Characterization of a novel adenosine binding protein sensitive to cyclic AMP in rat brain cytosolic and particulate fractions

Lorenzen, Anna; Groβekatthöfer, Birte; Kerst, Birgit; Vogt, Heidrun; Fein, Thomas; Schwabe, Ulrich

doi:10.1016/s0006-2952(96)00465-0

Cited by 13 publications

(8 citation statements)

References 31 publications

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“…The potent, nonselective adenosine receptor agonist NECA (22), which has been found to also bind with high affinity to nonadenosine receptor binding sites (Lorenzen et al, 1993(Lorenzen et al, , 1996Yoshioka et al, 2001), showed no significant binding affinity for the adenine receptor. As pointed out above, adenosine might be partially degraded to adenine during the assay, whereas NECA is enzymatically more stable.…”

Section: Discussionmentioning

confidence: 99%

Evidence for the Functional Expression and Pharmacological Characterization of Adenine Receptors in Native Cells and Tissues

Gorzalka

Vittori²,

Volpini³

et al. 2004

Mol Pharmacol

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An orphan G protein-coupled receptor from rat has recently been discovered to be activated by the nucleobase adenine (Proc Natl Acad Sci USA 99: [8573][8574][8575][8576][8577][8578] 2002). In the present study, we show for the first time that the adenine receptor is expressed in membrane preparations of native tissues and cell lines in high density, including rat brain cortex, rat brain striatum, and the mouse neuroblastoma ϫ rat glioma hybrid cell line 3 H]adenine competition experiments at rat brain cortex. Only minor substitution of the adenine structure was tolerated, the most potent compounds of the present series being 2-fluoroadenine (K i value of 620 nM), 8-thioadenine (K i value of 2.77 M), N 6 -methyladenine (K i value of 3.64 M), and 7-methyladenine (K i value of 4.13 M), all of which were partial agonists (40 -60% intrinsic activity). Adenine dose dependently inhibited forskolin-stimulated adenylate cyclase in membrane preparations of NG108-15 cells as well as in intact cells, showing that the receptor is functional in NG108-15 cells. Reverse transcriptasepolymerase chain reaction experiments followed by sequencing indicate that the NG108-15 cells express the murine ortholog of the adenine receptor. Moreover, preliminary radioligand binding studies with [3 H]adenine at membranes of human astrocytoma 1321N1 cells suggest that a human ortholog of the rat adenine receptor exists.Adenine was recently identified as the endogenous ligand of an orphan rat G protein-coupled receptor by a "reverse pharmacology" approach. mRNA localization studies revealed the highest expression in the small neurons of the dorsal root ganglia, suggesting a role in nociception (Bender et al., 2002). Potent neurotrophic effects of adenine have recently been described; in primary cultures of rat cerebellar Purkinje cells, the number of cells was greatly increased by the addition of adenine, although relatively high concentrations (1-2 mM) were required . Adenine was the most potent compound in a series of purine and pyrimidine derivatives that showed protective effects on Purkinje cells and prevented cell death, whereas other adeninebased purines, including adenosine, AMP, ADP, and ATP, did not promote Purkinje cell survival .Radioligand binding studies had previously been performed at membrane preparations of CHO cells stably transfected with the rat adenine receptor using [ 3 H]adenine as a radioligand (Bender et al., 2002). The recombinant rat adenine receptor expressed in CHO cells was found to be coupled to inhibition of adenylate cyclase via G i protein (Bender et al., 2002). However, the postulated adenine receptor has not been demonstrated to be present in native tissues on the protein level, and therefore it has not yet been officially accepted as a novel receptor by the International Union of Pharmacology committee on receptor nomenclature. A mouse S.G. and C.E.M. were supported by the Deutsche Forschungsgemeinschaft (GRK-804/1).Article, publication date, and citation information can be found at

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Section: Discussionmentioning

confidence: 99%

Evidence for the Functional Expression and Pharmacological Characterization of Adenine Receptors in Native Cells and Tissues

Gorzalka

Vittori²,

Volpini³

et al. 2004

Mol Pharmacol

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“…[3H]CGS 21680 is relatively selective for A 2A receptors, but may also bind to other sites in cerebral cortex 273,296. [3H]NECA binds to other non-receptor elements, which also recognise adenosine 307. XAC-BY630 has been described as a fluorescent antagonist for labelling A 1 adenosine receptors in living cells, although activity at other adenosine receptors was not examined 272.…”

