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. 1994 Jan;111(1):191–198. doi: 10.1111/j.1476-5381.1994.tb14043.x

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

M C Peakman 1, S J Hill 1
PMCID: PMC1910019  PMID: 8012696

Abstract

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

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Alexander S. P., Kendall D. A., Hill S. J. Differences in the adenosine receptors modulating inositol phosphates and cyclic AMP accumulation in mammalian cerebral cortex. Br J Pharmacol. 1989 Dec;98(4):1241–1248. doi: 10.1111/j.1476-5381.1989.tb12670.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bazil C. W., Minneman K. P. An investigation of the low intrinsic activity of adenosine and its analogs at low affinity (A2) adenosine receptors in rat cerebral cortex. J Neurochem. 1986 Aug;47(2):547–553. doi: 10.1111/j.1471-4159.1986.tb04534.x. [DOI] [PubMed] [Google Scholar]
  3. Bruns R. F., Fergus J. H., Badger E. W., Bristol J. A., Santay L. A., Hartman J. D., Hays S. J., Huang C. C. Binding of the A1-selective adenosine antagonist 8-cyclopentyl-1,3-dipropylxanthine to rat brain membranes. Naunyn Schmiedebergs Arch Pharmacol. 1987 Jan;335(1):59–63. doi: 10.1007/BF00165037. [DOI] [PubMed] [Google Scholar]
  4. Bruns R. F., Fergus J. H., Badger E. W., Bristol J. A., Santay L. A., Hays S. J. PD 115,199: an antagonist ligand for adenosine A2 receptors. Naunyn Schmiedebergs Arch Pharmacol. 1987 Jan;335(1):64–69. doi: 10.1007/BF00165038. [DOI] [PubMed] [Google Scholar]
  5. Bruns R. F., Lu G. H., Pugsley T. A. Characterization of the A2 adenosine receptor labeled by [3H]NECA in rat striatal membranes. Mol Pharmacol. 1986 Apr;29(4):331–346. [PubMed] [Google Scholar]
  6. Daly J. W., Butts-Lamb P., Padgett W. Subclasses of adenosine receptors in the central nervous system: interaction with caffeine and related methylxanthines. Cell Mol Neurobiol. 1983 Mar;3(1):69–80. doi: 10.1007/BF00734999. [DOI] [PubMed] [Google Scholar]
  7. Donaldson J., Brown A. M., Hill S. J. Influence of rolipram on the cyclic 3',5'-adenosine monophosphate response to histamine and adenosine in slices of guinea-pig cerebral cortex. Biochem Pharmacol. 1988 Feb 15;37(4):715–723. doi: 10.1016/0006-2952(88)90146-3. [DOI] [PubMed] [Google Scholar]
  8. Doore B. J., Bashor M. M., Spitzer N., Mawe R. C., Saier M. H., Jr Regulation of adenosine 3' :5'-monophosphate efflux from rat glioma cells in culture*. J Biol Chem. 1975 Jun 10;250(11):4371–4372. [PubMed] [Google Scholar]
  9. Ebersolt C., Perez M., Bockaert J. Neuronal, glial and meningeal localizations of neurotransmitter-sensitive adenylate cyclases in cerebral cortex of mice. Brain Res. 1981 May 25;213(1):139–150. doi: 10.1016/0006-8993(81)91254-3. [DOI] [PubMed] [Google Scholar]
  10. Ebersolt C., Perez M., Vassent G., Bockaert J. Characteristics of the beta 1-and beta 2-adrenergic-sensitive adenylate cyclases in glial cell primary cultures and their comparison with beta 2-adrenergic-sensitive adenylate cyclase of meningeal cells. Brain Res. 1981 May 25;213(1):151–161. doi: 10.1016/0006-8993(81)91255-5. [DOI] [PubMed] [Google Scholar]
  11. Ebersolt C., Premont J., Prochiantz A., Perez M., Bockaert J. Inhibition of brain adenylate cyclase by A1 adenosine receptors: pharmacological characteristics and locations. Brain Res. 1983 May 9;267(1):123–129. doi: 10.1016/0006-8993(83)91045-4. [DOI] [PubMed] [Google Scholar]
  12. Elfman L., Lindgren E., Walum E., Fredholm B. B. Adenosine analogues stimulate cyclic AMP-accumulation in cultured neuroblastoma and glioma cells. Acta Pharmacol Toxicol (Copenh) 1984 Oct;55(4):297–302. doi: 10.1111/j.1600-0773.1984.tb01985.x. [DOI] [PubMed] [Google Scholar]
