Addicting drugs utilize a synergistic molecular mechanism in common requiring adenosine and Gi-beta gamma dimers.
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The mesolimbic dopamine system and cAMP-dependent/protein kinase A (PKA) pathways are strongly implicated in addictive behaviors. Here we determine the role of dopamine D2 receptors (D2) in PKA signaling responses to delta-opioid (DOR) and cannabinoid (CB1) receptors. We find in NG108-15/D2 cells and in cultured primary neurons that a brief exposure to saturating concentrations of DOR and CB1 agonists increases cAMP, promotes PKA C alpha translocation and increases cAMP-dependent gene expression. Activation of PKA signaling is mediated by Gi-beta gamma dimers. Importantly, subthreshold concentrations of DOR or CB1 agonists with D2 agonists, which are without effect when added separately, together activate cAMP/PKA signaling synergistically. There is also synergy between DOR or CB1 with ethanol, another addicting agent. In all instances, synergy requires adenosine activation of adenosine A2 receptors and is mediated by beta gamma dimers. Synergy by this molecular mechanism appears to confer hypersensitivity to opioids and cannabinoids while simultaneously increasing the sensitivity of D2 signaling when receptors are expressed on the same cells. This mechanism may account, in part, for drug-induced activation of medium spiny neurons in the nucleus accumbens. (+info)
Comparative pharmacological studies on the A2 adenosine receptor agonist 5'-n-ethyl-carboxamidoadenosine and its F19 isotope labelled derivative.
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Adenosine receptors are expressed in various mammalian tissues where they mediate the effects of adenosine on cellular functions through a number of signalling mechanisms. 18F-NECA is the positron-emitting derivative of the A(2)-receptor agonist NECA (5'-n-ethyl-carboxamidoadenosine) and is a radioligand for PET imaging of adenosine receptors. Contractility and relaxation studies were performed on guinea pig atrial myocardium, pulmonary artery, and thoracic aorta to compare the pharmacological effects of NECA and F-NECA (a non-emitting derivative) on tissues. Furthermore, the effect of NECA and F-NECA on the potassium conductance was investigated in DDT1 MF-2 smooth muscle cells with the patch-clamp technique. Both NECA and F-NECA reduced the contractile force in atrial myocardium and evoked phasic contraction in pulmonary artery (A(1) adenosine-receptor-mediated actions) in a dose dependent manner; however, the apparent affinity was lower for F-NECA. No difference was found in relaxation induced by these compounds in 1 microM noradrenaline-precontracted aorta and pulmonary artery (in the presence of DPCPX, an A(1) adenosine receptor antagonist, tissue containing A(2B) adenosine receptors). NECA (5 microM) and F-NECA (5 microM) also decreased the peak current and accelerated activation and inactivation properties of the potassium channels, but F-NECA was less effective. These results suggest that while NECA and F-NECA are equivalent agonists of vascular A(2B) receptors, they mediate different changes of some parameters. When evaluating the data obtained by the use of radiolabelled ligands, one has to take into consideration the possible physiological effects of the ligands besides its binding properties to tissues. (+info)
A putative osmoreceptor system that controls neutrophil function through the release of ATP, its conversion to adenosine, and activation of A2 adenosine and P2 receptors.
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We have previously shown that hypertonic stress (HS) can suppress chemoattractant-induced neutrophil responses via cyclic adenosine monophosphate and enhance these responses through p38 mitogen-activated protein kinase (MAPK) activation. The underlying mechanisms are unknown. Here, we report that HS dose-dependently releases adenosine 5'-triphosphate (ATP) from neutrophils and that extracellular ATP is rapidly converted to adenosine or activates p38 MAPK and enhances N-formyl-methionyl-leucyl-phenylalanine-induced superoxide formation. In contrast, adenosine suppresses superoxide formation. Adenosine deaminase treatment abolished the suppressive effect of HS, indicating that HS inhibits neutrophils through adenosine generation. Neutrophils express mRNA, encoding all known P1 adenosine receptors (A1, A2a, A2b, and A3) and the nucleotide receptors P2Y2, P2Y4, P2Y6, P2Y11, and P2X7. A2 receptor agonists mimicked the suppressive effects of HS; the A2 receptor antagonists 8-(p-sulfophenyl)theophylline, 3,7-dimethyl-1-(2-propynyl)xanthine, 1,3,7-trimethyl-8-(3-chlorostyryl)xanthine, and 3-propylxanthine, but not A1 and A3 receptor antagonists, decreased the suppressive effect of HS, indicating that HS suppresses neutrophils via A2 receptor activation. Antagonists of P2 receptors counteracted the enhancing effects of ATP, suggesting that HS costimulates neutrophils by means of P2 receptor activation. We conclude that hypertonic stress regulates neutrophil function via a single molecule (ATP) and its metabolite (adenosine), using positive- and negative-feedback mechanisms through the activation of P2 and A2 receptors, respectively. (+info)
A2A adenosine receptor activation improves survival in mouse models of endotoxemia and sepsis.
