effects of a given gene deletion using an agonist acting on the system disrupted cannot be generalized to a

  19. Down-regulation of the A3 adenosine receptor in human mast cells upregulates mediators of angiogenesis and remodeling. (United States)

    Rudich, Noam; Dekel, Ornit; Sagi-Eisenberg, Ronit


    Adenosine activated mast cells have been long implicated in allergic asthma and studies in rodent mast cells have assigned the A3 adenosine receptor (A3R) a primary role in mediating adenosine responses. Here we analyzed the functional impact of A3R activation on genes that are implicated in tissue remodeling in severe asthma in the human mast cell line HMC-1 that shares similarities with lung derived human mast cells. Quantitative real time PCR demonstrated upregulation of IL6, IL8, VEGF, amphiregulin and osteopontin. Moreover, further upregulation of these genes was noted upon the addition of dexamethasone. Unexpectedly, activated A3R down regulated its own expression and knockdown of the receptor replicated the pattern of agonist induced gene upregulation. This study therefore identifies the human mast cell A3R as regulator of tissue remodeling gene expression in human mast cells and demonstrates a heretofore-unrecognized mode of feedback regulation that is exerted by this receptor.

  20. Functional expression of adenosine A2A and A3 receptors in the mouse dendritic cell line XS-106. (United States)

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


    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

  1. 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

  2. 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.


    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.

  3. Small-Animal PET Study of Adenosine A(1) Receptors in Rat Brain : Blocking Receptors and Raising Extracellular Adenosine

    NARCIS (Netherlands)

    Paul, Soumen; Khanapur, Shivashankar; Rybczynska, Anna A.; Kwizera, Chantal; Sijbesma, Jurgen W. A.; Ishiwata, Kiichi; Willemsen, Antoon T. M.; Elsinga, Philip H.; Dierckx, Rudi A. J. O.; van Waarde, Aren


    Activation of adenosine A(1) receptors (A(1)R) in the brain causes sedation, reduces anxiety, inhibits seizures, and promotes neuroprotection. Cerebral A(1)R can be visualized using 8-dicyclopropylmethyl-1-C-11-methyl-3-propyl-xanthine (C-11-MPDX) and PET. This study aims to test whether C-11-MPDX c

  4. Regulation of membrane cholecystokinin-2 receptor by agonists enables classification of partial agonists as biased agonists. (United States)

    Magnan, Rémi; Masri, Bernard; Escrieut, Chantal; Foucaud, Magali; Cordelier, Pierre; Fourmy, Daniel


    Given the importance of G-protein-coupled receptors as pharmacological targets in medicine, efforts directed at understanding the molecular mechanism by which pharmacological compounds regulate their presence at the cell surface is of paramount importance. In this context, using confocal microscopy and bioluminescence resonance energy transfer, we have investigated internalization and intracellular trafficking of the cholecystokinin-2 receptor (CCK2R) in response to both natural and synthetic ligands with different pharmacological features. We found that CCK and gastrin, which are full agonists on CCK2R-induced inositol phosphate production, rapidly and abundantly stimulate internalization. Internalized CCK2R did not rapidly recycle to plasma membrane but instead was directed to late endosomes/lysosomes. CCK2R endocytosis involves clathrin-coated pits and dynamin and high affinity and prolonged binding of β-arrestin1 or -2. Partial agonists and antagonists on CCK2R-induced inositol phosphate formation and ERK1/2 phosphorylation did not stimulate CCK2R internalization or β-arrestin recruitment to the CCK2R but blocked full agonist-induced internalization and β-arrestin recruitment. The extreme C-terminal region of the CCK2R (and more precisely phosphorylatable residues Ser(437)-Xaa(438)-Thr(439)-Thr(440)-Xaa(441)-Ser(442)-Thr(443)) were critical for β-arrestin recruitment. However, this region and β-arrestins were dispensable for CCK2R internalization. In conclusion, this study allowed us to classify the human CCK2R as a member of class B G-protein-coupled receptors with regard to its endocytosis features and identified biased agonists of the CCK2R. These new important insights will allow us to investigate the role of internalized CCK2R·β-arrestin complexes in cancers expressing this receptor and to develop new diagnosis and therapeutic strategies targeting this receptor.

  5. Creatine, similarly to ketamine, affords antidepressant-like effects in the tail suspension test via adenosine A₁ and A2A receptor activation. (United States)

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


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

  6. Adenosine activates ATP-sensitive potassium channels in arterial myocytes via A2 receptors and cAMP-dependent protein kinase. (United States)

    Kleppisch, T; Nelson, M T


    The mechanism by which the endogenous vasodilator adenosine causes ATP-sensitive potassium (KATP) channels in arterial smooth muscle to open was investigated by the whole-cell patch-clamp technique. Adenosine induced voltage-independent, potassium-selective currents, which were inhibited by glibenclamide, a blocker of KATP currents. Glibenclamide-sensitive currents were also activated by the selective adenosine A2-receptor agonist 2-p-(2-carboxethyl)-phenethylamino-5'-N- ethylcarboxamidoadenosine hydrochloride (CGS-21680), whereas 2-chloro-N6-cyclopentyladenosine (CCPA), a selective adenosine A1-receptor agonist, failed to induce potassium currents. Glibenclamide-sensitive currents induced by adenosine and CGS-21680 were largely reduced by blockers of the cAMP-dependent protein kinase (Rp-cAMP[S], H-89, protein kinase A inhibitor peptide). Therefore, we conclude that adenosine can activate KATP currents in arterial smooth muscle through the following pathway: (i) Adenosine stimulates A2 receptors, which activates adenylyl cyclase; (ii) the resulting increase intracellular cAMP stimulates protein kinase A, which, probably through a phosphorylation step, opens KATP channels. PMID:8618917

  7. Adenosine A(3) receptor-induced CCL2 synthesis in cultured mouse astrocytes

    NARCIS (Netherlands)

    Wittendorp, MC; Boddeke, HWGM; Biber, K


    During neuropathological conditions, high concentrations of adenosine are released, stimulating adenosine receptors in neurons and glial cells. It has recently been shown that stimulation of adenosine receptors in glial cells induces the release of neuroprotective substances such as NGF, S-100beta,

  8. P2X receptors regulate adenosine diphosphate release from hepatic cells. (United States)

    Chatterjee, Cynthia; Sparks, Daniel L


    Extracellular nucleotides act as paracrine regulators of cellular signaling and metabolic pathways. Adenosine polyphosphate (adenosine triphosphate (ATP) and adenosine diphosphate (ADP)) release and metabolism by human hepatic carcinoma cells was therefore evaluated. Hepatic cells maintain static nanomolar concentrations of extracellular ATP and ADP levels until stress or nutrient deprivation stimulates a rapid burst of nucleotide release. Reducing the levels of media serum or glucose has no effect on ATP levels, but stimulates ADP release by up to 10-fold. Extracellular ADP is then metabolized or degraded and media ADP levels fall to basal levels within 2-4 h. Nucleotide release from hepatic cells is stimulated by the Ca(2+) ionophore, ionomycin, and by the P2 receptor agonist, 2'3'-O-(4-benzoyl-benzoyl)-adenosine 5'-triphosphate (BzATP). Ionomycin (10 μM) has a minimal effect on ATP release, but doubles media ADP levels at 5 min. In contrast, BzATP (10-100 μM) increases both ATP and ADP levels by over 100-fold at 5 min. Ion channel purinergic receptor P2X7 and P2X4 gene silencing with small interference RNA (siRNA) and treatment with the P2X inhibitor, A438079 (100 μM), decrease ADP release from hepatic cells, but have no effect on ATP. P2X inhibitors and siRNA have no effect on BzATP-stimulated nucleotide release. ADP release from human hepatic carcinoma cells is therefore regulated by P2X receptors and intracellular Ca(2+) levels. Extracellular ADP levels increase as a consequence of a cellular stress response resulting from serum or glucose deprivation.

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


    Björklund, Olga


    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...

  10. Adenosine A3 Receptor: A promising therapeutic target in cardiovascular disease. (United States)

    Nishat, Shamama; Khan, Luqman A; Ansari, Zafar M; Basir, Seemi F


    Cardiovascular complications are one of the major factors for early mortality in the present worldwide scenario and have become a major challenge in both developing and developed nations. It has thus become of immense importance to look for different therapeutic possibilities and treatments for the growing burden of cardiovascular diseases. Recent advancements in research have opened various means for better understanding of the complication and treatment of the disease. Adenosine receptors have become tool of choice in understanding the signaling mechanism which might lead to the cardiovascular complications. Adenosine A3 receptor is one of the important receptor which is extensively studied as a therapeutic target in cardiovascular disorder. Recent studies have shown that A3AR is involved in the amelioration of cardiovascular complications by altering the expression of A3R. This review focuses towards the therapeutic potential of A3AR involved in cardiovascular disease and it might help in better understanding of mechanism by which this receptor may prove useful in improving the complications arising due to various cardiovascular diseases. Understanding of A3AR signaling may also help to develop newer agonists and antagonists which might be prove helpful in the treatment of cardiovascular disorder.

  11. Adenosine receptor control of cognition in normal and disease. (United States)

    Chen, Jiang-Fan


    Adenosine and adenosine receptors (ARs) are increasingly recognized as important therapeutic targets for controlling cognition under normal and disease conditions for its dual roles of neuromodulation as well as of homeostatic function in the brain. This chapter first presents the unique ability of adenosine, by acting on the inhibitory A1 and facilitating A2A receptor, to integrate dopamine, glutamate, and BNDF signaling and to modulate synaptic plasticity (e.g., long-term potentiation and long-term depression) in brain regions relevant to learning and memory, providing the molecular and cellular bases for adenosine receptor (AR) control of cognition. This led to the demonstration of AR modulation of social recognition memory, working memory, reference memory, reversal learning, goal-directed behavior/habit formation, Pavlovian fear conditioning, and effort-related behavior. Furthermore, human and animal studies support that AR activity can also, through cognitive enhancement and neuroprotection, reverse cognitive impairments in animal models of Alzheimer's disease (AD), Parkinson's disease (PD), Huntington's disease, and schizophrenia. Lastly, epidemiological evidence indicates that regular human consumption of caffeine, the most widely used psychoactive drug and nonselective AR antagonists, is associated with the reduced cognitive decline in aging and AD patients, and with the reduced risk in developing PD. Thus, there is a convergence of the molecular studies revealing AR as molecular targets for integrating neurotransmitter signaling and controlling synaptic plasticity, with animal studies demonstrating the strong procognitive impact upon AR antagonism in normal and disease brains and with epidemiological and clinical evidences in support of caffeine and AR drugs for therapeutic modulation of cognition. Since some of adenosine A2A receptor antagonists are already in phase III clinical trials for motor benefits in PD patients with remarkable safety profiles

  12. Adenosine A3 receptor activation is neuroprotective against retinal neurodegeneration. (United States)

    Galvao, Joana; Elvas, Filipe; Martins, Tiago; Cordeiro, M Francesca; Ambrósio, António Francisco; Santiago, Ana Raquel


    Death of retinal neural cells, namely retinal ganglion cells (RGCs), is a characteristic of several retinal neurodegenerative diseases. Although the role of adenosine A3 receptor (A3R) in neuroprotection is controversial, A3R activation has been reported to afford protection against several brain insults, with few studies in the retina. In vitro models (retinal neural and organotypic cultures) and animal models [ischemia-reperfusion (I-R) and partial optic nerve transection (pONT)] were used to study the neuroprotective properties of A3R activation against retinal neurodegeneration. The A3R selective agonist (2-Cl-IB-MECA, 1 μM) prevented apoptosis (TUNEL(+)-cells) induced by kainate and cyclothiazide (KA + CTZ) in retinal neural cultures (86.5 ± 7.4 and 37.2 ± 6.1 TUNEL(+)-cells/field, in KA + CTZ and KA + CTZ + 2-Cl-IB-MECA, respectively). In retinal organotypic cultures, 2-Cl-IB-MECA attenuated NMDA-induced cell death, assessed by TUNEL (17.3 ± 2.3 and 8.3 ± 1.2 TUNEL(+)-cells/mm(2) in NMDA and NMDA+2-Cl-IB-MECA, respectively) and PI incorporation (ratio DIV4/DIV2 3.3 ± 0.3 and 1.3 ± 0.1 in NMDA and NMDA+2-Cl-IB-MECA, respectively) assays. Intravitreal 2-Cl-IB-MECA administration afforded protection against I-R injury decreasing the number of TUNEL(+) cells by 72%, and increased RGC survival by 57%. Also, intravitreal administration of 2-Cl-IB-MECA inhibited apoptosis (from 449.4 ± 37.8 to 207.6 ± 48.9 annexin-V(+)-cells) and RGC loss (from 1.2 ± 0.6 to 8.1 ± 1.7 cells/mm) induced by pONT. This study demonstrates that 2-Cl-IB-MECA is neuroprotective to the retina, both in vitro and in vivo. Activation of A3R may have great potential in the management of retinal neurodegenerative diseases characterized by RGC death, as glaucoma and diabetic retinopathy, and ischemic diseases.

  13. Combining GLP-1 receptor agonists with insulin

    DEFF Research Database (Denmark)

    Holst, Jens Juul; Vilsbøll, T


    physicians and patients regarding the initiation and intensification of insulin therapy, in part due to concerns about the associated weight gain and increased risk of hypoglycaemia. Glucagon-like peptide-1 receptor agonists (GLP-1RAs) increase insulin release and suppress glucagon secretion in a glucose...... potential of GLP-1RA-insulin combination therapy, typically showing beneficial effects on glycaemic control and body weight, with a low incidence of hypoglycaemia and, in established insulin therapy, facilitating reductions in insulin dose. In this review, the physiological and pharmacological rationale...

  14. 腺苷受体激动剂降低低氧大鼠肺动脉高压及对诱导型一氧化氮合酶-一氧化氮和肾素-血管紧张素的影响%Adenosine receptors agonists mitigated PAH of rats induced by chronic hypoxia through reduction of renin activity/angiotensin Ⅱ levels and increase of inducible nitric oxide synthase-nitric oxide levels

    Institute of Scientific and Technical Information of China (English)

    谭建新; 黄秀兰; 王波; 方兴; 黄迪南


    Objective Recent studies showed that adenosine played important roles in vasodilation.This study aimed to investigate the effects of adenosine,its A1 and A2b receptor agonists on pulmonary artery hypertension (PAH) induced by chronic hypoxia in rats by continuously subcutaneous administration with an osmotic pump for 14 days,and to see if rennin angiotensin system and inducible nitric oxygen synthase (iNOS)/nitric oxide (NO) mediate the effects.Method Fifty-six male SD rats were randomly assigned to seven groups.Each group included eight rats.They were normoxic group,hypoxic group,adenosine-treated group [adenosine was administered at a dose of 150 μg(kg · min) under the hypoxic condition],adenosine A1 receptor agonist CPA-treated group [CPA was administered at a dose of 20 μg/(kg · min)under the hypoxic condition],CPA plus selective adenosine A1 antagonist DPCPX-treated group [CPA and DPCPX were administered simultaneously under the hypoxic condition,the dose of CPA was the same as the above,and the dose of DPCPX was 25 μg/(kg · min)],adenosine A2b receptor agonist NECA-treated group [NECA was administered at a dose of 30 μg/(kg · min) under the hypoxic condition],NECA plus selective adenosine A2b receptor antagonist MRS-treated group [NECA and MRS1754 were administered simultaneously under the hypoxic condition,the dose of NECA was the same as the above,and the dose of MRS1754 was 50 μg/(kg · min)].Osmotic pumps containing adenosine or selective adenosine A1 receptor agonist (CPA),or nonselective but potent adenosine A2b receptor agonist (NECA) were placed subcutaneously 7 days after hypoxia and continuously administered the agents for 14 days.Mean pulmonary artery pressure (mPAP) was detected after administration of the agents.Then blood samples were taken from heart for measurement of renin activity,angiotensin Ⅱ (Ang Ⅱ) and endothelin-1 (ET-1) concentration by radioimmunoassay,NO by measuring nitrate.Small pulmonary arteries were prepared for

  15. Adenosine receptors in post-mortem human brain. (United States)

    James, S; Xuereb, J H; Askalan, R; Richardson, P J


    1. Adenosine A2-like binding sites were characterized in post-mortem human brain membranes by examining several compounds for their ability to displace [3H]-CGS 21680 (2[p-(2 carboxyethyl)-phenethylamino]-5'-N-ethylcarboxamido adenosine) binding. 2. Two A2-like binding sites were identified in the striatum. 3. The more abundant striatal site was similar to the A2a receptor previously described in rat striatum, both in its pharmacological profile and striatal localization. 4. The less abundant striatal site had a pharmacological profile similar to that of the binding site characterized in the other brain regions examined. This was intermediate in character between A1 and A2 and may represent another adenosine receptor subtype. 5. The co-purification of [3H]-CGS 21680 binding during immunoisolation of human striatal cholinergic membranes was used to assess the possible cholinergic localization of A2-like binding sites in the human striatum. Only the more abundant striatal site co-purified with cholinergic membranes. This suggests that this A2a-like site is present on cholinergic neurones in the human striatum.

  16. Methylthioadenosine reprograms macrophage activation through adenosine receptor stimulation.

    Directory of Open Access Journals (Sweden)

    Peter A Keyel

    Full Text Available Regulation of inflammation is necessary to balance sufficient pathogen clearance with excessive tissue damage. Central to regulating inflammation is the switch from a pro-inflammatory pathway to an anti-inflammatory pathway. Macrophages are well-positioned to initiate this switch, and as such are the target of multiple therapeutics. One such potential therapeutic is methylthioadenosine (MTA, which inhibits TNFα production following LPS stimulation. We found that MTA could block TNFα production by multiple TLR ligands. Further, it prevented surface expression of CD69 and CD86 and reduced NF-KB signaling. We then determined that the mechanism of this action by MTA is signaling through adenosine A2 receptors. A2 receptors and TLR receptors synergized to promote an anti-inflammatory phenotype, as MTA enhanced LPS tolerance. In contrast, IL-1β production and processing was not affected by MTA exposure. Taken together, these data demonstrate that MTA reprograms TLR activation pathways via adenosine receptors to promote resolution of inflammation.

  17. GABAB and adenosine receptors mediate enhancement of the K+ current, IAHP, by reducing adenylyl cyclase activity in rat CA3 hippocampal neurons. (United States)

    Gerber, U; Gähwiler, B H


    1. Gamma-aminobuturic acid-B (GABAB) and adenosine A1 receptors, which are expressed in hippocampal pyramidal cells, are linked to pertussis toxin-sensitive G-proteins known to be coupled negatively to the enzyme adenylyl cyclase. This study investigates the electrophysiological consequences of adenylyl cyclase inhibition in response to stimulation of these receptors. 2. Single-electrode voltage-clamp recordings were obtained from CA3 pyramidal cells in rat hippocampal slice cultures in presence of tetrodotoxin. The calcium-dependent potassium current (IAHP), which is very sensitive to intracellular levels of adenosine 3',5'-cyclic monophosphate (cAMP), was used as an electrophysiological indicator of adenylyl cyclase activity. 3. Application of baclofen (10 microM), a selective agonist at GABAB receptors, or adenosine (50 microM) each resulted in a transient decrease followed by a significant enhancement in the amplitude of evoked IAHP. The initial reduction in amplitude of IAHP probably reflects inadequacies in voltage clamp of electronically distant dendritic sites, due to the shunting caused by concomitant activation of potassium conductance by baclofen/adenosine. Comparable increases in membrane conductance in response to the GABAA agonist, muscimol, caused a similar reduction in IAHP. The enhancement of IAHP is consistent with an inhibition of constitutively active adenylyl cyclase. 4. The receptor mediating the responses to adenosine was identified as belonging to the A1 subtype on the basis of its sensitivity to the selective antagonist 8-cyclopentyl-1,3-dipropylxanthine.(ABSTRACT TRUNCATED AT 250 WORDS)

  18. Extended N(6) substitution of rigid C2-arylethynyl nucleosides for exploring the role of extracellular loops in ligand recognition at the A3 adenosine receptor. (United States)

    Tosh, Dilip K; Paoletta, Silvia; Chen, Zhoumou; Moss, Steven M; Gao, Zhan-Guo; Salvemini, Daniela; Jacobson, Kenneth A


    2-Arylethynyl-(N)-methanocarba adenosine 5'-methyluronamides containing rigid N(6)-(trans-2-phenylcyclopropyl) and 2-phenylethynyl groups were synthesized as agonists for probing structural features of the A3 adenosine receptor (AR). Radioligand binding confirmed A3AR selectivity and N(6)-1S,2R stereoselectivity for one diastereomeric pair. The environment of receptor-bound, conformationally constrained N(6) groups was explored by docking to an A3AR homology model, indicating specific hydrophobic interactions with the second extracellular loop able to modulate the affinity profile. 2-Pyridylethynyl derivative 18 was administered orally in mice to reduce chronic neuropathic pain in the chronic constriction injury model.

  19. Estrogen receptor beta agonists in neurobehavioral investigations. (United States)

    Choleris, Elena; Clipperton, Amy E; Phan, Anna; Kavaliers, Martin


    Neurobehavioral investigations into the functions of estrogen receptor (ER)alpha and ERbeta have utilized 'knockout' mice, phytoestrogens and, more recently, ER-specific agonists. Feeding, sexual, aggressive and social behavior, anxiety, depression, drug abuse, pain perception, and learning (and associated synaptic plasticity) are affected by ERalpha and ERbeta in a manner that is dependent upon the specific behavior studied, gender and developmental stage. Overall, ERalpha and ERbeta appear to function together to foster sociosexual behavior while inhibiting behaviors that, if occurring at the time of behavioral estrous, may compete with reproduction (eg, feeding). Recently developed pharmacological tools have limited selectivity and availability to the research community at large, as they are not commercially available. The development of highly selective, commercially available ERbeta-specific antagonists would greatly benefit preclinical and applied research.

  20. Stimulation of NTS A1 adenosine receptors differentially resets baroreflex control of regional sympathetic outputs. (United States)

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


    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

  1. Arginine 199 and leucine 208 have key roles in the control of adenosine A2A receptor signalling function.

    Directory of Open Access Journals (Sweden)

    Nicolas Bertheleme

    Full Text Available One successful approach to obtaining high-resolution crystal structures of G-protein coupled receptors is the introduction of thermostabilising mutations within the receptor. This technique allows the generation of receptor constructs stabilised into different conformations suitable for structural studies. Previously, we functionally characterised a number of mutants of the adenosine A2A receptor, thermostabilised either in an agonist or antagonist conformation, using a yeast cell growth assay and demonstrated that there is a correlation between thermostability and loss of constitutive activity. Here we report the functional characterisation of 30 mutants intermediate between the Rag23 (agonist conformation mutant and the wild-type receptor using the same yeast signalling assay with the aim of gaining greater insight into the role individual amino acids have in receptor function. The data showed that R199 and L208 have important roles in receptor function; substituting either of these residues for alanine abolishes constitutive activity. In addition, the R199A mutation markedly reduces receptor potency while L208A reduces receptor efficacy. A184L and L272A mutations also reduce constitutive activity and potency although to a lesser extent than the R199A and L208A. In contrast, the F79A mutation increases constitutive activity, potency and efficacy of the receptor. These findings shed new light on the role individual residues have on stability of the receptor and also provide some clues as to the regions of the protein responsible for constitutive activity. Furthermore, the available adenosine A2A receptor structures have allowed us to put our findings into a structural context.

  2. Adenosine transiently modulates stimulated dopamine release in the caudate-putamen via A1 receptors. (United States)

    Ross, Ashley E; Venton, B Jill


    Adenosine modulates dopamine in the brain via A1 and A2A receptors, but that modulation has only been characterized on a slow time scale. Recent studies have characterized a rapid signaling mode of adenosine that suggests a possible rapid modulatory role. Here, fast-scan cyclic voltammetry was used to characterize the extent to which transient adenosine changes modulate stimulated dopamine release (5 pulses at 60 Hz) in rat caudate-putamen brain slices. Exogenous adenosine was applied and dopamine concentration monitored. Adenosine only modulated dopamine when it was applied 2 or 5 s before stimulation. Longer time intervals and bath application of 5 μM adenosine did not decrease dopamine release. Mechanical stimulation of endogenous adenosine 2 s before dopamine stimulation also decreased stimulated dopamine release by 41 ± 7%, similar to the 54 ± 6% decrease in dopamine after exogenous adenosine application. Dopamine inhibition by transient adenosine was recovered within 10 min. The A1 receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine blocked the dopamine modulation, whereas dopamine modulation was unaffected by the A2A receptor antagonist SCH 442416. Thus, transient adenosine changes can transiently modulate phasic dopamine release via A1 receptors. These data demonstrate that adenosine has a rapid, but transient, modulatory role in the brain. Here, transient adenosine was shown to modulate phasic dopamine release on the order of seconds by acting at the A1 receptor. However, sustained increases in adenosine did not regulate phasic dopamine release. This study demonstrates for the first time a transient, neuromodulatory function of rapid adenosine to regulate rapid neurotransmitter release.

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


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


    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 ...

  4. Pharmacological characterisation of the adenosine receptor mediating increased ion transport in the mouse isolated trachea and the effect of allergen challenge. (United States)

    Kornerup, Kristin N; Page, Clive P; Moffatt, James D


    The effect of adenosine on transepithelial ion transport was investigated in isolated preparations of murine trachea mounted in Ussing chambers. The possible regulation of adenosine receptors in an established model of allergic airway inflammation was also investigated. Mucosally applied adenosine caused increases in short-circuit current (I(SC)) that corresponded to approximately 50% of the response to the most efficacious secretogogue, ATP (delta I(SC) 69.5 +/- 6.7 microA cm2). In contrast, submucosally applied adenosine caused only small (<20%) increases in I(SC), which were not investigated further. The A1-selective (N6-cyclopentyladenosine, CPA, 1 nM-10 microM), A2A-selective (2-p-(2-carboxyethyl)phenethylamino-5'-N-ethylcarboxoamido adenosine; CGS 21680; 0.1-100 microM) and A3-selective (1-deoxy-1-[6-[[(3-iodophenyl)-methyl]amino]-9H-purin-9-yl]-N-methyl-beta-D-ribofuranuronamide; IB-MECA; 30 nM-100 microM) adenosine receptor agonists were either equipotent or less potent than adenosine, suggesting that these receptors do not mediate the response to adenosine. The A1 receptor selective antagonist 8-cyclopentyl-1,3-dipropylxanthine (DPCPX; 10 nM-1 microM) caused a rightward shift of the adenosine concentration-effect curve only at 1 microM. The mixed A2A/A2B receptor antagonist 4-(2-[7-amino-2-(2-furyl)[1,2,4]triazolo[2,3-a][1,3,5]triazin-5-ylamino]ethyl)phenol (ZM 241385) also caused rightward shift of the adenosine concentration-effect curve, again only at micromolar concentrations, suggestive of the involvement of A2B receptors. In preparations from animals sensitised to ovalbumin and challenged over 3 days with aerosol ovalbumin, a decrease in baseline I(SC) was observed and responses to ATP were diminished. Similarly, the amplitude of responses to adenosine were attenuated although there was no change in potency. These results suggest that the A2B receptor mediates the I(SC) response to adenosine in the mouse trachea. This receptor does not appear to be

  5. Activation of adenosine receptor potentiates the anticonvulsant effect of phenytoin against amygdala kindled seizures. (United States)

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


    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.

  6. 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


    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.

  7. 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


    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

  8. Role of adenosine A2b receptor overexpression in tumor progression. (United States)

    Sepúlveda, Cesar; Palomo, Iván; Fuentes, Eduardo


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

  9. Histamine H3-receptor inverse agonists as novel antipsychotics. (United States)

    Ito, Chihiro


    Schizophrenia (SZ) that is resistant to treatment with dopamine (DA) D2 antagonists may involve changes other than those in the dopaminergic system. Recently, histamine (HA), which regulates arousal and cognitive functions, has been suggested to act as a neurotransmitter in the central nervous system. Four HA receptors-H1, H2, H3, and H4-have been identified. Our recent basic and clinical studies revealed that brain HA improved the symptoms of SZ. The H3 receptor is primarily localized in the central nervous system, and it acts not only as a presynaptic autoreceptor that modulates the HA release but also as a presynaptic heteroreceptor that regulates the release of other neurotransmitters such as monoamines and amino acids. H3-receptor inverse agonists have been considered to improve cognitive functions. Many atypical antipsychotics are H3-receptor antagonists. Imidazole-containing H3-receptor inverse agonists inhibit not only cytochrome P450 but also hERG potassium channels (encoded by the human ether-a-go-go-related gene). Several imidazole H3-receptor inverse agonists also have high affinity for H4 receptors, which are expressed at high levels in mast cells and leukocytes. Clozapine is an H4-receptor agonist; this agonist activity may be related to the serious side effect of agranulocytosis caused by clozapine. Therefore, selective non-imidazole H3-receptor inverse agonists can be considered as novel antipsychotics that may improve refractory SZ.

  10. Unique interaction pattern for a functionally biased ghrelin receptor agonist

    DEFF Research Database (Denmark)

    Sivertsen, Bjørn Behrens; Lang, Manja; Frimurer, Thomas M.


    /13) pathway. The recognition pattern of wFw-Isn-NH(2) with the ghrelin receptor also differed significantly from that of all previously characterized unbiased agonists. Most importantly, wFw-Isn-NH(2) was not dependent on GluIII:09 (Glu3.33), which otherwise is an obligatory TM III anchor point residue...... orientation as compared with, for example, the wFw peptide agonists. It is concluded that the novel peptide-mimetic ligand wFw-Isn-NH(2) is a biased ghrelin receptor agonist and that the selective signaling pattern presumably is due to its unique receptor recognition pattern lacking interaction with key...

  11. Toll-like receptor 2 agonists inhibit human fibrocyte differentiation


    Maharjan Anu S; Pilling Darrell; Gomer Richard H


    Abstract Background In healing wounds, some monocytes enter the wound and differentiate into fibroblast-like cells called fibrocytes. Since Toll-like receptors (TLRs) are present on monocytes, and pathogens that can infect a wound have and/or release TLR agonists, we examined whether TLR agonists affect fibrocyte differentiation. Results When human peripheral blood mononuclear cells (PBMCs) were cultured with TLR3, TLR4, TLR5, TLR7, TLR8 or TLR9 agonists, there was no significant effect on fi...

  12. Adenosine in the tuberomammillary nucleus inhibits the histaminergic system via A1 receptors and promotes non-rapid eye movement sleep. (United States)

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


    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.

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

    Directory of Open Access Journals (Sweden)

    Sergi eFerre


    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.

  14. Nicotine receptor partial agonists for smoking cessation

    Directory of Open Access Journals (Sweden)

    Kate Cahill

    Full Text Available BACKGROUND: Nicotine receptor partial agonists may help people to stop smoking by a combination of maintaining moderate levels of dopamine to counteract withdrawal symptoms (acting as an agonist and reducing smoking satisfaction (acting as an antagonist. OBJECTIVES: The primary objective of this review is to assess the efficacy and tolerability of nicotine receptor partial agonists, including cytisine, dianicline and varenicline for smoking cessation. SEARCH METHODS: We searched the Cochrane Tobacco Addiction Group's specialised register for trials, using the terms ('cytisine' or 'Tabex' or 'dianicline' or 'varenicline' or 'nicotine receptor partial agonist' in the title or abstract, or as keywords. The register is compiled from searches of MEDLINE, EMBASE, PsycINFO and Web of Science using MeSH terms and free text to identify controlled trials of interventions for smoking cessation and prevention. We contacted authors of trial reports for additional information where necessary. The latest update of the specialized register was in December 2011. We also searched online clinical trials registers. SELECTION CRITERIA: We included randomized controlled trials which compared the treatment drug with placebo. We also included comparisons with bupropion and nicotine patches where available. We excluded trials which did not report a minimum follow-up period of six months from start of treatment. DATA COLLECTION AND ANALYSIS: We extracted data on the type of participants, the dose and duration of treatment, the outcome measures, the randomization procedure, concealment of allocation, and completeness of follow-up. The main outcome measured was abstinence from smoking at longest follow-up. We used the most rigorous definition of abstinence, and preferred biochemically validated rates where they were reported. Where appropriate we pooled risk ratios (RRs, using the Mantel-Haenszel fixed-effect model. MAIN RESULTS: Two recent cytisine trials (937 people

  15. [Effects of GLP-1 receptor agonists on carbohydrate metabolism control]. (United States)

    Fernández-García, José Carlos; Colomo, Natalia; Tinahones, Francisco José


    Glucagon-like peptide-1 (GLP-1) receptor agonists are a new group of drugs for the treatment of type 2 diabetes mellitus (DM2). In the present article, we review the available evidence on the efficacy of GLP-1 receptor agonists as glucose-lowering agents, their place in therapeutic algorithms, and the clinical factors associated with a favorable treatment response. Finally, we describe the clinical characteristics of patients who may benefit from these drugs.

  16. Toll-like receptor agonists in cancer therapy


    Adams, Sylvia


    Toll-like receptors (TLRs) are pattern-recognition receptors related to the Drosophila Toll protein. TLR activation alerts the immune system to microbial products and initiates innate and adaptive immune responses. The naturally powerful immunostimulatory property of TLR agonists can be exploited for active immunotherapy against cancer. Antitumor activity has been demonstrated in several cancers, and TLR agonists are now undergoing extensive clinical investigation. This review discusses recen...

  17. Evidence for an A1-adenosine receptor in the guinea-pig atrium. (United States)

    Collis, M. G.


    1 The purpose of this study was to determine whether the adenosine receptor that mediates a decrease in the force of contraction of the guinea-pig atrium is of the A1- or A2-sub-type. 2 Concentration-response curves to adenosine and a number of 5'- and N6-substituted analogues were constructed and the order of potency of the purines was: 5'-N-cyclopropylcarboxamide adenosine (NCPCA) = 5'-N-ethylcarboxamide adenosine (NECA) greater than N6cyclohexyladenosine (CHA) greater than L-N6-phenylisopropyl adenosine (L-PIA) = 2-chloroadenosine- greater than adenosine greater than D-N6-phenylisopropyl adenosine (D-PIA). 3 The difference in potency between the stereoisomers D- and L-PIA was over 100 fold. 4 The adenosine transport inhibitor, dipyridamole, potentiated submaximal responses to adenosine but had no significant effect on those evoked by the other purines. 5 Theophylline antagonized responses evoked by all purines, and with D-PIA revealed a positive inotropic effect that was abolished by atenolol. 6 The results indicate the existence of an adenosine A1-receptor in the guinea-pig atrium. PMID:6297647

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



    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...

  19. The cardiovascular effects of peroxisome proliferator-activated receptor agonists. (United States)

    Friedland, Sayuri N; Leong, Aaron; Filion, Kristian B; Genest, Jacques; Lega, Iliana C; Mottillo, Salvatore; Poirier, Paul; Reoch, Jennifer; Eisenberg, Mark J


    Although peroxisome proliferator-activated receptor agonists are prescribed to improve cardiovascular risk factors, their cardiovascular safety is controversial. We therefore reviewed the literature to identify landmark randomized controlled trials evaluating the effect of peroxisome proliferator-activated receptor gamma agonists (pioglitazone and rosiglitazone), alpha agonists (fenofibrate and gemfibrozil), and pan agonists (bezafibrate, muraglitazar, ragaglitazar, tesaglitazar, and aleglitazar) on cardiovascular outcomes. Pioglitazone may modestly reduce cardiovascular events but also may increase the risk of bladder cancer. Rosiglitazone increases the risk of myocardial infarction and has been withdrawn in European and restricted in the United States. Fibrates improve cardiovascular outcomes only in select subgroups: fenofibrate in diabetic patients with metabolic syndrome, gemfibrozil in patients with dyslipidemia, and bezafibrate in patients with diabetes or metabolic syndrome. The cardiovascular safety of the new pan agonist aleglitazar, currently in phase II trials, remains to be determined. The heterogenous effects of peroxisome proliferator-activated receptor agonists to date highlight the importance of postmarketing surveillance. The critical question of why peroxisome proliferator-activated receptor agonists seem to improve cardiovascular risk factors without significantly improving cardiovascular outcomes requires further investigation.

  20. 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


    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.


    Directory of Open Access Journals (Sweden)

    Ni Luh Putu Ayu Maha Iswari


    Full Text Available Melatonin is a hormone that has an important role in the mechanism of sleep. Hypnotic effects of melatonin and melatonin receptor agonist are mediated via MT1 and MT2 receptors, especially in circadian rhythm pacemaker, suprachiasmatic nucleus, which is worked on the hypothalamic sleep switch. This mechanism is quite different with the GABAergic drugs such as benzodiazepine. Agonist melatonin triggers the initiation of sleep and normalize circadian rhythms so that makes it easier to maintain sleep. The main disadvantage of melatonin in helping sleep maintenance on primary insomnia is that the half life is very short. The solution to this problem is the use of prolonged-release melatonin and melatonin receptor agonist agents such as ramelteon. Melatoninergic agonist does not cause withdrawal effects, dependence, as well as cognitive and psychomotor disorders as often happens on the use of benzodiazepine.  

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


    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.

  3. Evidence for an A2/Ra adenosine receptor in the guinea-pig trachea (United States)

    Brown, C.M.; Collis, M.G.


    1 An attempt was made to determine whether the extracellular adenosine receptor that mediates relaxation in the guinea-pig trachea is of the A1/Ri or A2/Ra subtype. 2 Dose-response curves to adenosine and a number of 5′- and N6-substituted analogues were constructed for the isolated guinea-pig trachea, contracted with carbachol. 3 The 5′-substituted analogues of adenosine were the most potent compounds tested, the order of potency being 5′-N-cyclopropylcarboxamide adenosine (NCPCA) > 5′-N-ethylcarboxamide adenosine (NECA) > 2-chloroadenosine > L-N6-phenylisopropyladenosine (L-PIA) > adenosine > D-N6-phenylisopropyladenosine (D-PIA). 4 The difference in potency between the stereoisomers D- and L-PIA on the isolated trachea was at the most five fold. 5 Responses to low doses of adenosine and its analogues were attenuated after treatment with either theophylline or 8-phenyltheophylline. The responses to 2-chloroadenosine were affected to a lesser extent than were those to the other purines. 6 Adenosine transport inhibitors, dipyridamole and dilazep, potentiated responses to adenosine, did not affect those to NCPCA, NECA, L-PIA and D-PIA but significantly reduced the responses to high doses of 2-chloroadenosine. 7 Relaxations evoked by 9-β-D-xylofuranosyladenosine which can activate intracellular but not extracellular adenosine receptors, were attenuated by dipyridamole but unaffected by 8-phenyltheophylline. 8 The results support the existence of an extracellular A2/Ra subtype of adenosine receptor and an intracellular purine-sensitive site, both of which mediate relaxation. PMID:6286021

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


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


    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.

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

    Directory of Open Access Journals (Sweden)

    Moidunny Shamsudheen


    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

  6. Enhanced A3 adenosine receptor selectivity of multivalent nucleoside-dendrimer conjugates

    Directory of Open Access Journals (Sweden)

    Shainberg Asher


    Full Text Available Abstract Background An approach to use multivalent dendrimer carriers for delivery of nucleoside signaling molecules to their cell surface G protein-coupled receptors (GPCRs was recently introduced. Results A known adenosine receptor (AR agonist was conjugated to polyamidoamine (PAMAM dendrimer carriers for delivery of the intact covalent conjugate to on the cell surface. Depending on the linking moiety, multivalent conjugates of the N6-chain elongated functionalized congener ADAC (N6-[4-[[[4-[[[(2-aminoethylamino]carbonyl]methyl]anilino]carbonyl]methyl]phenyl]-adenosine achieved unanticipated high selectivity in binding to the cytoprotective human A3 AR, a class A GPCR. The key to this selectivity of > 100-fold in both radioreceptor binding (Ki app = 2.4 nM and functional assays (EC50 = 1.6 nM in inhibition of adenylate cyclase was maintaining a free amino group (secondary in an amide-linked chain. Attachment of neutral amide-linked chains or thiourea-containing chains preserved the moderate affinity and efficacy at the A1 AR subtype, but there was no selectivity for the A3 AR. Since residual amino groups on dendrimers are associated with cytotoxicity, the unreacted terminal positions of this A3 AR-selective G2.5 dendrimer were present as carboxylate groups, which had the further benefit of increasing water-solubility. The A3 AR selective G2.5 dendrimer was also visualized binding the membrane of cells expressing the A3 receptor but did not bind cells that did not express the receptor. Conclusion This is the first example showing that it is feasible to modulate and even enhance the pharmacological profile of a ligand of a GPCR based on conjugation to a nanocarrier and the precise structure of the linking group, which was designed to interact with distal extracellular regions of the 7 transmembrane-spanning receptor. This ligand tool can now be used in pharmacological models of tissue rescue from ischemia and to probe the existence of A3 AR

  7. Topological sub-structural molecular design (TOPS-MODE): a useful tool to explore key fragments of human A3 adenosine receptor ligands. (United States)

    Saíz-Urra, Liane; Teijeira, Marta; Rivero-Buceta, Virginia; Helguera, Aliuska Morales; Celeiro, Maria; Terán, Ma Carmen; Besada, Pedro; Borges, Fernanda


    Adenosine regulates tissue function by activating four G-protein-coupled adenosine receptors (ARs). Selective agonists and antagonists for A3 ARs have been investigated for the treatment of a variety of immune disorders, cancer, brain, and heart ischemic conditions. We herein present a QSAR study based on a Topological sub-structural molecular design (TOPS-MODE) approach, intended to predict the A3 ARs of a diverse dataset of 124 (94 training set/ 30 prediction set) adenosine derivatives. The final model showed good fit and predictive capability, displaying 85.1 % of the experimental variance. The TOPS-MODE approach afforded a better understanding and interpretation of the developed model based on the useful information extracted from the analysis of the contribution of different molecular fragments to the affinity.

  8. Allosteric modulators affect the internalization of human adenosine A1 receptors.

    NARCIS (Netherlands)

    Klaasse, E.C.; Hout, G. van den; Roerink, S.F.; Grip, W.J. de; IJzerman, A.P.; Beukers, M.W.


    To study the effect of allosteric modulators on the internalization of human adenosine A(1) receptors, the receptor was equipped with a C-terminal yellow fluorescent protein tag. The introduction of this tag did not affect the radioligand binding properties of the receptor. CHO cells stably expressi

  9. In silico discovery of novel Retinoic Acid Receptor agonist structures

    Directory of Open Access Journals (Sweden)

    Samuels Herbert H


    Full Text Available Abstract Background Several Retinoic Acid Receptors (RAR agonists have therapeutic activity against a variety of cancer types; however, unacceptable toxicity profiles have hindered the development of drugs. RAR agonists presenting novel structural and chemical features could therefore open new avenues for the discovery of leads against breast, lung and prostate cancer or leukemia. Results We have analysed the induced fit of the active site residues upon binding of a known ligand. The derived binding site models were used to dock over 150,000 molecules in silico (or virtually to the structure of the receptor with the Internal Coordinates Mechanics (ICM program. Thirty ligand candidates were tested in vitro. Conclusions Two novel agonists resulting from the predicted receptor model were active at 50 nM. One of them displays novel structural features which may translate into the development of new ligands for cancer therapy.

  10. Principles of agonist recognition in Cys-loop receptors

    Directory of Open Access Journals (Sweden)

    Timothy eLynagh


    Full Text Available Cys-loop receptors are ligand-gated ion channels that are activated by a structurally diverse array of neurotransmitters, including acetylcholine, serotonin, glycine and GABA. After the term chemoreceptor emerged over 100 years ago, there was some wait until affinity labeling, molecular cloning, functional studies and X-ray crystallography experiments identified the extracellular interface of adjacent subunits as the principal site of agonist binding. The question of how subtle differences at and around agonist-binding sites of different Cys-loop receptors can accommodate transmitters as chemically diverse as glycine and serotonin has been subject to intense research over the last three decades. This review outlines the functional diversity and current structural understanding of agonist-binding sites, including those of invertebrate Cys-loop receptors. Together, this provides a framework to understand the atomic determinants involved in how these valuable therapeutic targets recognize and bind their ligands.

  11. Equilibrium and kinetic selectivity profiling on the human adenosine receptors. (United States)

    Guo, Dong; Dijksteel, Gabrielle S; van Duijl, Tirsa; Heezen, Maxime; Heitman, Laura H; IJzerman, Adriaan P


    Classical evaluation of target selectivity is usually undertaken by measuring the binding affinity of lead compounds against a number of potential targets under equilibrium conditions, without considering the kinetics of the ligand-receptor interaction. In the present study we propose a combined strategy including both equilibrium- and kinetics-based selectivity profiling. The adenosine receptor (AR) was chosen as a prototypical drug target. Six in-house AR antagonists were evaluated in a radioligand displacement assay for their affinity and in a competition association assay for their binding kinetics on three AR subtypes. One of the compounds with a promising kinetic selectivity profile was also examined in a [(35)S]-GTPγS binding assay for functional activity. We found that XAC and LUF5964 were kinetically more selective for the A1R and A3R, respectively, although they are non-selective in terms of their affinity. In comparison, LUF5967 displayed a strong equilibrium-based selectivity for the A1R over the A2AR, yet its kinetic selectivity thereon was less pronounced. In a GTPγS assay, LUF5964 exhibited insurmountable antagonism on the A3R while having a surmountable effect on the A1R, consistent with its kinetic selectivity profile. This study provides evidence that equilibrium and kinetic selectivity profiling can both be important in the early phases of the drug discovery process. Our proposed combinational strategy could be considered for future medicinal chemistry efforts and aid the design and discovery of different or even better leads for clinical applications.

  12. De novo analysis of receptor binding affinity data of xanthine adenosine receptor antagonists. (United States)

    Dalpiaz, A; Gardenghi, A; Borea, P A


    The receptor binding affinity data to adenosine A1 and A2 receptors of a wide series of xanthine derivatives have been analyzed by means of the Free-Wilson model. The analysis of the individual group contribution shows, for both A1 and A2 receptors, the primary importance of the presence of bulky substituents at position 8 for an optimum receptor binding. Moreover, considering the different aij contributions of bulky substituents at position 8 for affinity to A1 with respect to A2 receptors, this position appears to be the most important for the synthesis of highly A1 selective xanthine derivatives. Moreover the analysis of group contributions for other substitution positions of the xanthine moiety allows to state that suitable substitutions at positions 3 and 7 could confer some degree of A2 selectivity.

  13. Synthesis and pharmacological evaluation of dual acting ligands targeting the adenosine A2A and dopamine D2 receptors for the potential treatment of Parkinson's disease. (United States)

    Jörg, Manuela; May, Lauren T; Mak, Frankie S; Lee, Kiew Ching K; Miller, Neil D; Scammells, Peter J; Capuano, Ben


    A relatively new strategy in drug discovery is the development of dual acting ligands. These molecules are potentially able to interact at two orthosteric binding sites of a heterodimer simultaneously, possibly resulting in enhanced subtype selectivity, higher affinity, enhanced or modified physiological response, and reduced reliance on multiple drug administration regimens. In this study, we have successfully synthesized a series of classical heterobivalent ligands as well as a series of more integrated and "drug-like" dual acting molecules, incorporating ropinirole as a dopamine D2 receptor agonist and ZM 241385 as an adenosine A2A receptor antagonist. The best compounds of our series maintained the potency of the original pharmacophores at both receptors (adenosine A2A and dopamine D2). In addition, the integrated dual acting ligands also showed promising results in preliminary blood-brain barrier permeability tests, whereas the classical heterobivalent ligands are potentially more suited as pharmacological tools.