  13. Furlong T. J., Pierce K. D., Selbie L. A., Shine J. Molecular characterization of a human brain adenosine A2 receptor. Brain Res Mol Brain Res. 1992 Sep;15(1-2):62–66. doi: 10.1016/0169-328x(92)90152-2. [DOI] [PubMed] [Google Scholar]
  14. Geiger J. D., Johnston M. E., Yago V. Pharmacological characterization of rapidly accumulated adenosine by dissociated brain cells from adult rat. J Neurochem. 1988 Jul;51(1):283–291. doi: 10.1111/j.1471-4159.1988.tb04868.x. [DOI] [PubMed] [Google Scholar]
  15. Gurden M. F., Coates J., Ellis F., Evans B., Foster M., Hornby E., Kennedy I., Martin D. P., Strong P., Vardey C. J. Functional characterization of three adenosine receptor types. Br J Pharmacol. 1993 Jul;109(3):693–698. doi: 10.1111/j.1476-5381.1993.tb13629.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Hill S. J., Kendall D. A. Studies on the adenosine-receptor mediating the augmentation of histamine-induced inositol phospholipid hydrolysis in guinea-pig cerebral cortex. Br J Pharmacol. 1987 Jul;91(3):661–669. doi: 10.1111/j.1476-5381.1987.tb11260.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Hutchison A. J., Webb R. L., Oei H. H., Ghai G. R., Zimmerman M. B., Williams M. CGS 21680C, an A2 selective adenosine receptor agonist with preferential hypotensive activity. J Pharmacol Exp Ther. 1989 Oct;251(1):47–55. [PubMed] [Google Scholar]
  18. Jacobson K. A., van Galen P. J., Williams M. Adenosine receptors: pharmacology, structure-activity relationships, and therapeutic potential. J Med Chem. 1992 Feb 7;35(3):407–422. doi: 10.1021/jm00081a001. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Jarvis M. F., Schulz R., Hutchison A. J., Do U. H., Sills M. A., Williams M. [3H]CGS 21680, a selective A2 adenosine receptor agonist directly labels A2 receptors in rat brain. J Pharmacol Exp Ther. 1989 Dec;251(3):888–893. [PubMed] [Google Scholar]
  20. Kodama T., Matsukado Y., Shimizu H. The cyclic AMP system of human brain. Brain Res. 1973 Feb 14;50(1):135–146. doi: 10.1016/0006-8993(73)90600-8. [DOI] [PubMed] [Google Scholar]
  21. Libert F., Schiffmann S. N., Lefort A., Parmentier M., Gérard C., Dumont J. E., Vanderhaeghen J. J., Vassart G. The orphan receptor cDNA RDC7 encodes an A1 adenosine receptor. EMBO J. 1991 Jul;10(7):1677–1682. doi: 10.1002/j.1460-2075.1991.tb07691.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Libert F., Van Sande J., Lefort A., Czernilofsky A., Dumont J. E., Vassart G., Ensinger H. A., Mendla K. D. Cloning and functional characterization of a human A1 adenosine receptor. Biochem Biophys Res Commun. 1992 Sep 16;187(2):919–926. doi: 10.1016/0006-291x(92)91285-x. [DOI] [PubMed] [Google Scholar]
  23. Lohse M. J., Klotz K. N., Lindenborn-Fotinos J., Reddington M., Schwabe U., Olsson R. A. 8-Cyclopentyl-1,3-dipropylxanthine (DPCPX)--a selective high affinity antagonist radioligand for A1 adenosine receptors. Naunyn Schmiedebergs Arch Pharmacol. 1987 Aug;336(2):204–210. doi: 10.1007/BF00165806. [DOI] [PubMed] [Google Scholar]
  24. Lohse M. J., Klotz K. N., Schwabe U., Cristalli G., Vittori S., Grifantini M. 2-Chloro-N6-cyclopentyladenosine: a highly selective agonist at A1 adenosine receptors. Naunyn Schmiedebergs Arch Pharmacol. 1988 Jun;337(6):687–689. doi: 10.1007/BF00175797. [DOI] [PubMed] [Google Scholar]
  25. Lupica C. R., Cass W. A., Zahniser N. R., Dunwiddie T. V. Effects of the selective adenosine A2 receptor agonist CGS 21680 on in vitro electrophysiology, cAMP formation and dopamine release in rat hippocampus and striatum. J Pharmacol Exp Ther. 1990 Mar;252(3):1134–1141. [PubMed] [Google Scholar]
  26. Maenhaut C., Van Sande J., Libert F., Abramowicz M., Parmentier M., Vanderhaegen J. J., Dumont J. E., Vassart G., Schiffmann S. RDC8 codes for an adenosine A2 receptor with physiological constitutive activity. Biochem Biophys Res Commun. 1990 Dec 31;173(3):1169–1178. doi: 10.1016/s0006-291x(05)80909-x. [DOI] [PubMed] [Google Scholar]