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BACKGROUND: Sepsis is currently treated with antibiotics and various adjunctive therapies that are not very effective. METHODS: Mouse survival (4-5 days) and peritoneal and blood bacteria counts were determined after challenge with intraperitoneal lipopolysaccharide (LPS) or live Escherichia coli. RESULTS: The A(2A) adenosine receptor (AR) agonist 4-[3-[6-amino-9-(5-ethylcarbamoyl-3, 4-dihydroxy-tetrahydro-furan-2-yl)-9H-purin-2-yl]-prop-2-ynyl]-cyclohexanecarboxy lic acid methyl ester (ATL146e; 0.05-50 mu g/kg) protected mice from challenge with LPS, and protection occurred when treatment was delayed up to 24 h after challenge. Deletion of the A (2A) AR gene, Adora2a, inhibited protection by ATL146e. A putative A (3)AR agonist, N(6)-3-iodobenzyladenosine-5'-N-methyluronamide (IB-MECA; 500 mu g/kg but not 5 or 50 mu g/kg) protected mice from challenge with LPS. The protective effects of both ATL146e and IB-MECA were counteracted by the A(2A) AR selective antagonist 4-(2-[7-amino-2-[2-furyl][1,2,4]triazolo[2,3-a][1,3,5]triazin-5-yl-amino]ethyl)-p henol. In the live E. coli model, treatment with ATL146e (50 mu g/kg initiated 8 h after infection) increased survival in mice treated with ceftriaxone (5 days) from 40% to 100%. Treatment with ATL146e did not affect peritoneal numbers of live E. coli at the time of death or 120 h after infection but did increase numbers of peritoneal neutrophils and decreased the number of live E. coli in blood. CONCLUSIONS: AR agonists increase mouse survival in endotoxemia and sepsis via A(2A) AR-mediated mechanisms and reduce the number of live bacteria in blood. (+info)
Adenosine acts through A2 receptors to inhibit IL-2-induced tyrosine phosphorylation of STAT5 in T lymphocytes: role of cyclic adenosine 3',5'-monophosphate and phosphatases.
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Adenosine is a purine nucleoside with immunosuppressive activity that acts through cell surface receptors (A(1), A(2a), A(2b), A(3)) on responsive cells such as T lymphocytes. IL-2 is a major T cell growth and survival factor that is responsible for inducing Jak1, Jak3, and STAT5 phosphorylation, as well as causing STAT5 to translocate to the nucleus and bind regulatory elements in the genome. In this study, we show that adenosine suppressed IL-2-dependent proliferation of CTLL-2 T cells by inhibiting STAT5a/b tyrosine phosphorylation that is associated with IL-2R signaling without affecting IL-2-induced phosphorylation of Jak1 or Jak3. The inhibitory effect of adenosine on IL-2-induced STAT5a/b tyrosine phosphorylation was reversed by the protein tyrosine phosphatase inhibitors sodium orthovanadate and bpV(phen). Adenosine dramatically increased Src homology region 2 domain-containing phosphatase-2 (SHP-2) tyrosine phosphorylation and its association with STAT5 in IL-2-stimulated CTLL-2 T cells, implicating SHP-2 in adenosine-induced STAT5a/b dephosphorylation. The inhibitory effect of adenosine on IL-2-induced STAT5a/b tyrosine phosphorylation was reproduced by A(2) receptor agonists and was blocked by selective A(2a) and A(2b) receptor antagonists, indicating that adenosine was mediating its effect through A(2) receptors. Inhibition of STAT5a/b phosphorylation was reproduced with cell-permeable 8-bromo-cAMP or forskolin-induced activation of adenylyl cyclase, and blocked by the cAMP/protein kinase A inhibitor Rp-cAMP. Forskolin and 8-bromo-cAMP also induced SHP-2 tyrosine phosphorylation. Collectively, these findings suggest that adenosine acts through A(2) receptors and associated cAMP/protein kinase A-dependent signaling pathways to activate SHP-2 and cause STAT5 dephosphorylation that results in reduced IL-2R signaling in T cells. (+info)
Luminal adenosine stimulates chloride secretion through A1 receptor in mouse jejunum.