  14. Glucagon-like peptide 1 receptor agonist (GLP-1 RA)

    DEFF Research Database (Denmark)

    von Scholten, Bernt Johan; Hansen, Tine Willum; Goetze, Jens Peter;


    AIMS: In a short-term study including 31 patients with type 2 diabetes, glucagon-like peptide 1 receptor agonist (GLP-1 RA) treatment was associated with a significant reversible decline in GFR. Twenty-three patients re-initiated GLP-1 RA treatment after the primary study, and the aim was to inve......AIMS: In a short-term study including 31 patients with type 2 diabetes, glucagon-like peptide 1 receptor agonist (GLP-1 RA) treatment was associated with a significant reversible decline in GFR. Twenty-three patients re-initiated GLP-1 RA treatment after the primary study, and the aim...

  15. Rational design of sulfonated A3 adenosine receptor-selective nucleosides as pharmacological tools to study chronic neuropathic pain. (United States)

    Paoletta, Silvia; Tosh, Dilip K; Finley, Amanda; Gizewski, Elizabeth T; Moss, Steven M; Gao, Zhan-Guo; Auchampach, John A; Salvemini, Daniela; Jacobson, Kenneth A


    (N)-Methanocarba(bicyclo[3.1.0]hexane)adenosine derivatives were probed for sites of charged sulfonate substitution, which precludes diffusion across biological membranes, e.g., blood-brain barrier. Molecular modeling predicted that sulfonate groups on C2-phenylethynyl substituents would provide high affinity at both mouse (m) and human (h) A3 adenosine receptors (ARs), while a N(6)-p-sulfophenylethyl substituent would determine higher hA3AR vs mA3AR affinity. These modeling predictions, based on steric fitting of the binding cavity and crucial interactions with key residues, were confirmed by binding/efficacy studies of synthesized sulfonates. N(6)-3-Chlorobenzyl-2-(3-sulfophenylethynyl) derivative 7 (MRS5841) bound selectively to h/m A3ARs (Ki(hA3AR) = 1.9 nM) as agonist, while corresponding p-sulfo isomer 6 (MRS5701) displayed mixed A1/A3AR agonism. Both nucleosides administered ip reduced mouse chronic neuropathic pain that was ascribed to either A3AR or A1/A3AR using A3AR genetic deletion. Thus, rational design methods based on A3AR homology models successfully predicted sites for sulfonate incorporation, for delineating adenosine's CNS vs peripheral actions.

  16. Stimulation of NTS A1 adenosine receptors evokes counteracting effects on hindlimb vasculature. (United States)

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


    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

  17. Impairment of adenosine A3 receptor activity disrupts neutrophil migratory capacity and impacts innate immune function in vivo. (United States)

    Butler, Matt; Sanmugalingam, Devika; Burton, Victoria J; Wilson, Tammy; Pearson, Ruth; Watson, Robert P; Smith, Philip; Parkinson, Scott J


    Adenosine possesses potent anti-inflammatory properties which are partly mediated by G(i) -coupled adenosine A3 receptors (A3Rs). A3R agonists have shown clinical benefit in a number of inflammatory conditions although some studies in A3R-deficient mice suggest a pro-inflammatory role. We hypothesised that, in addition to cell signalling effects, A3R compounds might inhibit neutrophil chemotaxis by disrupting the purinergic feedback loop controlling leukocyte migration. Human neutrophil activation triggered rapid upregulation of surface A3R expression which was disrupted by pre-treatment with either agonist (Cl-IB-MECA) or antagonist (MRS1220). Both compounds reduced migration velocity and neutrophil transmigration capacity without impacting the response to chemokines per se. Similar effects were observed in murine neutrophils, while cells from A3R-deficient mice displayed a constitutively impaired migratory phenotype indicating compound-induced desensitisation and genetic ablation had the same functional outcome. In a dextran sodium sulphate-induced colitis model, A3R-deficient mice exhibited reduced colon pathology and decreased tissue myeloperoxidase levels at day 8 - consistent with reduced neutrophil recruitment. However, A3R-deficient mice were unable to resolve the dextran sodium sulphate-induced inflammation and had elevated numbers of tissue-associated bacteria by day 21. Our data indicate that A3Rs play a role in neutrophil migration and disrupting this function has the potential to adversely affect innate immune responses.

  18. Striatal adenosine A2A and cannabinoid CB1 receptors form functional heteromeric complexes that mediate the motor effects of cannabinoids. (United States)

    Carriba, Paulina; Ortiz, Oskar; Patkar, Kshitij; Justinova, Zuzana; Stroik, Jessica; Themann, Andrea; Müller, Christa; Woods, Anima S; Hope, Bruce T; Ciruela, Francisco; Casadó, Vicent; Canela, Enric I; Lluis, Carme; Goldberg, Steven R; Moratalla, Rosario; Franco, Rafael; Ferré, Sergi


    The mechanism of action responsible for the motor depressant effects of cannabinoids, which operate through centrally expressed cannabinoid CB1 receptors, is still a matter of debate. In the present study, we report that CB1 and adenosine A2A receptors form heteromeric complexes in co-transfected HEK-293T cells and rat striatum, where they colocalize in fibrilar structures. In a human neuroblastoma cell line, CB1 receptor signaling was found to be completely dependent on A2A receptor activation. Accordingly, blockade of A2A receptors counteracted the motor depressant effects produced by the intrastriatal administration of a cannabinoid CB1 receptor agonist. These biochemical and behavioral findings demonstrate that the profound motor effects of cannabinoids depend on physical and functional interactions between striatal A2A and CB1 receptors.

  19. 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


    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.

  20. Adenosine receptor signaling modulates permeability of the blood-brain barrier. (United States)

    Carman, Aaron J; Mills, Jeffrey H; Krenz, Antje; Kim, Do-Geun; Bynoe, Margaret S


    The blood-brain barrier (BBB) is comprised of specialized endothelial cells that form the capillary microvasculature of the CNS and is essential for brain function. It also poses the greatest impediment in the treatment of many CNS diseases because it commonly blocks entry of therapeutic compounds. Here we report that adenosine receptor (AR) signaling modulates BBB permeability in vivo. A(1) and A(2A) AR activation facilitated the entry of intravenously administered macromolecules, including large dextrans and antibodies to β-amyloid, into murine brains. Additionally, treatment with an FDA-approved selective A(2A) agonist, Lexiscan, also increased BBB permeability in murine models. These changes in BBB permeability are dose-dependent and temporally discrete. Transgenic mice lacking A(1) or A(2A) ARs showed diminished dextran entry into the brain after AR agonism. Following treatment with a broad-spectrum AR agonist, intravenously administered anti-β-amyloid antibody was observed to enter the CNS and bind β-amyloid plaques in a transgenic mouse model of Alzheimer's disease (AD). Selective AR activation resulted in cellular changes in vitro including decreased transendothelial electrical resistance, increased actinomyosin stress fiber formation, and alterations in tight junction molecules. These results suggest that AR signaling can be used to modulate BBB permeability in vivo to facilitate the entry of potentially therapeutic compounds into the CNS. AR signaling at brain endothelial cells represents a novel endogenous mechanism of modulating BBB permeability. We anticipate these results will aid in drug design, drug delivery and treatment options for neurological diseases such as AD, Parkinson's disease, multiple sclerosis and cancers of the CNS.

  1. Melatonin receptor agonists: new options for insomnia and depression treatment. (United States)

    Spadoni, Gilberto; Bedini, Annalida; Rivara, Silvia; Mor, Marco


    The circadian nature of melatonin (MLT) secretion, coupled with the localization of MLT receptors to the suprachiasmatic nucleus, has led to numerous studies of the role of MLT in modulation of the sleep-wake cycle and circadian rhythms in humans. Although much more needs to be understood about the various functions exerted by MLT and its mechanisms of action, three therapeutic agents (ramelteon, prolonged-release MLT, and agomelatine) are already in use, and MLT receptor agonists are now appearing as new promising treatment options for sleep and circadian-rhythm related disorders. In this review, emphasis has been placed on medicinal chemistry strategies leading to MLT receptor agonists, and on the evidence supporting therapeutic efficacy of compounds undergoing clinical evaluation. A wide range of clinical trials demonstrated that ramelteon, prolonged-release MLT and tasimelteon have sleep-promoting effects, providing an important treatment option for insomnia and transient insomnia, even if the improvements of sleep maintenance appear moderate. Well-documented effects of agomelatine suggest that this MLT agonist offers an attractive alternative for the treatment of depression, combining efficacy with a favorable side effect profile. Despite a large number of high affinity nonselective MLT receptor agonists, only limited data on MT₁ or MT₂ subtype-selective compounds are available up to now. Administration of the MT₂-selective agonist IIK7 to rats has proved to decrease NREM sleep onset latency, suggesting that MT₂ receptor subtype is involved in the acute sleep-promoting action of MLT; rigorous clinical studies are needed to demonstrate this hypothesis. Further clinical candidates based on selective activation of MT₁ or MT₂ receptors are expected in coming years.

  2. Evidence that the positive inotropic effects of the alkylxanthines are not due to adenosine receptor blockade. (United States)

    Collis, M. G.; Keddie, J. R.; Torr, S. R.


    We investigated the possibility that the positive inotropic effects of the alkylxanthines are due to adenosine receptor blockade. The potency of 8-phenyltheophylline, theophylline and enprofylline as adenosine antagonists was assessed in vitro, using the guinea-pig isolated atrium, and in vivo, using the anaesthetized dog. The order of potency of the alkylxanthines as antagonists of the negative inotropic response to 2-chloroadenosine in vitro, and of the hypotensive response to adenosine in vivo was 8-phenyltheophylline greater than theophylline greater than enprofylline. The order of potency of the alkylxanthines as positive inotropic and chronotropic agents in the anaesthetized dog was enprofylline greater than theophylline greater than 8-phenyltheophylline. The results of this study indicate that the inotropic effects of the alkylxanthines in the anaesthetized dog are not due to adenosine receptor blockade. PMID:6322898

  3. Toll-like receptor 2 agonists inhibit human fibrocyte differentiation

    Directory of Open Access Journals (Sweden)

    Maharjan Anu S


    Full Text Available Abstract Background In healing wounds, some monocytes enter the wound and differentiate into fibroblast-like cells called fibrocytes. Since Toll-like receptors (TLRs are present on monocytes, and pathogens that can infect a wound have and/or release TLR agonists, we examined whether TLR agonists affect fibrocyte differentiation. Results When human peripheral blood mononuclear cells (PBMCs were cultured with TLR3, TLR4, TLR5, TLR7, TLR8 or TLR9 agonists, there was no significant effect on fibrocyte differentiation, even though enhanced extracellular tumor necrosis factor (TNF-α accumulation and/or increased cell surface CD86 or major histocompatibility complex (MHC class II levels were observed. However, all TLR2 agonists tested inhibited fibrocyte differentiation without any significant effect on cell survival. Adding TLR2 agonists to purified monocytes had no effect on fibrocyte differentiation. However, some TLR2 agonists caused PBMCs to secrete a factor that inhibits the differentiation of purified monocytes into fibrocytes. This factor is not interferon (IFN-α, IFN-γ, interleukin (IL-12, aggregated immunoglobulin G (IgG or serum amyloid P (SAP, factors known to inhibit fibrocyte differentiation. TLR2 agonist-treated PBMCs secrete low levels of IL-6, TNF-α, IFN-γ, granulocyte colony-stimulating factor and tumor growth factor β1, but combinations of these factors had no effect on fibrocyte differentiation from purified monocytes. Conclusions Our results indicate that TLR2 agonists indirectly inhibit fibrocyte differentiation and that, for some TLR2 agonists, this inhibition involves other cell types in the PBMC population secreting an unknown factor that inhibits fibrocyte differentiation. Together, these data suggest that the presence of some bacterial signals can inhibit fibrocyte differentiation and may thus slow wound closure.

  4. Principles of agonist recognition in Cys-loop receptors

    DEFF Research Database (Denmark)

    Lynagh, Timothy Peter; Pless, Stephan Alexander


    diverse as glycine and serotonin has been subject to intense research over the last three decades. This review outlines the functional diversity and current structural understanding of agonist-binding sites, including those of invertebrate Cys-loop receptors. Together, this provides a framework...

  5. GABAA receptor partial agonists and antagonists

    DEFF Research Database (Denmark)

    Krall, Jacob; Balle, Thomas; Krogsgaard-Larsen, Niels;


    A high degree of structural heterogeneity of the GABAA receptors (GABAARs) has been revealed and is reflected in multiple receptor subtypes. The subunit composition of GABAAR subtypes is believed to determine their localization relative to the synapses and adapt their functional properties...... to the local temporal pattern of GABA impact, enabling phasic or tonic inhibition. Specific GABAAR antagonists are essential tools for physiological and pharmacological elucidation of the different type of GABAAR inhibition. However, distinct selectivity among the receptor subtypes (populations) has been shown...

  6. Adenosine enhances sweet taste through A2B receptors in the taste bud. (United States)

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


    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.

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

    DEFF Research Database (Denmark)

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


    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....

  8. Autoradiographic localization of adenosine receptors in rat brain using (/sup 3/H)cyclohexyladenosine

    Energy Technology Data Exchange (ETDEWEB)

    Goodman, R.R.; Synder, S.H.


    Adenosine (A1) receptor binding sites have been localized in rat brain by an in vitro light microscopic autoradiographic method. The binding of (/sup 3/H)N6-cyclohexyladenosine to slide-mounted rat brain tissue sections has the characteristics of A1 receptors. It is saturable with high affinity and has appropriate pharmacology and stereospecificity. The highest densities of adenosine receptors occur in the molecular layer of the cerebellum, the molecular and polymorphic layers of the hippocampus and dentate gyrus, the medial geniculate body, certain thalamic nuclei, and the lateral septum. High densities also are observed in certain layers of the cerebral cortex, the piriform cortex, the caudate-putamen, the nucleus accumbens, and the granule cell layer of the cerebellum. Most white matter areas, as well as certain gray matter areas, such as the hypothalamus, have negligible receptor concentrations. These localizations suggest possible central nervous system sites of action of adenosine.

  9. Agonist discrimination between AMPA receptor subtypes

    DEFF Research Database (Denmark)

    Coquelle, T; Christensen, J K; Banke, T G


    The lack of subtype-selective compounds for AMPA receptors (AMPA-R) led us to search for compounds with such selectivity. Homoibotenic acid analogues were investigated at recombinant GluR1o, GluR2o(R), GluR3o and GluR1o + 3o receptors expressed in Sf9 insect cells and affinities determined in [3H...


    NARCIS (Netherlands)



    The cells of origin of the perforant pathway are destroyed in Alzheimer's disease (AD). In rat the adenosine A1-receptors are specifically localized on the perforant path terminals in the molecular layer of the dentate gyrus. In the present study the density of A1-receptors in the hippocampus of Alz

  11. 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;


    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...

  12. Receptors and Channels Targeted by Synthetic Cannabinoid Receptor Agonists and Antagonists


    Pertwee, R. G.


    It is widely accepted that non-endogenous compounds that target CB1 and/or CB2 receptors possess therapeutic potential for the clinical management of an ever growing number of disorders. Just a few of these disorders are already treated with Δ9-tetrahydrocannabinol or nabilone, both CB1/CB2 receptor agonists, and there is now considerable interest in expanding the clinical applications of such agonists and also in exploiting CB2-selective agonists, peripherally restricted CB1/CB2 receptor ago...

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

    Directory of Open Access Journals (Sweden)

    Gérard Jean-Louis


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

  14. Identification of human dopamine receptors agonists from Chinese herbs

    Institute of Scientific and Technical Information of China (English)

    Yi-lin ZHANG; Hai-qing ZHANG; Xiao-yu LIU; Shi-neng HUA; Lu-bing ZHOU; Jun YU; Xue-hai TAN


    Aim: To find human dopamine receptors, especially D1-like receptor specific ago-nists from Chinese herbs as potential antihypertension drug leads. Methods: Two D1-like receptor cell lines carrying a β-lactamase reporter gene, and a D2 receptor cell line coexpressing a promiscuous G protein G15 were constructed using HEK293 cells. A natural compound library made from fractionated samples of herbal ex-tracts was used for high-throughput screening (HTS) against one of the cell lines,HEK/D5R/CRE-blax. The interested hits were evaluated for their activities against various dopamine receptors. Results: Fourteen hits were identified from primary screening, of which 2 of the better hit samples, HD0522 and HD0059, were selected for further material and activity analysis, and to obtain 2 compounds that ap-peared as 2 single peaks in HPLC, HD0522H01 and HD0059H01. HD0059H01 could activate D1, D2, and D5 receptors, with EC50 values of 2.28 μg/mL, 0.85 μg/mL, and 1.41 μg/mL, respectively. HD0522H01 could only activate D1R and D5R with EC50 values of 2.95 μg/mL and 8.38 μg/mL. Conclusion: We established cell-based assays for 3 different human dopamine receptors and identified specific agonists HD0522H01 and HD0059H01 through HTS. The specific agonist to D1-like receptors, HD0522H01, may become a new natural product-based drug lead for antihypertension treatment.

  15. Receptor discrimination and control of agonist-antagonist binding. (United States)

    Tallarida, R J


    The law of mass action is the common model for the interaction of agonist and antagonist compounds with cellular receptors. Parameters of the interaction, obtained from functional and radioligand-binding studies, allow discrimination and subtyping of receptors and aid in understanding specific mechanisms. This article reviews the theory and associated mathematical models and graphical transformations of data that underlie the determination of receptor parameters. The main theory assumes that agonist and antagonist compounds bind to cells that have a fixed number of receptors and provides the framework for obtaining drug-receptor parameters from data and their graphical transformations. Conditions that produce a change in receptor number, a newer concept in pharmacology, can have an important effect on the parameter values derived in the usual way. This review concludes with a discussion of the quantitative study of receptor-mediated feedback control of endogenous ligands, a very new topic with potentially important implications for understanding antagonist effectiveness, loss of control, and chaos in regulated mass action binding.

  16. 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.

  17. Improving the developability profile of pyrrolidine progesterone receptor partial agonists

    Energy Technology Data Exchange (ETDEWEB)

    Kallander, Lara S.; Washburn, David G.; Hoang, Tram H.; Frazee, James S.; Stoy, Patrick; Johnson, Latisha; Lu, Qing; Hammond, Marlys; Barton, Linda S.; Patterson, Jaclyn R.; Azzarano, Leonard M.; Nagilla, Rakesh; Madauss, Kevin P.; Williams, Shawn P.; Stewart, Eugene L.; Duraiswami, Chaya; Grygielko, Eugene T.; Xu, Xiaoping; Laping, Nicholas J.; Bray, Jeffrey D.; Thompson, Scott K. (GSKPA)


    The previously reported pyrrolidine class of progesterone receptor partial agonists demonstrated excellent potency but suffered from serious liabilities including hERG blockade and high volume of distribution in the rat. The basic pyrrolidine amine was intentionally converted to a sulfonamide, carbamate, or amide to address these liabilities. The evaluation of the degree of partial agonism for these non-basic pyrrolidine derivatives and demonstration of their efficacy in an in vivo model of endometriosis is disclosed herein.

  18. Characterization of the A2B adenosine receptor from mouse, rabbit, and dog. (United States)

    Auchampach, John A; Kreckler, Laura M; Wan, Tina C; Maas, Jason E; van der Hoeven, Dharini; Gizewski, Elizabeth; Narayanan, Jayashree; Maas, Garren E


    We have cloned and pharmacologically characterized the A(2B) adenosine receptor (AR) from the dog, rabbit, and mouse. The full coding regions of the dog and mouse A(2B)AR were obtained by reverse transcriptase-polymerase chain reaction, and the rabbit A(2B)AR cDNA was obtained by screening a rabbit brain cDNA library. It is noteworthy that an additional clone was isolated by library screening that was identical in sequence to the full-length rabbit A(2B)AR, with the exception of a 27-base pair deletion in the region encoding amino acids 103 to 111 (A(2B)AR(103-111)). This 9 amino acid deletion is located in the second intracellular loop at the only known splice junction of the A(2B)AR and seems to result from the use of an additional 5' donor site found in the rabbit and dog but not in the human, rat, or mouse sequences. [(3)H]3-Isobutyl-8-pyrrolidinoxanthine and 8-[4-[((4-cyano-[2,6-(3)H]-phenyl)carbamoylmethyl)oxy]phenyl]-1,3-di(n-propyl)xanthine ([(3)H]MRS 1754) bound with high affinity to membranes prepared from human embryonic kidney (HEK) 293 cells expressing mouse, rabbit, and dog A(2B)ARs. Competition binding studies performed with a panel of agonist (adenosine and 2-amino-3,5-dicyano-4-phenylpyridine analogs) and antagonist ligands identified similar potency orders for the A(2B)AR orthologs, although most xanthine antagonists displayed lower binding affinity for the dog A(2B)AR compared with A(2B)ARs from rabbit and mouse. No specific binding could be detected with membranes prepared from HEK 293 cells expressing the rabbit A(2B)AR(103-111) variant. Furthermore, the variant failed to stimulate adenylyl cyclase or calcium mobilization. We conclude that significant differences in antagonist pharmacology of the A(2B)AR exist between species and that some species express nonfunctional variants of the A(2B)AR due to "leaky" splicing.

  19. A tail of two signals: the C terminus of the A(2A)-adenosine receptor recruits alternative signaling pathways. (United States)

    Gsandtner, Ingrid; Freissmuth, Michael


    G protein-coupled receptors are endowed with carboxyl termini that vary greatly in length and sequence. In most instances, the distal portion of the C terminus is dispensable for G protein coupling. This is also true for the A(2A)-adenosine receptor, where the last 100 amino acids are of very modest relevance to G(s) coupling. The C terminus was originally viewed mainly as the docking site for regulatory proteins of the beta-arrestin family. These beta-arrestins bind to residues that have been phosphorylated by specialized kinases (G protein-coupled receptor kinases) and thereby initiate receptor desensitization and endocytosis. More recently, it has become clear that many additional "accessory" proteins bind to C termini of G protein-coupled receptors. The article by Sun et al. in the current issue of Molecular Pharmacology identifies translin-associated protein-X as yet another interaction partner of the A(2A) receptor; translin-associated protein allows the A(2A) receptor to impinge on the signaling mechanisms by which p53 regulates neuronal differentiation, but the underlying signaling pathways are uncharted territory. With a list of five known interaction partners, the C terminus of the A(2A) receptor becomes a crowded place. Hence, there must be rules that regulate the interaction. This allows the C terminus to act as coincidence detector and as signal integrator. Despite our ignorance about the precise mechanisms, the article has exciting implications: the gene encoding for translin-associated protein-X maps to a locus implicated in some forms of schizophrenia; A(2A) receptor agonists are candidate drugs for the treatment of schizophrenic symptoms. It is of obvious interest to explore a possible link.

  20. Ramelteon: A melatonin receptor agonist for the treatment of insomnia

    Directory of Open Access Journals (Sweden)

    Devi V


    Full Text Available Ramelteon is a novel MT1 and MT2 melatonin receptor selective agonist recently approved for the treatment of insomnia characterized by difficulty in sleep onset. It is a nonscheduled drug since it lacks the potential for abuse and does not interact with neurotransmitter receptors most associated with these phenomena. Although the effects of ramelteon use> 5 weeks are unknown, the available data confirms its safety and efficacy for short-term use. Clinical use and future research should uncover more information about ramelteon′s properties.

  1. Estrogen Receptor Agonists and Antagonists in the Yeast Estrogen Bioassay. (United States)

    Wang, Si; Bovee, Toine F H


    Cell-based bioassays can be used to predict the eventual biological activity of a substance on a living organism. In vitro reporter gene bioassays are based on recombinant vertebrate cell lines or yeast strains and especially the latter are easy-to-handle, cheap, and fast. Moreover, yeast cells do not express estrogen, androgen, progesterone or glucocorticoid receptors, and are thus powerful tools in the development of specific reporter gene systems that are devoid of crosstalk from other hormone pathways. This chapter describes our experience with an in-house developed RIKILT yeast estrogen bioassay for testing estrogen receptor agonists and antagonists, focusing on the applicability of the latter.

  2. Structural determinants of agonist-specific kinetics at the ionotropic glutamate receptor 2

    DEFF Research Database (Denmark)

    Holm, Mai Marie; Lunn, Marie-Louise; Traynelis, Stephen F;


    Glutamate receptors (GluRs) are the most abundant mediators of the fast excitatory neurotransmission in the human brain. Agonists will, after activation of the receptors, induce different degrees of desensitization. The efficacy of agonists strongly correlates with the agonist-induced closure of ...

  3. 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


    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.

  4. Impact of efficacy at the μ-opioid receptor on antinociceptive effects of combinations of μ-opioid receptor agonists and cannabinoid receptor agonists. (United States)

    Maguire, David R; France, Charles P


    Cannabinoid receptor agonists, such as Δ(9)-tetrahydrocannabinol (Δ(9)-THC), enhance the antinociceptive effects of μ-opioid receptor agonists, which suggests that combining cannabinoids with opioids would improve pain treatment. Combinations with lower efficacy agonists might be preferred and could avoid adverse effects associated with large doses; however, it is unclear whether interactions between opioids and cannabinoids vary across drugs with different efficacy. The antinociceptive effects of μ-opioid receptor agonists alone and in combination with cannabinoid receptor agonists were studied in rhesus monkeys (n = 4) using a warm water tail withdrawal procedure. Etorphine, fentanyl, morphine, buprenorphine, nalbuphine, Δ(9)-THC, and CP 55,940 (2-[(1R,2R,5R)-5-hydroxy-2-(3-hydroxypropyl) cyclohexyl]-5-(2-methyloctan-2-yl)phenol) each increased tail withdrawal latency. Pretreatment with doses of Δ(9)-THC (1.0 mg/kg) or CP 55,940 (0.032 mg/kg) that were ineffective alone shifted the fentanyl dose-effect curve leftward 20.6- and 52.9-fold, respectively, and the etorphine dose-effect curve leftward 12.4- and 19.6-fold, respectively. Δ(9)-THC and CP 55,940 shifted the morphine dose-effect curve leftward only 3.4- and 7.9-fold, respectively, and the buprenorphine curve only 5.4- and 4.1-fold, respectively. Neither Δ(9)-THC nor CP 55,940 significantly altered the effects of nalbuphine. Cannabinoid receptor agonists increase the antinociceptive potency of higher efficacy opioid receptor agonists more than lower efficacy agonists; however, because much smaller doses of each drug can be administered in combinations while achieving adequate pain relief and that other (e.g., abuse-related) effects of opioids do not appear to be enhanced by cannabinoids, these results provide additional support for combining opioids with cannabinoids to treat pain.

  5. 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


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

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

    Directory of Open Access Journals (Sweden)

    Gemma eNavarro


    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.

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

    Directory of Open Access Journals (Sweden)

    Víctor Fernández-Dueñas


    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.

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

    Directory of Open Access Journals (Sweden)

    Siew Lee Cheong


    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.

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

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


    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.

  10. Adenosine A2B receptor: from cell biology to human diseases (United States)

    Sun, Ying; Huang, Pingbo


    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.

  11. Identification of agonists for a group of human odorant receptors

    Directory of Open Access Journals (Sweden)

    Daniela eGonzalez-Kristeller


    Full Text Available Olfaction plays a critical role in several aspects of the human life. Odorants are detected by hundreds of odorant receptors (ORs which belong to the superfamily of G protein-coupled receptors. These receptors are expressed in the olfactory sensory neurons of the nose. The information provided by the activation of different combinations of ORs in the nose is transmitted to the brain, leading to odorant perception and emotional and behavioral responses. There are ~400 intact human ORs, and to date only a small percentage of these receptors (~10% have known agonists. The determination of the specificity of the human ORs will contribute to a better understanding of how odorants are discriminated by the olfactory system. In this work, we aimed to identify human specific ORs, that is, ORs that are present in humans but absent from other species, and their corresponding agonists. To do this, we first selected 22 OR gene sequences from the human genome with no counterparts in the mouse, rat or dog genomes. Then we used a heterologous expression system to screen a subset of these human ORs against a panel of odorants of biological relevance, including foodborne aroma volatiles. We found that different types of odorants are able to activate some of these previously uncharacterized human ORs.

  12. Quantum dot-based screening system for discovery of g protein-coupled receptor agonists. (United States)

    Lee, Junghan; Kwon, Yong-Jun; Choi, Youngseon; Kim, Hi Chul; Kim, Keumhyun; Kim, JinYeop; Park, Sun; Song, Rita


    Cellular imaging has emerged as an important tool to unravel biological complexity and to accelerate the drug-discovery process, including cell-based screening, target identification, and mechanism of action studies. Recently, semiconductor nanoparticles known as quantum dots (QDs) have attracted great interest in cellular imaging applications due to their unique photophysical properties such as size, tunable optical property, multiplexing capability, and photostability. Herein, we show that QDs can also be applied to assay development and eventually to high-throughput/content screening (HTS/HCS) for drug discovery. We have synthesized QDs modified with PEG and primary antibodies to be used as fluorescent probes for a cell-based HTS system. The G protein-coupled receptor (GPCR) family is known to be involved in most major diseases. We therefore constructed human osteosarcoma (U2OS) cells that specifically overexpress two types of differently tagged GPCRs: influenza hemagglutinin (HA) peptide-tagged κ-opioid receptors (κ-ORs) and GFP-tagged A3 adenosine receptors (A3AR). In this study, we have demonstrated that 1) anti-HA antibody-conjugated QDs could specifically label HA-tagged κ-ORs, 2) subsequent treatment of QD-tagged GPCR agonists allowed agonist-induced translocation to be monitored in real time, 3) excellent emission spectral properties of QD permitted the simultaneous detection of two GPCRs in one cell, and 4) the robust imaging capabilities of the QD-antibody conjugates could lead to reproducible quantitative data from high-content cellular images. These results suggest that the present QD-based GPCR inhibitor screening system can be a promising platform for further drug screening applications.

  13. Differential role of nitric oxide in regional sympathetic responses to stimulation of NTS A2a adenosine receptors. (United States)

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


    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

  14. Angiotensin receptor agonistic autoantibodies and hypertension: preeclampsia and beyond. (United States)

    Xia, Yang; Kellems, Rodney E


    Hypertensive disorders are life-threatening diseases with high morbidity and mortality, affecting billions of individuals worldwide. A multitude of underlying conditions may contribute to hypertension, thus the need for a plethora of treatment options to identify the approach that best meets the needs of individual patients. A growing body of evidence indicates that (1) autoantibodies that bind to and activate the major angiotensin II type I (AT₁) receptor exist in the circulation of patients with hypertensive disorders, (2) these autoantibodies contribute to disease pathophysiology, (3) antibody titers correlate to the severity of the disease, and (4) efforts to block or remove these pathogenic autoantibodies have therapeutic potential. These autoantibodies, termed AT₁ agonistic autoantibodies have been extensively characterized in preeclampsia, a life-threatening hypertensive condition of pregnancy. As reviewed here, these autoantibodies cause symptoms of preeclampsia when injected into pregnant mice. Somewhat surprisingly, these auto antibodies also appear in 3 animal models of preeclampsia. However, the occurrence of AT₁ agonistic autoantibodies is not restricted to pregnancy. These autoantibodies are prevalent among kidney transplant recipients who develop severe transplant rejection and malignant hypertension during the first week after transplantation. AT₁ agonistic autoantibodies are also highly abundant among a group of patients with essential hypertension that are refractory to standard therapy. More recently these autoantibodies have been seen in patients with the autoimmune disease, systemic sclerosis. These 3 examples extend the clinical impact of AT₁ agonistic autoantibodies beyond pregnancy. Research reviewed here raises the intriguing possibility that preeclampsia and other hypertensive conditions are autoimmune diseases characterized by the presence of pathogenic autoantibodies that activate the major angiotensin receptor, AT₁. These

  15. N-methyl-D-aspartic acid receptor agonists

    DEFF Research Database (Denmark)

    Madsen, U; Frydenvang, Karla Andrea; Ebert, B


    (R,S)-2-Amino-2-(3-hydroxy-5-methyl-4-isoxazolyl)acetic acid [(R,S)-AMAA, 4] is a potent and selective agonist at the N-methyl-D-aspartic acid (NMDA) subtype of excitatory amino acid receptors. Using the Ugi "four-component condensation" method, the two diastereomers (2R)- and (2S)-2-[3-(benzyloxy......) showed peak affinity for [3H]AMPA receptor sites (IC50 = 72 +/- 13 microM) and was shown to be a more potent inhibitor of [3H]CPP binding (IC50 = 3.7 +/- 1.5 microM) than (S)-AMAA (9) (IC50 = 61 +/- 6.4 microM). Neither enantiomer of AMAA affected [3H]kainic acid receptor binding significantly...

  16. Nucleus accumbens neurotransmission and effort-related choice behavior in food motivation: effects of drugs acting on dopamine, adenosine, and muscarinic acetylcholine receptors. (United States)

    Nunes, Eric J; Randall, Patrick A; Podurgiel, Samantha; Correa, Mercè; Salamone, John D


    Mesolimbic dopamine (DA) is a critical component of the brain circuitry regulating behavioral activation and effort-related processes. Although nucleus accumbens (NAc) DA depletions or antagonism leave aspects of appetite and primary food motivation intact, rats with impaired DA transmission reallocate their instrumental behavior away from food-reinforced tasks with high response requirements, and instead select less effortful food-seeking behaviors. Previous work showed that adenosine A2A antagonists can reverse the effects of DA D2 antagonists on effort-related choice, and that stimulation of adenosine A2A receptors produces behavioral effects that are similar to those induced by DA antagonism. The present review summarizes the literature on the role of NAc DA and adenosine in effort-related processes, and also presents original data on the effects of local stimulation of muscarinic acetylcholine receptors in NAc core. Local injections of the muscarinic agonist pilocarpine directly into NAc core produces shifts in effort-related choice behavior similar to those induced by DA antagonism or A2A receptor stimulation, decreasing lever pressing but increasing chow intake in rats responding on a concurrent fixed ratio/chow feeding choice task. In contrast, injections into a neostriatal control site dorsal to the NAc were ineffective. The actions of pilocarpine on this task were attenuated by co-administration of the muscarinic antagonist scopolamine. Thus, drugs that act on DA, adenosine A2A, and muscarinic receptors regulate effort-related choice behavior, which may have implications for the treatment of psychiatric symptoms such as psychomotor slowing, fatigue or anergia that can be observed in depression and other disorders.

  17. Gi-protein-coupled 5-HT1B/D receptor agonist sumatriptan induces type I hyperalgesic priming. (United States)

    Araldi, Dioneia; Ferrari, Luiz F; Levine, Jon D


    We have recently described a novel form of hyperalgesic priming (type II) induced by agonists at two clinically important Gi-protein-coupled receptors (Gi-GPCRs), mu-opioid and A1-adenosine. Like mu-opioids, the antimigraine triptans, which act at 5-HT1B/D Gi-GPCRs, have been implicated in pain chronification. We determined whether sumatriptan, a prototypical 5-HT1B/D agonist, produces type II priming. Characteristic of hyperalgesic priming, intradermal injection of sumatriptan (10 ng) induced a change in nociceptor function such that a subsequent injection of prostaglandin-E2 (PGE2) induces prolonged mechanical hyperalgesia. However, onset to priming was delayed 3 days, characteristic of type I priming. Also characteristic of type I priming, a protein kinase Cε, but not a protein kinase A inhibitor attenuated the prolongation phase of PGE2 hyperalgesia. The prolongation of PGE2 hyperalgesia was also permanently reversed by intradermal injection of cordycepin, a protein translation inhibitor. Also, hyperalgesic priming did not occur in animals pretreated with pertussis toxin or isolectin B4-positive nociceptor toxin, IB4-saporin. Finally, as observed for other agonists that induce type I priming, sumatriptan did not induce priming in female rats. The prolongation of PGE2 hyperalgesia induced by sumatriptan was partially prevented by coinjection of antagonists for the 5-HT1B and 5-HT1D, but not 5-HT7, serotonin receptors and completely prevented by coadministration of a combination of the 5-HT1B and 5-HT1D antagonists. Moreover, the injection of selective agonists, for 5-HT1B and 5-HT1D receptors, also induced hyperalgesic priming. Our results suggest that sumatriptan, which signals through Gi-GPCRs, induces type I hyperalgesic priming, unlike agonists at other Gi-GPCRs, which induce type II priming.

  18. GLP-1 receptor agonist-induced polyarthritis: a case report. (United States)

    Ambrosio, Maria Luisa; Monami, Matteo; Sati, Lavinia; Marchionni, Niccolò; Di Bari, Mauro; Mannucci, Edoardo


    Occasional cases of bilateral, symmetrical, seronegative polyarthritis have been reported in patients treated with dipeptidyl peptidase-4 inhibitors (Crickx et al. in Rheumatol Int, 2013). We report here a similar case observed during treatment with a GLP-1 receptor agonist. A 42-year-old man with type 2 diabetes treated with metformin 1,500 mg/day and liraglutide 1.8 mg/day. After 6 months from the beginning of treatment, the patient complained of bilateral arthralgia (hands, feet, ankles, knees, and hips). Erythrocyte sedimentation rate (ESR), C-reactive protein (CRP), and leukocytes were increased. Rheumatoid factor, anticyclic citrullinated protein antibody, antinuclear antibodies, anti-Borrelia, and burgdorferi antibodies were all negative, and myoglobin and calcitonin were normal. Liraglutide was withdrawn, and the symptoms completely disappeared within 1 week, with normalization of ESR, CRP, fibrinogen, and leukocytes. Previously described cases of polyarthritis associated with DPP4 inhibitors had been attributed to a direct effect of the drugs on inflammatory cells expressing the enzyme. The present case, occurred during treatment with a GLP-1 receptor agonists, suggests a possibly different mechanism, mediated by GLP-1 receptor stimulation, which deserved further investigation.

  19. Activation of the adenosine A2A receptor exacerbates experimental autoimmune neuritis in Lewis rats in association with enhanced humoral immunity. (United States)

    Zhang, Min; Li, Xiao-Li; Li, Heng; Wang, Shan; Wang, Cong-Cong; Yue, Long-Tao; Xu, Hua; Zhang, Peng; Chen, Hui; Yang, Bing; Duan, Rui-Sheng


    Accumulated evidence demonstrated that Adenosine A2A receptor (A2AR) is involved in the inflammatory diseases. In the present study, we showed that a selective A2AR agonist, CGS21680, exacerbated experimental autoimmune neuritis in Lewis rats induced with bovine peripheral myelin. The exacerbation was accompanied with reduced CD4(+)Foxp3(+) T cells, increased CD4(+)CXCR5(+) T cells, B cells, dendritic cells and antigen-specific autoantibodies, which is possibly due to the inhibition of IL-2 induced by CGS21680. Combined with previous studies, our data indicate that the effects of A2AR stimulation in vivo are variable in different diseases. Caution should be taken in the use of A2AR agonists.

  20. 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.


    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.

  1. 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


    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.

  2. Caffeine promotes anti-tumor immune response during tumor initiation: Involvement of the adenosine A2A receptor. (United States)

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


    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.

  3. Apparent affinity of 1,3-dipropyl-8-cyclopentylxanthine for adenosine A1 and A2 receptors in isolated tissues from guinea-pigs. (United States)

    Collis, M. G.; Stoggall, S. M.; Martin, F. M.


    1. The classification of adenosine receptor subtypes (A1 and A2) in intact tissues has been based on the order of agonist potency. In this study the apparent affinity of 1,3-dipropyl-8-cyclopentylxanthine (CPX), an antagonist which has been reported to be A1 selective, and the non-selective antagonist 1,3-dimethyl-8-phenylxanthine (8PT) has been evaluated on isolated tissues from the guinea-pig. 2. The isolated tissues used were atria (bradycardic response, proposed A1 sub-type), aorta and trachea (relaxant response, proposed A2 sub-type). 3. Both the xanthines antagonized responses to adenosine in the three tissues but had little or no effect on responses to carbachol (atria), sodium nitrite (aorta) or isoprenaline (trachea). 4. pA2 values for 8PT were similar on the three tissues (6.3-6.7), however, the pA2 value for CPX on the atria (7.9-8.4) was greater than that on the aorta (6.6) or trachea (6.6). 5. These results support the suggestion that the adenosine receptors which mediate bradycardia in the atrium are of the A1 sub-type and that those which mediate relaxation in the aorta and trachea are of the A2 type. PMID:2790383

  4. Electrophysiologic effects of adenosine triphosphate on rabbit sinoatrial node pace maker cells via P1 receptors

    Institute of Scientific and Technical Information of China (English)

    RENLei-Ming; LIJun-Xia; SHIChen-Xia; ZHAODing


    AIM: To study the electrophysiologic effects of adenosine triphosphate (ATP) on rabbit sinoatrial node pacemakercells and the receptors related with the action of ATP. METHODS: Intracellular microelectrode method was usedto record the parameters of action potential (AP) in the rabbit sinoatrial nodes. RESULTS: ATP (0.1-3 mmol/L)decreased the rate of pacemaker firing (RPF) by 16 %-43 % and velocity of diastolic depolarization (VDD) by 33 %-67 %, increased the amplitude of AP (APA) by 6 %-9 % and maximal rate of depolarization (Vmax) by 30 %-76 %,shortened APD50 by 7 %-12 % and APD90 by 6.3 %-9 %, concentration-dependently. The effects of ATP, adenos-ine (Ado), and adenosine diphosphate at the same concentration on AP were not different from each other significantly.Neither uridine triphosphate nor, α,β-methylene ATP had significant electrophysiologic effects on the sinoatrialnode of rabbits. Both the electrophysiologic effects of ATP and Ado on pacemaker cells were inhibited by P1receptor antagonist aminophylline 0.1 mmol/L (P0.05). CONCLUSION: There are nofunctional P2X1 and P2Y2 receptors on pacemaker cells of the rabbit sinoatrial nodes, and the electrophysiologiceffects of ATP in the rabbit sinoatrial node pacemaker cells are mediated via P1 receptors by Ado degraded fromATP.

  5. Agonist signalling properties of radiotracers used for imaging of dopamine D-2/3 receptors

    NARCIS (Netherlands)

    van Wieringen, Jan-Peter; Michel, Martin C.; Janssen, Henk M.; Janssen, Anton G.; Elsinga, Philip H.; Booij, Jan


    Background: Dopamine D-2/3 receptor (D2/3R) agonist radiopharmaceuticals are considered superior to antagonists to detect dopamine release, e.g. induced by amphetamines. Agonists bind preferentially to the high-affinity state of the dopamine D2R, which has been proposed as the reason why agonists ar

  6. Kinetic analysis of antagonist-occupied adenosine-A3 receptors within membrane microdomains of individual cells provides evidence of receptor dimerization and allosterism. (United States)

    Corriden, Ross; Kilpatrick, Laura E; Kellam, Barrie; Briddon, Stephen J; Hill, Stephen J


    In our previous work, using a fluorescent adenosine-A3 receptor (A3AR) agonist and fluorescence correlation spectroscopy (FCS), we demonstrated high-affinity labeling of the active receptor (R*) conformation. In the current study, we used a fluorescent A3AR antagonist (CA200645) to study the binding characteristics of antagonist-occupied inactive receptor (R) conformations in membrane microdomains of individual cells. FCS analysis of CA200645-occupied A3ARs revealed 2 species, τD2 and τD3, that diffused at 2.29 ± 0.35 and 0.09 ± 0.03 μm(2)/s, respectively. FCS analysis of a green fluorescent protein (GFP)-tagged A3AR exhibited a single diffusing species (0.105 μm(2)/s). The binding of CA200645 to τD3 was antagonized by nanomolar concentrations of the A3 antagonist MRS 1220, but not by the agonist NECA (up to 300 nM), consistent with labeling of R. CA200645 normally dissociated slowly from the A3AR, but inclusion of xanthine amine congener (XAC) or VUF 5455 during washout markedly accelerated the reduction in the number of particles exhibiting τD3 characteristics. It is notable that this effect was accompanied by a significant increase in the number of particles with τD2 diffusion. These data show that FCS analysis of ligand-occupied receptors provides a unique means of monitoring ligand A3AR residence times that are significantly reduced as a consequence of allosteric interaction across the dimer interface

  7. 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


    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 receptoragonist 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

  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


    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. Binding Mode of Insulin Receptor and Agonist Peptide

    Institute of Scientific and Technical Information of China (English)


    Insulin is a protein hormone secreted by pancreatic β cells. One of its main functions is to keep the balance of glucose inside the body by regulating the absorption and metabolism of glucose in the periphery tissue, as well as the production and storage of hepatic glycogen. The insulin receptor is a transmembrane glycoprotein in which two α subunits with a molecular weight of 135 kD and twoβ subunits with a molecular weight of 95 kD are joined by a disulfide bond to form a β-α-α-β structure. The extracellular α subunit, especially, its three domains near the N-terminal are partially responsible for signal transduction or ligand-binding, as indicated by the experiments. The extracellular α subunits are involved in binding the ligands. The experimental results indicate that the three domains of the N-terminal of the α subunits are the main determinative parts of the insulin receptor to bind the insulin or mimetic peptide.We employed the extracellular domain (PDBID: 1IGR) of the insulin-like growth factor-1 receptor (IGF-1 R ) as the template to simulate and optimize the spatial structures of the three domains in the extracellular domain of the insulin receptor, which includes 468 residues. The work was accomplished by making use of the homology program in the Insight Ⅱ package on an Origin3800 server. The docking calculations of the insulin receptor obtained by homology with hexapeptides were carried out by means of the program Affinity. The analysis indicated that there were hydrogen bonding, and electrostatic and hydrophobic effects in the docking complex of the insulin receptor with hexapeptides.Moreover, we described the spatial orientation of a mimetic peptide with agonist activity in the docking complex. We obtained a rough model of binding of DLAPSQ or STIVYS with the insulin receptor, which provides the powerful theoretical support for designing the minimal insulin mimetic peptide with agonist activity, making it possible to develop oral small

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

    Fishman, Pnina; Cohen, Shira


    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.