Section: Commentsmentioning

confidence: 99%

The Concise Guide to PHARMACOLOGY 2013/14: G Protein‐Coupled Receptors

Alexander

Benson

Faccenda

et al. 2013

British J Pharmacology

555

336

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The Concise Guide to PHARMACOLOGY 2013/14 provides concise overviews of the key properties of over 2000 human drug targets with their pharmacology, plus links to an open access knowledgebase of drug targets and their ligands (www.guidetopharmacology.org), which provides more detailed views of target and ligand properties. The full contents can be found at http://onlinelibrary.wiley.com/doi/10.1111/bph.12444/full.G protein-coupled receptors are one of the seven major pharmacological targets into which the Guide is divided, with the others being G protein-coupled receptors, ligand-gated ion channels, ion channels, catalytic receptors, nuclear hormone receptors, transporters and enzymes. These are presented with nomenclature guidance and summary information on the best available pharmacological tools, alongside key references and suggestions for further reading. A new landscape format has easy to use tables comparing related targets.It is a condensed version of material contemporary to late 2013, which is presented in greater detail and constantly updated on the website www.guidetopharmacology.org, superseding data presented in previous Guides to Receptors and Channels. It is produced in conjunction with NC-IUPHAR and provides the official IUPHAR classification and nomenclature for human drug targets, where appropriate. It consolidates information previously curated and displayed separately in IUPHAR-DB and the Guide to Receptors and Channels, providing a permanent, citable, point-in-time record that will survive database updates.

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“…Table 1 also suggests that P3LP has a similar pharmacological profile to that of adenotin 1 and adenotin 2, other adenosine binding proteins abundantly present in brain and peripheral tissues that can bind [ 3 H]-NECA with relatively high affinity. 23,24 Although adenotin 2 has been reported to bind weakly with adenosine and AMP (Ki values were approximately 7 mol/L for each compound), 24 P3LP was shown to bind strongly with these compounds (Ki = 87 and 89 nmol/L, respectively). Adenotin 1 was reported to bind CGS 21680 strongly with a Ki of 20 nmol/L in bovine striatum.…”

Section: Discussionmentioning

confidence: 99%

Pharmacology Of A Unique Adenosine Binding Site In Rat Brain Using A Selective Ligand

Yoshioka

Matsuda

Nakata

2001

Clin Exp Pharma Physio

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1. In order to further characterize the adenosine binding sites that we previously purified and termed P3 purinergic receptor-like protein (P3LP), a reliable binding assay method was developed using [3H]-5'-N-ethylcarboxamidoadenosine (NECA) as a radioligand and the newly developed high-affinity selective ligand 9-(6,7-dideoxy-beta-D-allo-hept-5-ynofuranosyl) adenine (HAK2701) as a selective displacer. 2. Using this assay method, it was found that rat brain membranes possess high- and low-affinity [3H]-NECA binding sites. The high-affinity binding site showed KD and Bmax values of 19.7+/-2.5 nmol/L and 0.192+/-0.05 pmol/mg protein, respectively, and the KD value for the low-affinity binding site was 4260+/-330 nmol/L. The KD value for the high-affinity site agreed well with that of the [3H]-NECA binding site determined with the partially purified P3LP preparation described previously. 3. The distribution of P3LP in rat tissues was determined using the [3H]-NECA binding method described above. The highest level of P3LP was in the cerebellum followed by the olfactory bulb and the spinal cord. 4. The order of the affinity for various purinergic or related compounds to P3LP in rat brain preparations was also determined by the [3H]-NECA binding method to be HAK2701 > NECA = adenosine 5'-O-(2-thiodiphosphate) > cAMP = beta,gamma-methyleneadenosine 5'-triphosphate > diadenosine tetraphosphate > alpha,beta-methyleneadenosine 5'-triphosphate > 5'-deoxy-5'-methylthioadenosine > N6-cyclopentyladenosine. 5. These studies reveal that the [3H]-NECA binding assay in combination with HAK2701 is successful in the characterization of P3LP, especially the membrane-bound form.

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Characterization of a novel adenosine binding protein sensitive to cyclic AMP in rat brain cytosolic and particulate fractions

Cited by 13 publications

References 31 publications

Evidence for the Functional Expression and Pharmacological Characterization of Adenine Receptors in Native Cells and Tissues

Evidence for the Functional Expression and Pharmacological Characterization of Adenine Receptors in Native Cells and Tissues

The Concise Guide to PHARMACOLOGY 2013/14: G Protein‐Coupled Receptors

Pharmacology Of A Unique Adenosine Binding Site In Rat Brain Using A Selective Ligand

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