  27. Mahan L. C., McVittie L. D., Smyk-Randall E. M., Nakata H., Monsma F. J., Jr, Gerfen C. R., Sibley D. R. Cloning and expression of an A1 adenosine receptor from rat brain. Mol Pharmacol. 1991 Jul;40(1):1–7. [PubMed] [Google Scholar]
  28. McCarthy K. D., de Vellis J. Preparation of separate astroglial and oligodendroglial cell cultures from rat cerebral tissue. J Cell Biol. 1980 Jun;85(3):890–902. doi: 10.1083/jcb.85.3.890. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. McCrea K. E., Hill S. J. Salmeterol, a long-acting beta 2-adrenoceptor agonist mediating cyclic AMP accumulation in a neuronal cell line. Br J Pharmacol. 1993 Oct;110(2):619–626. doi: 10.1111/j.1476-5381.1993.tb13856.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Moore P. K., Oluyomi A. O., Babbedge R. C., Wallace P., Hart S. L. L-NG-nitro arginine methyl ester exhibits antinociceptive activity in the mouse. Br J Pharmacol. 1991 Jan;102(1):198–202. doi: 10.1111/j.1476-5381.1991.tb12153.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Murphy M. G., Moak C. M., Byczko Z., MacDonald W. F. Adenosine-dependent regulation of cyclic AMP accumulation in primary cultures of rat astrocytes and neurons. J Neurosci Res. 1991 Dec;30(4):631–640. doi: 10.1002/jnr.490300406. [DOI] [PubMed] [Google Scholar]
  32. Olah M. E., Ren H., Ostrowski J., Jacobson K. A., Stiles G. L. Cloning, expression, and characterization of the unique bovine A1 adenosine receptor. Studies on the ligand binding site by site-directed mutagenesis. J Biol Chem. 1992 May 25;267(15):10764–10770. [PMC free article] [PubMed] [Google Scholar]
  33. Pierce K. D., Furlong T. J., Selbie L. A., Shine J. Molecular cloning and expression of an adenosine A2b receptor from human brain. Biochem Biophys Res Commun. 1992 Aug 31;187(1):86–93. doi: 10.1016/s0006-291x(05)81462-7. [DOI] [PubMed] [Google Scholar]
  34. Rindler M. J., Bashor M. M., Spitzer N., Saier M. H., Jr Regulation of adenosine 3':5'-monophosphate efflux from animal cells. J Biol Chem. 1978 Aug 10;253(15):5431–5436. [PubMed] [Google Scholar]
  35. Rivkees S. A., Reppert S. M. RFL9 encodes an A2b-adenosine receptor. Mol Endocrinol. 1992 Oct;6(10):1598–1604. doi: 10.1210/mend.6.10.1333049. [DOI] [PubMed] [Google Scholar]
  36. Ruck A., Kendall D. A., Hill S. J. Alpha- and beta-adrenoceptor regulation of cyclic AMP accumulation in cultured rat astrocytes. A comparison of primary protoplasmic and mixed fibrous/protoplasmic astroglial cultures. Biochem Pharmacol. 1991 Jun 21;42(1):59–69. doi: 10.1016/0006-2952(91)90681-t. [DOI] [PubMed] [Google Scholar]
  37. Sattin A., Rall T. W. The effect of adenosine and adenine nucleotides on the cyclic adenosine 3', 5'-phosphate content of guinea pig cerebral cortex slices. Mol Pharmacol. 1970 Jan;6(1):13–23. [PubMed] [Google Scholar]
  38. Stiles G. L. Adenosine receptors. J Biol Chem. 1992 Apr 5;267(10):6451–6454. [PubMed] [Google Scholar]
  39. Woods M. D., Freshney R. I., Ball S. G., Vaughan P. F. Regulation of cyclic AMP formation in cultures of human foetal astrocytes by beta 2-adrenergic and adenosine receptors. J Neurochem. 1989 Sep;53(3):864–869. doi: 10.1111/j.1471-4159.1989.tb11784.x. [DOI] [PubMed] [Google Scholar]
  40. Woods M. D., Freshney R. I., Pilkington G., Vaughan P. F. Expression of receptors, linked to cyclic AMP formation by cultures of human fetal astrocytes. Biochem Soc Trans. 1988 Aug;16(4):432–433. doi: 10.1042/bst0160432. [DOI] [PubMed] [Google Scholar]
  41. van Calker D., Müller M., Hamprecht B. Adenosine regulates via two different types of receptors, the accumulation of cyclic AMP in cultured brain cells. J Neurochem. 1979 Nov;33(5):999–1005. doi: 10.1111/j.1471-4159.1979.tb05236.x. [DOI] [PubMed] [Google Scholar]

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