  11. Primary Macrophage Chemotaxis Induced by Cannabinoid Receptor 2 Agonists Occurs Independently of the CB2 Receptor. (United States)

    Taylor, Lewis; Christou, Ivy; Kapellos, Theodore S; Buchan, Alice; Brodermann, Maximillian H; Gianella-Borradori, Matteo; Russell, Angela; Iqbal, Asif J; Greaves, David R


    Activation of CB2 has been demonstrated to induce directed immune cell migration. However, the ability of CB2 to act as a chemoattractant receptor in macrophages remains largely unexplored. Using a real-time chemotaxis assay and a panel of chemically diverse and widely used CB2 agonists, we set out to examine whether CB2 modulates primary murine macrophage chemotaxis. We report that of 12 agonists tested, only JWH133, HU308, L-759,656 and L-759,633 acted as macrophage chemoattractants. Surprisingly, neither pharmacological inhibition nor genetic ablation of CB2 had any effect on CB2 agonist-induced macrophage chemotaxis. As chemotaxis was pertussis toxin sensitive in both WT and CB2(-/-) macrophages, we concluded that a non-CB1/CB2, Gi/o-coupled GPCR must be responsible for CB2 agonist-induced macrophage migration. The obvious candidate receptors GPR18 and GPR55 could not mediate JWH133 or HU308-induced cytoskeletal rearrangement or JWH133-induced β-arrestin recruitment in cells transfected with either receptor, demonstrating that neither are the unidentified GPCR. Taken together our results conclusively demonstrate that CB2 is not a chemoattractant receptor for murine macrophages. Furthermore we show for the first time that JWH133, HU308, L-759,656 and L-759,633 have off-target effects of functional consequence in primary cells and we believe that our findings have wide ranging implications for the entire cannabinoid field.

  12. Carbamazepine-induced upregulation of adenosine A(1)-receptors in astrocyte cultures affects coupling to the phosphoinositol signaling pathway

    NARCIS (Netherlands)

    Biber, K; Fiebich, BL; Gebicke-Harter, P; van Calker, D


    The anticonvulsant and antibipolar drug carbamazepine (CBZ) is known to act as a specific antagonist at adenosine A(1)-receptors. After a 3-week application of CBZ, A(1)-receptors are upregulated in the rat brain. We have investigated the consequences of this upregulation for the A(1)-receptor-media

  13. Biased signaling by peptide agonists of protease activated receptor 2. (United States)

    Jiang, Yuhong; Yau, Mei-Kwan; Kok, W Mei; Lim, Junxian; Wu, Kai-Chen; Liu, Ligong; Hill, Timothy A; Suen, Jacky Y; Fairlie, David P


    Protease activated receptor 2 (PAR2) is associated with metabolism, obesity, inflammatory, respiratory and gastrointestinal disorders, pain, cancer and other diseases. The extracellular N-terminus of PAR2 is a common target for multiple proteases, which cleave it at different sites to generate different N-termini that activate different PAR2-mediated intracellular signaling pathways. There are no synthetic PAR2 ligands that reproduce the same signaling profiles and potencies as proteases. Structure-activity relationships here for 26 compounds spanned a signaling bias over 3 log units, culminating in three small ligands as biased agonist tools for interrogating PAR2 functions. DF253 (2f-LAAAAI-NH2) triggered PAR2-mediated calcium release (EC50 2 μM) but not ERK1/2 phosphorylation (EC50 > 100 μM) in CHO cells transfected with hPAR2. AY77 (Isox-Cha-Chg-NH2) was a more potent calcium-biased agonist (EC50 40 nM, Ca2+; EC50 2 μM, ERK1/2), while its analogue AY254 (Isox-Cha-Chg-A-R-NH2) was an ERK-biased agonist (EC50 2 nM, ERK1/2; EC50 80 nM, Ca2+). Signaling bias led to different functional responses in human colorectal carcinoma cells (HT29). AY254, but not AY77 or DF253, attenuated cytokine-induced caspase 3/8 activation, promoted scratch-wound healing and induced IL-8 secretion, all via PAR2-ERK1/2 signaling. Different ligand components were responsible for different PAR2 signaling and functions, clues that can potentially lead to drugs that modulate different pathway-selective cellular and physiological responses.

  14. Sumatriptan (5-HT1D receptor agonist) does not exacerbate symptoms in obsessive compulsive disorder

    NARCIS (Netherlands)

    Pian, KLH; Westerberg, HGM; van Megen, HJGM; den Boer, JA


    The non-selective serotonin (5-HT) receptor agonist meta-chlorophenylpiperazine (mCPP) has been reported to elicit symptoms in patients with obsessive compulsive disorder (OCD). MK-212, another nonselective 5-HT receptor agonist, does not seem to induce obsessive compulsive symptoms in OCD patients.

  15. The 5-HT(1F) receptor agonist lasmiditan as a potential treatment of migraine attacks

    DEFF Research Database (Denmark)

    Tfelt-Hansen, Peer C; Olesen, Jes


    Lasmiditan is a novel selective 5-HT(1F) receptor agonist. It is both scientifically and clinically relevant to review whether a 5-HT(1F) receptor agonist is effective in the acute treatment of migraine. Two RCTs in the phase II development of lasmiditan was reviewed. In the intravenous placebo...

  16. A Unified Model of the GABA(A) Receptor Comprising Agonist and Benzodiazepine Binding Sites

    DEFF Research Database (Denmark)

    Kongsbak, Kristine Grønning; Bergmann, Rikke; Sørensen, Pernille Louise


    We present a full-length a1b2c2 GABA receptor model optimized for agonists and benzodiazepine (BZD) allosteric modulators. We propose binding hypotheses for the agonists GABA, muscimol and THIP and for the allosteric modulator diazepam (DZP). The receptor model is primarily based on the glutamate...

  17. 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


    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.

  18. 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. (United States)

    Serpa, André; Correia, Sara; Ribeiro, Joaquim A; Sebastião, Ana M; Cascalheira, José F


    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 (Peffects 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.

  19. Nicotinic acetylcholine receptor agonist attenuates ILC2-dependent airway hyperreactivity (United States)

    Galle-Treger, Lauriane; Suzuki, Yuzo; Patel, Nisheel; Sankaranarayanan, Ishwarya; Aron, Jennifer L.; Maazi, Hadi; Chen, Lin; Akbari, Omid


    Allergic asthma is a complex and chronic inflammatory disorder that is associated with airway hyperreactivity (AHR) and driven by Th2 cytokine secretion. Type 2 innate lymphoid cells (ILC2s) produce large amounts of Th2 cytokines and contribute to the development of AHR. Here, we show that ILC2s express the α7-nicotinic acetylcholine receptor (α7nAChR), which is thought to have an anti-inflammatory role in several inflammatory diseases. We show that engagement of a specific agonist with α7nAChR on ILC2s reduces ILC2 effector function and represses ILC2-dependent AHR, while decreasing expression of ILC2 key transcription factor GATA-3 and critical inflammatory modulator NF-κB, and reducing phosphorylation of upstream kinase IKKα/β. Additionally, the specific α7nAChR agonist reduces cytokine production and AHR in a humanized ILC2 mouse model. Collectively, our data suggest that α7nAChR expressed by ILC2s is a potential therapeutic target for the treatment of ILC2-mediated asthma. PMID:27752043

  20. Toll-Like Receptor 9 Agonists for Cancer Therapy

    Directory of Open Access Journals (Sweden)

    Davide Melisi


    Full Text Available The immune system has acquired increasing importance as a key player in cancer maintenance and growth. Thus, modulating anti-tumor immune mediators has become an attractive strategy for cancer treatment. Toll-like receptors (TLRs have gradually emerged as potential targets of newer immunotherapies. TLR-9 is preferentially expressed on endosome membranes of B-cells and plasmacytoid dendritic cells (pDC and is known for its ability to stimulate specific immune reactions through the activation of inflammation-like innate responses. Several synthetic CpG oligonucleotides (ODNs have been developed as TLR-9 agonists with the aim of enhancing cancer immune surveillance. In many preclinical models, CpG ODNs were found to suppress tumor growth and proliferation both in monotherapy and in addition to chemotherapies or target therapies. TLR-9 agonists have been also tested in several clinical trials in patients with solid tumors. These agents showed good tolerability and usually met activity endpoints in early phase trials. However, they have not yet been demonstrated to significantly impact survival, neither as single agent treatments, nor in combination with chemotherapies or cancer vaccines. Further investigations in larger prospective studies are required.

  1. Crystal Structures of the Nuclear Receptor, Liver Receptor Homolog 1, Bound to Synthetic Agonists. (United States)

    Mays, Suzanne G; Okafor, C Denise; Whitby, Richard J; Goswami, Devrishi; Stec, Józef; Flynn, Autumn R; Dugan, Michael C; Jui, Nathan T; Griffin, Patrick R; Ortlund, Eric A


    Liver receptor homolog 1 (NR5A2, LRH-1) is an orphan nuclear hormone receptor that regulates diverse biological processes, including metabolism, proliferation, and the resolution of endoplasmic reticulum stress. Although preclinical and cellular studies demonstrate that LRH-1 has great potential as a therapeutic target for metabolic diseases and cancer, development of LRH-1 modulators has been difficult. Recently, systematic modifications to one of the few known chemical scaffolds capable of activating LRH-1 failed to improve efficacy substantially. Moreover, mechanisms through which LRH-1 is activated by synthetic ligands are entirely unknown. Here, we use x-ray crystallography and other structural methods to explore conformational changes and receptor-ligand interactions associated with LRH-1 activation by a set of related agonists. Unlike phospholipid LRH-1 ligands, these agonists bind deep in the pocket and do not interact with residues near the mouth nor do they expand the pocket like phospholipids. Unexpectedly, two closely related agonists with similar efficacies (GSK8470 and RJW100) exhibit completely different binding modes. The dramatic repositioning is influenced by a differential ability to establish stable face-to-face π-π-stacking with the LRH-1 residue His-390, as well as by a novel polar interaction mediated by the RJW100 hydroxyl group. The differing binding modes result in distinct mechanisms of action for the two agonists. Finally, we identify a network of conserved water molecules near the ligand-binding site that are important for activation by both agonists. This work reveals a previously unappreciated complexity associated with LRH-1 agonist development and offers insights into rational design strategies.

  2. Use of thrombopoietin receptor agonists in childhood immune thrombocytopenia

    Directory of Open Access Journals (Sweden)

    Angelica Maria Garzon


    Full Text Available Most children with immune thrombocytopenia (ITP will have spontaneous remission regardless of therapy, while about 20% will go on to have chronic ITP. In those children with chronic ITP who need treatment, standard therapies for acute ITP may have adverse effects that complicate their long term use. Thus, alternative treatment options are needed for children with chronic ITP. Thrombopoietin receptor agonists (TPO-RA have been shown to be safe and efficacious in adults with ITP, and represent a new treatment option for children with chronic ITP. One TPO-RA, eltrombopag, is now approved for children. Clinical trials in children are ongoing and data is emerging on safety and efficacy. This review will focus on the physiology of TPO-RA, their clinical use in children, as well as the long term safety issues that need to be considered when using these agents

  3. Adenosine A2A receptor deficiency up-regulates cystatin F expression in white matter lesions induced by chronic cerebral hypoperfusion. (United States)

    Duan, Wei; Ran, Hong; Zhou, Zhujuan; He, Qifen; Zheng, Jian


    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.

  4. 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.

  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;


    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. Combining a GLP-1 receptor agonist and basal insulin: study evidence and practical considerations. (United States)

    Carris, Nicholas W; Taylor, James R; Gums, John G


    Most patients with diabetes mellitus require multiple medications to achieve glycemic goals. Considering this and the increasing incidence of type 2 diabetes worldwide, the need for effective combination therapy is pressing. Basal insulin and glucagon-like peptide 1 (GLP-1) receptor agonists are frequently used to treat type 2 diabetes. Though both classes of medication are exclusively injectable, which may cause initial hesitation from providers, evidence for their combined use is substantial. This review summarizes the theoretical benefit, supporting evidence, and implementation of a combined basal insulin-GLP-1 receptor agonist regimen. Basal insulin added to a GLP-1 receptor agonist reduces hemoglobin A1c (HbA1c) without weight gain or significantly increased hypoglycemia. A GLP-1 receptor agonist added to basal insulin reduces HbA1c and body weight. Compared with the addition of meal-time insulin to basal insulin, a GLP-1 receptor agonist produces similar or greater reduction in HbA1c, weight loss instead of weight gain, and less hypoglycemia. Gastrointestinal adverse events are common with GLP-1 receptor agonists, especially during initiation and titration. However, combination with basal insulin is not expected to augment expected adverse events that come with using a GLP-1 receptor agonist. Basal insulin can be added to a GLP-1 receptor agonist with a slow titration to target goal fasting plasma glucose. In patients starting a GLP-1 receptor agonist, the dose of basal insulin should be decreased by 20 % in patients with an HbA1c ≤8 %. The evidence from 15 randomized prospective studies supports the combined use of a GLP-1 receptor agonist with basal insulin in a broad range of patients with uncontrolled type 2 diabetes.

  7. Interleukin-1beta but not tumor necrosis factor-alpha potentiates neuronal damage by quinolinic acid: protection by an adenosine A2A receptor antagonist. (United States)

    Stone, Trevor W; Behan, Wilhelmina M H


    Quinolinic acid is an agonist at glutamate receptors sensitive to N-methyl-D-aspartate (NMDA). It has been implicated in neural dysfunction associated with infections, trauma, and ischemia, although its neurotoxic potency is relatively low. This study was designed to examine the effects of a combination of quinolinic acid and the proinflammatory cytokines interleukin-1beta (IL-1beta) and tumor necrosis factor-alpha (TNF-alpha). Compounds were administered to the hippocampus of anesthetized male rats, animals being allowed to recover for 7 days before histological analysis of the hippocampus for neuronal damage estimated by counting of intact, healthy neurons. A low dose of quinolinic acid or IL-1beta produced no damage by itself, but the two together induced a significant loss of pyramidal neurons in the hippocampus. Higher doses produced almost total loss of pyramidal cells. Intrahippocampal TNF-alpha produced no effect alone but significantly reduced the neuronal loss produced by quinolinic acid. The adenosine A(2A) receptor antagonist ZM241385 reduced neuronal loss produced by the combinations of quinolinic acid and IL-1beta. The results suggest that simultaneous quinolinic acid and IL-1beta, both being induced by cerebral infection or injury, are synergistic in the production of neuronal damage and could together contribute substantially to traumatic, infective, or ischemic cerebral damage. Antagonism of adenosine A(2A) receptors protects neurons against the combination of quinolinic acid and IL-1beta.

  8. 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.


    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

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

    Directory of Open Access Journals (Sweden)

    Stefania Merighi


    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.

  10. Discovery of Potent and Highly Selective A2B Adenosine Receptor Antagonist Chemotypes. (United States)

    El Maatougui, Abdelaziz; Azuaje, Jhonny; González-Gómez, Manuel; Miguez, Gabriel; Crespo, Abel; Carbajales, Carlos; Escalante, Luz; García-Mera, Xerardo; Gutiérrez-de-Terán, Hugo; Sotelo, Eddy


    Three novel families of A2B adenosine receptor antagonists were identified in the context of the structural exploration of the 3,4-dihydropyrimidin-2(1H)-one chemotype. The most appealing series contain imidazole, 1,2,4-triazole, or benzimidazole rings fused to the 2,3-positions of the parent diazinone core. The optimization process enabled identification of a highly potent (3.49 nM) A2B ligand that exhibits complete selectivity toward A1, A2A, and A3 receptors. The results of functional cAMP experiments confirmed the antagonistic behavior of representative ligands. The main SAR trends identified within the series were substantiated by a molecular modeling study based on a receptor-driven docking model constructed on the basis of the crystal structure of the human A2A receptor.

  11. A2B adenosine receptors stimulate IL-6 production in primary murine microglia through p38 MAPK kinase pathway. (United States)

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


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

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

    Institute of Scientific and Technical Information of China (English)

    Zhe ZHANG; Hui-cai GUO; Li-nan ZHANG; Yong-li WANG


    Aim: The present study investigated the effect of adenosine on Na+-K+ pumps in acutely isolated guinea pig (C, avia sp.) ven-tricular myocytes.Methods: The whole-cell, patch-damp technique was used to record the Na+-K+ pump current (Ip) in acutely isolated guinea pig ventricular myocytes.Results: Adenosine inhibited the high DHO-affinity pump current (Ih) in a concentration-dependent manner, which was blocked by the selective adenosine A1 receptor antagonist DPCPX and the general protein kinase C (PKC) antagonists stau-rosporine, GF 109203X or the specific δ isoform antagonist rottlerin. In addition, the inhibitory action of adenosine was mimicked by a selective A1 receptor agonist CCPA and a specific activator peptide of PKC-δ, PP114. In contrast, the selec-tive A2A receptor agonist CGS21680 and A3 receptor agonist Cl-IB-MECA did not affect lb. Application of the selective A2A receptor antagonist SCH58261 and A3 receptor antagonist MRS1191 also failed to block the effect of adenosine. Further-more, H89, a selective protein kinase A (PKA) antagonist, did not exert any effect on adenosine-induced Ih inhibition.Conclusion: The present study provides the electrophysiological evidence that adenosine can induce significant inhibition of Ih via adenosine A1 receptors and the PKC-δ isoform.

  13. Dihydromorphine-peptide hybrids with delta receptor agonistic and mu receptor antagonistic actions

    Energy Technology Data Exchange (ETDEWEB)

    Smith, C.B.; Medzihradsky, F.; Woods, J.H.


    The actions of two morphine derivatives with short peptide side chains were evaluated upon the contraction of the isolated mouse vas deferens and upon displacement of /sup 3/H-etorphine from rat brain membranes. NIH-9833 (N-(6,14-endoetheno-7,8-dihydromorphine-7-alpha-carbonyl)-L-phenylalanyl-L-leucine ethyl ester HCl) was a potent agonist upon the vas deferens. Its EC50 for inhibition of the twitch was 1.2 +/- 0.1 nM. Both naltrexone (10/sup -7/ M) a relatively nonselective opioid antagonist, and ICI-174864 (10/sup -/' M) a highly selective delta receptor antagonist, blocked the actions of NIH-9833 which indicates that this drug is a delta receptor agonist. In contrast, NIH-9835 (N-(6,14-endoetheno-7,8-dihydromorphine-7-alpha-carbonyl)-L-glycyl-L-phenylalanyl-L-leucine ethyl ester HCl), which differs from NIH-9835 by the presence of a single amino acid residue, was devoid of opioid agonistic activity but was a potent antagonist of the inhibitory actions on the vas deferens of morphine and sufentanil. NIH-9833 and NIH-9835 were potent displacers of /sup 3/H-etorphine from rat cerebral membranes with EC50's of 0.58 nM and 1.7 nM, respectively. The observation that addition of a single glycyl group changes a dihydromorphine-peptide analog from a potent delta receptor agonist to an equally potent mu receptor antagonist suggests that the two receptor sites might be structurally quite similar.

  14. Histamine H3 receptor activation counteracts adenosine A2A receptor-mediated enhancement of depolarization-evoked [3H]-GABA release from rat globus pallidus synaptosomes. (United States)

    Morales-Figueroa, Guadalupe-Elide; Márquez-Gómez, Ricardo; González-Pantoja, Raúl; Escamilla-Sánchez, Juan; Arias-Montaño, José-Antonio


    High levels of histamine H3 receptors (H3Rs) are found in the globus pallidus (GP), a neuronal nucleus in the basal ganglia involved in the control of motor behavior. By using rat GP isolated nerve terminals (synaptosomes), we studied whether H3R activation modified the previously reported enhancing action of adenosine A2A receptor (A2AR) stimulation on depolarization-evoked [(3)H]-GABA release. At 3 and 10 nM, the A2AR agonist CGS-21680 enhanced [(3)H]-GABA release induced by high K(+) (20 mM) and the effect of 3 nM CGS-21680 was prevented by the A2AR antagonist ZM-241385 (100 nM). The presence of presynaptic H3Rs was confirmed by the specific binding of N-α-[methyl-(3)H]-histamine to membranes from GP synaptosomes (maximum binding, Bmax, 1327 ± 79 fmol/mg protein; dissociation constant, Kd, 0.74 nM), which was inhibited by the H3R ligands immepip, clobenpropit, and A-331440 (inhibition constants, Ki, 0.28, 8.53, and 316 nM, respectively). Perfusion of synaptosomes with the H3R agonist immepip (100 nM) had no effect on K(+)-evoked [(3)H]-GABA release, but inhibited the stimulatory action of A2AR activation. In turn, the effect of immepip was blocked by the H3R antagonist clobenpropit, which had no significant effect of its own on K(+)-induced [(3)H]-GABA release. These data indicate that H3R activation selectively counteracts the facilitatory action of A2AR stimulation on GABA release from striato-pallidal projections.

  15. 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

  16. The effect of cannabidiol on ischemia/reperfusion-induced ventricular arrhythmias: the role of adenosine A1 receptors. (United States)

    Gonca, Ersöz; Darıcı, Faruk


    Cannabidiol (CBD) is a nonpsychoactive phytocannabinoid with anti-inflammatory activity mediated by enhancing adenosine signaling. As the adenosine A1 receptor activation confers protection against ischemia/reperfusion (I/R)-induced ventricular arrhythmias, we hypothesized that CBD may have antiarrhythmic effect through the activation of adenosine A1 receptor. Cannabidiol has recently been shown to suppress ischemia-induced ventricular arrhythmias. We aimed to research the effect of CBD on the incidence and the duration of I/R-induced ventricular arrhythmias and to investigate the role of adenosine A1 receptor activation in the possible antiarrhythmic effect of CBD. Myocardial ischemia and reperfusion was induced in anesthetized male rats by ligating the left anterior descending coronary artery for 6 minutes and by loosening the bond at the coronary artery, respectively. Cannabidiol alone was given in a dose of 50 µg/kg, 10 minutes prior to coronary artery occlusion and coadministrated with adenosine A1 receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine (DPCPX) in a dose of 100 µg/kg, 15 minutes prior to coronary artery occlusion to investigate whether the antiarrhythmic effect of CBD is modified by the activation of adenosine A1 receptors. The experimental groups were as follows: (1) vehicle control (n = 10), (2) CBD (n = 9), (3) DPCPX (n = 7), and (4) CBD + DPCPX group (n = 7). Cannabidiol treatment significantly decreased the incidence and the duration of ventricular tachycardia, total length of arrhythmias, and the arrhythmia scores compared to control during the reperfusion period. The DPCPX treatment alone did not affect the incidence and the duration of any type of arrhythmias. However, DPCPX aborted the antiarrhythmic effect of CBD when it was combined with it. The present results demonstrated that CBD has an antiarrhythmic effect against I/R-induced arrhythmias, and the antiarrhythmic effect of CBD may be mediated through the activation of adenosine

  17. Development of novel silicon-containing inverse agonists of retinoic acid receptor-related orphan receptors. (United States)

    Toyama, Hirozumi; Nakamura, Masaharu; Nakamura, Masahiko; Matsumoto, Yotaro; Nakagomi, Madoka; Hashimoto, Yuichi


    Retinoic acid receptor (RAR)-related orphan receptors (RORs) regulate a variety of physiological processes, including hepatic gluconeogenesis, lipid metabolism, circadian rhythm and immune function. The RAR agonist: all-trans retinoic acid was reported to be an RORβ inverse agonist, but no information is available regarding ROR activity of its synthetic analogue Am580. Therefore, we screened Am580 and some related tetramethyltetrahydronaphthalene derivatives and carried out structural development studies, including substitution of carbon atoms with silicon, with the aim of creating a potent ROR transcriptional inhibitor. The phenyl amide disila compound 22 showed the most potent ROR-inhibitory activity among the compounds examined. Its activity towards RORα, RORβ and RORγ was increased compared to that of Am580. The IC₅₀ values for RORα, RORβ and RORγ are 1.3, >10 and 4.5 μM, respectively.

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

    Directory of Open Access Journals (Sweden)

    Zhenzhen Hu


    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.

  19. Adenosine receptors as markers of brain iron deficiency: Implications for Restless Legs Syndrome. (United States)

    Quiroz, César; Gulyani, Seema; Ruiqian, Wan; Bonaventura, Jordi; Cutler, Roy; Pearson, Virginia; Allen, Richard P; Earley, Christopher J; Mattson, Mark P; Ferré, Sergi


    Deficits of sensorimotor integration with periodic limb movements during sleep (PLMS) and hyperarousal and sleep disturbances in Restless Legs Syndrome (RLS) constitute two pathophysiologically distinct but interrelated clinical phenomena, which seem to depend mostly on alterations in dopaminergic and glutamatergic neurotransmission, respectively. Brain iron deficiency is considered as a main pathogenetic mechanism in RLS. Rodents with brain iron deficiency represent a valuable pathophysiological model of RLS, although they do not display motor disturbances. Nevertheless, they develop the main neurochemical dopaminergic changes found in RLS, such as decrease in striatal dopamine D2 receptor density. On the other hand, brain iron deficient mice exhibit the characteristic pattern of hyperarousal in RLS, providing a tool to find the link between brain iron deficiency and sleep disturbances in RLS. The present study provides evidence for a role of the endogenous sleep-promoting factor adenosine. Three different experimental preparations, long-term (22 weeks) severe or moderate iron-deficient (ID) diets (3- or 7-ppm iron diet) in mice and short-term (3 weeks) severe ID diet (3-ppm iron diet) in rats, demonstrated a significant downregulation (Western blotting in mouse and radioligand binding saturation experiments in rat brain tissue) of adenosine A1 receptors (A1R) in the cortex and striatum, concomitant to striatal D2R downregulation. On the other hand, the previously reported upregulation of adenosine A2A receptors (A2AR) was only observed with severe ID in both mice and rats. The results suggest a key role for A1R downregulation in the PLMS and hyperarousal in RLS.

  20. The effects of caffeine on sleep in Drosophila require PKA activity, but not the adenosine receptor. (United States)

    Wu, Mark N; Ho, Karen; Crocker, Amanda; Yue, Zhifeng; Koh, Kyunghee; Sehgal, Amita


    Caffeine is one of the most widely consumed stimulants in the world and has been proposed to promote wakefulness by antagonizing function of the adenosine A2A receptor. Here, we show that chronic administration of caffeine reduces and fragments sleep in Drosophila and also lengthens circadian period. To identify the mechanisms underlying these effects of caffeine, we first generated mutants of the only known adenosine receptor in flies (dAdoR), which by sequence is most similar to the mammalian A2A receptor. Mutants lacking dAdoR have normal amounts of baseline sleep, as well as normal homeostatic responses to sleep deprivation. Surprisingly, these mutants respond normally to caffeine. On the other hand, the effects of caffeine on sleep and circadian rhythms are mimicked by a potent phosphodiesterase inhibitor, IBMX (3-isobutyl-1-methylxanthine). Using in vivo fluorescence resonance energy transfer imaging, we find that caffeine induces widespread increase in cAMP levels throughout the brain. Finally, the effects of caffeine on sleep are blocked in flies that have reduced neuronal PKA activity. We suggest that chronic administration of caffeine promotes wakefulness in Drosophila, at least in part, by inhibiting cAMP phosphodiesterase activity.

  1. The A2B adenosine receptor protects against inflammation and excessive vascular adhesion (United States)

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


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

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

    DEFF Research Database (Denmark)

    Ilie, A; Ciocan, D; Constantinescu, A O


    . Several lines of evidence suggest that BS reflects an impairment of neocortical connectivity. Here we tested in vivo whether synaptic depression by adenosine A1 receptor (A1R) activation contributes to BS patterns following GCI. Male Wistar rats were subjected to 1, 5 or 10 min of GCI using a "four...... of post-ischemic BS patterns following brief ischemic episodes. It is likely that synaptic depression by post-ischemic A1R activation functionally disrupts the connectivity within the cortical networks to an extent that promotes BS patterns....

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

    DEFF Research Database (Denmark)

    Novak, Ivana; Hede, Susanne; Hansen, Mette


    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......, plasma membrane of many PANC-1 cells, but only a few CFPAC-1 cells. Taken together, our data indicate that A(2A) receptors open Cl(-) channels in pancreatic ducts cells with functional CFTR. We propose that adenosine can stimulate pancreatic secretion and, thereby, is an active player in the acini...

  4. Adenosine A2A receptor antagonists exert motor and neuroprotective effects by distinct cellular mechanisms


    Yu, Liqun; Shen, Hai-Ying; Coelho, Joana E.; Araújo, Inês M.; HUANG, QING-YUAN; Day, Yuan-Ji; Rebola, Nelson; Canas, Paula M.; Rapp, Erica Kirsten; Ferrara, Jarrod; Taylor, Darcie; Müller, Christa E.; Linden, Joel; Cunha, Rodrigo A.; Chen, Jiang-Fan


    To investigate whether the motor and neuroprotective effects of adenosine A2A receptor (A2AR) antagonists are mediated by distinct cell types in the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) model of Parkinson's disease.We used the forebrain A2AR knock-out mice coupled with flow cytometric analyses and intracerebroventricular injection to determine the contribution of A2ARs in forebrain neurons and glial cells to A2AR antagonist-mediated motor and neuroprotective effects.The selecti...

  5. Cannabinoid receptor interacting protein suppresses agonist-driven CB1 receptor internalization and regulates receptor replenishment in an agonist-biased manner. (United States)

    Blume, Lawrence C; Leone-Kabler, Sandra; Luessen, Deborah J; Marrs, Glen S; Lyons, Erica; Bass, Caroline E; Chen, Rong; Selley, Dana E; Howlett, Allyn C


    Cannabinoid receptor interacting protein 1a (CRIP1a) is a CB1 receptor (CB1 R) distal C-terminus-associated protein that modulates CB1 R signaling via G proteins, and CB1 R down-regulation but not desensitization (Blume et al. [2015] Cell Signal., 27, 716-726; Smith et al. [2015] Mol. Pharmacol., 87, 747-765). In this study, we determined the involvement of CRIP1a in CB1 R plasma membrane trafficking. To follow the effects of agonists and antagonists on cell surface CB1 Rs, we utilized the genetically homogeneous cloned neuronal cell line N18TG2, which endogenously expresses both CB1 R and CRIP1a, and exhibits a well-characterized endocannabinoid signaling system. We developed stable CRIP1a-over-expressing and CRIP1a-siRNA-silenced knockdown clones to investigate gene dose effects of CRIP1a on CB1 R plasma membrane expression. Results indicate that CP55940 or WIN55212-2 (10 nM, 5 min) reduced cell surface CB1 R by a dynamin- and clathrin-dependent process, and this was attenuated by CRIP1a over-expression. CP55940-mediated cell surface CB1 R loss was followed by a cycloheximide-sensitive recovery of surface receptors (30-120 min), suggesting the requirement for new protein synthesis. In contrast, WIN55212-2-mediated cell surface CB1 Rs recovered only in CRIP1a knockdown cells. Changes in CRIP1a expression levels did not affect a transient rimonabant (10 nM)-mediated increase in cell surface CB1 Rs, which is postulated to be as a result of rimonabant effects on 'non-agonist-driven' internalization. These studies demonstrate a novel role for CRIP1a in agonist-driven CB1 R cell surface regulation postulated to occur by two mechanisms: 1) attenuating internalization that is agonist-mediated, but not that in the absence of exogenous agonists, and 2) biased agonist-dependent trafficking of de novo synthesized receptor to the cell surface.

  6. Adenosine Receptors as a Biological Pathway for the Anti-Inflammatory and Beneficial Effects of Low Frequency Low Energy Pulsed Electromagnetic Fields

    Directory of Open Access Journals (Sweden)

    Katia Varani


    Full Text Available Several studies explored the biological effects of low frequency low energy pulsed electromagnetic fields (PEMFs on human body reporting different functional changes. Much research activity has focused on the mechanisms of interaction between PEMFs and membrane receptors such as the involvement of adenosine receptors (ARs. In particular, PEMF exposure mediates a significant upregulation of A2A and A3ARs expressed in various cells or tissues involving a reduction in most of the proinflammatory cytokines. Of particular interest is the observation that PEMFs, acting as modulators of adenosine, are able to increase the functionality of the endogenous agonist. By reviewing the scientific literature on joint cells, a double role for PEMFs could be hypothesized in vitro by stimulating cell proliferation, colonization of the scaffold, and production of tissue matrix. Another effect could be obtained in vivo after surgical implantation of the construct by favoring the anabolic activities of the implanted cells and surrounding tissues and protecting the construct from the catabolic effects of the inflammatory status. Moreover, a protective involvement of PEMFs on hypoxia damage in neuron-like cells and an anti-inflammatory effect in microglial cells have suggested the hypothesis of a positive impact of this noninvasive biophysical stimulus.

  7. Adenosine Receptors as a Biological Pathway for the Anti-Inflammatory and Beneficial Effects of Low Frequency Low Energy Pulsed Electromagnetic Fields (United States)

    Vincenzi, Fabrizio; Ravani, Annalisa; Pasquini, Silvia; Merighi, Stefania; Setti, Stefania; Cadossi, Matteo; Cadossi, Ruggero


    Several studies explored the biological effects of low frequency low energy pulsed electromagnetic fields (PEMFs) on human body reporting different functional changes. Much research activity has focused on the mechanisms of interaction between PEMFs and membrane receptors such as the involvement of adenosine receptors (ARs). In particular, PEMF exposure mediates a significant upregulation of A2A and A3ARs expressed in various cells or tissues involving a reduction in most of the proinflammatory cytokines. Of particular interest is the observation that PEMFs, acting as modulators of adenosine, are able to increase the functionality of the endogenous agonist. By reviewing the scientific literature on joint cells, a double role for PEMFs could be hypothesized in vitro by stimulating cell proliferation, colonization of the scaffold, and production of tissue matrix. Another effect could be obtained in vivo after surgical implantation of the construct by favoring the anabolic activities of the implanted cells and surrounding tissues and protecting the construct from the catabolic effects of the inflammatory status. Moreover, a protective involvement of PEMFs on hypoxia damage in neuron-like cells and an anti-inflammatory effect in microglial cells have suggested the hypothesis of a positive impact of this noninvasive biophysical stimulus. PMID:28255202

  8. Scanning mutagenesis in a yeast system delineates the role of the NPxxY(x)(5,6)F motif and helix 8 of the adenosine A(2B) receptor in G protein coupling. (United States)

    Liu, Rongfang; Nahon, Dennis; le Roy, Beau; Lenselink, Eelke B; IJzerman, Adriaan P


    The adenosine receptor subfamily includes four subtypes: the A1, A2A, A2B and A3 receptors, which all belong to the superfamily of G protein-coupled receptors (GPCRs). The adenosine A2B receptor is the least investigated of the adenosine receptors, and the molecular mechanisms of its activation have hardly been explored. We used a single-GPCR-one-G protein yeast screening method in combination with mutagenesis studies, molecular modeling and bio-informatics to investigate the importance of the different amino acid residues of the NPxxY(x)6F motif and helix 8 in the human adenosine A2B receptor (hA2BR) activation. A scanning mutagenesis protocol was employed, yielding 11 single mutations and one double mutation of the NPxxY(x)6F motif and 16 single mutations of helix 8. The amino acid residues P287(7.50), Y290(7.53), R293(7.56) and I304(8.57) were found to be essential, since mutation of these amino acid residues to alanine led to a complete loss of function. Western blot analysis showed that mutant receptor R293(7.56)A was not expressed, whereas the other proteins were. Amino acid residues that are also important in receptor activation are: N286(7.49), V289(7.52), Y292(7.55), N294(8.47), F297(8.50), R298(8.51), H302(8.55) and R307(8.60). The mutation Y290(7.53)F lost 50% of efficacy, while F297(8.50)A behaved similar to wild type receptor. The double mutation, Y290(7.53)F/F297(8.50)Y, lost around 70% of efficacy and displayed a lower potency for the reference agonist 5'-(N-ethylcarboxamido)adenosine (NECA). This study provides new insight into the molecular interplay and impact of TM7 and helix 8 for hA2B receptor activation, which may be extrapolated to other adenosine receptors and possibly to other GPCRs.

  9. Regulation of TrkB receptor translocation to lipid rafts by adenosine A2A receptors and its functional implications for BDNF-induced regulation of synaptic plasticity


    Assaife-Lopes, Natália; Sousa, Vasco C.; Pereira, Daniela B.; Ribeiro, Joaquim A.; Sebastião, Ana M.


    Brain-derived neurotrophic factor (BDNF) signalling is critical for neuronal development and transmission. Recruitment of TrkB receptors to lipid rafts has been shown to be necessary for the activation of specific signalling pathways and modulation of neurotransmitter release by BDNF. Since TrkB receptors are known to be modulated by adenosine A2A receptor activation, we hypothesized that activation of A2A receptors could influence TrkB receptor localization among different membrane microdoma...

  10. Small-molecule agonists for the glucagon-like peptide 1 receptor

    DEFF Research Database (Denmark)

    Knudsen, Lotte Bjerre; Kiel, Dan; Teng, Min;


    described and none in the B family of the G protein coupled receptors to which the GLP-1 receptor belongs. We have discovered a series of small molecules known as ago-allosteric modulators selective for the human GLP-1 receptor. These compounds act as both allosteric activators of the receptor...... and independent agonists. Potency of GLP-1 was not changed by the allosteric agonists, but affinity of GLP-1 for the receptor was increased. The most potent compound identified stimulates glucose-dependent insulin release from normal mouse islets but, importantly, not from GLP-1 receptor knockout mice. Also......The peptide hormone glucagon-like peptide (GLP)-1 has important actions resulting in glucose lowering along with weight loss in patients with type 2 diabetes. As a peptide hormone, GLP-1 has to be administered by injection. Only a few small-molecule agonists to peptide hormone receptors have been...

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


    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.

  12. Stereostructure-activity studies on agonists at the AMPA and kainate subtypes of ionotropic glutamate receptors

    DEFF Research Database (Denmark)

    Johansen, Tommy N; Greenwood, Jeremy R; Frydenvang, Karla Andrea;


    -methyl-4-isoxazolyl)propionic acid (AMPA) receptor subtype of ionotropic Glu receptors in the presence or absence of an agonist has provided important information about ligand-receptor interaction mechanisms. The availability of these binding domain crystal structures has formed the basis for rational...... design of ligands, especially for the AMPA and kainate subtypes of ionotropic Glu receptors. This mini-review will focus on structure-activity relationships on AMPA and kainate receptor agonists with special emphasis on stereochemical and three-dimensional aspects....

  13. Trialkyltin rexinoid-X receptor agonists selectively potentiate thyroid hormone induced programs of xenopus laevis metamorphosis

    NARCIS (Netherlands)

    Mengeling, Brenda J.; Murk, Albertinka J.; Furlow, J.D.


    The trialkyltins tributyltin (TBT) and triphenyltin (TPT) can function as rexinoid-X receptor (RXR) agonists. We recently showed that RXR agonists can alter thyroid hormone (TH) signaling in a mammalian pituitary TH-responsive reporter cell line, GH3.TRE-Luc. The prevalence of TBT and TPT in the

  14. Identification of diarylsulfonamides as agonists of the free fatty acid receptor 4 (FFA4/GPR120). (United States)

    Sparks, Steven M; Chen, Grace; Collins, Jon L; Danger, Dana; Dock, Steven T; Jayawickreme, Channa; Jenkinson, Stephen; Laudeman, Christopher; Leesnitzer, M Anthony; Liang, Xi; Maloney, Patrick; McCoy, David C; Moncol, David; Rash, Vincent; Rimele, Thomas; Vulimiri, Padmaja; Way, James M; Ross, Sean


    The exploration of a diarylsulfonamide series of free fatty acid receptor 4 (FFA4/GPR120) agonists is described. This work led to the identification of selective FFA4 agonist 8 (GSK137647A) and selective FFA4 antagonist 39. The in vitro profile of compounds 8 and 39 is presented herein.

  15. Adenosine receptors located in the NTS contribute to renal sympathoinhibition during hypotensive phase of severe hemorrhage in anesthetized rats. (United States)

    Scislo, Tadeusz J; O'Leary, Donal S


    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.

  16. The importance of the adenosine A(2A) receptor-dopamine D(2) receptor interaction in drug addiction. (United States)

    Filip, M; Zaniewska, M; Frankowska, M; Wydra, K; Fuxe, K


    Drug addiction is a serious brain disorder with somatic, psychological, psychiatric, socio-economic and legal implications in the developed world. Illegal (e.g., psychostimulants, opioids, cannabinoids) and legal (alcohol, nicotine) drugs of abuse create a complex behavioral pattern composed of drug intake, withdrawal, seeking and relapse. One of the hallmarks of drugs that are abused by humans is that they have different mechanisms of action to increase dopamine (DA) neurotransmission within the mesolimbic circuitry of the brain and indirectly activate DA receptors. Among the DA receptors, D(2) receptors are linked to drug abuse and addiction because their function has been proven to be correlated with drug reinforcement and relapses. The recognition that D(2) receptors exist not only as homomers but also can form heteromers, such as with the adenosine (A)(2A) receptor, that are pharmacologically and functionally distinct from their constituent receptors, has significantly expanded the range of potential drug targets and provided new avenues for drug design in the search for novel drug addiction therapies. The aim of this review is to bring current focus on A(2A) receptors, their physiology and pharmacology in the central nervous system, and to discuss the therapeutic relevance of these receptors to drug addiction. We concentrate on the contribution of A(2A) receptors to the effects of different classes of drugs of abuse examined in preclinical behavioral experiments carried out with pharmacological and genetic tools. The consequences of chronic drug treatment on A(2A) receptor-assigned functions in preclinical studies are also presented. Finally, the neurochemical mechanism of the interaction between A(2A) receptors and drugs of abuse in the context of the heteromeric A(2A)-D(2) receptor complex is discussed. Taken together, a significant amount of experimental analyses provide evidence that targeting A(2A) receptors may offer innovative translational strategies

  17. 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)


    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.)

  18. Desensitization of functional µ-opioid receptors increases agonist off-rate. (United States)

    Williams, John T


    Desensitization of µ-opioid receptors (MORs) develops over 5-15 minutes after the application of some, but not all, opioid agonists and lasts for tens of minutes after agonist removal. The decrease in function is receptor selective (homologous) and could result from 1) a reduction in receptor number or 2) a decrease in receptor coupling. The present investigation used photolysis of two caged opioid ligands to examine the kinetics of MOR-induced potassium conductance before and after MOR desensitization. Photolysis of a caged antagonist, carboxynitroveratryl-naloxone (caged naloxone), blocked the current induced by a series of agonists, and the time constant of decline was significantly decreased after desensitization. The increase in the rate of current decay was not observed after partial blockade of receptors with the irreversible antagonist, β-chlornaltrexamine (β-CNA). The time constant of current decay after desensitization was never more rapid than 1 second, suggesting an increased agonist off-rate rather than an increase in the rate of channel closure downstream of the receptor. The rate of G protein-coupled K(+) channel (GIRK) current activation was examined using photolysis of a caged agonist, carboxynitrobenzyl-tyrosine-[Leu(5)]-enkephalin. After acute desensitization or partial irreversible block of MORs with β-CNA, there was an increase in the time it took to reach a peak current. The decrease in the rate of agonist-induced GIRK conductance was receptor selective and dependent on receptor number. The results indicate that opioid receptor desensitization reduced the number of functional receptor and that the remaining active receptors have a reduced agonist affinity.

  19. The antilipolytic agent 3,5-dimethylpyrazole inhibits insulin release in response to both nutrient secretagogues and cyclic adenosine monophosphate agonists in isolated rat islets. (United States)

    Masiello, P; Novelli, M; Bombara, M; Fierabracci, V; Vittorini, S; Prentki, M; Bergamini, E


    This study intended to test the hypothesis that intracellular lipolysis in the pancreatic beta cells is implicated in the regulation of insulin secretion stimulated by nutrient secretagogues or cyclic adenosine monophosphate (cAMP) agonists. Indeed, although lipid signaling molecules were repeatedly reported to influence beta-cell function, the contribution of intracellular triglycerides to the generation of these molecules has remained elusive. Thus, we have studied insulin secretion of isolated rat pancreatic islets in response to various secretagogues in the presence or absence of 3,5-dimethylpyrazole (DMP), a water-soluble and highly effective antilipolytic agent, as previously shown in vivo. In vitro exposure of islets to DMP resulted in an inhibition (by approximately 50%) of the insulin release stimulated not only by high glucose, but also by another nutrient secretagogue, 2-ketoisocaproate, as well as the cAMP agonists 3-isobutyl-1-methylxanthine and glucagon. The inhibitory effect of DMP, which was not due to alteration of islet glucose oxidation, could be reversed upon addition of sn-1,2-dioctanoylglycerol, a synthetic diglyceride, which activates protein kinase C. The results provide direct pharmacologic evidence supporting the concept that endogenous beta-cell lipolysis plays an important role in the generation of lipid signaling molecules involved in the control of insulin secretion in response to both fuel stimuli and cAMP agonists.

  20. Fluorescence characteristics of hydrophobic partial agonist probes of the cholecystokinin receptor. (United States)

    Harikumar, Kaleeckal G; Pinon, Delia I; Miller, Laurence J


    Fluorescence spectroscopic studies are powerful tools for the evaluation of receptor structure and the dynamic changes associated with receptor activation. Here, we have developed two chemically distinct fluorescent probes of the cholecystokinin (CCK) receptor by attaching acrylodan or a nitrobenzoxadiazole moiety to the amino terminus of a partial agonist CCK analogue. These two probes were able to bind to the CCK receptor specifically and with high affinity, and were able to elicit only submaximal intracellular calcium responses typical of partial agonists. The fluorescence characteristics of these probes were compared with those previously reported for structurally-related full agonist and antagonist probes. Like the previous probes, the partial agonist probes exhibited longer fluorescence lifetimes and increased anisotropy when bound to the receptor than when free in solution. The receptor-bound probes were not easily quenched by potassium iodide, suggesting that the fluorophores were protected from the extracellular aqueous milieu. The fluorescence characteristics of the partial agonist probes were quite similar to those of the analogous full agonist probes and quite distinct from the analogous antagonist probes. These data suggest that the partially activated conformational state of this receptor is more closely related to its fully active state than to its inactive state.