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Adenosine is known to stimulate chloride secretion by mouse jejunum. Whereas the receptor on the basolateral side is believed to be A2B, the receptor involved in the luminal effect of adenosine has not been identified. We found that jejuna expressed mRNA for all adenosine receptor subtypes. In this study, we investigated the stimulation of chloride secretion by adenosine in jejuna derived from mice lacking the adenosine receptors of A1 (A1R) and A2A (A(2A)R) or control littermates. The jejunal epithelium was mounted in a Ussing chamber, and a new method on the basis of impedance analysis was used to calculate the short-circuit current (I(sc)) values. Chloride secretion was assessed by the I(sc) after inhibition of the sodium-glucose cotransporter by adding phloridzin to the apical bathing solution. The effect of apical adenosine on chloride secretion was lost in jejuna from mice lacking the A1R. There was no difference in the response to basolaterally applied adenosine or to apical forskolin. Furthermore, in jejuna from control mice, the effect of apical adenosine was also abolished in the presence of 8-cyclopentyl-1,3-dipropylxanthine, a specific A1R antagonist. Responses to adenosine were identical in jejuna from control and A(2A)R knockout mice. This study demonstrates that A1R (and not A(2A)R) mediates the enhancement of chloride secretion induced by luminal adenosine in mice jejunum. (+info)
Adenosine produces nitric oxide and prevents mitochondrial oxidant damage in rat cardiomyocytes.
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OBJECTIVE: To examine if adenosine prevents oxidant-induced mitochondrial dysfunction by producing nitric oxide (NO) in cardiomyocytes. METHODS AND RESULTS: Adenosine significantly enhanced the fluorescence of DAF-FM, a dye specific for NO, implying that adenosine induces synthesis of NO. Adenosine-induced NO production was blocked by both the nonspecific NOS inhibitor N(G)-nitro-l-arginine methyl ester (l-NAME) and N(5)-(1-Iminoethyl)-l-ornithine dihydrochloride (l-NIO), an inhibitor of endothelial NOS (eNOS), but not by N(6)-(1-Iminoethyl)-l-lysine hydrochloride (l-NIL), an inhibitor of inducible NOS (iNOS), indicating that adenosine activates eNOS. Adenosine also enhances eNOS phosphorylation and its activity. The adenosine A(2) receptor antagonist 8-(3-chlorostyryl)caffeine but not the A(1) antagonist 8-cyclopentyl-1,3-dipropylxanthine prevented the increase in NO production. CGS21680, an adenosine A(2) receptor agonist, markedly increased NO, further supporting the involvement of A(2) receptors. Adenosine-induced NO production was blocked by 4-Amino-5-(4-chlorophenyl)-7-(t-butyl)pyrazolo(3,4-d)pyrimidine (PP2), a selective Src tyrosine kinase inhibitor, suggesting that Src tyrosine kinase is crucial for adenosine-induced NO production. Adenosine-induced NO production was partially reversed by both wortmannin and Akt inhibitor indicating an involvement of PI3-kinase/Akt. Pretreatment of cells with adenosine prevented H(2)O(2)-induced depolarization of mitochondrial membrane potential (DeltaPsi(m)). The protective effect was blocked by l-NAME and l-NIO but not by l-NIL, indicating that eNOS plays a role in the action of adenosine. The protective effect of adenosine was further suppressed by KT5823, a specific inhibitor of protein kinase G (PKG), indicating the PKG may serve as a downstream target of adenosine. CONCLUSION: Adenosine protects mitochondria from oxidant damage through a pathway involving A(2) receptors, eNOS, NO, PI3-kinase/Akt, and Src tyrosine kinase. (+info)
S-adenosylmethionine (SAMe) modulates interleukin-10 and interleukin-6, but not TNF, production via the adenosine (A2) receptor.
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S-adenosylmethionine (SAMe) is the first product in methionine metabolism and serves as a precursor for glutathione (GSH) as well as a methyl donor in most transmethylation reactions. The administration of exogenous SAMe has beneficial effects in many types of liver diseases. One mechanism for the hepatoprotective action is its ability to regulate the immune system by modulating cytokine production from LPS stimulated monocytes. In the present study, we investigated possible mechanism(s) by which exogenous SAMe supplementation modulated production of TNF, IL-10 and IL-6 in LPS stimulated RAW 264.7 cells, a murine monocyte cell line. Our results demonstrated that exogenous SAMe supplementation inhibited TNF production but enhanced both IL-10 and IL-6 production. SAMe increased intracellular GSH level, however, N-acetylcysteine (NAC), the GSH pro-drug, decreased the production of all three cytokines. Importantly, SAMe increased intracellular adenosine levels, and exogenous adenosine supplementation had effects similar to SAMe on TNF, IL-10 and IL-6 production. 3-Deaza-adenosine (DZA), a specific inhibitor of S-adenosylhomocysteine (SAH) hydrolase, blocked the elevation of IL-10 and IL-6 production induced by SAMe, which was rescued by the addition of exogenous adenosine. Furthermore, the enhancement of LPS-stimulated IL-10 and IL-6 production by both SAMe and adenosine was inhibited by ZM241385, a specific antagonist of the adenosine (A(2)) receptor. Our results suggest that increased adenosine levels with subsequent binding to the A(2) receptor account, at least in part, for SAMe modulation of IL-10 and IL-6, but not TNF production, from LPS stimulated monocytes. (+info)