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On the high affinity binding site for [ 3 H]‐1,3‐dipropyl‐8‐cyclopentylxanthine in frog brain membranes
Author(s) -
Oliveira J.C.,
Sebastião A.M.,
Ribeiro J.A.
Publication year - 1993
Publication title -
british journal of pharmacology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.432
H-Index - 211
eISSN - 1476-5381
pISSN - 0007-1188
DOI - 10.1111/j.1476-5381.1993.tb13600.x
Subject(s) - gtp' , guanosine , binding site , chemistry , membrane , g protein , medicine , radioligand , adenosine , endocrinology , stereochemistry , receptor , biology , biochemistry , enzyme
1 Radioligand binding properties of the adenosine receptor ligands, [ 3 H]‐1,3‐dipropyl‐8‐cyclopentylxanthine ([ 3 H]‐DPCPX), and [ 3 H]‐ R ‐phenylisopropyladenosine ([ 3 H]‐ R ‐PIA) were investigated in frog brain membranes. 2 The specific binding of the adenosine antagonist, [ 3 H]‐DPCPX to frog brain membranes showed one binding site with K d and B max values of 43.8 n m and 0.238 ± 0.016 pmol mg −1 protein, respectively. Guanosine 5′‐triphosphate (GTP, 100 μ m ) decreased to 72 ± 7% and Mg 2+ (8 m m ) increased to 121 ± 3% [ 3 H]‐DPCPX (40 n m ) binding to frog brain membranes. 3 [ 3 H]‐DPCPX saturation binding experiments performed in the presence of Mg 2+ (8 m m ), or in the presence of GTP showed that Mg 2+ ions decreased the K d value of [ 3 H]‐DPCPX to 14 n m , and GTP increased this value to 65.6 n m . B max values were not significantly ( P > 0.05) modified (0.261 ± 0.018 pmol mg −1 protein, with Mg 2+ , and 0.266 ± 0.026 pmol mg −1 protein, in presence of GTP) by the presence of Mg 2+ or GTP. 4 The specific binding of [ 3 H]‐ R ‐PIA (15 n m ) was decreased to 37 ± 6% by GTP (100 μ m ) and increased to 123 ± 4% by Mg 2+ (8 m m ). [ 3 H]‐ R ‐PIA saturation binding experiments performed in the presence of Mg 2+ (8 m m ) showed one binding site with K d and B max values of 0.9 n m and 0.229 ± 0.008 pmol mg −1 of protein, respectively. 5 The concentration‐inhibition curves of adenosine agonists and antagonists versus [ 3 H]‐DPCPX binding showed the following order of potencies: CPA > R ‐PIA ≃ NECA > S‐PIA > > CGS 21680, for the agonists, and XAC ≃ DPCPX > > XCC > PACPX, for the antagonists. 6 The present results suggest that the adenosine binding site in the frog brain membranes is G‐protein coupled, but that the antagonist affinities and the pharmacological profile is different from the A 1 or A 2 adenosine receptors.