  1. Ion Fluxes through KCa2 (SK) and Cav1 (L-type) Channels Contribute to Chronoselectivity of Adenosine A1 Receptor-Mediated Actions in Spontaneously Beating Rat Atria (United States)

    Bragança, Bruno; Oliveira-Monteiro, Nádia; Ferreirinha, Fátima; Lima, Pedro A.; Faria, Miguel; Fontes-Sousa, Ana P.; Correia-de-Sá, Paulo


    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 and 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. PMID:27014060

  2. Adenosine A2 receptor activation ameliorates mitochondrial oxidative stress upon reperfusion through the posttranslational modification of NDUFV2 subunit of complex I in the heart. (United States)

    Xu, Jingman; Bian, Xiyun; Liu, Yuan; Hong, Lan; Teng, Tianming; Sun, Yuemin; Xu, Zhelong


    While it is well known that adenosine receptor activation protects the heart from ischemia/reperfusion injury, the precise mitochondrial mechanism responsible for the action remains unknown. This study probed the mitochondrial events associated with the cardioprotective effect of 5'-(N-ethylcarboxamido) adenosine (NECA), an adenosine A2 receptor agonist. Isolated rat hearts were subjected to 30min ischemia followed by 10min of reperfusion, whereas H9c2 cells experienced 20min ischemia and 10min reperfusion. NECA prevented mitochondrial structural damage, decreases in respiratory control ratio (RCR), and collapse of mitochondrial membrane potential (ΔΨm). Both the adenosine A2A receptor antagonist SCH58261 and A2B receptor antagonist MRS1706 inhibited the action of NECA. NECA reduced mitochondrial proteins carbonylation, H2O2, and superoxide generation at reperfusion, but did not change superoxide dismutase (SOD) activity. In support, the protective effects of NECA and Peg-SOD on ΔΨm upon reperfusion were additive, implying that NECA's protection is attributable to the reduced superoxide generation but not to the enhancement of the superoxide-scavenging capacity. NECA increased the mitochondrial Src tyrosine kinase activity and suppressed complex I activity at reperfusion in a Src-dependent manner. NECA also reduced mitochondrial superoxide through Src tyrosine kinase. Studies with liquid chromatography-mass spectrometer (LC-MS) identified Tyr118 of the NDUFV2 subunit of complex 1 as a likely site of the tyrosine phosphorylation. Furthermore, the complex I activity of cells transfected with the Y118F mutant was increased, suggesting that this site might be a negative regulator of complex I activity. In support, NECA failed to suppress complex I activity at reperfusion in cells transfected with the Y118F mutant of NDUFV2. In conclusion, NECA prevents mitochondrial oxidative stress by decreasing mitochondrial superoxide generation through inhibition of complex I

  3. Combined sodium ion sensitivity in agonist binding and internalization of vasopressin V1b receptors. (United States)

    Koshimizu, Taka-Aki; Kashiwazaki, Aki; Taniguchi, Junichi


    Reducing Na(+) in the extracellular environment may lead to two beneficial effects for increasing agonist binding to cell surface G-protein coupled receptors (GPCRs): reduction of Na(+)-mediated binding block and reduce of receptor internalization. However, such combined effects have not been explored. We used Chinese Hamster Ovary cells expressing vasopressin V1b receptors as a model to explore Na(+) sensitivity in agonist binding and receptor internalization. Under basal conditions, a large fraction of V1b receptors is located intracellularly, and a small fraction is in the plasma membrane. Decreases in external Na(+) increased cell surface [(3)H]AVP binding and decreased receptor internalization. Substitution of Na(+) by Cs(+) or NH4(+) inhibited agonist binding. To suppress receptor internalization, the concentration of NaCl, but not of CsCl, had to be less than 50 mM, due to the high sensitivity of the internalization machinery to Na(+) over Cs(+). Iso-osmotic supplementation of glucose or NH4Cl maintained internalization of the V1b receptor, even in a low-NaCl environment. Moreover, iodide ions, which acted as a counter anion, inhibited V1b agonist binding. In summary, we found external ionic conditions that could increase the presence of high-affinity state receptors at the cell surface with minimum internalization during agonist stimulations.

  4. Differential modulation of Beta-adrenergic receptor signaling by trace amine-associated receptor 1 agonists.

    Directory of Open Access Journals (Sweden)

    Gunnar Kleinau

    Full Text Available Trace amine-associated receptors (TAAR are rhodopsin-like G-protein-coupled receptors (GPCR. TAAR are involved in modulation of neuronal, cardiac and vascular functions and they are potentially linked with neurological disorders like schizophrenia and Parkinson's disease. Subtype TAAR1, the best characterized TAAR so far, is promiscuous for a wide set of ligands and is activated by trace amines tyramine (TYR, phenylethylamine (PEA, octopamine (OA, but also by thyronamines, dopamine, and psycho-active drugs. Unfortunately, effects of trace amines on signaling of the two homologous β-adrenergic receptors 1 (ADRB1 and 2 (ADRB2 have not been clarified yet in detail. We, therefore, tested TAAR1 agonists TYR, PEA and OA regarding their effects on ADRB1/2 signaling by co-stimulation studies. Surprisingly, trace amines TYR and PEA are partial allosteric antagonists at ADRB1/2, whereas OA is a partial orthosteric ADRB2-antagonist and ADRB1-agonist. To specify molecular reasons for TAAR1 ligand promiscuity and for observed differences in signaling effects on particular aminergic receptors we compared TAAR, tyramine (TAR octopamine (OAR, ADRB1/2 and dopamine receptors at the structural level. We found especially for TAAR1 that the remarkable ligand promiscuity is likely based on high amino acid similarity in the ligand-binding region compared with further aminergic receptors. On the other hand few TAAR specific properties in the ligand-binding site might determine differences in ligand-induced effects compared to ADRB1/2. Taken together, this study points to molecular details of TAAR1-ligand promiscuity and identified specific trace amines as allosteric or orthosteric ligands of particular β-adrenergic receptor subtypes.

  5. Intracerebroventricular administration of inosine is anticonvulsant against quinolinic acid-induced seizures in mice: an effect independent of benzodiazepine and adenosine receptors. (United States)

    Ganzella, Marcelo; Faraco, Rafael Berger; Almeida, Roberto Farina; Fernandes, Vinícius Fornari; Souza, Diogo Onofre


    Inosine (INO) has an anticonvulsant effect against seizures induced by antagonists of GABAergic system. Quinolinic acid (QA) is an agonist NMDA receptors implicated in the neurobiology of seizures. In the present study, we investigated the anticonvulsant effect of intracerebroventricular (i.c.v.) INO administration against QA-induced seizures in adult mice. We also investigated whether the benzodiazepines (BZ) or adenosine (ADO) receptors were involved in the INO effects. Animals were pretreated with an i.c.v. injection of either vehicle or INO before an i.c.v. administration of 4 μl QA (36.8 nmol). All animals pretreated with vehicle followed by QA presented seizures. INO protected against QA-induced seizures in a time and dose dependent manner (up to 60% at 400 nmol, 5 min before QA injection). Diazepam (DZ) and ADO (i.c.v.) also exhibited anticonvulsant effect against QA induced seizures. Additionally, i.p. administration of either flumazenil, a BZ receptor antagonist, or caffeine, an ADO receptor antagonist, did not change the anticonvulsant potency of INO i.c.v. injection, but completely abolished the DZ and ADO anticonvulsant effects, respectively. In conclusion, this study demonstrated that INO exert anticonvulsant effect against hyperactivity of the glutamatergic system independently of BZ or ADO receptors activation.

  6. Selected C8 two-chain linkers enhance the adenosine A1/A2A receptor affinity and selectivity of caffeine. (United States)

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


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

  7. Rational design of orally-active, pyrrolidine-based progesterone receptor partial agonists

    Energy Technology Data Exchange (ETDEWEB)

    Thompson, Scott K.; Washburn, David G.; Frazee, James S.; Madauss, Kevin P.; Hoang, Tram H.; Lapinski, Leahann; Grygielko, Eugene T.; Glace, Lindsay E.; Trizna, Walter; Williams, Shawn P.; Duraiswami, Chaya; Bray, Jeffrey D.; Laping, Nicholas J.; (GSKNC); (GSKPA)


    Using the X-ray crystal structure of an amide-based progesterone receptor (PR) partial agonist bound to the PR ligand binding domain, a novel PR partial agonist class containing a pyrrolidine ring was designed. Members of this class of N-alkylpyrrolidines demonstrate potent and highly selective partial agonism of the progesterone receptor, and one of these analogs was shown to be efficacious upon oral dosing in the OVX rat model of estrogen opposition.

  8. In vitro and in vivo efficacy of a potent opioid receptor agonist, biphalin, compared to subtype-selective opioid receptor agonists for stroke treatment. (United States)

    Yang, Li; Islam, Mohammad R; Karamyan, Vardan T; Abbruscato, Thomas J


    To meet the challenge of identification of new treatments for stroke, this study was designed to evaluate a potent, nonselective opioid receptor (OR) agonist, biphalin, in comparison to subtype selective OR agonists, as a potential neuroprotective drug candidate using in vitro and in vivo models of ischemic stroke. Our in vitro approach included mouse primary neuronal cells that were challenged with glutamate and hypoxic/aglycemic (H/A) conditions. We observed that 10nM biphalin, exerted a statistically significant neuroprotective effect after glutamate challenge, compared to all selective opioid agonists, according to lactate dehydrogenase (LDH) and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assays. Moreover, 10nM biphalin provided superior neuroprotection after H/A-reoxygenation compared to selective opioid agonists in all cases. Our in vitro investigations were supported by in vivo studies which indicate that the nonselective opioid agonist, biphalin, achieves enhanced neuroprotective potency compared to any of the selective opioid agonists, evidenced by reduced edema and infarct ratios. Reduction of edema and infarction was accompanied by neurological improvement of the animals in two independent behavioral tests. Collectively these data strongly suggest that concurrent agonist stimulation of mu, kappa and delta ORs with biphalin is neuroprotective and superior to neuroprotection by activation of any single OR subtype.

  9. Modulatory effects by CB1 receptors on rat spinal locomotor networks after sustained application of agonists or antagonists. (United States)

    Veeraraghavan, P; Nistri, A


    Sustained administration of cannabinoid agonists acting on neuronal CB1 receptors (CB1Rs) are proposed for treating spasticity and chronic pain. The impact of CB1Rs on mammalian locomotor networks remains, however, incompletely understood. To clarify how CB1Rs may control synaptic activity and locomotor network function, we used the rat spinal cord in vitro which is an advantageous model to investigate locomotor circuit mechanisms produced by the local central pattern generator. Neither the CB1 agonist anandamide (AEA) nor the CB1R antagonist AM-251 evoked early (3-24h largely impaired locomotor network activity induced by DR stimuli or neurochemicals, and depressed disinhibited bursting without changing reflex amplitude or inducing neurotoxicity even if CB1R immunoreactivity was lowered in the central region. Since CB1R activation usually inhibits cyclic adenosine monophosphate (cAMP) synthesis, we investigated how a 24-h application of AEA or AM-251 affected basal or forskolin-stimulated cAMP levels. While AEA decreased them in an AM-251-sensitive manner, AM-251 per se did not change resting or stimulated cAMP. Our data suggest that CB1Rs may control the circuit gateway regulating the inflow of sensory afferent inputs into the locomotor circuits, indicating a potential site of action for restricting peripheral signals disruptive for locomotor activity.

  10. 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.

  11. Development of selective agonists and antagonists of P2Y receptors



    Although elucidation of the medicinal chemistry of agonists and antagonists of the P2Y receptors has lagged behind that of many other members of group A G protein-coupled receptors, detailed qualitative and quantitative structure–activity relationships (SARs) were recently constructed for several of the subtypes. Agonists selective for P2Y1, P2Y2, and P2Y6 receptors and nucleotide antagonists selective for P2Y1 and P2Y12 receptors are now known. Selective nonnucleotide antagonists were report...

  12. A molecular characterization of the agonist binding site of a nematode cys-loop GABA receptor (United States)

    Kaji, Mark D; Kwaka, Ariel; Callanan, Micah K; Nusrat, Humza; Desaulniers, Jean-Paul; Forrester, Sean G


    Background and Purpose Cys-loop GABA receptors represent important targets for human chemotherapeutics and insecticides and are potential targets for novel anthelmintics (nematicides). However, compared with insect and mammalian receptors, little is known regarding the pharmacological characteristics of nematode Cys-loop GABA receptors. Here we have investigated the agonist binding site of the Cys-loop GABA receptor UNC-49 (Hco-UNC-49) from the parasitic nematode Haemonchus contortus. Experimental Approach We used two-electrode voltage-clamp electrophysiology to measure channel activation by classical GABA receptor agonists on Hco-UNC-49 expressed in Xenopus laevis oocytes, along with site-directed mutagenesis and in silico homology modelling. Key Results The sulphonated molecules P4S and taurine had no effect on Hco-UNC-49. Other classical Cys-loop GABAA receptor agonists tested on the Hco-UNC-49B/C heteromeric channel had a rank order efficacy of GABA > trans-4-aminocrotonic acid > isoguvacine > imidazole-4-acetic acid (IMA) > (R)-(−)-4-amino-3-hydroxybutyric acid [R(−)-GABOB] > (S)-(+)-4-amino-3-hydroxybutyric acid [S(+)-GABOB] > guanidinoacetic acid > isonipecotic acid > 5-aminovaleric acid (DAVA) (partial agonist) > β-alanine (partial agonist). In silico ligand docking revealed some variation in binding between agonists. Mutagenesis of a key serine residue in binding loop C to threonine had minimal effects on GABA and IMA but significantly increased the maximal response to DAVA and decreased twofold the EC50 for R(−)- and S(+)-GABOB. Conclusions and Implications The pharmacological profile of Hco-UNC-49 differed from that of vertebrate Cys-loop GABA receptors and insect resistance to dieldrin receptors, suggesting differences in the agonist binding pocket. These findings could be exploited to develop new drugs that specifically target GABA receptors of parasitic nematodes. PMID:25850584

  13. In vivo delta opioid receptor internalization controls behavioral effects of agonists.

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    Amynah A A Pradhan

    Full Text Available BACKGROUND: GPCRs regulate a remarkable diversity of biological functions, and are thus often targeted for drug therapies. Stimulation of a GPCR by an extracellular ligand triggers receptor signaling via G proteins, and this process is highly regulated. Receptor activation is typically accompanied by desensitization of receptor signaling, a complex feedback regulatory process of which receptor internalization is postulated as a key event. The in vivo significance of GPCR internalization is poorly understood. In fact, the majority of studies have been performed in transfected cell systems, which do not adequately model physiological environments and the complexity of integrated responses observed in the whole animal. METHODS AND FINDINGS: In this study, we used knock-in mice expressing functional fluorescent delta opioid receptors (DOR-eGFP in place of the native receptor to correlate receptor localization in neurons with behavioral responses. We analyzed the pain-relieving effects of two delta receptor agonists with similar signaling potencies and efficacies, but distinct internalizing properties. An initial treatment with the high (SNC80 or low (AR-M100390 internalizing agonist equally reduced CFA-induced inflammatory pain. However, subsequent drug treatment produced highly distinct responses. Animals initially treated with SNC80 showed no analgesic response to a second dose of either delta receptor agonist. Concomitant receptor internalization and G-protein uncoupling were observed throughout the nervous system. This loss of function was temporary, since full DOR-eGFP receptor responses were restored 24 hours after SNC80 administration. In contrast, treatment with AR-M100390 resulted in retained analgesic response to a subsequent agonist injection, and ex vivo analysis showed that DOR-eGFP receptor remained G protein-coupled on the cell surface. Finally SNC80 but not AR-M100390 produced DOR-eGFP phosphorylation, suggesting that the two

  14. Antinociception by systemically-administered acetaminophen (paracetamol) involves spinal serotonin 5-HT7 and adenosine A1 receptors, as well as peripheral adenosine A1 receptors. (United States)

    Liu, Jean; Reid, Allison R; Sawynok, Jana


    Acetaminophen (paracetamol) is a widely used analgesic, but its sites and mechanisms of action remain incompletely understood. Recent studies have separately implicated spinal adenosine A(1) receptors (A(1)Rs) and serotonin 5-HT(7) receptors (5-HT(7)Rs) in the antinociceptive effects of systemically administered acetaminophen. In the present study, we determined whether these two actions are linked by delivering a selective 5-HT(7)R antagonist to the spinal cord of mice and examining nociception using the formalin 2% model. In normal and A(1)R wild type mice, antinociception by systemic (i.p.) acetaminophen 300mg/kg was reduced by intrathecal (i.t.) delivery of the selective 5-HT(7)R antagonist SB269970 3μg. In mice lacking A(1)Rs, i.t. SB269970 did not reverse antinociception by systemic acetaminophen, indicating a link between spinal 5-HT(7)R and A(1)R mechanisms. We also explored potential roles of peripheral A(1)Rs in antinociception by acetaminophen administered both locally and systemically. In normal mice, intraplantar ( acetaminophen 200μg produced antinociception in the formalin test, and this was blocked by co-administration of the selective A(1)R antagonist DPCPX 4.5μg. Acetaminophen administered into the contralateral hindpaw had no effect, indicating a local peripheral action. When acetaminophen was administered systemically, its antinociceptive effect was reversed by DPCPX in normal mice; this was also observed in A(1)R wild type mice, but not in those lacking A(1)Rs. In summary, we demonstrate a link between spinal 5-HT(7)Rs and A(1)Rs in the spinal cord relevant to antinociception by systemic acetaminophen. Furthermore, we implicate peripheral A(1)Rs in the antinociceptive effects of locally- and systemically-administered acetaminophen.

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

    Directory of Open Access Journals (Sweden)

    Joana E Coelho


    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.

  16. Repetitive systemic morphine alters activity-dependent plasticity of Schaffer-collateral-CA1 pyramidal cell synapses: involvement of adenosine A1 receptors and adenosine deaminase. (United States)

    Sadegh, Mehdi; Fathollahi, Yaghoub


    The effectiveness of O-pulse stimulation (TPS) for the reversal of O-pattern primed bursts (PB)-induced long-term potentiation (LTP) were examined at the Schaffer-collateral-CA1 pyramidal cell synapses of hippocampal slices derived from rats chronically treated with morphine (M-T). The results showed that slices derived from both control and M-T rats had normal field excitatory postsynaptic potential (fEPSP)-LTP, whereas PS-LTP in slices from M-T rats was significantly greater than that from control slices. When morphine was applied in vitro to slices derived from rats chronically treated with morphine, the augmentation of PS-LTP was not seen. TPS given 30 min after LTP induction failed to reverse the fEPSP- or PS-LTP in both groups of slices. However, TPS delivered in the presence of long-term in vitro morphine caused the PS-LTP reversal. This effect was blocked by the adenosine A1 receptor (A1R) antagonist CPX (200 nM) and furthermore was enhanced by the adenosine deaminase (ADA) inhibitor EHNA (10 μM). Interestingly, TPS given 30 min after LTP induction in the presence of EHNA (10 μM) can reverse LTP in morphine-exposed control slices in vitro. These results suggest adaptive changes in the hippocampus area CA1 in particular in adenosine system following repetitive systemic morphine. Chronic in vivo morphine increases A1R and reduces ADA activity in the hippocampus. Consequently, adenosine can accumulate because of a stimulus train-induced activity pattern in CA1 area and takes the opportunity to work as an inhibitory neuromodulator and also to enable CA1 to cope with chronic morphine. In addition, adaptive mechanisms are differentially working in the dendrite layer rather than the somatic layer of hippocampal CA1.

  17. Adenosine receptors: regulatory players in the preservation of mitochondrial function induced by ischemic preconditioning of rat liver. (United States)

    Duarte, Filipe V; Amorim, João A; Varela, Ana T; Teodoro, João S; Gomes, Ana P; Cunha, Rodrigo A; Palmeira, Carlos M; Rolo, Anabela P


    Although adenosine A1 receptors (A1R) have been associated to ischemic preconditioning (IPC), direct evidence for their ability to preserve mitochondrial function upon hepatic preconditioning is still missing and could represent a novel strategy to boost the quality of liver transplants. We tested if the A1R antagonist 8-cyclopentyl-1,3-dipropylxanthine (DPCPX) prevented IPC in the liver and if the A1R agonist 2-chloro-N(6)-cyclopentyladenosine (CCPA) might afford a pharmacological preconditioning. Livers underwent a 120 min of 70% warm ischemia and 16 h of reperfusion (I/R), and the IPC group underwent a 5-min ischemic episode followed by a 10-min period of reperfusion before I/R. DPCPX or CCPA was administered intraperitoneally 2 h before IPC or I/R. The control of mitochondrial function emerged as the central element affected by IPC and controlled by endogenous A1R activation. Thus, livers from IPC- or CCPA-treated rats displayed an improved oxidative phosphorylation with higher state 3 respiratory rate, higher respiratory control ratio, increased ATP content, and decreased lag phase. IPC and CCPA also prevented the I/R-induced susceptibility to calcium-induced mitochondrial permeability transition, the rate of reactive oxygen species (ROS) generation, and the decreased mitochondrial content of phospho-Ser(9) GSK-3β. DPCPX abrogated these effects of IPC. These implicate the control of GSK-3β activity by Akt-mediated Ser(9)-GSK-3β phosphorylation preserving the efficiency of oxidative phosphorylation and ROS-mediated cell death in the ability of A1R activation to mimic IPC in the liver. In conclusion, the parallel between IPC and A1R-mediated preconditioning also paves the way to consider a putative therapeutic use of the later in liver transplants.

  18. 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


    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.

  19. AMP-guided tumour-specific nanoparticle delivery via adenosine A1 receptor. (United States)

    Dai, Tongcheng; Li, Na; Han, Fajun; Zhang, Hua; Zhang, Yuanxing; Liu, Qin


    Active targeting-ligands have been increasingly used to functionalize nanoparticles for tumour-specific clinical applications. Here we utilize nucleotide adenosine 5'-monophosphate (AMP) as a novel ligand to functionalize polymer-based fluorescent nanoparticles (NPs) for tumour-targeted imaging. We demonstrate that AMP-conjugated NPs (NPs-AMP) efficiently bind to and are following internalized into colon cancer cell CW-2 and breast cancer cell MDA-MB-468 in vitro. RNA interference and inhibitor assays reveal that the targeting effects mainly rely on the specific binding of AMP to adenosine A1 receptor (A1R), which is greatly up-regulated in cancer cells than in matched normal cells. More importantly, NPs-AMP specifically accumulate in the tumour site of colon and breast tumour xenografts and are further internalized into the tumour cells in vivo via tail vein injection, confirming that the high in vitro specificity of AMP can be successfully translated into the in vivo efficacy. Furthermore, NPs-AMP exhibit an active tumour-targeting behaviour in various colon and breast cancer cells, which is positively related to the up-regulation level of A1R in cancer cells, suggesting that AMP potentially suits for more extensive A1R-overexpressing cancer models. This work establishes AMP to be a novel tumour-targeting ligand and provides a promising strategy for future diagnostic or therapeutic applications.

  20. Chronic sleep restriction induces long-lasting changes in adenosine and noradrenaline receptor density in the rat brain (United States)



    SUMMARY Although chronic sleep restriction frequently produces long-lasting behavioural and physiological impairments in humans, the underlying neural mechanisms are unknown. Here we used a rat model of chronic sleep restriction to investigate the role of brain adenosine and noradrenaline systems, known to regulate sleep and wakefulness, respectively. The density of adenosine A1 and A2a receptors and β-adrenergic receptors before, during and following 5 days of sleep restriction was assessed with autoradiography. Rats (n = 48) were sleep-deprived for 18 h day–1 for 5 consecutive days (SR1–SR5), followed by 3 unrestricted recovery sleep days (R1–R3). Brains were collected at the beginning of the light period, which was immediately after the end of sleep deprivation on sleep restriction days. Chronic sleep restriction increased adenosine A1 receptor density significantly in nine of the 13 brain areas analysed with elevations also observed on R3 (+18 to +32%). In contrast, chronic sleep restriction reduced adenosine A2a receptor density significantly in one of the three brain areas analysed (olfactory tubercle which declined 26–31% from SR1 to R1). A decrease in b-adrenergic receptors density was seen in substantia innominata and ventral pallidum which remained reduced on R3, but no changes were found in the anterior cingulate cortex. These data suggest that chronic sleep restriction can induce long-term changes in the brain adenosine and noradrenaline receptors, which may underlie the long-lasting neurocognitive impairments observed in chronic sleep restriction. PMID:25900125

  1. The stimulatory adenosine receptor ADORA2B regulates serotonin (5-HT synthesis and release in oxygen-depleted EC cells in inflammatory bowel disease.

    Directory of Open Access Journals (Sweden)

    Rikard Dammen

    Full Text Available OBJECTIVE: We recently demonstrated that hypoxia, a key feature of IBD, increases enterochromaffin (EC cell 5-HT secretion, which is also physiologically regulated by the ADORA2B mechanoreceptor. Since hypoxia is associated with increased extracellular adenosine, we wanted to examine whether this nucleotide amplifies HIF-1α-mediated 5-HT secretion. DESIGN: The effects of hypoxia were studied on IBD mucosa, isolated IBD-EC cells, isolated normal EC cells and the EC cell tumor derived cell line KRJ-1. Hypoxia (0.5% O2 was compared to NECA (adenosine agonist, MRS1754 (ADORA2B receptor antagonist and SCH442146 (ADORA2A antagonist on HIF signaling and 5-HT secretion. Antisense approaches were used to mechanistically evaluate EC cells in vitro. PCR and western blot were used to analyze transcript and protein levels of HIF-1α signaling and neuroendocrine cell function. An animal model of colitis was evaluated to confirm hypoxia:adenosine signaling in vivo. RESULTS: HIF-1α is upregulated in IBD mucosa and IBD-EC cells, the majority (~90% of which express an activated phenotype in situ. Hypoxia stimulated 5-HT release maximally at 30 mins, an effect amplified by NECA and selectively inhibited by MRS1754, through phosphorylation of TPH-1 and activation of VMAT-1. Transient transfection with Renilla luciferase under hypoxia transcriptional response element (HRE control identified that ADORA2B activated HIF-1α signaling under hypoxic conditions. Additional signaling pathways associated with hypoxia:adenosine included MAP kinase and CREB. Antisense approaches mechanistically confirmed that ADORA2B signaling was linked to these pathways and 5-HT release under hypoxic conditions. Hypoxia:adenosine activation which could be reversed by 5'-ASA treatment was confirmed in a TNBS-model. CONCLUSION: Hypoxia induced 5-HT synthesis and secretion is amplified by ADORA2B signaling via MAPK/CREB and TPH-1 activation. Targeting ADORA2s may decrease EC cell 5-HT

  2. GABAB receptor-positive modulators: enhancement of GABAB receptor agonist effects in vivo. (United States)

    Koek, Wouter; France, Charles P; Cheng, Kejun; Rice, Kenner C


    In vivo effects of GABA(B) receptor-positive modulators suggest that they have therapeutic potential for treating central nervous system disorders such as anxiety, depression, and drug abuse. Although these effects generally are thought to be mediated by positive modulation of GABA(B) receptors, such modulation has been examined primarily in vitro. The present study was aimed at further examining the in vivo positive modulatory properties of the GABA(B) receptor-positive modulators, 2,6-di-tert-butyl-4-(3-hydroxy-2,2-dimethylpropyl) phenol (CGP7930) and (R,S)-5,7-di-tert-butyl-3-hydroxy-3-trifluoromethyl-3H-benzofuran-2-one (rac-BHFF). Both compounds enhanced loss of righting induced by baclofen in mice. However, CGP7930 was less effective and rac-BHFF was less potent for enhancing loss of righting induced by γ-hydroxybutyrate (GHB), which, like baclofen, has GABA(B) receptor agonist properties. In contrast with baclofen- and GHB-induced loss of righting, the hypothermic effects of baclofen and GHB were not enhanced by rac-BHFF but were enhanced by CGP7930 only at doses that produced hypothermia when given alone. CGP7930-induced hypothermia was not attenuated by the GABA(B) receptor antagonist 3-aminopropyl(diethoxymethyl)phosphinic acid (CGP35348), at doses that blocked baclofen-induced hypothermia, and was not increased by the nitric-oxide synthase inhibitor N(ω)-nitro-L-arginine methyl ester, at doses that increased the hypothermic effects of baclofen and GHB. The results provide evidence that CGP7930 and rac-BHFF act in vivo as positive modulators at GABA(B) receptors mediating loss of righting, but not at GABA(B) receptors mediating hypothermia. Conceivably, CGP7930, but not rac-BHFF, acts as an allosteric agonist at these latter receptors. Taken together, the results provide further evidence of pharmacologically distinct GABA(B) receptor subtypes, possibly allowing for a more selective therapeutic interference with the GABA(B) system.

  3. 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


    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.

  4. Synthesis and pharmacological characterization of novel xanthine carboxylate amides as A2A adenosine receptor ligands exhibiting bronchospasmolytic activity. (United States)

    Yadav, Rakesh; Bansal, Ranju; Rohilla, Suman; Kachler, Sonja; Klotz, Karl-Norbert


    The carboxylate amides of 8-phenyl-1,3-dimethylxanthine described herein represent a new series of selective ligands of the adenosine A2A receptors exhibiting bronchospasmolytic activity. The effects of location of 8-phenyl substitutions on the adenosine receptor (AR) binding affinities of the newly synthesized xanthines have also been studied. The compounds displayed moderate to potent binding affinities toward various adenosine receptor subtypes when evaluated through radioligand binding studies. However, most of the compounds showed the maximum affinity for the A2A subtype, some with high selectivity versus all other subtypes. Xanthine carboxylate amide 13b with a diethylaminoethylamino moiety at the para-position of the 8-phenylxanthine scaffold was identified as the most potent A2A adenosine receptor ligand with Ki=0.06μM. Similarly potent and highly A2A-selective are the isovanillin derivatives 16a and 16d. In addition, the newly synthesized xanthine derivatives showed good in vivo bronchospasmolytic activity when tested in guinea pigs.

  5. Early Cessation of Adenosine Diphosphate Receptor Inhibitors Among Acute Myocardial Infarction Patients Treated With Percutaneous Coronary Intervention

    DEFF Research Database (Denmark)

    Fosbøl, Emil L; Ju, Christine; Anstrom, Kevin J


    BACKGROUND: Guidelines recommend the use of adenosine diphosphate receptor inhibitor (ADPri) therapy for 1 year postacute myocardial infarction; yet, early cessation of therapy occurs frequently in clinical practice. METHODS AND RESULTS: We examined 11 858 acute myocardial infarction patients tre...... adverse cardiovascular event risk. CLINICAL TRIAL REGISTRATION: URL: Unique identifier: NCT01088503....

  6. Effects of IL-4 and IL-13 on adenosine receptor expression and responsiveness of the human mast cell line 1

    NARCIS (Netherlands)

    Versluis, Mieke; Postma, Dirkje S.; Timens, Wim; Hylkema, Machteld N.


    Background: Inhalation of adenosine-5'-monophosphate (AMP) causes bronchoconstriction in asthma but not in healthy subjects. Bronchoconstriction upon AMP inhalation is thought to occur by histamine release and subsequent binding to receptors on airway smooth muscle cells. Methods: To explain enhance

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

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


    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.

  8. Dissociated sterol-based liver X receptor agonists as therapeutics for chronic inflammatory diseases. (United States)

    Yu, Shan; Li, Sijia; Henke, Adam; Muse, Evan D; Cheng, Bo; Welzel, Gustav; Chatterjee, Arnab K; Wang, Danling; Roland, Jason; Glass, Christopher K; Tremblay, Matthew


    Liver X receptor (LXR), a nuclear hormone receptor, is an essential regulator of immune responses. Activation of LXR-mediated transcription by synthetic agonists, such as T0901317 and GW3965, attenuates progression of inflammatory disease in animal models. However, the adverse effects of these conventional LXR agonists in elevating liver lipids have impeded exploitation of this intriguing mechanism for chronic therapy. Here, we explore the ability of a series of sterol-based LXR agonists to alleviate inflammatory conditions in mice without hepatotoxicity. We show that oral treatment with sterol-based LXR agonists in mice significantly reduces dextran sulfate sodium colitis-induced body weight loss, which is accompanied by reduced expression of inflammatory markers in the large intestine. The anti-inflammatory property of these agonists is recapitulated in vitro in mouse lamina propria mononuclear cells, human colonic epithelial cells, and human peripheral blood mononuclear cells. In addition, treatment with LXR agonists dramatically suppresses inflammatory cytokine expression in a model of traumatic brain injury. Importantly, in both disease models, the sterol-based agonists do not affect the liver, and the conventional agonist T0901317 results in significant liver lipid accumulation and injury. Overall, these results provide evidence for the development of sterol-based LXR agonists as novel therapeutics for chronic inflammatory diseases.-Yu, S., Li, S., Henke, A., Muse, E. D., Cheng, B., Welzel, G., Chatterjee, A. K., Wang, D., Roland, J., Glass, C. K., Tremblay, M. Dissociated sterol-based liver X receptor agonists as therapeutics for chronic inflammatory diseases.

  9. Selective Estrogen Receptor Modulators: Cannabinoid Receptor Inverse Agonists with Differential CB1 and CB2 Selectivity (United States)

    Franks, Lirit N.; Ford, Benjamin M.; Prather, Paul L.


    Selective estrogen receptor modulators (SERMs) are used to treat estrogen receptor (ER)-positive breast cancer and osteoporosis. Interestingly, tamoxifen and newer classes of SERMs also exhibit cytotoxic effects in cancers devoid of ERs, indicating a non-estrogenic mechanism of action. Indicative of a potential ER-independent target, reports demonstrate that tamoxifen binds to cannabinoid receptors (CBRs) with affinity in the low μM range and acts as an inverse agonist. To identify cannabinoids with improved pharmacological properties relative to tamoxifen, and further investigate the use of different SERM scaffolds for future cannabinoid drug development, this study characterized the affinity and activity of SERMs in newer structural classes at CBRs. Fourteen SERMs from five structurally distinct classes were screened for binding to human CBRs. Compounds from four of five SERM classes examined bound to CBRs. Subsequent studies fully characterized CBR affinity and activity of one compound from each class. Ospemifine (a triphenylethylene) selectively bound to CB1Rs, while bazedoxifine (an indole) bound to CB2Rs with highest affinity. Nafoxidine (a tetrahydronaphthalene) and raloxifene (RAL; a benzothiaphene) bound to CB1 and CB2Rs non-selectively. All four compounds acted as inverse agonists at CB1 and CB2Rs, reducing basal G-protein activity with IC50 values in the nM to low μM range. Ospemifine, bazedoxifene and RAL also acted as inverse agonists to elevate basal intracellular cAMP levels in intact CHO-hCB2 cells. The four SERMs examined also acted as CB1 and CB2R antagonists in the cAMP assay, producing rightward shifts in the concentration-effect curve of the CBR agonist CP-55,940. In conclusion, newer classes of SERMs exhibit improved pharmacological characteristics (e.g., in CBR affinity and selectivity) relative to initial studies with tamoxifen, and thus suggest that different SERM scaffolds may be useful for development of safe and selective drugs acting

  10. Ontogeny of catecholamine and adenosine receptor-mediated cAMP signaling of embryonic red blood cells: role of cGMP-inhibited phosphodiesterase 3 and hemoglobin. (United States)

    Baumann, R; Blass, C; Götz, R; Dragon, S


    We have previously shown that the cAMP signaling pathway controls major aspects of embryonic red blood cell (RBC) function in avian embryos (Glombitza et al, Am J Physiol 271:R973, 1996; and Dragon et al, Am J Physiol 271:R982, 1996) that are important for adaptation of the RBC gas transport properties to the progressive hypercapnia and hypoxia of later stages of avian embryonic development. Data about the ontogeny of receptor-mediated cAMP signaling are lacking. We have analyzed the response of primitive and definitive chick embryo RBC harvested from day 3 to 18 of development towards forskolin, beta-adrenergic, and A2 receptor agonists. The results show a strong response of immature definitive and primitive RBC to adenosine A2 and beta-adrenergic receptor agonists, which is drastically reduced in the last stage of development, coincident with the appearance of mature, transcriptionally inactive RBC. Modulation of cGMP-inhibited phosphodiesterase 3 (PDE3) has a controlling influence on cAMP accumulation in definitive RBC. Under physiological conditions, PDE3 is inhibited due to activation of soluble guanylyl cyclase (sGC). Inhibition of sGC with the specific inhibitor ODQ decreases receptor-mediated stimulation of cAMP production; this effect is reversed by the PDE3 inhibitor milrinone. sGC is acitivated by nitric oxide (NO), but we found no evidence for production of NO by erythrocyte NO-synthase. However, embryonic hemoglobin releases NO in an oxygen-linked manner that may activate guanylyl cyclase.

  11. Attenuated renovascular constrictor responses to angiotensin II in adenosine 1 receptor knockout mice

    DEFF Research Database (Denmark)

    Hansen, Pernille B; Hashimoto, Seiji; Briggs, Josie


    In the present experiments we examined the renovascular constrictor effects of ANG II in the chronic and complete absence of A1 adenosine receptors (A1AR) using mice with targeted deletion of the A1AR gene. Glomerular filtration rate (GFR) was not different between A1AR +/+ and A1AR -/- mice under...... and increased renal vascular resistance significantly more in A1AR +/+ than in A1AR -/- mice. Perfused afferent arterioles isolated from A1AR +/+ mice constricted in response to bath ANG II with an EC50 of 1.5 +/- 0.4 x 10(-10) mol/l, whereas a right shift in the dose-response relationship with an EC50 of 7.......3 +/- 1.2 x 10(-10) mol/l (P resistance...

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

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


    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.

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


    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.

  14. Glutamate-induced depression of EPSP-spike coupling in rat hippocampal CA1 neurons and modulation by adenosine receptors. (United States)

    Ferguson, Alexandra L; Stone, Trevor W


    The presence of high concentrations of glutamate in the extracellular fluid following brain trauma or ischaemia may contribute substantially to subsequent impairments of neuronal function. In this study, glutamate was applied to hippocampal slices for several minutes, producing over-depolarization, which was reflected in an initial loss of evoked population potential size in the CA1 region. Orthodromic population spikes recovered only partially over the following 60 min, whereas antidromic spikes and excitatory postsynaptic potentials (EPSPs) showed greater recovery, implying a change in EPSP-spike coupling (E-S coupling), which was confirmed by intracellular recording from CA1 pyramidal cells. The recovery of EPSPs was enhanced further by dizocilpine, suggesting that the long-lasting glutamate-induced change in E-S coupling involves NMDA receptors. This was supported by experiments showing that when isolated NMDA-receptor-mediated EPSPs were studied in isolation, there was only partial recovery following glutamate, unlike the composite EPSPs. The recovery of orthodromic population spikes and NMDA-receptor-mediated EPSPs following glutamate was enhanced by the adenosine A1 receptor blocker DPCPX, the A2A receptor antagonist SCH58261 or adenosine deaminase, associated with a loss of restoration to normal of the glutamate-induced E-S depression. The results indicate that the long-lasting depression of neuronal excitability following recovery from glutamate is associated with a depression of E-S coupling. This effect is partly dependent on activation of NMDA receptors, which modify adenosine release or the sensitivity of adenosine receptors. The results may have implications for the use of A1 and A2A receptor ligands as cognitive enhancers or neuroprotectants.

  15. In vivo and in vitro evaluation of novel μ-opioid receptor agonist compounds. (United States)

    Nikaido, Yoshiaki; Kurosawa, Aya; Saikawa, Hitomi; Kuroiwa, Satoshi; Suzuki, Chiharu; Kuwabara, Nobuo; Hoshino, Hazime; Obata, Hideaki; Saito, Shigeru; Saito, Tamio; Osada, Hiroyuki; Kobayashi, Isao; Sezutsu, Hideki; Takeda, Shigeki


    Opioids are the most effective and widely used drugs for pain treatment. Morphine is an archetypal opioid and is an opioid receptor agonist. Unfortunately, the clinical usefulness of morphine is limited by adverse effects such as analgesic tolerance and addiction. Therefore, it is important to study the development of novel opioid agonists as part of pain control. The analgesic effects of opioids are mediated by three opioid receptors, namely opioid μ-, δ-, and κ-receptors. They belong to the G protein-coupled receptor superfamily and are coupled to Gi proteins. In the present study, we developed a ligand screening system to identify novel opioid μ-receptor agonists that measures [(35)S]GTPγS binding to cell membrane fractions prepared from the fat body of transgenic silkworms expressing μ-receptor-Gi1α fusion protein. We screened the RIKEN Natural Products Depository (NPDepo) chemical library, which contains 5848 compounds, and analogs of hit compounds. We successfully identified a novel, structurally unique compound, that we named GUM1, with agonist activity for the opioid μ-receptor (EC50 of 1.2 µM). The Plantar Test (Hargreaves' Method) demonstrated that subcutaneous injection of 3mg/kg of GUM1 into wild-type rats significantly extended latency time. This extension was also observed in a rat model of morphine tolerance and was inhibited by pre-treatment of naloxone. The unique molecular skeleton of GUM1 makes it an attractive molecule for further ligand-opioid receptor binding studies.

  16. Optimisation of in silico derived 2-aminobenzimidazole hits as unprecedented selective kappa opioid receptor agonists

    DEFF Research Database (Denmark)

    Sasmal, Pradip K; Krishna, C Vamsee; Sudheerkumar Adabala, S


    Kappa opioid receptor (KOR) is an important mediator of pain signaling and it is targeted for the treatment of various pains. Pharmacophore based mining of databases led to the identification of 2-aminobenzimidazole derivative as KOR agonists with selectivity over the other opioid receptors DOR...

  17. Tweaking agonist efficacy at N-methyl-D-aspartate receptors by site-directed mutagenesis

    DEFF Research Database (Denmark)

    Hansen, Kasper B; Clausen, Rasmus P; Bjerrum, Esben J


    The structural basis for partial agonism at N-methyl-D-aspartate (NMDA) receptors is currently unresolved. We have characterized several partial agonists at the NR1/NR2B receptor and investigated the mechanisms underlying their reduced efficacy by introducing mutations in the glutamate binding si...

  18. Unraveling the high- and low-sensitivity agonist responses of nicotinic acetylcholine receptors

    DEFF Research Database (Denmark)

    Harpsøe, Kasper; Ahring, Philip K; Christensen, Jeppe K;


    interfaces, the (a4)(3)(ß2)(2) receptor contains a third low-sensitivity agonist binding site in the a4a4 interface. Occupation of this site is required for full activation and is responsible for the widened dynamic response range of this receptor subtype. By site-directed mutagenesis, we show that three...... residues, which differ between the a4ß2 and a4a4 sites, control agonist sensitivity. The results presented here provide a basic insight into the function of pentameric ligand-gated ion channels, which enables modulation of the receptors with hitherto unseen precision; it becomes possible to rationally...

  19. The discovery of diazepinone-based 5-HT3 receptor partial agonists. (United States)

    Manning, David D; Guo, Cheng; Zhang, Zhenjun; Ryan, Kristen N; Naginskaya, Jennifer; Choo, Sok Hui; Masih, Liaqat; Earley, William G; Wierschke, Jonathan D; Newman, Amy S; Brady, Catherine A; Barnes, Nicholas M; Guzzo, Peter R


    Serotonin type 3 (5-HT3) receptor partial agonists have been targeted as potential new drugs for the symptomatic relief of irritable bowel syndrome (IBS). Multiple diazepinone-based compounds have been discovered, which exhibit nanomolar binding affinity for the h5-HT3A receptor and display a range of intrinsic activities (IA=7-87% of 5-HT Emax) in HEK cells heterologously expressing the h5-HT3A receptor. Favorable physicochemical properties and in vitro ADME profile coupled with oral activity in the murine von Bezold-Jarisch reflex model demonstrates the series has promise for producing low to moderate IA partial agonists suitable for an IBS indication.

  20. γ-Hydroxybutyric acid (GHB is not an agonist of extrasynaptic GABAA receptors.

    Directory of Open Access Journals (Sweden)

    William M Connelly

    Full Text Available γ-Hydroxybutyric acid (GHB is an endogenous compound and a drug used clinically to treat the symptoms of narcolepsy. GHB is known to be an agonist of GABAB receptors with millimolar affinity, but also binds with much higher affinity to another site, known as the GHB receptor. While a body of evidence has shown that GHB does not bind to GABAA receptors widely, recent evidence has suggested that the GHB receptor is in fact on extrasynaptic α4β1δ GABAA receptors, where GHB acts as an agonist with an EC50 of 140 nM. We investigated three neuronal cell types that express a tonic GABAA receptor current mediated by extrasynaptic receptors: ventrobasal (VB thalamic neurons, dentate gyrus granule cells and striatal medium spiny neurons. Using whole-cell voltage clamp in brain slices, we found no evidence that GHB (10 µM induced any GABAA receptor mediated current in these cell types, nor that it modulated inhibitory synaptic currents. Furthermore, a high concentration of GHB (3 mM was able to produce a GABAB receptor mediated current, but did not induce any other currents. These results suggest either that GHB is not a high affinity agonist at native α4β1δ receptors, or that these receptors do not exist in classical areas associated with extrasynaptic currents.

  1. γ-Hydroxybutyric acid (GHB) is not an agonist of extrasynaptic GABAA receptors. (United States)

    Connelly, William M; Errington, Adam C; Crunelli, Vincenzo


    γ-Hydroxybutyric acid (GHB) is an endogenous compound and a drug used clinically to treat the symptoms of narcolepsy. GHB is known to be an agonist of GABAB receptors with millimolar affinity, but also binds with much higher affinity to another site, known as the GHB receptor. While a body of evidence has shown that GHB does not bind to GABAA receptors widely, recent evidence has suggested that the GHB receptor is in fact on extrasynaptic α4β1δ GABAA receptors, where GHB acts as an agonist with an EC50 of 140 nM. We investigated three neuronal cell types that express a tonic GABAA receptor current mediated by extrasynaptic receptors: ventrobasal (VB) thalamic neurons, dentate gyrus granule cells and striatal medium spiny neurons. Using whole-cell voltage clamp in brain slices, we found no evidence that GHB (10 µM) induced any GABAA receptor mediated current in these cell types, nor that it modulated inhibitory synaptic currents. Furthermore, a high concentration of GHB (3 mM) was able to produce a GABAB receptor mediated current, but did not induce any other currents. These results suggest either that GHB is not a high affinity agonist at native α4β1δ receptors, or that these receptors do not exist in classical areas associated with extrasynaptic currents.

  2. Evidence that the angiotensin at 2-receptor agonist compound 21 is also a low affinity thromboxane TXA2-receptor antagonist

    DEFF Research Database (Denmark)

    Fredgart, M.; Leurgans, T.; Stenelo, M.;


    Objective: The objective of this study was to test whether Compound 21 (C21), a high-affinity, non-peptide angiotensinAT2-receptor agonist, is also an antagonist of thromboxane A2 (TXA2) receptors thus reducing both vasoconstriction and platelet aggregation. Design and method: Binding of C21 to t...

  3. The effects of the 5-HT(6) receptor agonist EMD and the 5-HT(7) receptor agonist AS19 on memory formation. (United States)

    Meneses, A; Perez-Garcia, G; Liy-Salmeron, G; Flores-Galvez, D; Castillo, C; Castillo, E


    Growing evidence indicates that 5-hydrohytryptamine (5-HT) receptors mediate learning and memory. Particularly interesting are 5-HT(6) and 5-HT(7) receptors, which are localized in brain areas involved in memory formation. Interestingly, recently selective 5-HT(6) and 5-HT(7) receptor agonists and antagonists have become available. Previous evidence indicates that 5-HT(6) or 5-HT(7) receptors antagonists had no effects, improved memory formation and/or reversed amnesia. Herein, the effects of EMD (a 5-HT(6) receptor agonist) and AS19 (a 5-HT(7) receptor agonist) in the associative learning task of autoshaping were studied. Post-training systemic administration of EMD (1-10 mg/kg) or AS19 (1-10 mg/kg) were tested in short-term memory (STM) and long-term memory (LTM). Results showed that only EMD 5.0mg/kg impaired both STM and LTM. AS19 at 1-10 mg/kg significantly impaired STM but not LTM. In those groups used to test only LTM, EMD impaired it; while AS19 improved LTM. Moreover, in the interaction experiments, the STM EMD-impairment effect was partially reversed by the selective 5-HT(6) receptor antagonist SB-399885 (10 mg/kg). The STM AS19-impairment effect (5.0 mg/kg) was not altered by the selective 5-HT(1A) antagonist WAY 100635 (0.3 mg/kg) but reversed by the selective 5-HT(7) receptor antagonist SB-269970 (10.0 mg/kg). The AS19-SB-269970 combination impaired LTM. Taken together these data suggest that the stimulation of 5-HT(6) impaired both STM and LTM. 5-HT(7) receptors stimulation impaired STM but improved LTM. And these results are discussed in the context of their possible neural bases.

