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Bovine Brain Coated Vesicles Contain Adenosine A 1 Receptors. Presence of Adenylate Cyclase Coupled to the Receptor
Author(s) -
GonzalezCalero Gloria,
Martin Mairena,
Cubero Ana,
Andrés Antonio
Publication year - 1990
Publication title -
journal of neurochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.75
H-Index - 229
eISSN - 1471-4159
pISSN - 0022-3042
DOI - 10.1111/j.1471-4159.1990.tb08827.x
Subject(s) - adenosine , vesicle , adenosine receptor , chemistry , cyclase , adenylate kinase , forskolin , agonist , gtp' , adenosine a2b receptor , receptor , biochemistry , binding site , stereochemistry , membrane , enzyme
Clathrin‐coated vesicles purified from bovine brain express adenosine A 1 receptor binding activity. N 6 ‐Cyclohexyl[ 3 H]adenosine ([ 3 H]CHA), an agonist for the A 1 receptor, binds specifically to coated vesicles. High and low agonist affinity states of the receptor for the radioligand [ 3 H]CHA with K D values of 0.18 and 4.4 n M , respectively, were detected. The high purity of coated vesicles was established by assays for biochemical markers and by electron microscopy. Binding competition experiments using agonists N 6 CHA, N ‐cyclopentyladenosine, 5′‐( N ‐ethylcarboxamido)adenosine, and N 6 ‐[( R )‐ and N 6 ‐[( S )‐phenylisopropyl]adenosine and antagonists (theophylline, 3‐isobutyl‐1‐methylxanthine, and caffeine) confirmed the typical adenosine A 1 nature of the binding site. This binding site presents stereospecificity for N 6 ‐phenylisopropyladenosine, showing 33 times more affinity for N 6 ‐[( R )‐ than for N 6 ‐[( S )‐phenylisopropyl]adenosine. The specific binding of [ 3 H]CHA in coated vesicles is regulated by guanine nucleotides. [ 3 H]CHA specific binding was decreased by 70% in the presence of the hydrolysis‐resistant GTP analogue guanyl‐5‐ylimidodiphosphate. Bovine brain coated vesicles present adenylate cyclase activity. This activity was modulated by forskolin and CHA. The results of this study support the evidence that adenosine A 1 receptors present in coated vesicles are coupled to adenylate cyclase activity through a G i protein.