  4. Agonist high- and low-affinity states of dopamine D-2 receptors : methods of detection and clinical implications

    NARCIS (Netherlands)

    van Wieringen, Jan-Peter; Booij, Jan; Shalgunov, Vladimir; Elsinga, Philip; Michel, Martin C.


    Dopamine D-2 receptors, similar to other G-protein-coupled receptors, exist in a high- and low-affinity state for agonists. Based upon a review of the methods for detecting D-2 receptor agonist high-affinity states, we discuss alterations of such states in animal models of disease and the implicatio

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

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


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

  6. Quantitative encoding of a partial agonist effect on individual opioid receptors by multi-site phosphorylation and threshold detection


    Lau, Elaine K.; Trester-Zedlitz, Michelle; Trinidad, Jonathan C.; Kotowski, Sarah J.; Krutchinsky, Andrew N.; Burlingame, Alma L; von Zastrow, Mark


    Many drugs act as partial agonists of seven-transmembrane signaling receptors when compared to endogenous ligands. Partial agonism is well described as a 'macroscopic' property manifest at the level of physiological systems or cell populations, but it is not known whether partial agonists encode discrete regulatory information at the 'microscopic' level of individual receptors. We addressed this question by focusing on morphine, a partial agonist drug for µ-type opioid peptide receptors, and ...

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

    Directory of Open Access Journals (Sweden)

    Dae-Sil Lee


    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.

  8. Inhibition of AMPA Receptors by Polyamine Toxins is Regulated by Agonist Efficacy and Stargazin

    DEFF Research Database (Denmark)

    Poulsen, Mette H; Lucas, Simon; Strømgaard, Kristian;


    The α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid receptors (AMPARs) are glutamate-gated cation channels mediating the majority of fast excitatory synaptic transmission in the central nervous system (CNS). Polyamine toxins derived from spiders and wasps are use- and voltage-dependent...... explored the effect of the TARP γ-2 (also known as stargazin) on the inhibitory potency of three structurally different polyamine toxins at Ca(2+)-permeable homomeric GluA1 AMPARs expressed in oocytes. We find that polyamine toxin IC50 is differentially affected by presence of stargazin depending...... on the efficacy of the agonists used to activate GluA1. Co-assembly of GluA1 receptors with stargazin increases the potency of the polyamine toxins when activated by the weak partial agonist kainate, but has no effect in presence of full-agonist L-glutamate (Glu) and partial agonist (RS)-willardiine....

  9. Antidiabetic properties of the histamine H3 receptor protean agonist proxyfan. (United States)

    Henry, Melanie B; Zheng, Shuqin; Duan, Chenxia; Patel, Bhuneshwari; Vassileva, Galya; Sondey, Christopher; Lachowicz, Jean; Hwa, Joyce J


    Proxyfan is a histamine H3 receptor protean agonist that can produce a spectrum of pharmacological effects including agonist, inverse agonist, and antagonist. We have discovered that proxyfan (10 mg/kg orally) significantly improved glucose excursion after an ip glucose tolerance test in either lean or high-fat/cholesterol diet-induced obese mice. It also reduced plasma glucose levels comparable to that of metformin (300 mg/kg orally) in a nongenetic type 2 diabetes mouse model. The dose-dependent decrease in glucose excursion correlated with inhibition of ex vivo H3 receptor binding in the cerebral cortex. In addition, glucose levels were significantly reduced compared with vehicle-treated mice after intracerebroventricular administration of proxyfan, suggesting the involvement of central H3 receptors. Proxyfan-induced reduction of glucose excursion was not observed in the H3 receptor knockout mice, suggesting that proxyfan mediates this effect through H3 receptors. Proxyfan reduced glucose excursion by significantly increasing plasma insulin levels in a glucose-independent manner. However, no difference in insulin sensitivity was observed in proxyfan-treated mice. The H1 receptor antagonist chlorpheniramine and the H2 receptor antagonist zolantidine had modest effects on glucose excursion, and neither inhibited the glucose excursion reduced by proxyfan. The H3 receptor antagonist/inverse agonist, thioperamide, had weaker effects on glucose excursion compared with proxyfan, whereas the H3 receptor agonist imetit did not affect glucose excursion. In conclusion, these findings demonstrate, for the first time, that manipulation of central histamine H3 receptor by proxyfan can significantly improve glucose excursion by increasing plasma insulin levels via a glucose-independent mechanism.

  10. Effects of glucagon-like peptide-1 receptor agonists on renal function

    Institute of Scientific and Technical Information of China (English)

    Theodosios; D; Filippatos; Moses; S; Elisaf


    Glucagon-like peptide-1(GLP-1)receptor agonists result in greater improvements in glycemic control than placebo and promote weight loss with minimal hypoglycemia in patients with type 2 diabetes mellitus.A number of case reports show an association of GLP-1receptor agonists,mainly exenatide,with the development of acute kidney injury.The present review aims to present the available data regarding the effects of GLP-1 receptor agonists on renal function,their use in subjects with chronic renal failure and their possible association with acute kidney injury.Based on the current evidence,exenatide is eliminated by renal mechanisms and should not be given in patients with severe renal impairment or end stage renal disease.Liraglutide is not eliminated by renal or hepatic mechanisms,but it should be used with caution since there are only limited data in patients with renal or hepatic impairment.There is evidence from animal studies that GLP-1 receptor agonists exert protective role in diabetic nephropathy with mechanisms that seem to be independent of their glucose-lowering effect.Additionally,there is evidence that GLP-1 receptor agonists influence water and electrolyte balance.These effects may represent new ways to improve or even prevent diabetic nephropathy.

  11. 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


    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.

  12. Prospects for Creation of Cardioprotective and Antiarrhythmic Drugs Based on Opioid Receptor Agonists


    Maslov, Leonid N; Khaliulin, Igor; Oeltgen, Peter R; Naryzhnaya, Natalia V.; Pei, Jian‐Ming; Brown, Stephen A; Lishmanov, Yury B.; Downey, James M


    Abstract It has now been demonstrated that the μ, δ1, δ2, and κ1 opioid receptor (OR) agonists represent the most promising group of opioids for the creation of drugs enhancing cardiac tolerance to the detrimental effects of ischemia/reperfusion (I/R). Opioids are able to prevent necrosis and apoptosis of cardiomyocytes during I/R and improve cardiac contractility in the reperfusion period. The OR agonists exert an infarct‐reducing effect with prophylactic administration and prevent reperfusi...

  13. Behavioural effects of the benzodiazepine receptor partial agonist RO 16-6028 in mice. (United States)

    Belzung, C; Misslin, R; Vogel, E


    The imidazo-diazepinone RO 16-6028 is a benzodiazepine receptor partial agonist which exhibits some anti-conflict effects in the two-chambered light/dark test without significantly affecting the behaviour of mice confronted with the staircase test. In addition, this drug slightly reduced locomotion and more markedly rearing in a free exploration procedure. These results indicate that RO 16-6028 appears to produce some anxiolytic and sedative properties like full agonists, but with weaker magnitude. This could be related to the benzodiazepine partial agonistic profile of the compound.

  14. Ligand-based receptor tyrosine kinase partial agonists: New paradigm for cancer drug discovery? (United States)

    Riese, David J.


    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

  15. Free Fatty Acid Receptor 1 (FFA1/GPR40) Agonists

    DEFF Research Database (Denmark)

    Christiansen, Elisabeth; Due-Hansen, Maria E; Urban, Christian;


    FFA1 (GPR40) is a new target for treatment of type 2 diabetes. We recently identified the potent FFA1 agonist TUG-469 (5). Inspired by the structurally related TAK-875, we explored the effects of a mesylpropoxy appendage on 5. The appendage significantly lowers lipophilicity and improves metaboli...

  16. GLP-1 receptor agonists or DPP-4 inhibitors: how to guide the clinician? (United States)

    Scheen, André J


    Pharmacological treatment of type 2 diabetes has been enriched during recent years, with the launch of incretin therapies targeting glucagon-like peptide-1 (GLP-1). Such medications comprise either GLP-1 receptor agonists, with short (one or two daily injections: exenatide, liraglutide, lixisenatide) or long duration (one injection once weekly: extended-released exenatide, albiglutide, dulaglutide, taspoglutide); or oral compounds inhibiting dipeptidyl peptidase-4 (DPP-4), the enzyme that inactives GLP-1, also called gliptins (sitagliptin, vildagliptin, saxagliptin, linagliptin, alogliptin). Although both pharmacological approaches target GLP-1, important differences exist concerning the mode of administration (subcutaneous injection versus oral ingestion), the efficacy (better with GLP-1 agonists), the effects on body weight and systolic blood pressure (diminution with agonists versus neutrality with gliptins), the tolerance profile (nausea and possibly vomiting with agonists) and the cost (higher with GLP-1 receptor agonists). Both agents may exert favourable cardiovascular effects. Gliptins may represent a valuable alternative to a sulfonylurea or a glitazone after failure of monotherapy with metformin while GLP-1 receptor agonists may be considered as a good alternative to insulin (especially in obese patients) after failure of a dual oral therapy. However, this scheme is probably too restrictive and modalities of using incretins are numerous, in almost all stages of type 2 diabetes. Physicians may guide the pharmacological choice based on clinical characteristics, therapeutic goals and patient's preference.

  17. Alpha/sub 1/ receptor coupling events initiated by methoxy-substituted tolazoline partial agonists

    Energy Technology Data Exchange (ETDEWEB)

    Wick, P.; Keung, A.; Deth, R.


    A series of mono- and dimethyoxy substituted tolazoline derivatives, known to be partial agonists at the alpha/sub 1/ receptor, were compared with the ..cap alpha../sub 1/ selective full agonist phenylephrine (PE) on isolated strips of rabbit aorta Agonist activity was evaluated in contraction, /sup 45/Ca influx, /sup 45/Ca efflux, and /sup 32/P-Phospholipid labelling studies. Maximum contractile responses for the 2-, 3-, and 3, 5- methoxy substituted tolazoline derivatives (10/sup -5/M) were 53.8, 67.6 and 99.7% of the PE (10/sup -5/M) response respectively. These same partial agonists caused a stimulation of /sup 45/Ca influx to the extent of 64, 86, and 95% of the PE response respectively. In /sup 45/Ca efflux studies, (a measure of the intracellular Ca/sup +2/ release) the tolazolines caused: 30%, 63%, and 78% of the PE stimulated level. /sup 32/P-Phosphatidic acid (PA) labelling was measured as an index of PI turnover after ..cap alpha../sub 1/ receptor stimulation. Compared to PE, the 2-, 3-, and 3,5- methoxy substituted tolazoline derivatives caused 22, 46, and 72% PA labelling. The above values are all in reasonable accord with the rank order or agonist activity shown in maximum contractile responses. The results of this investigation suggest that partial agonists stimulate ..cap alpha.. receptor coupling events at a level which is quantitatively comparable to their potencies in causing contraction of arterial smooth muscle.

  18. Bitter Taste Receptor Agonists Mitigate Features of Allergic Asthma in Mice (United States)

    Sharma, Pawan; Yi, Roslyn; Nayak, Ajay P.; Wang, Nadan; Tang, Francesca; Knight, Morgan J.; Pan, Shi; Oliver, Brian; Deshpande, Deepak A.


    Asthma is characterized by airway inflammation, mucus secretion, remodeling and hyperresponsiveness (AHR). Recent research has established the bronchodilatory effect of bitter taste receptor (TAS2R) agonists in various models. Comprehensive pre-clinical studies aimed at establishing effectiveness of TAS2R agonists in disease models are lacking. Here we aimed to determine the effect of TAS2R agonists on features of asthma. Further, we elucidated a mechanism by which TAS2R agonists mitigate features of asthma. Asthma was induced in mice using intranasal house dust mite or aerosol ova-albumin challenge, and chloroquine or quinine were tested in both prophylactic and treatment models. Allergen challenge resulted in airway inflammation as evidenced by increased immune cells infiltration and release of cytokines and chemokines in the lungs, which were significantly attenuated in TAS2R agonists treated mice. TAS2R agonists attenuated features of airway remodeling including smooth muscle mass, extracellular matrix deposition and pro-fibrotic signaling, and also prevented mucus accumulation and development of AHR in mice. Mechanistic studies using human neutrophils demonstrated that inhibition of immune cell chemotaxis is a key mechanism by which TAS2R agonists blocked allergic airway inflammation and exerted anti-asthma effects. Our comprehensive studies establish the effectiveness of TAS2R agonists in mitigating multiple features of allergic asthma.

  19. Treatment of Type 2 Diabetes by Free Fatty Acid Receptor Agonists


    Kenneth R. Watterson; Hudson, Brian D.; Ulven, Trond; Milligan, Graeme


    Dietary free fatty acids (FFAs), such as ω-3 fatty acids, regulate metabolic and anti-inflammatory processes, with many of these effects attributed to FFAs interacting with a family of G protein-coupled receptors. Selective synthetic ligands for Free Fatty Acid receptors (FFA1-4) have consequently been developed as potential treatments for type 2 diabetes (T2D). In particular, clinical studies show that Fasiglifam, an agonist of the long chain FFA receptor, FFA1, improved glycaemic control an...

  20. Studies Toward the Pharmacophore of Salvinorin A, a Potent Kappa Opioid Receptor Agonist


    Munro, Thomas A.; Mark A. Rizzacasa; Roth, Bryan L.; Toth, Beth A.; Yan, Feng


    Salvinorin A (1), from the sage Salvia divinorum, is a potent and selective kappa opioid receptor (KOR) agonist. We screened other salvinorins and derivatives for binding affinity and functional activity at opioid receptors. Our results suggest that the methyl ester and furan ring are required for activity, but that the lactone and ketone functionalities are not. Other salvinorins showed negligible binding affinity at the KOR. None of the compounds bound to mu or delta opioid receptors.

  1. Studies toward the pharmacophore of salvinorin A, a potent kappa opioid receptor agonist. (United States)

    Munro, Thomas A; Rizzacasa, Mark A; Roth, Bryan L; Toth, Beth A; Yan, Feng


    Salvinorin A (1), from the sage Salvia divinorum, is a potent and selective kappa opioid receptor (KOR) agonist. We screened other salvinorins and derivatives for binding affinity and functional activity at opioid receptors. Our results suggest that the methyl ester and furan ring are required for activity but that the lactone and ketone functionalities are not. Other salvinorins showed negligible binding affinity at the KOR. None of the compounds bound to mu or delta opioid receptors.

  2. PET imaging to measure therapy-related occupancy and disease-induced changes of expression of adenosine A1 receptors in the rodent brain

    NARCIS (Netherlands)

    Paul, Souman


    Rol van adenosine A1 receptor in de vroege fase van encefalitis Adenosine A1 receptoren (A1R) spelen een belangrijke rol bij de bescherming van hersencellen tijdens de vroege fase van hersenontsteking (encefalitis) bij ratten en mogelijk ook bij mensen. Dat concludeert Souman Paul in zijn proefschri

  3. The Quintiles Prize Lecture 2004: The identification of the adenosine A2B receptor as a novel therapeutic target in asthma (United States)

    Holgate, Stephen T


    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

  4. The Quintiles Prize Lecture 2004. The identification of the adenosine A2B receptor as a novel therapeutic target in asthma. (United States)

    Holgate, Stephen T


    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.

  5. Modification of formalin-induced nociception by different histamine receptor agonists and antagonists. (United States)

    Farzin, Davood; Nosrati, Farnaz


    The present study evaluated the effects of different histamine receptor agonists and antagonists on the nociceptive response in the mouse formalin test. Intracerebroventricular (20-40 microg/mouse i.c.v.) or subcutaneous (1-10 mg/kg s.c.) injection of HTMT (H(1) receptor agonist) elicited a dose-related hyperalgesia in the early and late phases. Conversely, intraperitoneal (20 and 30 mg/kg i.p.) injection of dexchlorpheniramine (H(1) receptor antagonist) was antinociceptive in both phases. At a dose ineffective per se, dexchlorpheniramine (10 mg/kg i.p.) antagonized the hyperalgesia induced by HTMT (40 mug/mouse i.c.v. or 10 mg/kg s.c.). Dimaprit (H(2) receptor agonist, 30 mg/kg i.p.) and ranitidine (H(2) receptor antagonist, 20 and 40 mg/kg i.p.) reduced the nociceptive responses in the early and late phases. No significant change in the antinociceptive activity was found following the combination of dimaprit (30 mg/kg i.p.) with ranitidine (10 mg/kg i.p.). The antinociceptive effect of dimaprit (30 mg/kg i.p.) was prevented by naloxone (5 mg/kg i.p.) in the early phase or by imetit (H(3) receptor agonist, 25 mg/kg i.p.) in both early and late phases. The histamine H(3) receptor agonist imetit was hyperalgesic following i.p. administration of 50 mg/kg. Imetit-induced hyperalgesia was completely prevented by treatment with a dose ineffective per se of thioperamide (H(3) receptor antagonist, 5 mg/kg i.p.). The results suggest that histamine H(1) and H(3) receptor activations increase sensitivity to nociceptive stimulus in the formalin test.

  6. In pursuit of alpha4beta2 nicotinic receptor partial agonists for smoking cessation: carbon analogs of (-)-cytisine. (United States)

    Coe, Jotham W; Vetelino, Michael G; Bashore, Crystal G; Wirtz, Michael C; Brooks, Paige R; Arnold, Eric P; Lebel, Lorraine A; Fox, Carol B; Sands, Steven B; Davis, Thomas I; Schulz, David W; Rollema, Hans; Tingley, F David; O'Neill, Brian T


    The preparation and biological activity of analogs of (-)-cytisine, an alpha4beta2 nicotinic receptor partial agonist, are discussed. All-carbon-containing phenyl ring replacements of the pyridone ring system, generated via Heck cyclization protocols, exhibited weaker affinity and lower efficacy partial agonist profiles relative to (-)-cytisine. In vivo, selected compounds exhibit lower efficacy partial agonist profiles than that of (-)-cytisine.

  7. Phosphorylation and chronic agonist treatment atypically modulate GABAB receptor cell surface stability. (United States)

    Fairfax, Benjamin P; Pitcher, Julie A; Scott, Mark G H; Calver, Andrew R; Pangalos, Menelas N; Moss, Stephen J; Couve, Andrés


    GABA(B) receptors are heterodimeric G protein-coupled receptors that mediate slow synaptic inhibition in the central nervous system. The dynamic control of the cell surface stability of GABA(B) receptors is likely to be of fundamental importance in the modulation of receptor signaling. Presently, however, this process is poorly understood. Here we demonstrate that GABA(B) receptors are remarkably stable at the plasma membrane showing little basal endocytosis in cultured cortical and hippocampal neurons. In addition, we show that exposure to baclofen, a well characterized GABA(B) receptor agonist, fails to enhance GABA(B) receptor endocytosis. Lack of receptor internalization in neurons correlates with an absence of agonist-induced phosphorylation and lack of arrestin recruitment in heterologous systems. We also demonstrate that chronic exposure to baclofen selectively promotes endocytosis-independent GABA(B) receptor degradation. The effect of baclofen can be attenuated by activation of cAMP-dependent protein kinase or co-stimulation of beta-adrenergic receptors. Furthermore, we show that increased degradation rates are correlated with reduced receptor phosphorylation at serine 892 in GABA(B)R2. Our results support a model in which GABA(B)R2 phosphorylation specifically stabilizes surface GABA(B) receptors in neurons. We propose that signaling pathways that regulate cAMP levels in neurons may have profound effects on the tonic synaptic inhibition by modulating the availability of GABA(B) receptors.

  8. CB1 and CB2 receptor agonists promote analgesia through synergy in a murine model of tumor pain. (United States)

    Khasabova, Iryna A; Gielissen, James; Chandiramani, Anisha; Harding-Rose, Catherine; Odeh, Desiree Abu; Simone, Donald A; Seybold, Virginia S


    In light of the adverse side-effects of opioids, cannabinoid receptor agonists may provide an effective alternative for the treatment of cancer pain. This study examined the potency and efficacy of synthetic CB1 and CB2 receptor agonists in a murine model of tumor pain. Intraplantar injection of the CB1 receptor agonist arachidonylcyclopropylamide (ED(50) of 18.4 μg) reduced tumor-related mechanical hyperalgesia by activation of peripheral CB1 but not CB2 receptors. Similar injection of the CB2 receptor agonist AM1241 (ED50 of 19.5 μg) reduced mechanical hyperalgesia by activation of peripheral CB2 but not CB1 receptors. Both agonists had an efficacy comparable with that of morphine (intraplantar), but their analgesic effects were independent of opioid receptors. Isobolographic analysis of the coinjection of arachidonylcyclopropylamide and AM1241 determined that the CB1 and CB2 receptor agonists interacted synergistically to reduce mechanical hyperalgesia in the tumor-bearing paw. These data extend our previous findings that the peripheral cannabinoid receptors are a promising target for the management of cancer pain and mixed cannabinoid receptor agonists may have a therapeutic advantage over selective agonists.

  9. Identification of adiponectin receptor agonist utilizing a fluorescence polarization based high throughput assay.

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    Yiyi Sun

    Full Text Available Adiponectin, the adipose-derived hormone, plays an important role in the suppression of metabolic disorders that can result in type 2 diabetes, obesity, and atherosclerosis. It has been shown that up-regulation of adiponectin or adiponectin receptor has a number of therapeutic benefits. Given that it is hard to convert the full size adiponectin protein into a viable drug, adiponectin receptor agonists could be designed or identified using high-throughput screening. Here, we report on the development of a two-step screening process to identify adiponectin agonists. First step, we developed a high throughput screening assay based on fluorescence polarization to identify adiponectin ligands. The fluorescence polarization assay reported here could be adapted to screening against larger small molecular compound libraries. A natural product library containing 10,000 compounds was screened and 9 hits were selected for validation. These compounds have been taken for the second-step in vitro tests to confirm their agonistic activity. The most active adiponectin receptor 1 agonists are matairesinol, arctiin, (--arctigenin and gramine. The most active adiponectin receptor 2 agonists are parthenolide, taxifoliol, deoxyschizandrin, and syringin. These compounds may be useful drug candidates for hypoadiponectin related diseases.

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

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    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

  11. Potency and characterization of estrogen-receptor agonists in United Kingdom estuarine sediments. (United States)

    Thomas, Kevin V; Balaam, Jan; Hurst, Mark; Nedyalkova, Zoya; Mekenyan, Ovanes


    The activity of estrogen-receptor (ER) agonists in sediments collected from the United Kingdom (UK) estuaries was assessed using the in vitro recombinant yeast estrogen screen (YES assay). The YES assay was successfully used to determine the in vitro ER agonist potency of pore waters and solvent extracts of sediments collected from UK estuaries. Estrogen-receptor agonists were detected in 66% of the pore water samples and in 91% of the sediment solvent extracts tested. The pore waters tested had ER agonist potencies from less than 2 to 68 ng 17beta-estradiol (E2) L(-1), whereas sediment extracts had potencies from less than 0.2 to 13 microg E2 kg(-1). A toxicity identification evaluation approach using bioassay-directed fractionation was used in an attempt to identify the ER agonists in extracts of sediments collected from the Tyne and Tees estuaries (UK). Gas chromatography-mass spectrometry was used to provide lists of compounds in the fractions obtained that were evaluated for known ER agonist activity using published data and an ER quantitative structure-activity relationship model. Toxicity identification evaluation characterization failed to identify any ER agonists in pore water extracts; however, three compounds in sediment solvent extracts were identified as ER agonists. Nonylphenol, cinnarizine, and cholesta-4,6-dien-3-one were identified in the sample collected from the Tyne estuary. Important ER agonist substances that contaminate marine sediments remain unidentified. The present study as well as previous work on effluents point toward the involvement of natural products in the estrogenic burdens of marine sediments. Further work is required to establish the relative contribution of natural products and anthropogenic chemicals to current environmental impacts in the context of the Oslo and Paris Commission strategy to eliminate hazardous substances by 2020.

  12. Structural determinants of agonist-specific kinetics at the ionotropic glutamate receptor 2. (United States)

    Holm, Mai Marie; Lunn, Marie-Louise; Traynelis, Stephen F; Kastrup, Jette S; Egebjerg, Jan


    Glutamate receptors (GluRs) are the most abundant mediators of the fast excitatory neurotransmission in the human brain. Agonists will, after activation of the receptors, induce different degrees of desensitization. The efficacy of agonists strongly correlates with the agonist-induced closure of the ligand-binding domain. However, the differences in desensitization properties are less well understood. By using high-resolution x-ray structure of the GluR2 flop (GluR2o) ligand-binding core protein in complex with the partial glutamate receptor agonist (S)-2-amino-3-(3-hydroxy-5-tert-butyl-4-isothiazolyl)propionic acid [(S)-thio-ATPA], we show that (S)-thio-ATPA induces an 18 degrees closure of the binding core similar to another partial agonist, (S)-2-amino-3-(4-bromo-3-hydroxy-5-isoxazolyl)propionic acid [(S)-Br-HIBO]. Despite the similar closure of the ligand-binding domain, we find in electrophysiological studies that (S)-thio-ATPA induced a 6.4-fold larger steady-state current than (RS)-Br-HIBO, and rapid agonist applications show that (S)-thio-ATPA induces a 3.6-fold higher steady-state/peak ratio and a 2.2-fold slower desensitization time constant than (RS)-Br-HIBO. Structural comparisons reveal that (S)-Br-HIBO, but not (S)-thio-ATPA, induces a twist of the ligand-binding core compared with the apostructure, and the agonist-specific conformation of Leu-650 correlates with the different kinetic profiles pointing at a key role in defining the desensitization kinetics. We conclude that, especially for intermediate efficacious agonists, the desensitization properties are influenced by additional ligand-induced factors beyond domain closure.

  13. Pre-synaptic adenosine A2A receptors control cannabinoid CB1 receptor-mediated inhibition of striatal glutamatergic neurotransmission. (United States)

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


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

  14. Ameliorative effect of adenosine on hypoxia-reoxygenation injury in LLC-PK1, a porcine kidney cell line. (United States)

    Yonehana, T; Gemba, M


    We studied the effects of adenosine on injury caused by hypoxia and reoxygenation in LLC-PK1 cells. Lactate dehydrogenase and gamma-glutamyltranspeptidase were released from cells exposed to hypoxia for 6 hr and then reoxygenation for 1 hr. The addition of adenosine at 100 microM to the medium before hypoxia began significantly decreased enzyme leakage into medium during both hypoxia and reoxygenation. The adenosine A1-receptor agonist, R(-)-N6-(2-phenylisopropyl)adenosine (R-PIA), at the concentration of 100 microM, did not affect enzyme release, but the adenosine A2-receptor agonist 2-p-[2-car-boxyethyl]phenethyl-amino-5'-N-ethylcarboxamido-adenosi ne hydrochloride (CGS 21680) at the concentration of 100 nM, suppressed the injury caused by hypoxia and reoxygenation. There were decreases in cAMP contents and ATP levels in LLC-PK1 cells injured by hypoxia and reoxygenation. Adenosine (100 microM) restored ATP levels in the cells during reoxygenation. With adenosine, the intracellular cAMP level was increased prominently during reoxygenation. These results suggest that adenosine protects LLC-PK1 cells from injury caused by hypoxia and reoxygenation by increasing the intracellular cAMP level via adenosine A2 receptor.

  15. A selective adenosine A2A receptor antagonist ameliorated hyperlocomotion in an animal model of lateral fluid percussion brain injury. (United States)

    Mullah, Saad Habib-E-Rasul; Inaji, Motoki; Nariai, Tadashi; Ishibashi, Satoru; Ohno, Kikuo


    Increased concentration of extracellular adenosine after brain injury is supposed to be one of the causes of secondary brain damage. The purpose of the present study is to examine whether or not administration of adenosine A2A receptor antagonist may be efficacious in ameliorating neurological symptoms by blocking secondary brain damage through cascades initiated by adenosine A2a receptor.Mongolian gerbils were divided into four groups: the trauma-medication (T-M), trauma-saline (T-S), sham-medication (S-M), and sham-saline (S-S) groups. Trauma groups received lateral fluid percussion injury. Medication groups received i.p. injection of SCH58261 (selective adenosine A2A receptor antagonist) until the fifth post-injury day. Open-field locomotion test and grabbing test were conducted before and 1, 3, 5, 7, and 9 days after injury.The total distance of movement in the T-S group was significantly greater than in the other three groups at all time points. In the T-M group, administration of SCH58261 significantly blocked hyperlocomotion, which was observed in the T-S group. There was no significant difference in the total distance among the T-M, S-M, and S-S groups. In the grabbing test, grabbing time was significantly increased in the T-S group 3, 5, 7, and 9 days after the operation. SCH58261 also improved grabbing time in the T-M group.Adenosine A2A antagonist successfully suppressed the trauma-induced hyperlocomotion, presumably by blocking secondary brain damage.

  16. Effects of Glucagon-Like Peptide-1 Receptor Agonists on Body Weight: A Meta-Analysis


    Matteo Monami; Ilaria Dicembrini; Niccolò Marchionni; Rotella, Carlo M.; Edoardo Mannucci


    Glucagon-Like Peptide-1 receptor agonists (GLP-1RAs), approved as glucose-lowering drugs for the treatment of type 2 diabetes, have also been shown to reduce body weight. An extensive Medline, Cochrane database, and Embase search for “exenatide,” “liraglutide,” “albiglutide,” “semaglutide,” and “lixisenatide” was performed, collecting all randomized clinical trials on humans up to December 15, 2011, with a duration of at least 24 weeks, comparing GLP-1 receptor agonists with either placebo or...

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

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    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.

  18. Identification of human dopamine D1-like receptor agonist using a cell-based functional assay

    Institute of Scientific and Technical Information of China (English)

    Nan JIANG; Ke-qing OU-YANG; Shao-xi CAI; Ying-he HU; Zhi-liang XU


    Aim: To establish a cell-based assay to screen human dopamine D1 and D5 receptor agonists against compounds from a natural product compound library.Methods: Synthetic responsive elements 6×cAMP response elements (CRE) and a mini promoter containing a TATA box were inserted into the pGL3 basic vector to generate the reporter gene construct pCRE/TA/Luci. CHO cells were co-transfected with the reporter gene construct and human D1 or D5 receptor cDNA in mammalian expression vectors. Stable cell lines were established for agonist screening. A natural product compound library from over 300 herbs has been established. The extracts from these herbs were used for human D1 and D5 receptor agonist screenings. Results: A number of extracts were identified that activated both D1 and D5 receptors. One of the herb extracts, SBG492, demonstrated distinct pharmacological characteristics with human D1 and D5 receptors.The EC50 values of SBG492 were 342.7 μg/mL for the D1 receptor and 31.7 μg/mL for the D5 receptor. Conclusion: We have established a cell-based assay for high-throughput drug screening to identify D 1-like receptor agonists from natural products. Several extracts that can active D1-like receptors were discovered.These compounds could be useful tools for studies on the functions of these receptors in the brain and could potentially be developed into therapeutic drugs for the treatment of central nervous system diseases.

  19. Structural Requirements of N-Substituted Spiropiperidine Analogues as Agonists of Nociceptin/Orphanin FQ Receptor

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    Ling Yang


    Full Text Available The nociceptin/orphanin FQ (NOP receptor is involved in a wide range of biological functions, including pain, anxiety, depression and drug abuse. Especially, its agonists have great potential to be developed into anxiolytics. In this work, both the ligand- and receptor-based three-dimensional quantitative structure–activity relationship (3D-QSAR studies were carried out using comparative molecular field analysis (CoMFA and comparative molecular similarity indices analysis (CoMSIA techniques on 103 N-substituted spiropiperidine analogues as NOP agonists. The resultant optimal ligand-based CoMSIA model exhibited Q2 of 0.501, R2ncv of 0.912 and its predictive ability was validated by using an independent test set of 26 compounds which gave R2pred value of 0.818. In addition, docking analysis and molecular dynamics simulation (MD were also applied to elucidate the probable binding modes of these agonists. Interpretation of the 3D contour maps, in the context of the topology of the active site of NOP, provided insight into the NOP-agonist interactions. The information obtained from this work can be used to accurately predict the binding affinity of related agonists and also facilitate the future rational design of novel agonists with improved activity.

  20. Rodent antinociception following acute treatment with different histamine receptor agonists and antagonists. (United States)

    Farzin, Davood; Asghari, Ladan; Nowrouzi, Mahvash


    The effects of different histamine receptor agonists and antagonists on the nociceptive threshold were investigated in mice by two different kinds of noxious stimuli: thermal (hot plate) and chemical (acetic acid-induced abdominal writhing). Intracerebroventricular (icv) injection of the histamine H(1) receptor agonist, HTMT (6-[2-(4-imidazolyl)ethylamino]-N-(4-trifluoromethylphenyl) heptanecarboxamide) (50 microg/mouse), produced a hypernociception in the hot plate and writhing tests. Conversely, intraperitoneal (ip) injection of dexchlorpheniramine (30 and 40 mg/kg) and diphenhydramine (20 and 40 mg/kg) increased the pain threshold in both tests. The histamine H(2) receptor agonist, dimaprit (50 and 100 microg/mouse icv), or antagonist, ranitidine (50 and 100 microg/mouse icv), raised the pain threshold in both hot plate and writhing tests. In the mouse hot plate test, the histamine H(3) receptor agonist, imetit (50 mg/kg ip), reduced the pain threshold, while the histamine H(3) receptor antagonist, thioperamide (10 and 20 mg/kg ip), produced an antinociception. The hypernociceptive effects of HTMT and imetit were antagonized by dexchlorpheniramine (20 mg/kg ip) and thioperamide (5 mg/kg ip), respectively. The results suggest that histaminergic mechanisms may be involved in the modulation of nociceptive stimuli.

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

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    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)


    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

  2. Pharmacological profiles of alpha 2 adrenergic receptor agonists identified using genetically altered mice and isobolographic analysis. (United States)

    Fairbanks, Carolyn A; Stone, Laura S; Wilcox, George L


    Endogenous, descending noradrenergic fibers impose analgesic control over spinal afferent circuitry mediating the rostrad transmission of pain signals. These fibers target alpha 2 adrenergic receptors (alpha(2)ARs) on both primary afferent terminals and secondary neurons, and their activation mediates substantial inhibitory control over this transmission, rivaling that of opioid receptors which share a similar pattern of distribution. The terminals of primary afferent nociceptive neurons and secondary spinal dorsal horn neurons express alpha(2A)AR and alpha(2C)AR subtypes, respectively. Spinal delivery of these agents serves to reduce their side effects, which are mediated largely at supraspinal sites, by concentrating the drugs at the spinal level. Targeting these spinal alpha(2)ARs with one of five selective therapeutic agonists, clonidine, dexmedetomidine, brimonidine, ST91 and moxonidine, produces significant antinociception that can work in concert with opioid agonists to yield synergistic antinociception. Application of several genetically altered mouse lines had facilitated identification of the primary receptor subtypes that likely mediate the antinociceptive effects of these agents. This review provides first an anatomical description of the localization of the three subtypes in the central nervous system, second a detailed account of the pharmacological history of each of the six primary agonists, and finally a comprehensive report of the specific interactions of other GPCR agonists with each of the six principal alpha(2)AR agonists featured.

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

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    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.


    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)

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


    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.

  5. Structural complexes of the agonist, inverse agonist and antagonist bound C5a receptor: insights into pharmacology and signaling. (United States)

    Rana, Soumendra; Sahoo, Amita Rani; Majhi, Bharat Kumar


    The C5a receptor (C5aR) is a pharmacologically important G-protein coupled receptor (GPCR) that interacts with (h)C5a, by recruiting both the "orthosteric" sites (site1 at the N-terminus and site2 at the ECS, extra cellular surface) on C5aR in a two site-binding model. However, the complex pharmacological landscape and the distinguishing chemistry operating either at the "orthosteric" site1 or at the functionally important "orthosteric" site2 of C5aR are still not clear, which greatly limits the understanding of C5aR pharmacology. One of the major bottlenecks is the lack of an experimental structure or a refined model structure of C5aR with appropriately defined active sites. The study attempts to understand the pharmacology at the "orthosteric" site2 of C5aR rationally by generating a highly refined full-blown model structure of C5aR through advanced molecular modeling techniques, and further subjecting it to automated docking and molecular dynamics (MD) studies in the POPC bilayer. The first series of structural complexes of C5aR respectively bound to a linear native peptide agonist ((h)C5a-CT), a small molecule inverse agonist (NDT) and a cyclic peptide antagonist (PMX53) are reported, apparently establishing the unique pharmacological landscape of the "orthosteric" site2, which also illustrates an energetically distinct but coherent competitive chemistry ("cation-π" vs. "π-π" interactions) involved in distinguishing the established ligands known for targeting the "orthosteric" site2 of C5aR. Over a total of 1 μs molecular dynamics (MD) simulation in the POPC bilayer, it is evidenced that while the agonist prefers a "cation-π" interaction, the inverse agonist prefers a "cogwheel/L-shaped" interaction in contrast to the "edge-to-face/T-shaped" type π-π interactions demonstrated by the antagonist by engaging the F275(7.28) of the C5aR. In the absence of a NMR or crystallographically guided model structure of C5aR, the computational model complexes not only

  6. Ascorbic acid enables reversible dopamine receptor /sup 3/H-agonist binding

    Energy Technology Data Exchange (ETDEWEB)

    Leff, S.; Sibley, D.R.; Hamblin, M.; Creese, I.


    The effects of ascorbic acid on dopaminergic /sup 3/H-agonist receptor binding were studied in membrane homogenates of bovine anterior pituitary and caudate, and rat striatum. In all tissues virtually no stereospecific binding (defined using 1uM (+)butaclamol) of the /sup 3/H-agonists N-propylnorapomorphine (NPA), apomorphine, or dopamine could be demonstrated in the absence of ascorbic acid. Although levels of total /sup 3/H-agonist binding were three to five times greater in the absence than in the presence of 0.1% ascorbic acid, the increased binding was entirely non-stereospecific. Greater amounts of dopamine-inhibitable /sup 3/H-NPA binding could be demonstrated in the absence of 0.1% ascorbic acid, but this measure of ''specific binding'' was demonstrated not to represent dopamine receptor binding since several other catecholamines and catechol were equipotent with dopamine and more potent than the dopamine agonist (+/-)amino-6,7-dihydroxy-1,2,3,4-tetrahydronapthalene (ADTN) in inhibiting this binding. High levels of dopamine-displaceable /sup 3/H-agonist binding were detected in fresh and boiled homogenates of cerebellum, an area of brain which receives no dopaminergic innervation, further demonstrating the non-specific nature of /sup 3/H-agonist binding in the absence of ascorbic acid. These studies emphasize that under typical assay conditions ascorbic acid is required in order to demonstrate reversible and specific /sup 3/H-agonist binding to dopamine receptors.



    Vincenzi, Fabrizio


    Several studies have suggested the possible involvement of A2A adenosine receptors in the pathogenesis of neuronal disorders, including Huntington’s disease. Huntington’s disease is an inherited neurodegenerative disease clinically characterized by motor, cognitive and behavioural impairments. The genetic cause of the disease is the expanded CAG triplet in a gene coding for huntingtin, a protein involved in several physiological processes. Huntington’s disease affects primarly ...

  8. Effects of caffeine on behavioral and inflammatory changes elicited by copper in zebrafish larvae: Role of adenosine receptors. (United States)

    Cruz, Fernanda Fernandes; Leite, Carlos Eduardo; Kist, Luiza Wilges; de Oliveira, Giovanna Medeiros; Bogo, Maurício Reis; Bonan, Carla Denise; Campos, Maria Martha; Morrone, Fernanda Bueno


    This study investigated the effects of caffeine in the behavioral and inflammatory alterations caused by copper in zebrafish larvae, attempting to correlate these changes with the modulation of adenosine receptors. To perform a survival curve, 7dpf larvae were exposed to 10μM CuSO4, combined to different concentrations of caffeine (100μM, 500μM and 1mM) for up to 24h. The treatment with copper showed lower survival rates only when combined with 500μM and 1mM of caffeine. We selected 4 and 24h as treatment time-points. The behavior evaluation was done by analyzing the traveled distance, the number of entries in the center, and the length of permanence in the center and the periphery of the well. The exposure to 10μM CuSO4 plus 500μM caffeine at 4 and 24h changed the behavioral parameters. To study the inflammatory effects of caffeine, we assessed the PGE2 levels by using UHPLC-MS/MS, and TNF, COX-2, IL-6 and IL-10 gene expression by RT-qPCR. The expression of adenosine receptors was also evaluated with RT-qPCR. When combined to copper, caffeine altered inflammatory markers depending on the time of exposure. Adenosine receptors expression was significantly increased, especially after 4h exposure to copper and caffeine together or separately. Our results demonstrated that caffeine enhances the inflammation induced by copper by decreasing animal survival, altering inflammatory markers and promoting behavioral changes in zebrafish larvae. We also conclude that alterations in adenosine receptors are related to those effects.

  9. THIP and isoguvacine are partial agonists of GABA-stimulated benzodiazepine receptor binding. (United States)

    Karobath, M; Lippitsch, M


    The effects of THIP and isoguvacine on 3H-flunitrazepam binding to washed membranes prepared from the cerebral cortex of adult rats have been examined. THIP, which has only minimal stimulatory effects on benzodiazepine (BZ) receptor binding, has been found to inhibit the stimulation induced by small concentrations (2 microM) of exogenous GABA. While isoguvacine stimulates BZ receptor binding, although to a smaller extent than GABA, it also antagonizes the stimulation of BZ receptor binding induced by GABA. Thus THIP and isoguvacine exhibit the properties of a partial agonist of GABA-stimulated BZ receptor binding.

  10. Adenosine A2A receptors and uric acid mediate protective effects of inosine against TNBS-induced colitis in rats. (United States)

    Rahimian, Reza; Fakhfouri, Gohar; Daneshmand, Ali; Mohammadi, Hamed; Bahremand, Arash; Rasouli, Mohammad Reza; Mousavizadeh, Kazem; Dehpour, Ahmad Reza


    Inflammatory bowel disease comprises chronic recurrent inflammation of gastrointestinal tract. This study was conducted to investigate inosine, a potent immunomodulator, in 2,4,6-trinitrobenzene sulphonic acid (TNBS)-induced chronic model of experimental colitis, and contribution of adenosine A(2A) receptors and the metabolite uric acid as possible underlying mechanisms. Experimental colitis was rendered in rats by a single colonic administration of 10 mg of TNBS. Inosine, potassium oxonate (a hepatic uricase inhibitor), SCH-442416 (a selective adenosine A(2A) receptor antagonist), inosine+potassium oxonate, or inosine+SCH-442416 were given twice daily for 7 successive days. At the end of experiment, macroscopic and histopathologic scores, colonic malondialdehyde (MDA), Tumor Necrosis Factor-alpha (TNF-α) and Interleukin-1beta (IL-1β) levels, and myeloperoxidase (MPO) activity were assessed. Plasma uric acid level was measured throughout the experiment. Both macroscopic and histological features of colonic injury were markedly ameliorated by either inosine, oxonate or inosine+oxonate. Likewise, the elevated amounts of MPO and MDA abated as well as those of TNF-α and IL-1β (Pacid levels were significantly higher in inosine or oxonate groups compared to control. Inosine+oxonate resulted in an even more elvelated uric acid level than each treatment alone (Pacid and adenosine A(2A) receptors contribute to these salutary properties.

  11. Topiramate selectively protects against seizures induced by ATPA, a GluR5 kainate receptor agonist. (United States)

    Kaminski, Rafal M; Banerjee, Madhumita; Rogawski, Michael A


    Although the mechanism of action of topiramate is not fully understood, its anticonvulsant properties may result, at least in part, from an interaction with AMPA/kainate receptors. We have recently shown that topiramate selectively inhibits postsynaptic responses mediated by GluR5 kainate receptors. To determine if this action of topiramate is relevant to the anticonvulsant effects of the drug in vivo, we determined the protective activity of topiramate against seizures induced by intravenous infusion of various ionotropic glutamate receptor agonists in mice. Topiramate (25-100 mg/kg, i.p.) produced a dose-dependent elevation in the threshold for clonic seizures induced by infusion of ATPA, a selective agonist of GluR5 kainate receptors. Topiramate was less effective in protecting against clonic seizures induced by kainate, a mixed agonist of AMPA and kainate receptors. Topiramate did not affect clonic seizures induced by AMPA or NMDA. In contrast, the thresholds for tonic seizures induced by higher doses of these various glutamate receptor agonists were all elevated by topiramate. Unlike topiramate, carbamazepine elevated the threshold for AMPA- but not ATPA-induced clonic seizures. Our results are consistent with the possibility that the effects of topiramate on clonic seizure activity are due to functional blockade of GluR5 kainate receptors. Protection from tonic seizures may be mediated by other actions of the drug. Together with our in vitro cellular electrophysiological results, the present observations strongly support a unique mechanism of action of topiramate, which involves GluR5 kainate receptors.

  12. The protease-activated receptor-2 agonist induces gastric mucus secretion and mucosal cytoprotection


    Kawabata, Atsufumi; Kinoshita, Mitsuhiro; Nishikawa, Hiroyuki; Kuroda, Ryotaro; Nishida, Minoru; Araki, Hiromasa; Arizono, Naoki; Oda, Yasuo; Kakehi, Kazuaki


    Protease-activated receptor-2 (PAR-2), a receptor activated by trypsin/tryptase, modulates smooth muscle tone and exocrine secretion in the salivary glands and pancreas. Given that PAR-2 is expressed throughout the gastrointestinal tract, we investigated effects of PAR-2 agonists on mucus secretion and gastric mucosal injury in the rat. PAR-2–activating peptides triggered secretion of mucus in the stomach, but not in the duodenum. This mucus secretion was abolished by pretreatment with capsai...

  13. Treatment potential of the GLP-1 receptor agonists in type 2 diabetes mellitus

    DEFF Research Database (Denmark)

    Østergaard, L; Frandsen, Christian S; Madsbad, S


    Over the last decade, the discovery of glucagon-like peptide 1 receptor agonists (GLP-1 RAs) has increased the treatment options for patients with type 2 diabetes mellitus (T2DM). GLP-1 RAs mimic the effects of native GLP-1, which increases insulin secretion, inhibits glucagon secretion, increases...

  14. Dopamine D1 receptor-agonist interactions: A mutagenesis and homology modeling study. (United States)

    Mente, Scot; Guilmette, Edward; Salafia, Michelle; Gray, David


    The dopamine D1 receptor is a G protein-coupled receptor that regulates intracellular signaling via agonist activation. Although the number of solved GPCR X-ray structures has been steadily increasing, still no structure of the D1 receptor exists. We have used site-directed mutagenesis of 12 orthosteric vicinity residues of possible importance to G protein-coupled activation to examine the function of prototypical orthosteric D1 agonists and partial agonists. We find that residues from four different regions of the D1 receptor make significant contributions to agonist function. All compounds studied, which are catechol-amines, are found to interact with the previously identified residues: the conserved D103(3.32), as well as the trans-membrane V serine residues. Additional key interactions are found for trans-membrane VI residues F288(6.51), F289(6.52) and N292(6.55), as well as the extra-cellular loop residue L190(ECL2). Molecular dynamics simulations of a D1 homology model have been used to help put the ligand-residue interactions into context. Finally, we considered the rescaling of fold-shift data as a method to account for the change in the size of the mutated side-chain and found that this rescaling helps to relate the calculated ligand-residue energies with observed experimental fold-shifts.

  15. In utero Exposure to beta-2-Adrenergic Receptor Agonist Drugs and Risk for Autism Spectrum Disorders

    DEFF Research Database (Denmark)

    Gidaya, Nicole B.; Lee, Brian K.; Burstyn, Igor


    OBJECTIVES: The purpose of this study was to investigate associations between use of β-2-adrenergic receptor (B2AR) agonist drugs during pregnancy and risk for autism spectrum disorders (ASD). METHODS: A case-control study was conducted by using Denmark’s health and population registers. Among...

  16. Impact of the AT(2) receptor agonist C21 on blood pressure and beyond

    DEFF Research Database (Denmark)

    Foulquier, Sébastien; Steckelings, Ulrike Muscha; Unger, Thomas


    It is now widely accepted that the angiotensin AT(2) receptor (AT(2)R) plays an important protective role during pathophysiologic conditions, acting as a repair system. The development of the first selective nonpeptide AT(2)R agonist C21 accelerated our understanding of AT(2)R-mediated protective...

  17. Dopamine receptor agonists modulate voluntary alcohol intake independently of individual levels of alcohol intake in rats

    NARCIS (Netherlands)

    Spoelder, Marcia; Baars, Annemarie M; Rotte, Marthe D; Vanderschuren, Louk J M J; Lesscher, Heidi M B


    RATIONALE: Individual susceptibility to alcohol use disorder has been related to functional changes in dopaminergic neurotransmission. OBJECTIVES: The aim of the current work was to assess the effects of selective dopamine D1 and D2 receptor agonists and antagonists on alcohol consumption in rats th

  18. Non-Acidic Free Fatty Acid Receptor 4 Agonists with Antidiabetic Activity

    DEFF Research Database (Denmark)

    Goncalves de Azavedo, Carlos M. B. P.; Watterson, Kenneth R; Wargent, Ed T;


    The free fatty acid receptor 4 (FFA4 or GPR120) has appeared as an interesting potential target for the treatment of metabolic disorders. At present, most FFA4 ligands are carboxylic acids that are assumed to mimic the endogenous long-chain fatty acid agonists. Here, we report preliminary structu...

  19. Interaction of a non-peptide agonist with angiotensin II AT1 receptor mutants

    DEFF Research Database (Denmark)

    Costa-Neto, Claudio M; Miyakawa, Ayumi A; Pesquero, João B;


    To identify residues of the rat AT1A angiotensin II receptor involved with signal transduction and binding of the non-peptide agonist L-162,313 (5,7-dimethyl-2-ethyl-3-[[4-[2(n-butyloxycarbonylsulfonamido)-5-isobutyl-3-thienyl]phenyl]methyl]imidazol[4,5,6]-pyridine) we have performed ligand bindi...

  20. Differential Effects of Cannabinoid Receptor Agonist on Social Discrimination and Contextual Fear in Amygdala and Hippocampus (United States)

    Segev, Amir; Akirav, Irit


    We examined whether the cannabinoid receptor agonist WIN55,212-2 (WIN; 5 [mu]g/side) microinjected into the hippocampus or the amygdala would differentially affect memory processes in a neutral vs. an aversive task. In the aversive contextual fear task, WIN into the basolateral amygdala impaired fear acquisition/consolidation, but not retrieval.…

  1. The impact of improved glycaemic control with GLP-1 receptor agonist therapy on diabetic retinopathy. (United States)

    Varadhan, Lakshminarayanan; Humphreys, Tracy; Walker, Adrian B; Varughese, George I


    Rapid improvement in glycaemic control with GLP-1 receptor agonist (RA) therapy has been reported to be associated with significant progression of diabetic retinopathy. This deterioration is transient, and continuing GLP-1 RA treatment is associated with reversal of this phenomenon. Pre-existent maculopathy, higher grade of retinopathy and longer duration of diabetes may be risk factors for persistent deterioration.

  2. [Cardiovascular effects of GLP-1 receptor agonist treatment: focus on liraglutide]. (United States)

    Haluzík, Martin; Trachta, Pavel; Mráz, Miloš


    Cardiovascular risk reduction is the major aim of type 2 diabetes mellitus treatment. The effects of various antidiabetics on the cardiovascular complications are currently under careful scrutiny. Incretin-based therapy that utilizes the effects of glucagon-like peptide 1 (GLP-1) or stimulation of its receptor by GLP-1 receptor agonists represents one of the most promising approaches from the potential cardiovascular risk reduction point of view. Experimental studies have shown that the GLP-1 and GLP-1 agonists treatment improves endothelial function, decrease blood pressure and protects myocardium during experimentally-induced ischemia. Clinical studies with GLP-1 receptor agonists consistently show that, in addition to good antidiabetic efficacy, its long-term administration decreases blood pressure, body weight and improves circulating lipid levels while slightly increasing heart rate. In this paper, we focus on the cardiovascular effects of GLP-1 receptor agonist liraglutide. Preliminary analyses of cardiovascular complications in phase III trials with liraglutide indicate its good cardiovascular safety. A possibility of cardioprotective effects of liraglutide remains still open and is currently studied within a prospective cardiovascular trial LEADER.

  3. Identification of the first surrogate agonists for the G protein-coupled receptor GPR132

    DEFF Research Database (Denmark)

    Shehata, Mohamed A.; Christensen, Hanna Belcik; Isberg, Vignir;


    -arrestin recruitment assay, and thereby identified the first disclosed surrogate GPR132 agonist 1 with a potency of 3.4 μM. This constitutes the first available pharmacological tool for the in vitro characterization of the orphan receptor GPR132. The testing of 32 analogs furthermore identified a number of compounds...

  4. Biostructural and pharmacological studies of bicyclic analogues of the 3-isoxazolol glutamate receptor agonist ibotenic acid

    DEFF Research Database (Denmark)

    Frydenvang, Karla Andrea; Pickering, Darryl S; Greenwood, Jeremy R;


    We describe an improved synthesis and detailed pharmacological characterization of the conformationally restricted analogue of the naturally occurring nonselective glutamate receptor agonist ibotenic acid (RS)-3-hydroxy-4,5,6,7-tetrahydroisoxazolo[5,4-c]pyridine-7-carboxylic acid (7-HPCA, 5) at A...

  5. Transient receptor potential vanilloid 1 agonists as candidates for anti-inflammatory and immunomodulatory agents

    NARCIS (Netherlands)

    Tsuji, F.; Murai, M.; Oki, K.; Seki, I.; Ueda, K.; Inoue, H.; Nagelkerken, L.; Sasano, M.; Aono, H.


    We recently demonstrated that SA13353 [1-[2-(1-adamantyl)ethyl]-1-pentyl-3-[3-(4-pyridyl)propyl]urea], a novel transient receptor potential vanilloid 1 (TRPV1) agonist, inhibits TNF-α production through the activation of capsaicin-sensitive afferent neurons. In the present study, we investigated the

  6. Molecular interaction of a potent nonpeptide agonist with the chemokine receptor CCR8

    DEFF Research Database (Denmark)

    Jensen, Pia C; Nygaard, Rie; Thiele, Stefanie;


    Most nonpeptide antagonists for CC-chemokine receptors share a common pharmacophore with a centrally located, positively charged amine that interacts with the highly conserved glutamic acid (Glu) located in position 6 of transmembrane helix VII (VII:06). We present a novel CCR8 nonpeptide agonist...

  7. Glucagon-like peptide-1 receptor agonist treatment reduces beta cell mass in normoglycaemic mice

    NARCIS (Netherlands)

    Ellenbroek, J.H.; Tons, H.A.; Westerouen van Meeteren, M.J.; de Graaf, N.; Hanegraaf, M.A.; Rabelink, T.J.; Carlotti, F.; de Koning, E.J.


    AIMS/HYPOTHESIS: Incretin-based therapies improve glycaemic control in patients with type 2 diabetes. In animal models of diabetes, glucagon-like peptide-1 receptor agonists (GLP-1RAs) increase beta cell mass. GLP-1RAs are also evaluated in non-diabetic individuals with obesity and cardiovascular di

  8. Novel non-indolic melatonin receptor agonists differentially entrain endogenous melatonin rhythm and increase its amplitude

    NARCIS (Netherlands)

    Drijfhout, W.J; de Vries, J.B; Homan, E.J; Brons, H.F; Copinga, S; Gruppen, G; Beresford, I.J M; Hagan, R.M; Grol, Cor; Westerink, B.H.C.


    In this study we have examined the ability of melatonin and four synthetic melatonin receptor agonists to entrain endogenous melatonin secretion in rats, free running in constant darkness. The circadian melatonin profile was measured by trans-pineal microdialysis, which not only reveals the time of

  9. Dimerization of the D1 dopamine receptors is related with agonist and inverse agonist-induced receptor internalization

    Institute of Scientific and Technical Information of China (English)

    Yi-minTAO; Xue-junXU; Min-huaHONG; JieCHEN; Zhi-qiangCHI; Jing-genLIU


    AIM: To examine the relationship between D1 dopamine receptor dimer formation and ligand-induced receptor internalization. METHODS: FLAG-tagged D 1 dopamine receptor was transiently expressed in Sf9 cells. The cells were treated with SKF38393 or (+)butaclamol 1 μmol/L for different periods timeor at different doses for 30 min respectively. Western blot assaywas performed to assess dimer formation and flow

  10. Sigma-1 receptor agonists directly inhibit Nav1.2/1.4 channels.

    Directory of Open Access Journals (Sweden)

    Xiao-Fei Gao

    Full Text Available (+-SKF 10047 (N-allyl-normetazocine is a prototypic and specific sigma-1 receptor agonist that has been used extensively to study the function of sigma-1 receptors. (+-SKF 10047 inhibits K(+, Na(+ and Ca2+ channels via sigma-1 receptor activation. We found that (+-SKF 10047 inhibited Na(V1.2 and Na(V1.4 channels independently of sigma-1 receptor activation. (+-SKF 10047 equally inhibited Na(V1.2/1.4 channel currents in HEK293T cells with abundant sigma-1 receptor expression and in COS-7 cells, which barely express sigma-1 receptors. The sigma-1 receptor antagonists BD 1063,BD 1047 and NE-100 did not block the inhibitory effects of (+-SKF-10047. Blocking of the PKA, PKC and G-protein pathways did not affect (+-SKF 10047 inhibition of Na(V1.2 channel currents. The sigma-1 receptor agonists Dextromethorphan (DM and 1,3-di-o-tolyl-guanidine (DTG also inhibited Na(V1.2 currents through a sigma-1 receptor-independent pathway. The (+-SKF 10047 inhibition of Na(V1.2 currents was use- and frequency-dependent. Point mutations demonstrated the importance of Phe(1764 and Tyr(1771 in the IV-segment 6 domain of the Na(V1.2 channel and Phe(1579 in the Na(V1.4 channel for (+-SKF 10047 inhibition. In conclusion, our results suggest that sigma-1 receptor agonists directly inhibit Na(V1.2/1.4 channels and that these interactions should be given special attention for future sigma-1 receptor function studies.

  11. Anticonvulsant effect of AMP by direct activation of adenosine A1 receptor. (United States)

    Muzzi, Mirko; Coppi, Elisabetta; Pugliese, Anna Maria; Chiarugi, Alberto


    Purinergic neurotransmission mediated by adenosine (Ado) type 1 receptors (A1Rs) plays pivotal roles in negative modulation of epileptic seizures, and Ado is thought to be a key endogenous anticonvulsant. Recent evidence, however, indicates that AMP, the metabolic precursor of Ado, also activate A1Rs. Here, we evaluated the antiepileptic effects of AMP adopting in vitro and in vivo models of epilepsy. We report that AMP reversed the increase in population spike (PS) amplitude and the decrease in PS latency induced by a Mg(2+)-free extracellular solution in CA1 neurons of mouse hippocampal slices. The AMP effects were inhibited by the A1R antagonist DPCPX, but not prevented by inhibiting conversion of AMP into Ado, indicating that AMP inhibited per se sustained hippocampal excitatory neurotransmission by directly activating A1Rs. AMP also reduced seizure severity and mortality in a model of audiogenic convulsion. Of note, the anticonvulsant effects of AMP were potentiated by preventing its conversion into Ado and inhibited by DPCPX. When tested in a model of kainate-induced seizure, AMP prolonged latency of convulsions but had no effects on seizure severity and mortality. Data provide the first evidence that AMP is an endogenous anticonvulsant acting at A1Rs.

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


    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.

  13. Effects of the potential 5-HT7 receptor agonist AS 19 in an autoshaping learning task. (United States)

    Perez-García, Georgina S; Meneses, A


    This work aimed to evaluate further the role of 5-HT7 receptors during memory formation in an autoshaping Pavlovian/instrumental learning task. Post-training administration of the potential 5-HT7 receptor agonist AS 19 or antagonist SB-269970 enhanced memory formation or had no effect, respectively. The AS 19 facilitatory effect was reversed by SB-269970, but not by the selective 5-HT1A antagonist WAY100635. Amnesia induced by scopolamine (cholinergic antagonist) or dizocilpine (NMDA antagonist) was also reversed by AS 19. Certainly, reservations regarding the selectivity of AS 19 for 5-HT7 and other 5-HT receptors in vivo are noteworthy and, therefore, its validity for use in animal models as a pharmacological tool. Having mentioned that, it should be noticed that together these data are providing further support to the notion of the 5-HT7 receptors role in memory formation. Importantly, this 5-HT7 receptor agonist AS 19 appears to represent a step forward respect to the notion that potent and selective 5-HT7 receptor agonists can be useful in the treatment of dysfunctional memory in aged-related decline and Alzheimer's disease.

  14. De novo analysis of receptor binding affinity data of 8-ethenyl-xanthine antagonists to adenosine A1 and A2a receptors. (United States)

    Dalpiaz, A; Gessi, S; Varani, K; Borea, P A


    The receptor binding affinity data to adenosine A1 and A2a receptors of a wide series of 8-ethenyl-xanthine derivatives has been analyzed by means of the Free-Wilson model. The analysis of the individual group contributions (aij) shows the importance of the presence of an ethenyl moiety at position 8 on the xanthine ring for obtaining selective A2a antagonists. The different aij values of the substituents for the adenosine. A1 receptor do not correlate with the corresponding ones for the A2a receptor, indicating the possibility to obtain A1 and A2a selective compounds. The presence of aromatic substituents at the 8-ethenyl group, such as 3,5-(OCH3)2-phenyl, permits to obtain strongly A2a selective compounds (affinity ratio of up to 100); moreover, it appears that 8-ethenyl-xanthinic derivatives cannot have high selectivity for the adenosine A1 receptor (affinity ratio < or = 10).

  15. Presynaptic facilitatory adenosine A2A receptors mediate fade induced by neuromuscular relaxants that exhibit anticholinesterase activity. (United States)

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


    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.

  16. Effect of Deletion of the Prostaglandin EP2 Receptor on the Anabolic Response to Prostaglandin E2 and a Selective EP2 Receptor Agonist


    Choudhary, Shilpa; Alander, Cynthia; Zhan, Peili; Gao, Qi; Pilbeam, Carol; Raisz, Lawrence


    Studies using prostaglandin E receptor (EP) agonists indicate that prostaglandin (PG) E2 can have anabolic effects through both EP4 and EP2 receptors. We previously found that the anabolic response to a selective EP4 receptor agonist (EP4A, Ono Pharmaceutical) was substantially greater than to a selective EP2 receptor agonist (EP2A) in cultured murine calvarial osteoblastic cells. To further define the role of the EP2 receptor in PG-mediated effects on bone cells, we examined the effects of E...

  17. (TH)205-501, a non-catechol dopaminergic agonist, labels selectively and with high affinity dopamine D2 receptors

    Energy Technology Data Exchange (ETDEWEB)

    Closse, A.; Frick, W.; Markstein, R.; Maurer, R.; Nordmann, R.


    (TH)205-501, a non dopaminergic agonist, is presented as a ligand with high affinity (Ksub(D) approx= 1 nM) and high selectivity for dopamine receptors. pKsubi values of dopaminergic agonists derived from competition isotherms in the (TH)205-501 binding assay correlate very well with their potency in the acetylcholine release assay, which is controlled by dopamine D2 receptors. There is however no correlation with their potency stimulating aldenylate cyclase, a process controlled by dopamine D1 receptors. Thus (TH)205-501 is the first agonist ligand selective for dopamine D2 receptors. (Author).

  18. Maternal caffeine intake during gestation and lactation down-regulates adenosine A1 receptor in rat brain from mothers and neonates. (United States)

    Lorenzo, A M; León, D; Castillo, C A; Ruiz, M A; Albasanz, J L; Martín, M


    Even though caffeine can be excreted in breast milk, few studies have analyzed the effect of maternal caffeine consumption during lactation on neonatal brain. In the present work pregnant rats were treated daily with 1 g/L of caffeine in their drinking water during pregnancy and/or lactation and the effect on adenosine A(1) receptor in brains from both lactating mothers and 15 days-old neonates was assayed using radioligand binding and real time PCR assays. Mothers receiving caffeine during gestational period developed motor activation in gestational days 8-10 which was associated with a significant decrease of total adenosine A(1) receptor number (84%). A similar decrease was detected in mothers treated with caffeine during lactation (76%) and throughout gestation and lactation (73%); this was accompanied by a significant decrease in mRNA level coding adenosine A(1) receptor (28%). In male neonates, adenosine A(1) receptor was also decreased after chronic caffeine exposure during gestation (80%), lactation (76%) and gestation plus lactation (80%). In female neonates, adenosine A(1) receptor tended to decrease in response to caffeine exposure although no significant variations were found. No variation in the level of mRNA coding adenosine A(1) receptor was detected in neonates in any case. Concerning adenosine A(2A) receptor, radioligand binding assays revealed that this receptor remains unaltered in maternal and neonatal brain in response to caffeine exposure. However, caffeine consumption during gestation and lactation evoked a significant decrease in mRNA level coding A(2A) receptor (32%) in mothers' brain.

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

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


    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.

  20. Computational Prediction and Biochemical Analyses of New Inverse Agonists for the CB1 Receptor. (United States)

    Scott, Caitlin E; Ahn, Kwang H; Graf, Steven T; Goddard, William A; Kendall, Debra A; Abrol, Ravinder


    Human cannabinoid type 1 (CB1) G-protein coupled receptor is a potential therapeutic target for obesity. The previously predicted and experimentally validated ensemble of ligand-free conformations of CB1 [Scott, C. E. et al. Protein Sci. 2013 , 22 , 101 - 113 ; Ahn, K. H. et al. Proteins 2013 , 81 , 1304 - 1317] are used here to predict the binding sites for known CB1-selective inverse agonists including rimonabant and its seven known derivatives. This binding pocket, which differs significantly from previously published models, is used to identify 16 novel compounds expected to be CB1 inverse agonists by exploiting potential new interactions. We show experimentally that two of these compounds exhibit inverse agonist properties including inhibition of basal and agonist-induced G-protein coupling activity, as well as an enhanced level of CB1 cell surface localization. This demonstrates the utility of using the predicted binding sites for an ensemble of CB1 receptor structures for designing new CB1 inverse agonists.

  1. Agonist-mediated assembly of the crustacean methyl farnesoate receptor (United States)

    Kakaley, Elizabeth K. Medlock; Wang, Helen Y.; LeBlanc, Gerald A.


    The methyl farnesoate receptor (MfR) orchestrates aspects of reproduction and development such as male sex determination in branchiopod crustaceans. Phenotypic endpoints regulated by the receptor have been well-documented, but molecular interactions involved in receptor activation remain elusive. We hypothesized that the MfR subunits, methoprene-tolerant transcription factor (Met) and steroid receptor coactivator (SRC), would be expressed coincident with the timing of sex programming of developing oocytes by methyl farnesoate in daphnids. We also hypothesized that methyl farnesoate activates MfR assembly. Met mRNA was expressed rhythmically during the reproductive cycle, with peak mRNA accumulation just prior period of oocytes programming of sex. Further, we revealed evidence that Met proteins self-associate in the absence of methyl farnesoate, and that the presence of methyl farnesoate stimulates dissociation of Met multimers with subsequent association with SRC. Results demonstrated that the Met subunit is highly dynamic in controlling the action of methyl farnesoate through temporal variation in its expression and availability for receptor assembly.

  2. Studies on the pharmacology of the novel histamine H3 receptor agonist Sch 50971. (United States)

    Hey, J A; Aslanian, R; Bolser, D C; Chapman, R W; Egan, R W; Rizzo, C A; Shih, N Y; Fernandez, X; McLeod, R L; West, R; Kreutner, W


    Experiments were performed to characterize the pharmacology of Sch 50971 ((+)-trans-4-(4(R)-methyl-3(R)-pyrolidinyl)-1H-imidazole dihydrochloride, CAS 167610-28-8), a novel histamine H3 receptor agonist. The activity of Sch 50971 was compared with that of (R)-alpha-methylhistamine (CAS 75614-87-8), a potent and moderately selective agonist of histamine H3 receptors, in a series of in vitro and in vivo assays. Sch 50971 is a high affinity, selective H3 receptor agonist in vitro and in vivo. Sch 50971 inhibits [3H]-N-alpha-methylhistamine (CAS 673-50-7) binding to the histamine H3 receptor in human brain (Ki = 5.0 nmol/l) and guinea pig brain (Ki = 2.5 nmol/l). Sch 50971 also inhibits electric field stimulated guinea pig ileum contractions (pD2 = 7.47) and decreases [3H]-norepinephrine (CAS 51-41-2) release (pD2 = 7.48) from guinea pig pulmonary artery by activation of presynaptic inhibitory H3 receptors. The in vitro effects of Sch 50971 are antagonized by low concentrations of a selective H3 antagonist, thioperamide (CAS 106243-16-7). Sch 50971 has low affinity (IC50's > 10 mumol/l) for histamine H1, dopamine D1 and D2, serotonin 5-HT2 and muscarinic cholinergic receptors. It also does not exhibit histamine H2-antagonist activity. In guinea pigs and cats, Sch 50971 exhibits in vivo H3 agonist activity. Sch 50971 inhibits sympathetic hypertension evoked by stimulation of the medulla oblongata in anesthetized guinea pigs (ED30 = 0.3 mg/kg i.v., ED30 = 1.0 mg/kg i.d.). Sch 50971 also inhibits the effects of sympathetic nerve stimulation on nasal resistance in cats. In these assays, Sch 50971 exhibits an efficacy and potency comparable to H3-agonist (R)-alpha-methylhistamine. However, under in vivo conditions, Sch 50971 does not exhibit histamine H1-mediated responses that are seen with (R)-alpha-methylhistamine at doses close to those that produce H3 effects. Therefore, Sch 50971 is a novel, potent and selective agonist of histamine H3 receptors with an improved in

  3. Characterization of the 5-HT7 receptor. Determination of the pharmacophore for 5-HT7 receptor agonism and CoMFA-based modeling of the agonist binding site

    NARCIS (Netherlands)

    Vermeulen, ES; Schmidt, AW; Sprouse, JS; Wikstrom, HV; Grol, CJ


    On the basis of a set of 20 diverse 5-HT7 receptor agonists, the pharmacophore for 5-HT7 receptor agonism was determined. Additionally two CoMFA models were developed, based on different alignments of the agonists. Both models show good correlations between experimental and predictive pK(i) values a

  4. Pungency of TRPV1 agonists is directly correlated with kinetics of receptor activation and lipophilicity. (United States)

    Ursu, Daniel; Knopp, Kelly; Beattie, Ruth E; Liu, Bin; Sher, Emanuele


    TRPV1 (transient receptor potential vanilloid 1) is a ligand-gated ion channel expressed predominantly in nociceptive primary afferents that plays a key role in pain processing. In vivo activation of TRPV1 receptors by natural agonists like capsaicin is associated with a sharp and burning pain, frequently described as pungency. To elucidate the mechanisms underlying pungency we investigated a series of TRPV1 agonists that included both pungent and non-pungent compounds covering a large range of potencies. Pungency of capsaicin, piperine, arvanil, olvanil, RTX (resiniferatoxin) and SDZ-249665 was evaluated in vivo, by determining the increase in the number of eye wipes caused by direct instillation of agonist solutions into the eye. Agonist-induced calcium fluxes were recorded using the FLIPR technique in a recombinant, TRPV1-expressing cell line. Current-clamp recordings were performed in rat DRG (dorsal root ganglia) neurons in order to assess the consequences of TRPV1 activation on neuronal excitability. Using the eye wipe assay the following rank of pungency was obtained: capsaicin>piperine>RTX>arvanil>olvanil>SDZ-249665. We found a strong correlation between kinetics of calcium flux, pungency and lipophilicity of TRPV1 agonists. Current-clamp recordings confirmed that the rate of receptor activation translates in the ability of agonists to generate action potentials in sensory neurons. We have demonstrated that the lipophilicity of the compounds is directly related to the kinetics of TRPV1 activation and that the latter influences their ability to trigger action potentials in sensory neurons and, ultimately, pungency.

  5. The use of anchored agonists of phagocytic receptors for cancer immunotherapy: B16-F10 murine melanoma model.

    Directory of Open Access Journals (Sweden)

    Tereza Janotová

    Full Text Available The application of the phagocytic receptor agonists in cancer immunotherapy was studied. Agonists (laminarin, molecules with terminal mannose, N-Formyl-methioninyl-leucyl-phenylalanine were firmly anchored to the tumor cell surface. When particular agonists of phagocytic receptors were used together with LPS (Toll-like receptor agonist, high synergy causing tumour shrinkage and a temporary or permanent disappearance was observed. Methods of anchoring phagocytic receptor agonists (charge interactions, anchoring based on hydrophobic chains, covalent bonds and various regimes of phagocytic agonist/LPS mixture applications were tested to achieve maximum therapeutic effect. Combinations of mannan/LPS and f-MLF/LPS (hydrophobic anchors in appropriate (pulse regimes resulted in an 80% and 60% recovery for mice, respectively. We propose that substantial synergy between agonists of phagocytic and Toll-like receptors (TLR is based on two events. The TLR ligand induces early and massive inflammatory infiltration of tumors. The effect of this cell infiltrate is directed towards tumor cells, bearing agonists of phagocytic receptors on their surface. The result of these processes was effective killing of tumor cells. This novel approach represents exploitation of innate immunity mechanisms for treating cancer.

  6. 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)


    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.)

  7. Conformational variability of the glycine receptor M2 domain in response to activation by different agonists. (United States)

    Pless, Stephan A; Dibas, Mohammed I; Lester, Henry A; Lynch, Joseph W


    Models describing the structural changes mediating Cys loop receptor activation generally give little attention to the possibility that different agonists may promote activation via distinct M2 pore-lining domain structural rearrangements. We investigated this question by comparing the effects of different ligands on the conformation of the external portion of the homomeric alpha1 glycine receptor M2 domain. Conformational flexibility was assessed by tethering a rhodamine fluorophore to cysteines introduced at the 19' or 22' positions and monitoring fluorescence and current changes during channel activation. During glycine activation, fluorescence of the label attached to R19'C increased by approximately 20%, and the emission peak shifted to lower wavelengths, consistent with a more hydrophobic fluorophore environment. In contrast, ivermectin activated the receptors without producing a fluorescence change. Although taurine and beta-alanine were weak partial agonists at the alpha1R19'C glycine receptor, they induced large fluorescence changes. Propofol, which drastically enhanced these currents, did not induce a glycine-like blue shift in the spectral emission peak. The inhibitors strychnine and picrotoxin elicited fluorescence and current changes as expected for a competitive antagonist and an open channel blocker, respectively. Glycine and taurine (or beta-alanine) also produced an increase and a decrease, respectively, in the fluorescence of a label attached to the nearby L22'C residue. Thus, results from two separate labeled residues support the conclusion that the glycine receptor M2 domain responds with distinct conformational changes to activation by different agonists.

  8. Highly selective and potent agonists of sphingosine-1-phosphate 1 (S1P1) receptor. (United States)

    Vachal, Petr; Toth, Leslie M; Hale, Jeffrey J; Yan, Lin; Mills, Sander G; Chrebet, Gary L; Koehane, Carol A; Hajdu, Richard; Milligan, James A; Rosenbach, Mark J; Mandala, Suzanne


    Novel series of sphingosine-1-phosphate (S1P) receptor agonists were developed through a systematic SAR aimed to achieve high selectivity for a single member of the S1P family of receptors, S1P1. The optimized structure represents a highly S1P1-selective and efficacious agonist: S1P1/S1P2, S1P1/S1P3, S1P1/S1P4>10,000-fold, S1P1/S1P5>600-fold, while EC50 (S1P1) <0.2 nM. In vivo experiments are consistent with S1P1 receptor agonism alone being sufficient for achieving desired lymphocyte-lowering effect.

  9. A Novel Non-Peptidic Agonist of the Ghrelin Receptor with Orexigenic Activity In vivo (United States)

    Pastor-Cavada, Elena; Pardo, Leticia M.; Kandil, Dalia; Torres-Fuentes, Cristina; Clarke, Sarah L.; Shaban, Hamdy; McGlacken, Gerard P.; Schellekens, Harriet


    Loss of appetite in the medically ill and ageing populations is a major health problem and a significant symptom in cachexia syndromes, which is the loss of muscle and fat mass. Ghrelin is a gut-derived hormone which can stimulate appetite. Herein we describe a novel, simple, non-peptidic, 2-pyridone which acts as a selective agonist for the ghrelin receptor (GHS-R1a). The small 2-pyridone demonstrated clear agonistic activity in both transfected human cells and mouse hypothalamic cells with endogenous GHS-R1a receptor expression. In vivo tests with the hit compound showed significant increased food intake following peripheral administration, which highlights the potent orexigenic effect of this novel GHS-R1a receptor ligand.

  10. Ah receptor agonist activity in frequently consumed food items

    NARCIS (Netherlands)

    Waard, de W.J.; Aarts, J.M.M.J.G.; Peijnenburg, A.A.C.M.; Kok, de T.M.C.M.; Schooten, van F.J.; Hoogenboom, L.A.P.


    The aryl hydrocarbon receptor (AhR) receives much attention for its role in the toxicity of dioxins and dioxin-like polychlorinated biphenyls. However, many other compounds have also been reported to bind and activate AhR, of which natural food components are of special interest from a human health

  11. Selective Orthosteric Free Fatty Acid Receptor 2 (FFA2) Agonists (United States)

    Schmidt, Johannes; Smith, Nicola J.; Christiansen, Elisabeth; Tikhonova, Irina G.; Grundmann, Manuel; Hudson, Brian D.; Ward, Richard J.; Drewke, Christel; Milligan, Graeme; Kostenis, Evi; Ulven, Trond


    Free fatty acid receptor 2 (FFA2; GPR43) is a G protein-coupled seven-transmembrane receptor for short-chain fatty acids (SCFAs) that is implicated in inflammatory and metabolic disorders. The SCFA propionate has close to optimal ligand efficiency for FFA2 and can hence be considered as highly potent given its size. Propionate, however, does not discriminate between FFA2 and the closely related receptor FFA3 (GPR41). To identify FFA2-selective ligands and understand the molecular basis for FFA2 selectivity, a targeted library of small carboxylic acids was examined using holistic, label-free dynamic mass redistribution technology for primary screening and the receptor-proximal G protein [35S]guanosine 5′-(3-O-thio)triphosphate activation, inositol phosphate, and cAMP accumulation assays for hit confirmation. Structure-activity relationship analysis allowed formulation of a general rule to predict selectivity for small carboxylic acids at the orthosteric binding site where ligands with substituted sp3-hybridized α-carbons preferentially activate FFA3, whereas ligands with sp2- or sp-hybridized α-carbons prefer FFA2. The orthosteric binding mode was verified by site-directed mutagenesis: replacement of orthosteric site arginine residues by alanine in FFA2 prevented ligand binding, and molecular modeling predicted the detailed mode of binding. Based on this, selective mutation of three residues to their non-conserved counterparts in FFA3 was sufficient to transfer FFA3 selectivity to FFA2. Thus, selective activation of FFA2 via the orthosteric site is achievable with rather small ligands, a finding with significant implications for the rational design of therapeutic compounds selectively targeting the SCFA receptors. PMID:21220428

  12. Novel histamine H3-receptor antagonists and partial agonists with a non-aminergic structure


    Nickel, Tobias; Bauer, Ulrich; Schlicker, Eberhard; Kathmann, Markus; Göthert, Manfred; Sasse, Astrid; Stark, Holger; Schunack, Walter


    We determined the affinities of eight novel histamine H3-receptor ligands (ethers and carbamates) for H3-receptor binding sites and their agonistic/antagonistic effects in two functional H3-receptor models. The compounds differ from histamine in that the ethylamine chain is replaced by a propyloxy chain; in the three ethers mentioned below (FUB 335, 373 and 407), R is n-pentyl, 3-methylbutyl and 3,3-dimethylbutyl, respectively.The compounds monophasically inhibited [3H]-Nα-methylhistamine bin...

  13. LP-211 is a brain penetrant selective agonist for the serotonin 5-HT7 receptor


    Hedlund, Peter B.; Leopoldo, Marcello; Caccia, Silvio; Sarkisyan, Gor; Fracasso, Claudia; Martelli, Giuliana; Lacivita, Enza; Berardi, Francesco; Perrone, Roberto


    We have determined the pharmacological profile of the new serotonin 5-HT7 receptor agonist N-(4-cyanophenylmethyl)-4-(2-diphenyl)-1-piperazinehexanamide (LP-211). Radioligand binding assays were performed on a panel of 5-HT receptor subtypes. The compound was also evaluated in vivo by examining its effect on body temperature regulation in mice lacking the 5-HT7 receptor (5-HT7−/−) and their 5-HT7+/+ sibling controls. Disposition studies were performed in mice of both genotypes. It was found t...

  14. Halogenated cytisine derivatives as agonists at human neuronal nicotinic acetylcholine receptor subtypes. (United States)

    Slater, Y E; Houlihan, L M; Maskell, P D; Exley, R; Bermúdez, I; Lukas, R J; Valdivia, A C; Cassels, B K


    Cytisine (cy) is a potent and competitive partial agonist at alpha4 subunit-containing nicotinic acetylcholine (nACh) receptors while at homomeric alpha7-nACh receptors it behaves as a full agonist with a relatively lower potency. In the present study, we assessed the effects of bromination or iodination of the pyridone ring of cy and N-methylcytisine (N-Me-cy) on the effects of these compounds on recombinant human (h) alpha7, halpha4beta2 and halpha4beta4 nACh receptors expressed in clonal cell lines and Xenopus oocytes. Halogenation at C(3) of cy or N-Me-cy usually brings about a marked increase in both affinity and efficacy at halpha7, halpha4beta2 and halpha4beta4 nACh, the extent of which depends on whether the halogen is bromine or iodine, and upon receptor subtype. The effects of halogenation at C(5) are strongly influenced by the specific halogen substituent so that bromination causes a decrease in both affinity and efficacy while iodination decreases affinity but its effects on efficacy range from a decrease (halpha7, halpha4beta4 nACh receptors) to a marked increase (halpha4beta2 nACh receptors). Based on these findings, which differ from those showing that neither the affinity nor efficacy of nicotine, 3-(2-azetidinylmethoxy)-pyridine or epibatidine are greatly affected by halogenation, dehalogenation or halogen exchange at equivalent positions, we suggest that cy, N-Me-cy and their halo-isosteres bind to neuronal nACh receptors in a different orientation allowing the halogen atom to interact with a hydrophobic halogen-accepting region within the predominantly hydrophobic agonist-binding pocket of the receptors.

  15. Metabotropic glutamate receptor agonists potentiate a slow afterdepolarization in CNS neurons (United States)

    Zheng, F.; Gallagher, J. P.


    We have previously reported that, in the rat dorsolateral septal nucleus (DLSN), metabotropic glutamate receptor (met-GluR) agonists evoked a slow depolarization accompanied by an increase in membrane conductance and burst firing. We have speculated that the burst firing elicited by met-GluR agonists may be due to activation or enhancement of a non-specific cation current, which exists in some DLSN neurons. Now we report that a slow afterdepolarization (sADP) mediated by a non-specific cation current was potentiated by both 1S,3R-ACPD and quisqualate. In addition, met-GluR agonists unmask a sADP in DLSN neurons which did not show a sADP under control conditions. Our data suggest that a non-specific cation current can be potentiated by activation of the met-GluR.

  16. Caffeine, Through Adenosine A3 Receptor-Mediated Actions, Suppresses Amyloid-β Protein Precursor Internalization and Amyloid-β Generation. (United States)

    Li, Shanshan; Geiger, Nicholas H; Soliman, Mahmoud L; Hui, Liang; Geiger, Jonathan D; Chen, Xuesong


    Intraneuronal accumulation and extracellular deposition of amyloid-β (Aβ) protein continues to be implicated in the pathogenesis of Alzheimer's disease (AD), be it familial in origin or sporadic in nature. Aβ is generated intracellularly following endocytosis of amyloid-β protein precursor (AβPP), and, consequently, factors that suppress AβPP internalization may decrease amyloidogenic processing of AβPP. Here we tested the hypothesis that caffeine decreases Aβ generation by suppressing AβPP internalization in primary cultured neurons. Caffeine concentration-dependently blocked low-density lipoprotein (LDL) cholesterol internalization and a specific adenosine A3 receptor (A3R) antagonist as well as siRNA knockdown of A3Rs mimicked the effects of caffeine on neuronal internalization of LDL cholesterol. Further implicating A3Rs were findings that a specific A3R agonist increased neuronal internalization of LDL cholesterol. In addition, caffeine as well as siRNA knockdown of A3Rs blocked the ability of LDL cholesterol to increase Aβ levels. Furthermore, caffeine blocked LDL cholesterol-induced decreases in AβPP protein levels in neuronal plasma membranes, increased surface expression of AβPP on neurons, and the A3R antagonist as well as siRNA knockdown of A3Rs mimicked the effects of caffeine on AβPP surface expression. Moreover, the A3R agonist decreased neuronal surface expression of AβPP. Our findings suggest that caffeine exerts protective effects against amyloidogenic processing of AβPP at least in part by suppressing A3R-mediated internalization of AβPP.

  17. Cannabidiol is a partial agonist at dopamine D2High receptors, predicting its antipsychotic clinical dose (United States)

    Seeman, P


    Although all current antipsychotics act by interfering with the action of dopamine at dopamine D2 receptors, two recent reports showed that 800 to 1000 mg of cannabidiol per day alleviated the signs and symptoms of schizophrenia, although cannabidiol is not known to act on dopamine receptors. Because these recent clinical findings may indicate an important exception to the general rule that all antipsychotics interfere with dopamine at dopamine D2 receptors, the present study examined whether cannabidiol acted directly on D2 receptors, using tritiated domperidone to label rat brain striatal D2 receptors. It was found that cannabidiol inhibited the binding of radio-domperidone with dissociation constants of 11 nm at dopamine D2High receptors and 2800 nm at dopamine D2Low receptors, in the same biphasic manner as a dopamine partial agonist antipsychotic drug such as aripiprazole. The clinical doses of cannabidiol are sufficient to occupy the functional D2High sites. it is concluded that the dopamine partial agonist action of cannabidiol may account for its clinical antipsychotic effects. PMID:27754480

  18. Biological effects of exogenous adenosine 5 prime -triphosphate on cultured mammalian cells: Evidence for a receptor mechanism and its regulation by desensitization

    Energy Technology Data Exchange (ETDEWEB)

    Gonzalez, F.A.


    Exogenous adenosine 5{prime}-triphosphate (ATP) mobilized intracellular calcium in human carcinoma A43l cells and in Swiss 3T3 and 3T6 mouse fibroblasts by increasing inositol trisphosphate similar to well down growth factors (platelet-derived growth factor (PDGF), epidermal growth factor (EGF), bradykinin (BK), serum). Calcium mobilization was examined by video imaging of fura-2 fluorescence is single cells, following the radioactive isotope {sup 45}Ca, and monitoring the decrease in fluorescence of cells loaded with chlortetracycline. Uridine 5{prime}-triphosphate, but not other nucleotides, mimicked ATP. Single-cell analysis revealed synchronous responses in 10 sec to ATP, BK or serum, while PDGF (3T3) and EGF (A431) produced slower signals with significant cell-to-cell variation. PDGF desensitized 3T3 cells to ATP and BK added 100 sec later but ATP or BK did not desensitized to PDGF. Homologous desensitization was seen with all agonists. Heterologous desensitization was also observed in A431 cells where ATP desensitized to serum, but serum did not to ATP. ATP-stimulated calcium entry was detected after 10 sec in A431 cells, but not in Swiss 3T6 cells. Entry started before significant efflux had occurred and did not fit the capacitance model of Putney. A 2-3 hr ATP pretreatment produced a homologous desensitization state that required 20 hr to disappear, probably due to down-regulation of the putative ATP receptors.

  19. 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:


    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.

  20. Ghrelin receptor inverse agonists: identification of an active peptide core and its interaction epitopes on the receptor

    DEFF Research Database (Denmark)

    Holst, Birgitte; Lang, Manja; Brandt, Erik


    (TM)-III, TM-VI and TM-VII]. In contrast, the inverse agonist peptides bind in a pocket that extends all the way from the extracellular end of TM-II (AspII:20) across between TM-III and TM-VI/VII to TM-V and TM-IV. The potency of the main inverse agonist could be improved up to 20-fold by a number...... of space-generating mutants located relatively deep in the binding pocket at key positions in TM-III, TM-IV and TM-V. It is proposed that the inverse agonists prevent the spontaneous receptor activation by inserting relatively deeply across the main ligand-binding pocket and sterically blocking...

  1. 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.

  2. Blunted dynamics of adenosine A2A receptors is associated with increased susceptibility to Candida albicans infection in the elderly (United States)

    Rodrigues, Lisa; Miranda, Isabel M.; Andrade, Geanne M.; Mota, Marta; Cortes, Luísa; Rodrigues, Acácio G.; Cunha, Rodrigo A.; Gonçalves, Teresa


    Opportunistic gut infections and chronic inflammation, in particular due to overgrowth of Candida albicans present in the gut microbiota, are increasingly reported in the elder population. In aged, adult and young mice, we now compared the relative intestinal over-colonization by ingested C. albicans and their translocation to other organs, focusing on the role of adenosine A2A receptors that are a main stop signal of inflammation. We report that elderly mice are more prone to over-colonization by C. albicans than adult and young mice. This fungal over-growth seems to be related with higher growth rate in intestinal lumen, independent of gut tissues invasion, but resulting in higher GI tract inflammation. We observed a particularly high colonization of the stomach, with increased rate of yeast-to-hypha transition in aged mice. We found a correlation between A2A receptor density and tissue damage due to yeast infection: comparing with young and adults, aged mice have a lower gut A2A receptor density and C. albicans infection failed to increase it. In conclusion, this study shows that aged mice have a lower ability to cope with inflammation due to C. albicans over-colonization, associated with an inability to adaptively adjust adenosine A2A receptors density. PMID:27590517

  3. Distinct Signaling Cascades Elicited by Different Formyl Peptide Receptor 2 (FPR2 Agonists

    Directory of Open Access Journals (Sweden)

    Fabio Cattaneo


    Full Text Available The formyl peptide receptor 2 (FPR2 is a remarkably versatile transmembrane protein belonging to the G-protein coupled receptor (GPCR family. FPR2 is activated by an array of ligands, which include structurally unrelated lipids and peptide/proteins agonists, resulting in different intracellular responses in a ligand-specific fashion. In addition to the anti-inflammatory lipid, lipoxin A4, several other endogenous agonists also bind FPR2, including serum amyloid A, glucocorticoid-induced annexin 1, urokinase and its receptor, suggesting that the activation of FPR2 may result in potent pro- or anti-inflammatory responses. Other endogenous ligands, also present in biological samples, include resolvins, amyloidogenic proteins, such as beta amyloid (Aβ-42 and prion protein (Prp106–126, the neuroprotective peptide, humanin, antibacterial peptides, annexin 1-derived peptides, chemokine variants, the neuropeptides, vasoactive intestinal peptide (VIP and pituitary adenylate cyclase activating polypeptide (PACAP-27, and mitochondrial peptides. Upon activation, intracellular domains of FPR2 mediate signaling to G-proteins, which trigger several agonist-dependent signal transduction pathways, including activation of phospholipase C (PLC, protein kinase C (PKC isoforms, the phosphoinositide 3-kinase (PI3K/protein kinase B (Akt pathway, the mitogen-activated protein kinase (MAPK pathway, p38MAPK, as well as the phosphorylation of cytosolic tyrosine kinases, tyrosine kinase receptor transactivation, phosphorylation and nuclear translocation of regulatory transcriptional factors, release of calcium and production of oxidants. FPR2 is an attractive therapeutic target, because of its involvement in a range of normal physiological processes and pathological diseases. Here, we review and discuss the most significant findings on the intracellular pathways and on the cross-communication between FPR2 and tyrosine kinase receptors triggered by different FPR2

  4. Classification of 5-HT1A receptor agonists and antagonists using GA-SVM method

    Institute of Scientific and Technical Information of China (English)

    Xue-lian ZHU; Hai-yan CAI; Zhi-jian XU; Yong WANG; He-yao WANG; Ao ZHANG; Wei-liang ZHU


    Aim:To construct a reliable computational model for the classification of agonists and antagonists of 5-HT1A receptor.Methods:Support vector machine (SVM),a well-known machine learning method,was employed to build a prediction model,and genetic algorithm (GA) was used to select the most relevant descriptors and to optimize two important parameters,C and r of the SVM model.The overall dataset used in this study comprised 284 ligands of the 5-HT1A receptor with diverse structures reported in the literatures.Results:A SVM model was successfully developed that could be used to predict the probability of a ligand being an agonist or antagonist of the 5-HT1A receptor.The predictive accuracy for training and test sets was 0.942 and 0.865,respectively.For compounds with probability estimate higher than 0.7,the predictive accuracy of the model for training and test sets was 0.954 and 0.927,respectively.To further validate our model,the receiver operating characteristic (ROC) curve was plotted,and the Area-Under-the-ROC-Curve (AUC) value was calculated to be 0.883 for training set and 0.906 for test set.Conclusion:A reliable SVM model was successfully developed that could effectively distinguish agonists and antagonists among the ligands of the 5-HT1A receptor.To our knowledge,this is the first effort for the classification of 5-HT1A receptor agonists and antagonists based on a diverse dataset.This method may be used to classify the ligands of other members of the GPCR family.

  5. Functional potencies of dopamine agonists and antagonists at human dopamine D₂ and D₃ receptors. (United States)

    Tadori, Yoshihiro; Forbes, Robert A; McQuade, Robert D; Kikuchi, Tetsuro


    We measured the functional agonist potencies of dopamine agonists including antiparkinson drugs, and functional antagonist potencies of antipsychotics at human dopamine D(2) and D(3) receptors. In vitro pharmacological assessment included inhibition of forskolin-stimulated cAMP accumulation and the reversal of dopamine-induced inhibition in clonal Chinese hamster ovary cells expressing low and high densities of human dopamine D(2L) and D(2S) receptors (hD(2L)-Low, hD(2L)-High, hD(2S)-Low and hD(2S)-High, respectively) and human dopamine D(3) Ser-9 and D(3) Gly-9 receptors (hD(3)-Ser-9 and hD(3)-Gly-9, respectively). Cabergoline, bromocriptine, pergolide, (±)-7-hydroxy-N,N-di-n-propyl-2-aminotetralin (7-OH-DPAT), talipexole, pramipexole, R-(+)-trans-3,4,4a,10b-tetrahydro-4-propyl-2H,5H-[1]benzopyrano[4,3-b]-1,4-oxazin-9-olhydrochloride (PD128907) and ropinirole behaved as dopamine D(2) and D(3) receptor full agonists and showed higher potencies in hD(2L)-High and hD(2S)-High compared to hD(2L)-Low and hD(2S)-Low. In hD(3)-Ser-9 and hD(3)-Gly-9 compared to hD(2L)-Low and hD(2S)-Low, dopamine, ropinirole, PD128907, and pramipexole potencies were clearly higher; talipexole and 7-OH-DPAT showed slightly higher potencies; pergolide showed slightly lower potency; and, cabergoline and bromocriptine potencies were lower. Aripiprazole acted as an antagonist in hD(2L)-Low; a low intrinsic activity partial agonist in hD(2S)-Low; a moderate partial agonist in hD(3)-Ser-9 and hD(3)-Gly-9; a robust partial agonist in hD(2L)-High; and a full agonist in hD(2S)-High. Amisulpride, sulpiride and perphenazine behaved as preferential antagonists; and chlorpromazine and asenapine behaved as modest preferential antagonists; whereas fluphenazine, haloperidol, and blonanserin behaved as non-preferential antagonists in hD(2S)-Low and hD(2S)-High compared to hD(3)-Ser-9 and hD(3)-Gly-9. These findings may help to elucidate the basis of therapeutic benefit observed with these drugs, with

  6. Metabotropic glutamate receptor agonists modify the pyloric output of the crustacean stomatogastric ganglion. (United States)

    Pérez-Acevedo, Nivia L; Krenz, Wulf D


    We have studied the effects of groups I, II, and III metabotropic glutamate receptor (mGluR) agonists and antagonists on pyloric activity in the stomatogastric ganglion (STG) of the Caribbean spiny lobster Panulirus argus. We have found that agonists for all three groups of mGluRs modify the pyloric output. The group I agonist, l-quisqualic acid (l-QA), activated the pyloric central pattern generator (CPG). When the pyloric rhythm was partially suppressed by sucrose-block of input fibers in the stomatogastric nerve (stn), l-QA accelerated the rhythmic activity. In addition, the number of spike discharges was increased in pyloric motoneurons: pyloric (PY), and lateral pyloric (LP). In completely blocked preparations, a slow pyloric rhythm was initiated by l-QA. Groups II and III agonists exerted an inhibitory effect on pyloric activity. The group II agonist, (2S,1'S,2'S)-2-(Carboxycyclopropyl)glycine (L-CCG-I), decreased both the frequency of the pyloric rhythm and the number of spike discharges in the motoneurons: ventricular dilator (VD), PY, and LP. The effects of L-CCG-I were dose-dependent. The group III agonist, l-(+)-2-Amino-4-phosphonobutyric acid (l-AP4), slightly decreased the frequency of the pyloric rhythm and suppressed spike discharges in the VD neuron. All effects of mGluR agonists were reversible. The effect of l-QA was blocked by the broad spectrum mGluR antagonist (S)-Methyl-4-carboxyphenylglycine (MCPG). The inhibitory effect of L-CCG-I was prevented by MCPG and by the group II/III mGluR antagonist (RS)-alpha-Methyl-4-phosphonophenylglycine (MPPG), and was partially blocked by the group II mGluR antagonist (RS)-1-amino-5-phosphonoindan-1-carboxylic acid (APICA). The inhibitory effect of l-AP4 was blocked by MPPG and partially blocked by APICA.

  7. Modulation Effect of Peroxisome Proliferator-Activated Receptor Agonists on Lipid Droplet Proteins in Liver. (United States)

    Zhu, Yun-Xia; Zhang, Ming-Liang; Zhong, Yuan; Wang, Chen; Jia, Wei-Ping


    Peroxisome proliferator-activated receptor (PPAR) agonists are used for treating hyperglycemia and type 2 diabetes. However, the mechanism of action of these agonists is still under investigation. The lipid droplet-associated proteins FSP27/CIDEC and LSDP5, regulated directly by PPARγ and PPARα, are associated with hepatic steatosis and insulin sensitivity. Here, we evaluated the expression levels of FSP27/CIDEC and LSDP5 and the regulation of these proteins by consumption of a high-fat diet (HFD) or administration of PPAR agonists. Mice with diet-induced obesity were treated with the PPARγ or PPARα agonist, pioglitazone or fenofibrate, respectively. Liver tissues from db/db diabetic mice and human were also collected. Interestingly, FSP27/CIEDC was expressed in mouse and human livers and was upregulated in obese C57BL/6J mice. Fenofibrate treatment decreased hepatic triglyceride (TG) content and FSP27/CIDEC protein expression in mice fed an HFD diet. In mice, LSDP5 was not detected, even in the context of insulin resistance or treatment with PPAR agonists. However, LSDP5 was highly expressed in humans, with elevated expression observed in the fatty liver. We concluded that fenofibrate greatly decreased hepatic TG content and FSP27/CIDEC protein expression in mice fed an HFD, suggesting a potential regulatory role for fenofibrate in the amelioration of hepatic steatosis.

  8. Differentiation of δ, μ, and κ opioid receptor agonists based on pharmacophore development and computed physicochemical properties (United States)

    Filizola, Marta; Villar, Hugo O.; Loew, Gilda H.


    Compounds that bind with significant affinity to the opioid receptor types, δ, μ, and κ, with different combinations of activation and inhibition at these three receptors could be promising behaviorally selective agents. Working on this hypothesis, the chemical moieties common to three different sets of opioid receptor agonists with significant affinity for each of the three receptor types δ, μ, or κ were identified. Using a distance analysis approach, common geometric arrangements of these chemical moieties were found for selected δ, μ, or κ opioid agonists. The chemical and geometric commonalities among agonists at each opioid receptor type were then compared with a non-specific opioid recognition pharmacophore recently developed. The comparison provided identification of the additional requirements for activation of δ, μ, and κ opioid receptors. The distance analysis approach was able to clearly discriminate κ-agonists, while global molecular properties for all compounds were calculated to identify additional requirements for activation of δ and μ receptors. Comparisons of the combined geometric and physicochemical properties calculated for each of the three sets of agonists allowed the determination of unique requirements for activation of each of the three opioid receptors. These results can be used to improve the activation selectivity of known opioid agonists and as a guide for the identification of novel selective opioid ligands with potential therapeutic usefulness.

  9. Peroxisome Proliferator-Activated Receptor Agonists Modulate Neuropathic Pain: a Link to Chemokines?

    Directory of Open Access Journals (Sweden)

    Caroline eFreitag


    Full Text Available Chronic pain presents a widespread and intractable medical problem. While numerous pharmaceuticals are used to treat chronic pain, drugs that are safe for extended use and highly effective at treating the most severe pain do not yet exist. Chronic pain resulting from nervous system injury (neuropathic pain is common in conditions ranging from multiple sclerosis to HIV-1 infection to type II diabetes. Inflammation caused by neuropathy is believed to contribute to the generation and maintenance of neuropathic pain. Chemokines are key inflammatory mediators, several of which (MCP-1, RANTES, MIP-1α, fractalkine, SDF-1 among others have been linked to chronic, neuropathic pain in both human conditions and animal models. The important roles chemokines play in inflammation and pain make them an attractive therapeutic target. Peroxisome proliferator-activated receptors are a family of nuclear receptors known for their roles in metabolism. Recent research has revealed that PPARs also play a role in inflammatory gene repression. PPAR agonists have wide-ranging effects including inhibition of chemokine expression and pain behavior reduction in animal models. Experimental evidence suggests a connection between PPAR agonists' pain ameliorating effects and suppression of inflammatory gene expression, including chemokines. In early clinical research, one PPARα agonist, palmitoylethanolamide, shows promise in relieving chronic pain. If this link can be better established, PPAR agonists may represent a new drug therapy for neuropathic pain.

  10. Preclinical evaluation of SMM-189, a cannabinoid receptor 2-specific inverse agonist. (United States)

    Presley, Chaela; Abidi, Ammaar; Suryawanshi, Satyendra; Mustafa, Suni; Meibohm, Bernd; Moore, Bob M


    Cannabinoid receptor 2 agonists and inverse agonists are emerging as new therapeutic options for a spectrum of autoimmune-related disease. Of particular interest, is the ability of CB2 ligands to regulate microglia function in neurodegenerative diseases and traumatic brain injury. We have previously reported the receptor affinity of 3',5'-dichloro-2,6-dihydroxy-biphenyl-4-yl)-phenyl-methanone (SMM-189) and the characterization of the beneficial effects of SMM-189 in the mouse model of mild traumatic brain injury. Herein, we report the further characterization of SMM-189 as a potent and selective CB2 inverse agonist, which acts as a noncompetitive inhibitor of CP 55,940. The ability of SMM-189 to regulate microglial activation, in terms of chemokine expression and cell morphology, has been determined. Finally, we have determined that SMM-189 possesses acceptable biopharmaceutical properties indicating that the triaryl class of CB2 inverse agonists are viable compounds for continued preclinical development for the treatment of neurodegenerative disorders and traumatic brain injury.

  11. Therapeutic Effects of Melatonin Receptor Agonists on Sleep and Comorbid Disorders

    Directory of Open Access Journals (Sweden)

    Moshe Laudon


    Full Text Available Several melatonin receptors agonists (ramelteon, prolonged-release melatonin, agomelatine and tasimelteon have recently become available for the treatment of insomnia, depression and circadian rhythms sleep-wake disorders. The efficacy and safety profiles of these compounds in the treatment of the indicated disorders are reviewed. Accumulating evidence indicates that sleep-wake disorders and co-existing medical conditions are mutually exacerbating. This understanding has now been incorporated into the new Diagnostic and Statistical Manual of Mental Disorders, 5th Edition (DSM-5. Therefore, when evaluating the risk/benefit ratio of sleep drugs, it is pertinent to also evaluate their effects on wake and comorbid condition. Beneficial effects of melatonin receptor agonists on comorbid neurological, psychiatric, cardiovascular and metabolic symptomatology beyond sleep regulation are also described. The review underlines the beneficial value of enhancing physiological sleep in comorbid conditions.

  12. Therapeutic effects of melatonin receptor agonists on sleep and comorbid disorders. (United States)

    Laudon, Moshe; Frydman-Marom, Anat


    Several melatonin receptors agonists (ramelteon, prolonged-release melatonin, agomelatine and tasimelteon) have recently become available for the treatment of insomnia, depression and circadian rhythms sleep-wake disorders. The efficacy and safety profiles of these compounds in the treatment of the indicated disorders are reviewed. Accumulating evidence indicates that sleep-wake disorders and co-existing medical conditions are mutually exacerbating. This understanding has now been incorporated into the new Diagnostic and Statistical Manual of Mental Disorders, 5th Edition (DSM-5). Therefore, when evaluating the risk/benefit ratio of sleep drugs, it is pertinent to also evaluate their effects on wake and comorbid condition. Beneficial effects of melatonin receptor agonists on comorbid neurological, psychiatric, cardiovascular and metabolic symptomatology beyond sleep regulation are also described. The review underlines the beneficial value of enhancing physiological sleep in comorbid conditions.

  13. Alpha-2 receptor agonists for the treatment of posttraumatic stress disorder. (United States)

    Belkin, Molly R; Schwartz, Thomas L


    Clonidine and guanfacine are alpha-2 receptor agonists that decrease sympathetic outflow from the central nervous system. Posttraumatic stress disorder (PTSD) is an anxiety disorder that is theorized to be related to a hyperactive sympathetic nervous system. Currently, the only US Food and Drug Administration (FDA)-approved medications for PTSD are the selective serotonin reuptake inhibitors (SSRIs) sertraline and paroxetine. Sometimes use of the SSRIs may not lead to full remission and symptoms of hyperarousal often persist. This article specifically reviews the literature on alpha-2 receptor agonist use for the treatment of PTSD and concludes that while the evidence base is limited, these agents might be considered useful when SSRIs fail to treat symptoms of agitation and hyperarousal in patients with PTSD.

  14. Immunoactive effects of cannabinoids: considerations for the therapeutic use of cannabinoid receptor agonists and antagonists. (United States)

    Greineisen, William E; Turner, Helen


    The active constituents of Cannabis sativa have been used for centuries as recreational drugs and medicinal agents. Today, marijuana is the most prevalent drug of abuse in the United States and, conversely, therapeutic use of marijuana constituents are gaining mainstream clinical and political acceptance. Given the documented contributions of endocannabinoid signaling to a range of physiological systems, including cognitive function, and the control of eating behaviors, it is unsurprising that cannabinoid receptor agonists and antagonists are showing significant clinical potential. In addition to the neuroactive effects of cannabinoids, an emerging body of data suggests that both endogenous and exogenous cannabinoids are potently immunoactive. The central premise of this review article is that the immunological effects of cannabinoids should be considered in the context of each prescribing decision. We present evidence that the immunological effects of cannabinoid receptor agonists and antagonists are highly relevant to the spectrum of disorders for which cannabinoid therapeutics are currently offered.

  15. Glucagon-like peptide-1 (GLP-1) receptor agonists, obesity and psoriasis: diabetes meets dermatology. (United States)

    Drucker, D J; Rosen, C F


    Type 2 diabetes mellitus is characterised by beta cell failure, which frequently develops in the setting of insulin resistance. Inflammation contributes to the pathophysiology of type 2 diabetes by impairing insulin action in peripheral tissues and via reduction of beta cell function. Inflammation may also play an important role in the development of complications that arise in patients with type 2 diabetes. Hence, the anti-inflammatory actions of commonly used glucose-lowering drugs may contribute, indirectly, to their mechanisms of action and therapeutic benefit. Herein we highlight the anti-inflammatory actions of glucagon-like peptide-1 (GLP-1), which exerts direct and indirect actions on immune function. The observations that GLP-1 receptor agonists exert anti-inflammatory actions in preclinical studies, taken together with case reports linking improvements in psoriasis with GLP-1 receptor agonist therapy, illustrates the emerging clinical implications of non-classical anti-inflammatory actions of incretin-based therapeutics.

  16. Future Treatment of Constipation-associated Disorders: Role of Relamorelin and Other Ghrelin Receptor Agonists (United States)

    Mosińska, Paula; Zatorski, Hubert; Storr, Martin; Fichna, Jakub


    There is an unmet need for effective pharmacological therapies for constipation, a symptom that significantly deteriorates patients’ quality of life and impacts health care. Ghrelin is an endogenous ligand for the growth hormone secretagogue receptor and has been shown to exert prokinetic effects on gastrointestinal (GI) motility via the vagus and pelvic nerves. The pharmacological potential of ghrelin is hampered by its short half-life. Ghrelin receptor (GRLN-R) agonists with enhanced pharmacokinetics were thus developed. Centrally penetrant GRLN-R agonists stimulate defecation and improve impaired lower GI transit in animals and humans. This review summarizes the current knowledge on relamorelin, a potent ghrelin mimetic, and other GRLN-R analogs which are in preclinical or clinical stages of development for the management of disorders with underlying GI hypomotility, like constipation. PMID:28238253

  17. Trafficking of α1B-adrenergic receptor mediated by inverse agonist in living cells

    Institute of Scientific and Technical Information of China (English)

    MingXU; Ying-huaGUAN; NingXU; Zhang-yiLIANG; Shu-yiWang; YaoSONG; Chi-deHAN; Xin-shengZHAO; You-yiZHANG


    AIM The project is aimed at understanding the action of inverse agonist at single molecule level and capturing the real time picture of molecular behavior of α1B-adrenergic receptor (AR) mediated by inverse agonist in living cells by single molecule detection (SMD). METHODS The location and distribution of α1B-AR was detected by laser confocal and whole cell 3H-prazosin binding assay. Dynamic imaging of BODIPY-FL-labeled prazosin (Praz), specific antagonist of (1-AR, was observed in α1B-AR stably expressed human embryonic kidney 293 (HEK293) living cells. The detection of real-time dynamic behaviors of AR was achieved by using fluorescence-labeled AR and its ligand combined with SMD techniques. RESULTS α1B-AR was predominantly distributed on the cell surface and 8.2% of the total receptors were located in cytosol.

  18. Treatment of type 2 diabetes with glucagon-like peptide-1 receptor agonists

    DEFF Research Database (Denmark)

    Hansen, K B; Knop, F K; Holst, Jens Juul;


    ongoing decline in pancreatic beta-cell function and, lastly, they have a number of side effects including hypoglycaemia and weight gain. Glucagon-like peptide-1 (GLP-1) receptor agonists are a new class of pharmacological agents, which improve glucose homeostasis in a multifaceted way. Their effects...... of hypoglycaemia with GLP-1 receptor agonists is low, the compounds have clinically relevant effects on body weight, and data are suggesting beneficial effects on cardiovascular risk factors. Exenatide was released in 2005 for the treatment of type 2 diabetes and liraglutide is expected to be approved by the Food...... and Drug Administration in US and the European Medical Agency in Europe for use in 2009. In this review, the available data on the two drugs are presented and discussed....

  19. Treatment of type 2 diabetes mellitus with agonists of the GLP-1 receptor or DPP-IV inhibitors

    DEFF Research Database (Denmark)

    Holst, Jens Juul


    analogues of the hormone (or agonists of the GLP-1 receptor) are in development, along with DPP-IV inhibitors, which have been demonstrated to protect the endogenous hormone and enhance its activity. Agonists include both albumin-bound analogues of GLP-1 and exendin-4, a lizard peptide. Clinical studies...

  20. Heterocyclic acetamide and benzamide derivatives as potent and selective beta3-adrenergic receptor agonists with improved rodent pharmacokinetic profiles. (United States)

    Goble, Stephen D; Wang, Liping; Howell, K Lulu; Bansal, Alka; Berger, Richard; Brockunier, Linda; DiSalvo, Jerry; Feighner, Scott; Harper, Bart; He, Jiafang; Hurley, Amanda; Hreniuk, Donna; Parmee, Emma; Robbins, Michael; Salituro, Gino; Sanfiz, Anthony; Streckfuss, Eric; Watkins, Eloisa; Weber, Ann E; Struthers, Mary; Edmondson, Scott D


    A series of amide derived beta(3)-adrenergic receptor (AR) agonists is described. The discovery and optimization of several series of compounds derived from 1, is used to lay the SAR foundation for second generation beta(3)-AR agonists for the treatment of overactive bladder.

  1. 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


    [(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

  2. A Robotic BG1Luc Reporter Assay to Detect Estrogen Receptor Agonists


    Stoner, Matthew A.; Yang, Chun Z.; Bittner, George D.


    Endocrine disrupting chemicals with estrogenic activity (EA) have been associated with various adverse health effects. US agencies (ICCVAM/NICEATM) tasked to assess in vitro transcription activation assays to detect estrogenic receptor (ER) agonists for EA have recently validated a BG1Luc assay in manual format, but prefer robotic formats. We have developed a robotic BG1Luc EA assay to detect EA that demonstrated 100% concordance with ICCVAM meta-analyses and ICCVAM BG1Luc results in manual f...

  3. An Alpha-1A Adrenergic Receptor Agonist Prevents Acute Doxorubicin Cardiomyopathy in Male Mice (United States)

    Montgomery, Megan D.; Chan, Trevor; Swigart, Philip M.; Myagmar, Bat-erdene; Dash, Rajesh; Simpson, Paul C.


    Alpha-1 adrenergic receptors mediate adaptive effects in the heart and cardiac myocytes, and a myocyte survival pathway involving the alpha-1A receptor subtype and ERK activation exists in vitro. However, data in vivo are limited. Here we tested A61603 (N-[5-(4,5-dihydro-1H-imidazol-2-yl)-2-hydroxy-5,6,7,8-tetrahydronaphthalen-1-yl]methanesulfonamide), a selective imidazoline agonist for the alpha-1A. A61603 was the most potent alpha-1-agonist in activating ERK in neonatal rat ventricular myocytes. A61603 activated ERK in adult mouse ventricular myocytes and protected the cells from death caused by the anthracycline doxorubicin. A low dose of A61603 (10 ng/kg/d) activated ERK in the mouse heart in vivo, but did not change blood pressure. In male mice, concurrent subcutaneous A61603 infusion at 10 ng/kg/d for 7 days after a single intraperitoneal dose of doxorubicin (25 mg/kg) increased survival, improved cardiac function, heart rate, and cardiac output by echocardiography, and reduced cardiac cell necrosis and apoptosis and myocardial fibrosis. All protective effects were lost in alpha-1A-knockout mice. In female mice, doxorubicin at doses higher than in males (35–40 mg/kg) caused less cardiac toxicity than in males. We conclude that the alpha-1A-selective agonist A61603, via the alpha-1A adrenergic receptor, prevents doxorubicin cardiomyopathy in male mice, supporting the theory that alpha-1A adrenergic receptor agonists have potential as novel heart failure therapies. PMID:28081170

  4. The Melatonin Receptor Agonist Ramelteon Effectively Treats Insomnia and Behavioral Symptoms in Autistic Disorder


    Kentaro Kawabe; Fumie Horiuchi; Yasunori Oka; Shu-ichi Ueno


    Children with autism spectrum disorders (ASD), including autistic disorder, frequently suffer from comorbid sleep problems. An altered melatonin rhythm is considered to underlie the impairment in sleep onset and maintenance in ASD. We report three cases with autistic disorder in whom nocturnal symptoms improved with ramelteon, a selective melatonin receptor agonist. Insomnia and behavior, assessed using the Clinical Global Impression-Improvement Scale, improved in two cases with 2 mg ramelteo...

  5. Alpha-2 receptor agonists for the treatment of posttraumatic stress disorder


    Belkin, Molly R; Schwartz, Thomas L.


    Clonidine and guanfacine are alpha-2 receptor agonists that decrease sympathetic outflow from the central nervous system. Posttraumatic stress disorder (PTSD) is an anxiety disorder that is theorized to be related to a hyperactive sympathetic nervous system. Currently, the only US Food and Drug Administration (FDA)-approved medications for PTSD are the selective serotonin reuptake inhibitors (SSRIs) sertraline and paroxetine. Sometimes use of the SSRIs may not lead to full remission and sympt...

  6. Lineage-dependent effects of aryl hydrocarbon receptor agonists contribute to liver tumorigenesis


    Harrill, Joshua A.; Bethany B Parks; Wauthier, Eliane; Rowlands, J. Craig; Reid, Lola M.; Thomas, Russell S.


    Rodent cancer bioassays indicate that the aryl hydrocarbon receptor (AHR) agonist, 2,3,7,8-tetracholorodibenzo-p-dioxin (TCDD), causes increases in both hepatocytic and cholangiocytic tumors. Effects of AHR activation have been evaluated on rodent hepatic stem cells (rHpSCs) versus their descendants, hepatoblasts (rHBs), two lineage stages of multipotent, hepatic precursors with overlapping but also distinct phenotypic traits. This was made possible by defining the first successful culture co...

  7. Effects of 5-HT4 receptor agonists and antagonists in learning. (United States)

    Meneses, A; Hong, E


    In the present work, the effects of pre- or post-training (ip) injection of BIMU1 and BIMU8 (5-HT4 agonists) were figured out in the autoshaping learning task. Furthermore, the post-training effects of these agonists after treatment with SDZ 205-557 and GR 125487D (5-HT4 antagonists) or p-Chloroamphetamine (PCA) were also explored. Animals were individually trained in a lever-press response on the autoshaping task and 24 hours later were tested. The results showed that pre-training injection of BIMU1 (5 20 mg/Kg) or BIMU8 (20 mg/Kg) increased the CR; in contrast, the post-training administration of BIMU1 (10-20 mg/Kg) or BIMU8 (5 and 20 mg/Kg) decreased it. Further experiments revealed that the post-training injections of SDZ 205-557 (1.0-10.0 mg/Kg) or GR 125487D (0.39-1.56 mg/Kg) by themselves did not alter the CR. When BIMU1 or BIMU8 was administered to rats pretreated with SDZ 205-557 (10 mg/Kg) or GR 125487D (0.78 mg/Kg), the decrement induced by 5-HT4 the agonists was reversed; in contrast, the administration of PCA failed to modify the CR or the agonist-induced responses. The findings showed that the pre-training stimulation of 5-HT4 receptors enhanced the acquisition of CR, while, post-training activation of 5-HT4 receptors, impaired the consolidation of learning. The latter effect was not altered by PCA pretreatment. The data show that 5-HT4 receptors are involved in the acquisition and consolidation of learning. It seems that postsynaptic 5-HT4 receptors are involved in the latter effect.

  8. Presynaptic Adenosine Receptor-Mediated Regulation of Diverse Thalamocortical Short-Term Plasticity in the Mouse Whisker Pathway. (United States)

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


    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.

  9. 1,2,4-Triazolo[1,5-a]quinoxaline derivatives: synthesis and biological evaluation as adenosine receptor antagonists. (United States)

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


    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.

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

    Directory of Open Access Journals (Sweden)

    Giovanni eFerrati


    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.

  11. Effects of dopamine D(2)-like receptor agonists in mice trained to discriminate cocaine from saline: influence of feeding condition. (United States)

    Collins, Gregory T; Jackson, Jonathan A; Koek, Wouter; France, Charles P


    In rats, the discriminative stimulus effects of direct- and indirect-acting dopamine receptor agonists are mediated by multiple dopamine receptor subtypes and the relative contribution of dopamine D2 and D3 receptors to these effects varies as a function of feeding condition. In these studies, free-fed and food-restricted mice were trained to discriminate 10.0mg/kg cocaine using a two-lever discrimination procedure in which responding was maintained by food. Both groups of mice acquired the discrimination; however, free-fed mice responded at lower rates than food-restricted mice. Dopamine D3 receptor agonists, pramipexole and quinpirole, increased cocaine-appropriate responding (>85%) in food-restricted, but not in free-fed mice. The dopamine D2 receptor agonist, sumanirole, and the nonselective dopamine receptor agonist, apomorphine, failed to increase cocaine-appropriate responding in either group. Free-fed mice were more sensitive than food-restricted mice to the rate-decreasing effects of dopamine receptor agonists and these effects could not be overcome by increasing the magnitude of reinforcement. Because feeding condition did not alter quinpirole-induced hypothermia, it is unlikely that differences in the discriminative stimulus or rate-decreasing effects of dopamine D2-like receptor agonists were due to differences in the pharmacokinetic properties of the drugs. Although these results suggest that the discriminative stimulus effects of cocaine are mediated by both dopamine D2 and D3 receptors in food-restricted mice, the increased sensitivity of free-fed mice to the rate-decreasing effects of dopamine D2-like receptor agonists limited conclusions about the impact of feeding conditions on the relative contribution of dopamine D2 and D3 receptors to the discriminative stimulus effects of cocaine.

  12. N-terminal galanin-(1-16) fragment is an agonist at the hippocampal galanin receptor

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    Fisone, G.; Berthold, M.; Bedecs, K.; Unden, A.; Bartfai, T.; Bertorelli, R.; Consolo, S.; Crawley, J.; Martin, B.; Nilsson, S.; (Univ. of Stockholm (Sweden))


    The galanin N-terminal fragment (galanin-(1-16)) has been prepared by solid-phase synthesis and by enzymic cleavage of galanin by endoproteinase Asp-N. This peptide fragment displaced {sup 125}I-labeled galanin in receptor autoradiography experiments on rat forebrain and spinal cord and in equilibrium binding experiments from high-affinity binding sites in the ventral hippocampus with an IC50 of approximately 3 nM. In tissue slices of the same brain area, galanin-(1-16), similarly to galanin, inhibited the muscarinic agonist-stimulated breakdown of inositol phospholipids. Upon intracerebroventricular administration, galanin-(1-16) (10 micrograms/15 microliters) also inhibited the scopolamine (0.3 mg/kg, s.c.)-evoked release of acetylcholine, as studied in vivo by microdialysis. Substitution of (L-Trp2) for (D-Trp2) resulted in a 500-fold loss in affinity as compared with galanin-(1-16). It is concluded that, in the ventral hippocampus, the N-terminal galanin fragment (galanin-(1-16)) is recognized by the galanin receptors controlling acetylcholine release and muscarinic agonist-stimulated inositol phospholipid breakdown as a high-affinity agonist and that amino acid residue (Trp2) plays an important role in the receptor-ligand interactions.

  13. Beta-Adrenergic Receptors and Mechanisms in Asthma: The New Long-Acting Beta-Agonists

    Directory of Open Access Journals (Sweden)

    Robert G Townley


    Full Text Available The objective is to review β-adrenergic receptors and mechanisms in the immediate and late bronchial reaction in asthma and the new long-acting β-agonist. This will be discussed in light of the controversy of the potential adverse effect of regular use of long-acting β-agonists. We studied the effect of formoterol on the late asthmatic response (LAR and airway inflammation in guinea-pigs. Formoterol suppressed the LAR, antigen-induced airway inflammation and hyperresponsiveness, although isoproterenol failed to inhibit these parameters. β-Adrenergic hyporesponsiveness, and cholinergic and a- adrenergic hyperresponsiveness have been implicated in the pathogenesis of asthma. A decrease in β-adrenoreceptor function can result either from exogenously administered β-agonist or from exposure to allergens resulting in a late bronchial reaction. There is increasing evidence that eosinophils, macrophages, and lymphocytes which are of primary importance in the late bronchial reaction are also modulated by β2- adrenoreceptors. In functional studies of guinea-pig or human isolated trachea and lung parenchyma, PAF and certain cytokines significantly reduced the potency of isoproterenol to reverse methacholine- or histamine-induced contraction. The effect of glucocorticoids on pulmonary β-adrenergic receptors and responses suggests an important role for glucocorticoids to increase β-adrenergic receptors and responsiveness.

  14. Headache, Raynaud's syndrome and serotonin receptor agonists in systemic lupus erythematosus. (United States)

    Bernatsky, S; Pineau, C A; Lee, J L; Clarke, A E


    There are potential concerns regarding serotonin receptor agonists in SLE patients with migraine, particularly patients with concomitant Raynaud's syndrome. We estimated the prevalence of lupus-related headache and Raynaud's syndrome in the Montreal General Hospital SLE clinic cohort and evaluated the relationship between these two variables in multivariable logistic regression models, controlling for age, sex, race, SLE duration and the presence of lupus anticoagulant and antibodies to cardiolipin and beta2 glycoprotein I. We also assessed, through chart review in those individuals with both Raynaud's syndrome and migraine, a history of serotonin receptor agonist use, and any associated worsening vasospasm. Based on Systemic Lupus Activity Measure (SLAM) scores, the cumulative incidence of lupus-related headache in our sample (n = 391) was 46.1%; the prevalence of Raynaud's syndrome was 49.4%. The adjusted odds ratio (OR) for lupus-related headache and Raynaud's syndrome was 1.7 (95% CI 1.1, 2.5). In addition, there was a strong independent relationship between headache and anti-beta2 glycoprotein I antibodies (adjusted OR 5.6 [95% CI 1.8, 17.0]). The data from our chart review suggest that careful use of serotonin receptor agonists in patients with both Raynaud's syndrome and migraines may be undertaken, although caution would necessitate that these agents not be used in individuals with very severe Raynaud's (eg, digital ulcerations, and so on).

  15. Selective Human Estrogen Receptor Partial Agonists (ShERPAs) for Tamoxifen-Resistant Breast Cancer. (United States)

    Xiong, Rui; Patel, Hitisha K; Gutgesell, Lauren M; Zhao, Jiong; Delgado-Rivera, Loruhama; Pham, Thao N D; Zhao, Huiping; Carlson, Kathryn; Martin, Teresa; Katzenellenbogen, John A; Moore, Terry W; Tonetti, Debra A; Thatcher, Gregory R J


    Almost 70% of breast cancers are estrogen receptor α (ERα) positive. Tamoxifen, a selective estrogen receptor modulator (SERM), represents the standard of care for many patients; however, 30-50% develop resistance, underlining the need for alternative therapeutics. Paradoxically, agonists at ERα such as estradiol (E2) have demonstrated clinical efficacy in patients with heavily treated breast cancer, although side effects in gynecological tissues are unacceptable. A drug that selectively mimics the actions of E2 in breast cancer therapy but minimizes estrogenic effects in other tissues is a novel, therapeutic alternative. We hypothesized that a selective human estrogen receptor partial agonist (ShERPA) at ERα would provide such an agent. Novel benzothiophene derivatives with nanomolar potency in breast cancer cell cultures were designed. Several showed partial agonist activity, with potency of 0.8-76 nM, mimicking E2 in inhibiting growth of tamoxifen-resistant breast cancer cell lines. Three ShERPAs were tested and validated in xenograft models of endocrine-independent and tamoxifen-resistant breast cancer, and in contrast to E2, ShERPAs did not cause significant uterine growth.

  16. A fluorinated quinuclidine benzamide named LMA 10203 acts as an agonist of insect nicotinic acetylcholine receptors. (United States)

    Mathé-Allainmat, Monique; Bodereau-Dubois, Béatrice; Lapied, Bruno; Lebreton, Jacques; Thany, Steeve H


    In the present study, we take advantage of the fact that cockroach dorsal unpaired median neurons express different nicotinic acetylcholine receptor subtypes to demonstrate that simple quinuclidine benzamides such as the 2-fluorinated benzamide LMA 10203, could act as an agonist of cockroach α-bungarotoxin-insensitive nicotinic acetylcholine receptor subtype, called nAChR2. Indeed, 1 mM LMA 10203 induced ionic currents which were partially blocked by 0.5 μM α-bungarotoxin and methyllycaconitine and completely blocked by 5 μM mecamylamine. Moreover, the current-voltage curve revealed that the ionic current induced by LMA 10203 increased from -30 mV to +20 mV confirming that it acted as an agonist of α-bungarotoxin-insensitive nAChR2. In addition, 1 mM LMA 10203 induced a depolarization of the sixth abdominal ganglion and this neuroexcitatory activity was completely blocked by 5 μM mecamylamine. These data suggest that nAChR2 was also expressed at the postsynaptic level on the synapse between the cercal afferent nerve and the giant interneurons. Interestingly, despite LMA 10203 being an agonist of cockroach nicotinic receptors, it had a poor insecticidal activity. We conclude that LMA 10203 could be used as an interesting compound to identify specific insect nAChR subtypes.

  17. The identification of GPR3 inverse agonist AF64394; the first small molecule inhibitor of GPR3 receptor function. (United States)

    Jensen, Thomas; Elster, Lisbeth; Nielsen, Søren Møller; Poda, Suresh Babu; Loechel, Frosty; Volbracht, Christiane; Klewe, Ib Vestergaard; David, Laurent; Watson, Stephen P


    The identification of the novel and selective GPR3 inverse agonist AF64394, the first small molecule inhibitor of GPR3 receptor function, is described. Structure activity relationships and syntheses based around AF64394 are reported.

  18. Estrogen Receptor β Agonists Differentially Affect the Growth of Human Melanoma Cell Lines.

    Directory of Open Access Journals (Sweden)

    Monica Marzagalli

    Full Text Available Cutaneous melanoma is an aggressive malignancy; its incidence is increasing worldwide and its prognosis remains poor. Clinical observations indicate that estrogen receptor β (ERβ is expressed in melanoma tissues and its expression decreases with tumor progression, suggesting its tumor suppressive function. These experiments were performed to investigate the effects of ERβ activation on melanoma cell growth.Protein expression was analyzed by Western blot and immunofluorescence assays. Cell proliferation was assessed by counting the cells by hemocytometer. ERβ transcriptional activity was evaluated by gene reporter assay. Global DNA methylation was analyzed by restriction enzyme assay and ERβ isoforms were identified by qRT-PCR. We demonstrated that ERβ is expressed in a panel of human melanoma cell lines (BLM, WM115, A375, WM1552. In BLM (NRAS-mutant cells, ERβ agonists significantly and specifically inhibited cell proliferation. ERβ activation triggered its cytoplasmic-to-nuclear translocation and transcriptional activity. Moreover, the antiproliferative activity of ERβ agonists was associated with an altered expression of G1-S transition-related proteins. In these cells, global DNA was found to be hypomethylated when compared to normal melanocytes; this DNA hypomethylation status was reverted by ERβ activation. ERβ agonists also decreased the proliferation of WM115 (BRAF V600D-mutant cells, while they failed to reduce the growth of A375 and WM1552 (BRAF V600E-mutant cells. Finally, we could observe that ERβ isoforms are expressed at different levels in the various cell lines. Specific oncogenic mutations or differential expression of receptor isoforms might be responsible for the different responses of cell lines to ERβ agonists.Our results demonstrate that ERβ is expressed in melanoma cell lines and that ERβ agonists differentially regulate the proliferation of these cells. These data confirm the notion that melanoma is a

  19. Inhibitory effects of peroxisome proliferator-activated receptor γ agonists on collagen IV production in podocytes. (United States)

    Li, Yanjiao; Shen, Yachen; Li, Min; Su, Dongming; Xu, Weifeng; Liang, Xiubin; Li, Rongshan


    Peroxisome proliferator-activated receptor-γ (PPAR-γ) agonists have beneficial effects on the kidney diseases through preventing microalbuminuria and glomerulosclerosis. However, the mechanisms underlying these effects remain to be fully understood. In this study, we investigate the effects of PPAR-γ agonist, rosiglitazone (Rosi) and pioglitazone (Pio), on collagen IV production in mouse podocytes. The endogenous expression of PPAR-γ was found in the primary podocytes and can be upregulated by Rosi and Pio, respectively, detected by RT-PCR and Western blot. PPAR-γ agonist markedly blunted the increasing of collagen IV expression and extraction in podocytes induced by TGF-β. In contrast, adding PPAR-γ antagonist, GW9662, to podocytes largely prevented the inhibition of collagen IV expression from Pio treatment. Our data also showed that phosphorylation of Smad2/3 enhanced by TGF-β in a time-dependent manner was significantly attenuated by adding Pio. The promoter region of collagen IV gene contains one putative consensus sequence of Smad-binding element (SBE) by promoter analysis, Rosi and Pio significantly ameliorated TGF-β-induced SBE4-luciferase activity. In conclusion, PPAR-γ activation by its agonist, Rosi or Pio, in vitro directly inhibits collagen IV expression and synthesis in primary mouse podocytes. The suppression of collagen IV production was related to the inhibition of TGF-β-driven phosphorylation of Smad2/3 and decreased response activity of SBEs of collagen IV in PPAR-γ agonist-treated mouse podocytes. This represents a novel mechanistic support regarding PPAR-γ agonists as podocyte protective agents.

  20. Time-Dependent Protection of CB2 Receptor Agonist in Stroke.

    Directory of Open Access Journals (Sweden)

    Seong-Jin Yu

    Full Text Available Recent studies have indicated that type 2 cannabinoid receptor (CB2R agonists reduce neurodegeneration after brain injury through anti-inflammatory activity. The purpose of this study was to examine the time-dependent interaction of CB2R and inflammation in stroke brain. Adult male rats were subjected to right middle cerebral artery occlusion (MCAo. CB2R mRNA expression was significantly elevated >20 fold on day 2, peaked >40-fold on day 5, and normalized on day 10 post-stroke. Inflammatory markers IBA1 and TLR4 were significantly upregulated 15 fold until day 5 after MCAo. Because of the delayed upregulation of CB2R and IBA1, we next treated animals daily with CB2R agonist AM1241 or anti-inflammatory PPAR-γ agonist pioglitazone from 2 to 5 days after MCAo. Delayed treatment with pioglitazone significantly reduced abnormal neurological scores and body asymmetry as well as brain infarction in stroke animals. No behavioral improvement or reduction in brain infarction was found in animals receiving AM1241. Pioglitazone, but not AM1241, significantly reduced IBA1 expression in the stroke cortex, suggesting that delayed treatment with AM1241 failed to alter ischemia-mediated IBA-1 upregulation. In contrast, pretreatment with AM1241 significantly reduced brain infarction and neurological deficits. In conclusion, our data support a time-dependent neuroprotection of CB2 agonist in an animal model of stroke. Delayed post- treatment with PPAR-γ agonist induced behavioral recovery and microglial suppression; early treatment with CB2R agonist suppressed neurodegeneration in stroke animals.

  1. High constitutive signaling of the ghrelin receptor--identification of a potent inverse agonist

    DEFF Research Database (Denmark)

    Holst, Birgitte; Cygankiewicz, Adam; Jensen, Tine Halkjaer;


    Ghrelin is a GH-releasing peptide that also has an important role as an orexigenic hormone-stimulating food intake. By measuring inositol phosphate turnover or by using a reporter assay for transcriptional activity controlled by cAMP-responsive elements, the ghrelin receptor showed strong, ligand...... and appetite control. It is suggested that inverse agonists for the ghrelin receptor could be particularly interesting for the treatment of obesity.......Ghrelin is a GH-releasing peptide that also has an important role as an orexigenic hormone-stimulating food intake. By measuring inositol phosphate turnover or by using a reporter assay for transcriptional activity controlled by cAMP-responsive elements, the ghrelin receptor showed strong, ligand...... found to be a high potency (EC50 = 5.2 nm) full inverse agonist as it decreased the constitutive signaling of the ghrelin receptor down to that observed in untransfected cells. The homologous motilin receptor functioned as a negative control as it did not display any sign of constitutive activity...

  2. Inhibition of GSK3 attenuates dopamine D1 receptor agonist-induced hyperactivity in mice. (United States)

    Miller, Jonathan S; Tallarida, Ronald J; Unterwald, Ellen M


    Recent evidence suggests a critical role for the intracellular signaling protein glycogen synthase kinase-3 (GSK3) in hyperactivity associated with dopaminergic transmission. Here, we investigated whether activation of GSK3 is necessary for the expression of behaviors specifically produced by dopamine D1 receptor activation. To assess the role of GSK3 in dopamine D1 receptor-induced hyperactivity, mice were pretreated with the selective GSK3 inhibitor SB 216763 (0.25-7.5mg/kg, i.p.) or its vehicle prior to administration of the dopamine D1 receptor full-agonist SKF-82958 (1.0mg/kg, i.p.) or saline control. Inhibition of GSK3 via SB 216763 dose-dependently reduced ambulatory and stereotypic activity produced by SKF-82958. These data implicate a role for GSK3 in the behavioral manifestations associated with dopamine D1 receptor activation.

  3. Conversion of agonist site to metal-ion chelator site in the beta(2)-adrenergic receptor

    DEFF Research Database (Denmark)

    Elling, C E; Thirstrup, K; Holst, Birgitte


    in the mutant receptors not by normal catecholamine ligands but instead either by free zinc ions or by zinc or copper ions in complex with small hydrophobic metal-ion chelators. Chelation of the metal ions by small hydrophobic chelators such as phenanthroline or bipyridine protected the cells from the toxic......Previously metal-ion sites have been used as structural and functional probes in seven transmembrane receptors (7TM), but as yet all the engineered sites have been inactivating. Based on presumed agonist interaction points in transmembrane III (TM-III) and -VII of the beta(2)-adrenergic receptor......, in this paper we construct an activating metal-ion site between the amine-binding Asp-113 in TM-III-or a His residue introduced at this position-and a Cys residue substituted for Asn-312 in TM-VII. No increase in constitutive activity was observed in the mutant receptors. Signal transduction was activated...

  4. Assessment of 5-HT7 Receptor Agonists Selectivity Using Nociceptive and Thermoregulation Tests in Knockout versus Wild-Type Mice

    Directory of Open Access Journals (Sweden)

    Alex Brenchat


    Full Text Available No study has ever examined the effect of 5-HT7 receptor agonists on nociception by using 5-HT7 receptor knockout mice. Basal sensitivity to noxious heat stimuli and formalin-induced nociception in both phase I and II of the formalin test did not differ in 5-HT7 receptor knockout mice and paired wild-type controls. Similarly, there was no significant difference in basal body temperature between both genotypes. Subcutaneous administration of 5-HT7 receptor agonists AS-19 (10 mg/kg, E-57431 (10 mg/kg, and E-55888 (20 mg/kg significantly reduced formalin-induced licking/biting behavior during the phase II of the test in wild-type but not in 5-HT7 receptor knockout mice. At these active analgesic doses, none of the three 5-HT7 receptor agonists modified the basal body temperature neither in wild-type nor in 5-HT7 receptor knockout mice. However, a significant decrease in body temperature was observed at a higher dose (20 mg/kg of AS-19 and E-57431 in both genotypes. Our data strongly suggest that the 5-HT7 receptor agonists AS-19, E-57431, and E-55888 produce antinociception in the formalin test by activating 5-HT7 receptors. These results also strengthen the idea that the 5-HT7 receptor plays a role in thermoregulation, but by acting in concert with other receptors.

  5. Development of a homogeneous calcium mobilization assay for high throughput screening of mas-related gene receptor agonists

    Institute of Scientific and Technical Information of China (English)

    Rui ZHANG; Pang-ke YAN; Cai-hong ZHOU; Jia-yu LIAO; Ming-wei WANG


    Aim: To develop homogeneous calcium mobilization assay for high-throughput screening (HTS) of mas-related gene (Mrg) receptor agonists. Methods: CHO-K1 cells stably expressing the full-length MrgD receptor and a calcium-sensitive dye were used to develop an HTS assay based on intracellular calcium influx. This method was applied to large-scale screening of a library containing 8000 synthetic compounds and natural product extracts, cAMP measurements were camed out to verify the bioactivities of the hits found by the calcium mobilization assay. Similar approaches were also employed in the identification of the MrgA1 recep-tor agonists following HTS of 16 000 samples. Results: EC50 values of the positive control compounds (β-alanine for MrgD receptor and dynorphin A for MrgA1 receptor) determined by the calcium mobilization assay were consistent with those reported in the literature, and the Z' factors were 0.65 and 0.50 for MrgD and MrgA1 receptor assay, respectively. About 31 compounds for the MrgD receptor and 48 compounds for the MrgA1 receptor showing ≥20% of the maximal agonist activities found in the controls were initially identified as hits. Secondary screen- ing confirmed that 2 compounds for each receptor possessed specific agonist activities. Intracellular cAMP level measurements indicated that the 2 confirmed hits displayed the functionality of the MrgD receptor agonists. Conclusion: A series of validation studies demonstrated that the homogeneous calcium mobili-zation assay developed was highly efficient, amenable to automation and a robust tool to screen potential MrgD and MrgA1 receptor agonists. Its application may be expanded to other G-protein coupled receptors that mobilize calcium influx upon activation.

  6. Quantitative encoding of a partial agonist effect on individual opioid receptors by multi-site phosphorylation and threshold detection (United States)

    Lau, Elaine K.; Trester-Zedlitz, Michelle; Trinidad, Jonathan C.; Kotowski, Sarah J.; Krutchinsky, Andrew N.; Burlingame, Alma L.; von Zastrow, Mark


    Many drugs act as partial agonists of seven-transmembrane signaling receptors when compared to endogenous ligands. Partial agonism is well described as a 'macroscopic' property manifest at the level of physiological systems or cell populations, but it is not known whether partial agonists encode discrete regulatory information at the 'microscopic' level of individual receptors. We addressed this question by focusing on morphine, a partial agonist drug for µ-type opioid peptide receptors, and combining quantitative mass spectrometry with cell biological analysis to investigate morphine's reduced efficacy for promoting receptor endocytosis when compared to a peptide full agonist. We show that these chemically distinct ligands produce a complex, and qualitatively similar mixture of phosphorylated opioid receptor forms in intact cells. Quantitatively, however, the agonists promote markedly disproportional production of multi-site phosphorylation involving a specific Ser/Thr motif, whose modification at more than one residue is essential for efficient recruitment of the adaptor protein β-arrestin to clathrin-coated pits that mediate subsequent endocytosis of MORs. These results reveal quantitative encoding of agonist-selective endocytosis at the level of individual opioid receptors, based on the conserved biochemical principles of multi-site phosphorylation and threshold detection. PMID:21868358

  7. Gold Nanoparticles Functionalized with Peptides for Specific Affinity Aggregation Assays of Estrogen Receptors and Their Agonists

    Directory of Open Access Journals (Sweden)

    Yoshiyuki Takatsuji


    Full Text Available Nuclear receptors regulate the transcription of genes and various functions such as development, differentiation, homeostasis, and behavior by formation of complexes with ligand and co-activator. Recent findings have shown that agonists of a ligand may have a toxic effect on cellular/tissular function through improper activation of nuclear receptors. In this study, a simple assay system of hetero-complexes of three different molecules (estrogen receptor, ligand, and co-activator peptide has been developed. This assay system employs functionalized gold nanoparticles (GNPs: 15 nm in diameter. The surfaces of the GNPs were modified by a 12- or 20-amino-acid peptide that contains the sequence of co-activator for activating nuclear receptor by an agonist ligand. Owing to the affinity of the peptide, the functionalized GNPs aggregate faster when the nuclear receptor and the agonist ligand are also present. The aggregation of GNPs can be identified by shifts in adsorption spectrum, which give information about the specificity of agonist ligands. Similarly, this spectrum shift can measure concentration of known agonist ligand. This simple agonist screening will be employed as high through-put analysis (HTA in the discovery of drugs that act through nuclear receptors.

  8. Comparative receptor surface analysis of agonists for tyramine receptor which inhibit sex-pheromone production in Plodia interpunctella. (United States)

    Hirashima, A; Eiraku, T; Kuwano, E; Eto, M


    The quantitative structure-activity relationship (QSAR) of a set of 29 agonists for tyramine (TA) receptor responsible for the inhibition of sex-pheromone production in Plodia interpunctella, was analyzed using comparative receptor surface analysis (CoRSA). Using the common steric and electrostatic features of the most active members of a series of compounds, CoRSA generated a virtual receptor model, represented as points on a surface complementary to the van der Waals or Wyvill steric surface of the aligned compounds. Three-dimensional energetics descriptors were calculated from receptor surface model (RSM)/ligand interaction and these three-dimensional descriptors were used in genetic partial least squares data analysis to generate a QSAR model, giving a 3D QSAR with r(2)=0.969 for calibration and CV- r(2)=0.635 for the leave-one-out cross validation.

  9. Inverse agonist histamine H3 receptor PET tracers labelled with carbon-11 or fluorine-18. (United States)

    Hamill, Terence G; Sato, Nagaaki; Jitsuoka, Makoto; Tokita, Shigeru; Sanabria, Sandra; Eng, Waisi; Ryan, Christine; Krause, Stephen; Takenaga, Norihiro; Patel, Shil; Zeng, Zhizhen; Williams, David; Sur, Cyrille; Hargreaves, Richard; Burns, H Donald


    Two histamine H3 receptor (H3R) inverse agonist PET tracers have been synthesized and characterized in preclinical studies. Each tracer has high affinity for the histamine H3 receptor, has suitable lipophilicity, and neither is a substrate for the P-glycoprotein efflux pump. A common phenolic precursor was used to synthesize each tracer with high specific activity and radiochemical purity by an alkylation reaction using either [(11)C]MeI or [(18)F]FCD(2)Br. Autoradiographic studies in rhesus monkey and human brain slices showed that each tracer had a widespread distribution with high binding densities in frontal cortex, globus pallidus and striatum, and lower uptake in cerebellum. The specificity of this expression pattern was demonstrated by the blockade of the autoradiographic signal by either the H3R agonist R-alpha-methylhistamine or a histamine H3R inverse agonist. In vivo PET imaging studies in rhesus monkey showed rapid uptake of each tracer into the brain with the same distribution seen in the autoradiographic studies. Each tracer could be blocked by pretreatment with a histamine H3R inverse agonist giving a good specific signal. Comparison of the in vitro metabolism of each compound showed slower metabolism in human liver microsomes than in rhesus monkey liver microsomes, with each compound having a similar clearance rate in humans. The in vivo metabolism of 1b in rhesus monkey showed that at 60 min, approximately 35% of the circulating counts were due to the parent. These tracers are very promising candidates as clinical PET tracers to both study the histamine H3R system and measure receptor occupancy of H3R therapeutic compounds.

  10. Collybolide is a novel biased agonist of κ-opioid receptors with potent antipruritic activity (United States)

    Gupta, Achla; Gomes, Ivone; Bobeck, Erin N.; Fakira, Amanda K.; Massaro, Nicholas P.; Sharma, Indrajeet; Cavé, Adrien; Hamm, Heidi E.; Parello, Joseph


    Among the opioid receptors, the κ-opioid receptor (κOR) has been gaining considerable attention as a potential therapeutic target for the treatment of complex CNS disorders including depression, visceral pain, and cocaine addiction. With an interest in discovering novel ligands targeting κOR, we searched natural products for unusual scaffolds and identified collybolide (Colly), a nonnitrogenous sesquiterpene from the mushroom Collybia maculata. This compound has a furyl-δ-lactone core similar to that of Salvinorin A (Sal A), another natural product from the plant Salvia divinorum. Characterization of the molecular pharmacological properties reveals that Colly, like Sal A, is a highly potent and selective κOR agonist. However, the two compounds differ in certain signaling and behavioral properties. Colly exhibits 10- to 50-fold higher potency in activating the mitogen-activated protein kinase pathway compared with Sal A. Taken with the fact that the two compounds are equipotent for inhibiting adenylyl cyclase activity, these results suggest that Colly behaves as a biased agonist of κOR. Behavioral studies also support the biased agonistic activity of Colly in that it exhibits ∼10-fold higher potency in blocking non–histamine-mediated itch compared with Sal A, and this difference is not seen in pain attenuation by these two compounds. These results represent a rare example of functional selectivity by two natural products that act on the same receptor. The biased agonistic activity, along with an easily modifiable structure compared with Sal A, makes Colly an ideal candidate for the development of novel therapeutics targeting κOR with reduced side effects. PMID:27162327

  11. Agonist action of taurine on glycine receptors in rat supraoptic magnocellular neurones: possible role in osmoregulation. (United States)

    Hussy, N; Deleuze, C; Pantaloni, A; Desarménien, M G; Moos, F


    1. To evaluate the implication of taurine in the physiology of supraoptic neurones, we (i) investigated the agonist properties of taurine on glycine and GABAA receptors of supraoptic magnocellular neurones acutely dissociated from adult rats, using whole-cell voltage clamp, (ii) studied the effects of taurine and strychnine in vivo by extracellular recordings of supraoptic vasopressin neurones in anaesthetized rats, and (iii) measured the osmolarity-dependent release of endogenous taurine from isolated supraoptic nuclei by HPLC. 2. GABA, glycine and taurine evoked rapidly activating currents that all reversed close to the equilibrium potential for Cl-, indicating activation of Cl(-)-selective channels. Glycine-activated currents were reversibly blocked by strychnine (IC50 of 35 nM with 100 microM glycine), but were unaffected by the GABAA antagonist gabazine (1-3 microM). GABA-activated currents were reversibly antagonized by 3 microM gabazine, but not by strychnine (up to 1 microM). 3. Responses to 1 mM taurine were blocked by strychnine but not by gabazine and showed no additivity with glycine-induced currents, indicating selective activation of glycine receptors. Responses to 10 mM taurine were partially antagonized by gabazine, the residual current being blocked by strychnine. Thus, taurine is also a weak agonist of GABAA receptors. 4. In the presence of gabazine, taurine activated glycine receptors with an EC50 of 406 microM. Taurine activated at most 70% of maximal glycine currents, suggesting that it is a partial agonist of glycine receptors. 5. In vivo, locally applied strychnine (300 nM) increased and taurine (1 mM) decreased the basal electrical activity of vasopressin neurones in normally hydrated rats. The effect of strychnine was markedly more pronounced in water-loaded rats. 6. Taurine, which is concentrated in supraoptic glial cells, could be released from isolated supraoptic nuclei upon hyposmotic stimulation. Decreases in osmolarity of 15 and 30

  12. Antinociceptive interactions between Mu-opioid receptor agonists and the serotonin uptake inhibitor clomipramine in rhesus monkeys: role of Mu agonist efficacy. (United States)

    Banks, Matthew L; Rice, Kenner C; Negus, S Stevens


    Mu-opioid agonists are effective analgesics but have undesirable effects such as sedation and abuse liability that limit their clinical effectiveness. Serotonergic systems also modulate nociception, and serotonin uptake inhibitors may be useful as adjuncts to enhance analgesic effects and/or attenuate undesirable effects of mu agonists. This study examined the effects of the serotonin uptake inhibitor clomipramine on behavioral effects produced in rhesus monkeys by mu agonists with varying efficacy at mu receptors (nalbuphine morphine > methadone. In the assay of capsaicin-induced allodynia, nalbuphine produced dose-dependent antiallodynia. Clomipramine alone was inactive, but as in the assay of thermal nociception, it produced a proportion-dependent enhancement in the effects of nalbuphine. These findings suggest that serotonin uptake inhibitors can selectively enhance the antinociceptive effects of mu agonists in nonhuman primates. These effects of serotonin uptake inhibitors may depend on the proportion of the serotonin uptake inhibitor and the efficacy of the mu agonist. The greatest enhancement was observed with intermediate proportions of clomipramine in combination with the low-efficacy mu agonist nalbuphine.

  13. Agonistic autoantibodies directed against G-protein-coupled receptors and their relationship to cardiovascular diseases. (United States)

    Wallukat, Gerd; Schimke, Ingolf


    Agonistic autoantibodies (AABs) against G-protein-coupled receptor (GPCR) are present mainly in diseases of the cardiovascular system or in diseases associated with cardiovascular disturbances. The increasing knowledge about the role of autoantibodies against G-protein-coupled receptor (GPCR-AABs) as pathogenic drivers, the resulting development of strategies aimed at their removal or neutralization, and the evidenced patient benefit associated with such therapies have created the need for a summary of GPCR-AAB-associated diseases. Here, we summarize the present knowledge about GPCR-AABs in cardiovascular diseases. The identity of the GPCR-AABs and their prevalence in each of several specific cardiovascular diseases are documented. The structure of GPCR is also briefly discussed. Using this information, differences between classic agonists and GPCR-AABs in their GPCR binding and activation are presented and the resulting pathogenic consequences are discussed. Furthermore, treatment strategies that are currently under study, most of which are aimed at the removal and in vivo neutralization of GPCR-AABs, are indicated and their patient benefits discussed. In this context, immunoadsorption using peptides/proteins or aptamers as binders are introduced. The use of peptides or aptamers for in vivo neutralization of GPCR-AABs is also described. Particular attention is given to the GPCR-AABs directed against the adrenergic beta1-, beta2-, and α1-receptor as well as the muscarinic receptor M2, angiotensin II-angiotensin receptor type I, endothelin1 receptor type A, angiotensin (1-7) Mas-receptor, and 5-hydroxytryptamine receptor 4. Among the diseases associated with GPCR-AABs, special focus is given to idiopathic dilated cardiomyopathy, Chagas' cardiomyopathy, malignant and pulmonary hypertension, and kidney diseases. Relationships of GPCR-AABs are indicated to glaucoma, peripartum cardiomyopathy, myocarditis, pericarditis, preeclampsia, Alzheimer's disease, Sj

  14. A robotic BG1Luc reporter assay to detect estrogen receptor agonists. (United States)

    Stoner, Matthew A; Yang, Chun Z; Bittner, George D


    Endocrine disrupting chemicals with estrogenic activity (EA) have been associated with various adverse health effects. US agencies (ICCVAM/NICEATM) tasked to assess in vitro transcription activation assays to detect estrogenic receptor (ER) agonists for EA have recently validated a BG1Luc assay in manual format, but prefer robotic formats. We have developed a robotic BG1Luc EA assay to detect EA that demonstrated 100% concordance with ICCVAM meta-analyses and ICCVAM BG1Luc results in manual format for 27 ICCVAM test substances, i.e. no false negatives or false positives. This robotic assay also consistently assessed other, more problematic ICCVAM test substances such as clomiphene citrate, L-thyroxin, and tamoxifen. Agonist responses using this robotic BG1Luc assay were consistently inhibited by the ER antagonist ICI 182,780, confirming that agonist responses were due to binding to ERs rather than to a non-specific agonist response. This robotic assay also detected EA in complex mixtures of substances such as extracts of personal care products, plastic resins or plastic consumer products. This robotic BG1Luc assay had at least as high accuracy and greater sensitivity and repeatability when compared to its manual version or to the other ICCVAM/OECD validated assays for EA (manual BG1Luc and CERI).

  15. Novel Zn2+ Modulated GPR39 Receptor Agonists Do Not Drive Acute Insulin Secretion in Rodents.

    Directory of Open Access Journals (Sweden)

    Ola Fjellström

    Full Text Available Type 2 diabetes (T2D occurs when there is insufficient insulin release to control blood glucose, due to insulin resistance and impaired β-cell function. The GPR39 receptor is expressed in metabolic tissues including pancreatic β-cells and has been proposed as a T2D target. Specifically, GPR39 agonists might improve β-cell function leading to more adequate and sustained insulin release and glucose control. The present study aimed to test the hypothesis that GPR39 agonism would improve glucose stimulated insulin secretion in vivo. A high throughput screen, followed by a medicinal chemistry program, identified three novel potent Zn2+ modulated GPR39 agonists. These agonists were evaluated in acute rodent glucose tolerance tests. The results showed a lack of glucose lowering and insulinotropic effects not only in lean mice, but also in diet-induced obese (DIO mice and Zucker fatty rats. It is concluded that Zn2+ modulated GPR39 agonists do not acutely stimulate insulin release in rodents.

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

    DEFF Research Database (Denmark)

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


    Cerebral ischemia induces a rapid suppression of spontaneous brain rhythms prior to major alterations in ionic homeostasis. It was found in vitro during ischemia that the rapidly formed adenosine, resulting from the intracellular breakdown of ATP, may inhibit synaptic transmission via the A(1) re...

  17. Quantitative Phosphoproteomics Unravels Biased Phosphorylation of Serotonin 2A Receptor at Ser280 by Hallucinogenic versus Nonhallucinogenic Agonists* (United States)

    Karaki, Samah; Becamel, Carine; Murat, Samy; Mannoury la Cour, Clotilde; Millan, Mark J.; Prézeau, Laurent; Bockaert, Joël; Marin, Philippe; Vandermoere, Franck


    The serotonin 5-HT2A receptor is a primary target of psychedelic hallucinogens such as lysergic acid diethylamine, mescaline, and psilocybin, which reproduce some of the core symptoms of schizophrenia. An incompletely resolved paradox is that only some 5-HT2A receptor agonists exhibit hallucinogenic activity, whereas structurally related agonists with comparable affinity and activity lack such a psychoactive activity. Using a strategy combining stable isotope labeling by amino acids in cell culture with enrichment in phosphorylated peptides by means of hydrophilic interaction liquid chromatography followed by immobilized metal affinity chromatography, we compared the phosphoproteome in HEK-293 cells transiently expressing the 5-HT2A receptor and exposed to either vehicle or the synthetic hallucinogen 1-[2,5-dimethoxy-4-iodophenyl]-2-aminopropane (DOI) or the nonhallucinogenic 5-HT2A agonist lisuride. Among the 5995 identified phosphorylated peptides, 16 sites were differentially phosphorylated upon exposure of cells to DOI versus lisuride. These include a serine (Ser280) located in the third intracellular loop of the 5-HT2A receptor, a region important for its desensitization. The specific phosphorylation of Ser280 by hallucinogens was further validated by quantitative mass spectrometry analysis of immunopurified receptor digests and by Western blotting using a phosphosite specific antibody. The administration of DOI, but not of lisuride, to mice, enhanced the phosphorylation of 5-HT2A receptors at Ser280 in the prefrontal cortex. Moreover, hallucinogens induced a less pronounced desensitization of receptor-operated signaling in HEK-293 cells and neurons than did nonhallucinogenic agonists. The mutation of Ser280 to aspartic acid (to mimic phosphorylation) reduced receptor desensitization by nonhallucinogenic agonists, whereas its mutation to alanine increased the ability of hallucinogens to desensitize the receptor. This study reveals a biased phosphorylation of

  18. Muscarinic receptor agonists stimulate matrix metalloproteinase 1-dependent invasion of human colon cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Raufman, Jean-Pierre, E-mail: [Division of Gastroenterology and Hepatology, University of Maryland School of Medicine, Baltimore, MD (United States); Cheng, Kunrong; Saxena, Neeraj; Chahdi, Ahmed; Belo, Angelica; Khurana, Sandeep; Xie, Guofeng [Division of Gastroenterology and Hepatology, University of Maryland School of Medicine, Baltimore, MD (United States)


    Highlights: Black-Right-Pointing-Pointer Muscarinic receptor agonists stimulated robust human colon cancer cell invasion. Black-Right-Pointing-Pointer Anti-matrix metalloproteinase1 antibody pre-treatment blocks cell invasion. Black-Right-Pointing-Pointer Bile acids stimulate MMP1 expression, cell migration and MMP1-dependent invasion. -- Abstract: Mammalian matrix metalloproteinases (MMPs) which degrade extracellular matrix facilitate colon cancer cell invasion into the bloodstream and extra-colonic tissues; in particular, MMP1 expression correlates strongly with advanced colon cancer stage, hematogenous metastasis and poor prognosis. Likewise, muscarinic receptor signaling plays an important role in colon cancer; muscarinic receptors are over-expressed in colon cancer compared to normal colon epithelial cells. Muscarinic receptor activation stimulates proliferation, migration and invasion of human colon cancer cells. In mouse intestinal neoplasia models genetic ablation of muscarinic receptors attenuates carcinogenesis. In the present work, we sought to link these observations by showing that MMP1 expression and activation plays a mechanistic role in muscarinic receptor agonist-induced colon cancer cell invasion. We show that acetylcholine, which robustly increases MMP1 expression, stimulates invasion of HT29 and H508 human colon cancer cells into human umbilical vein endothelial cell monolayers - this was abolished by pre-incubation with atropine, a non-selective muscarinic receptor inhibitor, and by pre-incubation with anti-MMP1 neutralizing antibody. Similar results were obtained using a Matrigel chamber assay and deoxycholyltaurine (DCT), an amidated dihydroxy bile acid associated with colon neoplasia in animal models and humans, and previously shown to interact functionally with muscarinic receptors. DCT treatment of human colon cancer cells resulted in time-dependent, 10-fold increased MMP1 expression, and DCT-induced cell invasion was also blocked by pre

  19. Adenosine A(2A) receptor gene (ADORA2A) variants may increase autistic symptoms and anxiety in autism spectrum disorder. (United States)

    Freitag, Christine M; Agelopoulos, Konstantin; Huy, Ellen; Rothermundt, Matthias; Krakowitzky, Petra; Meyer, Jobst; Deckert, Jürgen; von Gontard, Alexander; Hohoff, Christa


    Autism spectrum disorders (ASDs) are heterogeneous disorders presenting with increased rates of anxiety. The adenosine A(2A) receptor gene (ADORA2A) is associated with panic disorder and is located on chromosome 22q11.23. Its gene product, the adenosine A(2A) receptor, is strongly expressed in the caudate nucleus, which also is involved in ASD. As autistic symptoms are increased in individuals with 22q11.2 deletion syndrome, and large 22q11.2 deletions and duplications have been observed in ASD individuals, in this study, 98 individuals with ASD and 234 control individuals were genotyped for eight single-nucleotide polymorphisms in ADORA2A. Nominal association with the disorder was observed for rs2236624-CC, and phenotypic variability in ASD symptoms was influenced by rs3761422, rs5751876 and rs35320474. In addition, association of ADORA2A variants with anxiety was replicated for individuals with ASD. Findings point toward a possible mediating role of ADORA2A variants on phenotypic expression in ASD that need to be replicated in a larger sample.

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

    DEFF Research Database (Denmark)

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


    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......) diets for a minimum of 10 days before telemetric blood pressure and urinary excretion measurements in metabolic cages. On the NS diet, daytime and nighttime mean arterial blood pressure (MAP) was 7-10 mmHg higher in A1R-/- than in A1R+/+ mice. HS diet did not affect the MAP in A1R-/- mice......, but the daytime and nighttime MAP of the A1R+/+ mice increased by approximately 10 mmHg, to the same level as that in the A1R-/-. On the SD diet, day- and nighttime MAP decreased by approximately 6 mmHg in both A1R-/- and A1R+/+ mice, although the MAP remained higher in A1R-/- than in A1R+/+ mice. Although plasma...

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

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


    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.

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


    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. Therapeutic potential of histamine H3 receptor agonist for the treatment of obesity and diabetes mellitus. (United States)

    Yoshimoto, Ryo; Miyamoto, Yasuhisa; Shimamura, Ken; Ishihara, Akane; Takahashi, Kazuhiko; Kotani, Hidehito; Chen, Airu S; Chen, Howard Y; Macneil, Douglas J; Kanatani, Akio; Tokita, Shigeru


    Histamine H3 receptors (H3Rs) are located on the presynaptic membranes and cell soma of histamine neurons, where they negatively regulate the synthesis and release of histamine. In addition, H3Rs are also located on nonhistaminergic neurons, acting as heteroreceptors to regulate the releases of other amines such as dopamine, serotonin, and norepinephrine. The present study investigated the effects of H3R ligands on appetite and body-weight regulation by using WT and H3R-deficient mice (H3RKO), because brain histamine plays a pivotal role in energy homeostasis. The results showed that thioperamide, an H3R inverse agonist, increases, whereas imetit, an H3R agonist, decreases appetite and body weight in diet-induced obese (DiO) WT mice. Moreover, in DiO WT mice, but not in DiO H3RKO mice, imetit reduced fat mass, plasma concentrations of leptin and insulin, and hepatic triglyceride content. The anorexigenic effects of imetit were associated with a reduction in histamine release, but a comparable reduction in histamine release with alpha-fluoromethylhistidine, an inhibitor of histamine synthesis, increased appetite. Moreover, the anorexigenic effects of imetit were independent of the melanocortin system, because imetit comparably reduced appetite in melanocortin 3 and 4 receptor-deficient mice. The results provide roles of H3Rs in energy homeostasis and suggest a therapeutic potential for H3R agonists in the treatment of obesity and diabetes mellitus.

  4. A Novel Role of Serotonin Receptor 2B Agonist as an Anti-Melanogenesis Agent

    Directory of Open Access Journals (Sweden)

    Eun Ju Oh


    Full Text Available BW723C86, a serotonin receptor 2B agonist, has been investigated as a potential therapeutic for various conditions such as anxiety, hyperphagia and hypertension. However, the functional role of BW723C86 against melanogenesis remains unclear. In this study, we investigate the effect of serotonin receptor 2B (5-HTR2B agonist on melanogenesis and elucidate the mechanism involved. BW723C86 reduced melanin synthesis and intracellular tyrosinase activity in melan-A cells and normal human melanocytes. The expression of melanogenesis-related proteins (tyrosinase, TRP-1 and TRP-2 and microphthalmia-associated transcription factor (MITF in melan-A cells decreased after BW723C86 treatment. The promoter activity of MITF was also reduced by BW723C86 treatment. The reduced level of MITF was associated with inhibition of protein kinase A (PKA and cAMP response element-binding protein (CREB activation by BW723C86 treatment. These results suggest that the serotonin agonist BW723C86 could be a potential therapeutic agent for skin hyperpigmentation disorders.

  5. β2-Adrenergic receptor agonists activate CFTR in intestinal organoids and subjects with cystic fibrosis. (United States)

    Vijftigschild, Lodewijk A W; Berkers, Gitte; Dekkers, Johanna F; Zomer-van Ommen, Domenique D; Matthes, Elizabeth; Kruisselbrink, Evelien; Vonk, Annelotte; Hensen, Chantal E; Heida-Michel, Sabine; Geerdink, Margot; Janssens, Hettie M; van de Graaf, Eduard A; Bronsveld, Inez; de Winter-de Groot, Karin M; Majoor, Christof J; Heijerman, Harry G M; de Jonge, Hugo R; Hanrahan, John W; van der Ent, Cornelis K; Beekman, Jeffrey M


    We hypothesized that people with cystic fibrosis (CF) who express CFTR (cystic fibrosis transmembrane conductance regulator) gene mutations associated with residual function may benefit from G-protein coupled receptor (GPCR)-targeting drugs that can activate and enhance CFTR function.We used intestinal organoids to screen a GPCR-modulating compound library and identified β2-adrenergic receptor agonists as the most potent inducers of CFTR function.β2-Agonist-induced organoid swelling correlated with the CFTR genotype, and could be induced in homozygous CFTR-F508del organoids and highly differentiated primary CF airway epithelial cells after rescue of CFTR trafficking by small molecules. The in vivo response to treatment with an oral or inhaled β2-agonist (salbutamol) in CF patients with residual CFTR function was evaluated in a pilot study. 10 subjects with a R117H or A455E mutation were included and showed changes in the nasal potential difference measurement after treatment with oral salbutamol, including a significant improvement of the baseline potential difference of the nasal mucosa (+6.35 mV, pCFTR activation when administered ex vivo to organoids.This proof-of-concept study suggests that organoids can be used to identify drugs that activate CFTR function in vivo and to select route of administration.

  6. Renoprotective Effects of AVE0991, a Nonpeptide Mas Receptor Agonist, in Experimental Acute Renal Injury

    Directory of Open Access Journals (Sweden)

    Lívia Corrêa Barroso


    Full Text Available Renal ischemia and reperfusion (I/R is the major cause of acute kidney injury in hospitalized patients. Mechanisms underlying reperfusion-associated injury include recruitment and activation of leukocytes and release of inflammatory mediators. In this study, we investigated the renal effects of acute administration of AVE0991, an agonist of Mas, the angiotensin-(1–7 receptor, the angiotensin-(1–7 receptor, in a murine model of renal I/R. Male C57BL/6 wild-type or Mas−/− mice were subjected to 30 min of bilateral ischemia and 24 h of reperfusion. Administration of AVE0991 promoted renoprotective effects, as seen by improvement of function, decreased tissue injury, prevention of local and remote leucocyte infiltration, and release of the chemokine, CXCL1. I/R injury was similar in WT and Mas−/− mice, suggesting that endogenous activation of this receptor does not control renal damage under baseline conditions. In conclusion, pharmacological interventions using Mas receptor agonists may represent a therapeutic opportunity for the treatment of renal I/R injury.

  7. The effects of methylmercury on motor activity are sex- and age-dependent, and modulated by genetic deletion of adenosine receptors and caffeine administration. (United States)

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


    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

  8. Selective agonists for serotonin 7 (5-HT7) receptor and their applications in preclinical models: an overview. (United States)

    Di Pilato, Pantaleo; Niso, Mauro; Adriani, Walter; Romano, Emilia; Travaglini, Domenica; Berardi, Francesco; Colabufo, Nicola A; Perrone, Roberto; Laviola, Giovanni; Lacivita, Enza; Leopoldo, Marcello


    The serotonin 7 (5-HT7) receptor was the last serotonin receptor subtype to be discovered in 1993. This receptor system has been implicated in several central nervous system (CNS) functions, including circadian rhythm, rapid eye movement sleep, thermoregulation, nociception, memory and neuropsychiatric symptoms and pathologies, such as anxiety, depression and schizophrenia. In 1999, medicinal chemistry efforts led to the identification of SB-269970, which became the gold standard selective 5-HT7 receptor antagonist, and later of various selective agonists such as AS-19, LP-44, LP-12, LP-211 and E-55888. In this review, we summarize the preclinical pharmacological studies performed using these agonists, highlighting their strengths and weaknesses. The data indicate that 5-HT7 receptor agonists can have neuroprotective effects against N-methyl-d-aspartate-induced toxicity, modulate neuronal plasticity in rats, enhance morphine-induced antinociception and alleviate hyperalgesia consecutive to nerve lesion in neuropathic animals.

  9. Investigation of the histamine H3 receptor binding site. Design and synthesis of hybrid agonists with a lipophilic side chain. (United States)

    Ishikawa, Makoto; Watanabe, Takashi; Kudo, Toshiaki; Yokoyama, Fumikazu; Yamauchi, Miki; Kato, Kazuhiko; Kakui, Nobukazu; Sato, Yasuo


    As a part of our search for novel histamine H3 receptor agonists, we designed and synthesized hybrid compounds in which the lipophilic (4'-alkylphenylthio)ethyl moiety of a novel H3 receptor agonist, 4-(2-(4'-tert-butylphenylthio)ethyl)-1H-imidazole (1), was incorporated into N(alpha)-methylhistamine, immepip, and immethridine derivatives. These hybrid compounds were expected to interact concurrently with the histamine-binding site and a putative hydrophobic region in the H3 receptor. Among them, piperidine- and pyridine-type derivatives displayed partial agonist activity, and (S)-4-(1-(1H-imidazol-4-yl)-2-(4-(trifluoromethyl)phenylthio)ethyl)piperidine (36) was identified as a potent H3 agonist. We performed computational docking studies to examine the binding mode of the agonists. The results indicated that immepip interacts with the key residues, Asp114 and Glu206, in a different manner from histamine. The binding mode of 36 to these residues is similar to that of immepip, and the lipophilic tail of 36 has an additional interaction with a hydrophobic region in transmembrane helix 6 of the receptor. These results indicated that 36 served as a useful tool for studies on receptor-agonist interactions and drug design.

  10. Influence of different histamine receptor agonists and antagonists on apomorphine-induced licking behavior in rat. (United States)

    Farzin, D; Attarzadeh, M


    The effects of different histamine receptor agonists and antagonists on apomorphine-induced licking behavior in rats were investigated. Subcutaneous (s.c.) injection of various doses of apomorphine (0. 125-1.25 mg/kg) induced licking. The licking response was counted by direct observation and recorded for a 75-min period. Intracerebroventricular (i.c.v.) or intraperitoneal (i.p.) injection of the histamine H(1) or H(2) receptor agonist, HTMT (6-[2-(4-imidazolyl)ethylamino]-N-(4-trifluoromethylphenyl) heptanecarboxamide) (50 and 100 microg per rat), or dimaprit (10 and 15 mg/kg, i.p.), respectively, potentiated apomorphine-induced licking, while the histamine H(3) receptor agonist, imetit (5 and 10 mg/kg, i.p.), reduced the licking response induced by apomorphine. Pretreatment with various histamine receptor antagonists, dexchlorpheniramine (30 and 40 mg/kg, i.p.), diphenhydramine (20, 30 and 40 mg/kg, i.p.), famotidine (30 and 40 mg/kg, s.c.) and ranitidine (20, 30 and 40 mg/kg), reduced apomorphine-induced licking, while thioperamide (5 and 10 mg/kg, i.p.) potentiated the apomorphine effect. The effects of HTMT and dimaprit were blocked by dexchlorpheniramine (20 mg/kg, i.p.) and famotidine (20 mg/kg, s.c.), respectively. The inhibitory effect elicited by imetit on apomorphine-induced licking behavior was also abolished in animals treated with thioperamide (2.5 mg/kg, i.p.). The results suggest that histaminergic mechanisms may be involved in the modulation of apomorphine-induced licking behavior.

  11. Selective agonists of retinoic acid receptors: comparative toxicokinetics and embryonic exposure. (United States)

    Arafa, H M; Elmazar, M M; Hamada, F M; Reichert, U; Shroot, B; Nau, H


    Three biologically active synthetic retinoids were investigated that bind selectively to retinoic acid receptors RARs (alpha, beta and gamma). The retinoids were previously demonstrated to have different teratogenic effects in the mouse in terms of potency and regioselectivity. The teratogenic potency rank order (alpha >beta >gamma) was found to be more or less compatible with the receptor binding affinities and transactivation potencies of the retinoid ligands to their respective receptors. The RARalpha agonist (Am580; CD336) induced a wide spectrum of malformations; CD2019 (RARbeta agonist) and especially CD437 (RARgamma agonist) produced more restricted defects. In the current study we tried to address whether the differences in teratogenic effects are solely related to binding affinity and transactivation differences or also due to differences in embryonic exposure. Therefore, transplacental kinetics of the ligands were assessed following administration of a single oral dose of 15 mg/kg of either retinoid given to NMRI mice on day 11 of gestation. Am580 was rapidly transferred to the embryo resulting in the highest embryonic exposure [embryo to maternal plasma area under the time vs concentration curve (AUC)(0-24 h )ratio (E/M) was 1.7], in accordance with its highest teratogenic potency. The low placental transfer of CD2019 (E/M of 0.3) was compatible with its lower teratogenic potential. Of major interest was the finding that the CD437, though being least teratogenic, exhibited considerable embryonic exposure (E/M of 0.6). These findings suggest that both the embryonic exposure and receptor binding transactivation selectivity are crucial determinants of the teratogenicity of these retinoid ligands.

  12. Estradiol and Estrogen Receptor Agonists Oppose Oncogenic Actions of Leptin in HepG2 Cells.

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    Minqian Shen

    Full Text Available Obesity is a significant risk factor for certain cancers, including hepatocellular carcinoma (HCC. Leptin, a hormone secreted by white adipose tissue, precipitates HCC development. Epidemiology data show that men have a much higher incidence of HCC than women, suggesting that estrogens and its receptors may inhibit HCC development and progression. Whether estrogens antagonize oncogenic action of leptin is uncertain. To investigate potential inhibitory effects of estrogens on leptin-induced HCC development, HCC cell line HepG2 cells were treated with leptin in combination with 17 β-estradiol (E2, estrogen receptor-α (ER-α selective agonist PPT, ER-β selective agonist DPN, or G protein-coupled ER (GPER selective agonist G-1. Cell number, proliferation, and apoptosis were determined, and leptin- and estrogen-related intracellular signaling pathways were analyzed. HepG2 cells expressed a low level of ER-β mRNA, and leptin treatment increased ER-β expression. E2 suppressed leptin-induced HepG2 cell proliferation and promoted cell apoptosis in a dose-dependent manner. Additionally E2 reversed leptin-induced STAT3 and leptin-suppressed SOCS3, which was mainly achieved by activation of ER-β. E2 also enhanced ERK via activating ER-α and GPER and activated p38/MAPK via activating ER-β. To conclude, E2 and its receptors antagonize the oncogenic actions of leptin in HepG2 cells by inhibiting cell proliferation and stimulating cell apoptosis, which was associated with reversing leptin-induced changes in SOCS3/STAT3 and increasing p38/MAPK by activating ER-β, and increasing ERK by activating ER-α and GPER. Identifying roles of different estrogen receptors would provide comprehensive understanding of estrogenic mechanisms in HCC development and shed light on potential treatment for HCC patients.

  13. Effects of kappa opioid receptor-selective agonists on responses of pelvic nerve afferents to noxious colorectal distension. (United States)

    Su, X; Sengupta, J N; Gebhart, G F


    The aim of this study was to examine the effects of kappa-opioid receptor selective agonists on responses of mechanosensitive afferent fibers in the pelvic nerve. Single-fiber recordings were made from pelvic nerve afferents in the decentralized S1 dorsal root of the rat. A total of 572 afferent fibers in the S1 dorsal root were identified by electrical stimulation of the pelvic nerve; 252 (44%) responded to noxious colorectal distension (CRD; 80 mmHg). Of these 252 fibers that responded to CRD, 100 were studied further. All 100 fibers gave monotonic increases in firing to increasing pressures of CRD. Eighty-eight fibers had low thresholds for response (mean: 3 mmHg) and 12 fibers had high-thresholds for response (mean: 28 mmHg). Responses of 17 fibers also were tested after instillation of 5% mustard oil (MO) into the colon. The resting activity of 16/17 fibers significantly increased after MO instillation; 13 (77%) also exhibited sensitization of responses to graded CRD when tested 30 min after intracolonic MO instillation. The effects of kappa1-opioid receptor preferring agonists (U50,488H, U69,593 and U62,066), the kappa2-opioid receptor preferring agonist bremazocine, and the kappa3-opioid receptor preferring agonist naloxone benzoylhydrazone (nalBzoH) were tested on responses of 64 mechanosensitive afferent fibers to noxious CRD. All five agonists dose-dependently inhibited afferent fiber responses to noxious CRD. Doses producing inhibition to 50% of the control response to CRD did not differ among the five agonists, ranging from approximately 4 to 15 mg/kg. The effects of kappa1, kappa2, and kappa3 receptor agonists were attenuated by naloxone; two kappa-opioid receptor-selective antagonists were ineffective. There were no differences in the dose-response relationships of these drugs for fibers recorded from untreated and irritant-treated colons. Conduction velocities of the fibers remained unaffected after high doses of all tested agonists. In an in vitro

  14. Development of betulinic acid as an agonist of TGR5 receptor using a new in vitro assay

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    Lo SH


    Full Text Available Shih-Hsiang Lo,1,2 Kai-Chung Cheng,3 Ying-Xiao Li,3,4 Chin-Hong Chang,4,5 Juei-Tang Cheng,4,6 Kung-Shing Lee7,8 1Division of Cardiology, Department of Internal Medicine, Zhongxing Branch of Taipei City Hospital, 2Department of History and Geography, University of Taipei, Taipei, Taiwan; 3Department of Psychosomatic Internal Medicine, Kagoshima University Graduate School of Medical and Dental Sciences, Kagoshima, Japan; 4Department of Medical Research, 5Department of Neurosurgery, Chi-Mei Medical Center, Yong Kang, 6Institute of Medical Science, College of Health Science, Chang Jung Christian University, Tainan, 7Department of Surgery, Pingtung Hospital, 8Division of Neurosurgery, Department of Surgery, Kaohsiung Medical University Chung-Ho Memorial Hospital, School of Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan Background: G-protein-coupled bile acid receptor 1, also known as TGR5 is known to be involved in glucose homeostasis. In animal models, treatment with a TGR5 agonist induces incretin secretion to reduce hyperglycemia. Betulinic acid, a triterpenoid present in the leaves of white birch, has been introduced as a selective TGR5 agonist. However, direct activation of TGR5 by betulinic acid has not yet been reported. Methods: Transfection of TGR5 into cultured Chinese hamster ovary (CHO-K1 cells was performed to establish the presence of TGR5. Additionally, TGR5-specific small interfering RNA was employed to silence TGR5 in cells (NCI-H716 cells that secreted incretins. Uptake of glucose by CHO-K1 cells was evaluated using a fluorescent indicator. Amounts of cyclic adenosine monophosphate and glucagon-like peptide were quantified using enzyme-linked immunosorbent assay kits. Results: Betulinic acid dose-dependently increases glucose uptake by CHO-K1 cells transfected with TGR5 only, which can be considered an alternative method instead of radioligand binding assay. Additionally, signals coupled to TGR5 activation are also

  15. CPG-7909 (PF-3512676, ProMune): toll-like receptor-9 agonist in cancer therapy. (United States)

    Murad, Yanal M; Clay, Timothy M; Lyerly, H Kim; Morse, Michael A


    Stimulation of toll-like receptor (TLR)9 activates human plasmacytoid dendritic cells and B cells, and induces potent innate immune responses in preclinical tumor models and in patients. CpG oligodeoxynucleotides (ODNs) are TLR9 agonists that show promising results as vaccine adjuvants and in the treatment of cancers, infections, asthma and allergy. PF-3512676 (ProMune) was developed as a TLR9 agonist for the treatment of cancer as monotherapy and as an adjuvant in combination with chemo- and immunotherapy. Phase I and II trials have tested this drug in several hematopoietic and solid tumors. Pfizer has initiated Phase III trials to test PF-3512676 in combination with standard chemotherapy for non-small-cell lung cancer.

  16. Liver X Receptor Agonists Inhibit the Phospholipid Regulatory Gene CTP: Phosphoethanolamine Cytidylyltransferase-Pcyt2

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    Lin Zhu


    Full Text Available Metabolic pulse-chase experiments demonstrated that 25-hydroxycholesterol (25-OH, the endogenous activator of the liver X receptor (LXR, significantly reduced the biosynthesis of phosphatidylethanolamine via CDP-ethanolamine (Kennedy pathway at the step catalyzed by CTP: phosphoethanolamine cytidylyltransferase (Pcyt2. In the mouse embryonic fibroblasts C3H10T1/2, the LXR synthetic agonist TO901317 lowered Pcyt2 promoter-luciferase activity in a concentration-dependent manner. Furthermore, 25-OH and TO901317 reduced mouse Pcyt2 mRNA and protein levels by 35–60%. The inhibitory effects of oxysterols and TO901317 on the Pcyt2 promoter function, mRNA and protein expression were conserved in the human breast cancer cells MCF-7. These studies identify the Pcyt2 gene as a novel target whereby LXR agonists may indirectly modulate inflammatory responses and atherosclerosis.

  17. Isoform-Specific Biased Agonism of Histamine H3 Receptor Agonists. (United States)

    Riddy, Darren M; Cook, Anna E; Diepenhorst, Natalie A; Bosnyak, Sanja; Brady, Ryan; Mannoury la Cour, Clotilde; Mocaer, Elisabeth; Summers, Roger J; Charman, William N; Sexton, Patrick M; Christopoulos, Arthur; Langmead, Christopher J


    The human histamine H3 receptor (hH3R) is subject to extensive gene splicing that gives rise to a large number of functional and nonfunctional isoforms. Despite the general acceptance that G protein-coupled receptors can adopt different ligand-induced conformations that give rise to biased signaling, this has not been studied for the H3R; further, it is unknown whether splice variants of the same receptor engender the same or differential biased signaling. Herein, we profiled the pharmacology of histamine receptor agonists at the two most abundant hH3R splice variants (hH3R445 and hH3R365) across seven signaling endpoints. Both isoforms engender biased signaling, notably for 4-[3-(benzyloxy)propyl]-1H-imidazole (proxyfan) [e.g., strong bias toward phosphorylation of glycogen synthase kinase 3β (GSK3β) via the full-length receptor] and its congener 3-(1H-imidazol-4-yl)propyl-(4-iodophenyl)-methyl ether (iodoproxyfan), which are strongly consistent with the former's designation as a "protean" agonist. The 80 amino acid IL3 deleted isoform hH3R365 is more permissive in its signaling than hH3R445: 2-(1H-imidazol-5-yl)ethyl imidothiocarbamate (imetit), proxyfan, and iodoproxyfan were all markedly biased away from calcium signaling, and principal component analysis of the full data set revealed divergent profiles for all five agonists. However, most interesting was the identification of differential biased signaling between the two isoforms. Strikingly, hH3R365 was completely unable to stimulate GSK3β phosphorylation, an endpoint robustly activated by the full-length receptor. To the best of our knowledge, this is the first quantitative example of differential biased signaling via isoforms of the same G protein-coupled receptor that are simultaneously expressed in vivo and gives rise to the possibility of selective pharmacological targeting of individual receptor splice variants.



    McConalogue, K; Grady, E. F.; MINNIS, J.; Balestra, B; Tonini, M; Brecha, N C; Bunnett, N. W.; Sternini, C.


    The multiple effects of opiate alkaloids, important therapeutic drugs used for pain control, are mediated by the neuronal μ-opioid receptor. Among the side effects of these drugs is a profound impairment of gastrointestinal transit. Endomorphins are opioid peptides recently isolated from the nervous system, which have high affinity and selectivity for μ-opioid receptors. Since the μ-opioid receptor undergoes ligand-induced receptor endocytosis in an agonist-dependent manner, we compared the a...

  19. Delayed cardioprotection is mediated via a non-peptide delta opioid agonist, SNC-121, independent of opioid receptor stimulation. (United States)

    Patel, Hemal H; Hsu, Anna; Gross, Garrett J


    Acute cardioprotection is mediated primarily through delta opioid receptor stimulation independent of micro or kappa opioid receptor stimulation. Delayed cardioprotection is mediated by delta opioid receptor agonists but ambiguity remains about direct receptor involvement. Therefore, we investigated the potential of SNC-121, a non-peptide delta opioid agonist, to produce delayed cardioprotection and characterized the role of opioid receptors in this delayed response. All rats underwent 30 minutes of ischemia followed by 2 hours of reperfusion. SNC-121 induced a significant delayed cardioprotective effect. To determine the nature of this SNC-121-induced delayed cardioprotection, rats were treated with specific opioids receptor antagonists and underwent pertussis toxin (PT) treatment prior to opioid agonist stimulation. Control rats were injected with saline and allowed to recover for 24 hours. Pretreatment and early treatment with opioid receptor antagonists failed to inhibit the delayed protective effects of SNC-121, as did pretreatment with PT. Treatment with a free radical scavenger, 2-mercaptopropionyl glycine, at the time of opioid stimulation attenuated the delayed cardioprotective effects of SNC-121. These data suggest that delayed cardioprotection is stimulated via non-peptide delta opioid agonists by a mechanism unrelated to opioid receptor activation. The mechanism appears to be a non-opioid receptor mediated production of reactive oxygen species that triggers the signaling cascade leading to delayed cardioprotection.

  20. Estrogen receptor hormone agonists limit trauma hemorrhage shock-induced gut and lung injury in rats.

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    Danielle Doucet

    Full Text Available BACKGROUND: Acute lung injury (ALI and the development of the multiple organ dysfunction syndrome (MODS is a major cause of death in trauma patients. Earlier studies in trauma hemorrhagic shock (T/HS have documented that splanchnic ischemia leading to gut inflammation and loss of barrier function is an initial triggering event that leads to gut-induced ARDS and MODS. Since sex hormones have been shown to modulate the response to T/HS and proestrous (PE females are more resistant to T/HS-induced gut and distant organ injury, the goal of our study was to determine the contribution of estrogen receptor (ERalpha and ERbeta in modulating the protective response of female rats to T/HS-induced gut and lung injury. METHODS/PRINCIPAL FINDINGS: The incidence of gut and lung injury was assessed in PE and ovariectomized (OVX female rats subjected to T/HS or trauma sham shock (T/SS as well as OVX rats that were administered estradiol (E2 or agonists for ERalpha or ERbeta immediately prior to resuscitation. Marked gut and lung injury was observed in OVX rats subjected to T/HS as compared to PE rats or E2-treated OVX rats subjected to T/HS. Both ERalpha and ERbeta agonists were equally effective in limiting T/HS-induced morphologic villous injury and bacterial translocation, whereas the