title: Adenosine receptors in cortical-derived vesicles of the rat: studies on binding sites and accumulation of cyclic AMP.
abstract: A vesicular preparation derived from the cerebral cortex of the rat was used to obtain, under the same experimental condition, binding parameters and stimulation data for cyclic AMP. Two analogues of adenosine were employed in the binding studies: [3H]NECA, a mixed A1/A2 agonist and [3H]CHA, a more selective A1 agonist. The [3H]CHA seemed to bind to a single high affinity site (Kd = 1.31 nM, Bmax = 0.327 pmol bound); saturation data for [3H]NECA were resolved for the presence of a high and a low affinity binding site (Kd1 = 3.08 nM, Bmax1 0.115 pmol bound; Kd2 = 204 nM, Bmax2 1.59 pmol bound), but only when calcium ions were omitted from the incubation medium. At 0 degree C, [3H]NECA bound to a single, low affinity site; the presence of calcium ions (1 mM) significantly reduced the affinity of [3H]NECA (Kd 419 nM), with respect to the absence of calcium (Kd 208 nM), without affecting the Bmax value. The influence of calcium ions was also investigated on the binding of [3H]CHA and a reduction of the Bmax value (36%) was found. Regardless of the presence or the absence of calcium ions, NECA stimulated accumulation of cyclic AMP in a dose-dependent way with an EC50 of 2.79 microM; this value did not correlate with the Kd of the low affinity binding site for [3H]NECA. Thus, the purpose of establishing a correlation between binding sites for analogues of adenosine and the site in the cerebral cortex through which the accumulation of cyclic AMP is induced, was not achieved. It is concluded that the stimulatory effect of analogues of adenosine on adenylate cyclase might not be a receptor-mediated effect. The complex influence of calcium ions on affinity and binding capacity of analogues of adenosine is discussed.
authors.affiliation: Institute of Pharmacology and Pharmacognosy, Faculty of Pharmacy, University of Trieste, Italy.
authors.forename: C
authors.initials: C
authors.lastname: Florio
authors.forename: A M
authors.initials: AM
authors.lastname: Rosati
authors.forename: U
authors.initials: U
authors.lastname: Traversa
authors.forename: R
authors.initials: R
authors.lastname: Vertua
mesh_terms: D000241:Adenosine / Q000031:analogs & derivatives / Q000378:metabolism; D019830:Adenosine-5'-(N-ethylcarboxamide); D000262:Adenylyl Cyclases / Q000378:metabolism; D000818:Animals; D001665:Binding Sites; D001667:Binding, Competitive; D002118:Calcium / Q000502:physiology; D002540:Cerebral Cortex / Q000201:enzymology / Q000378:metabolism*; D000242:Cyclic AMP / Q000378:metabolism*; D066298:In Vitro Techniques; D007700:Kinetics; D008297:Male; D051381:Rats; D011919:Rats, Inbred Strains; D011983:Receptors, Purinergic / Q000378:metabolism*; D013574:Synaptosomes / Q000201:enzymology / Q000378:metabolism; D013696:Temperature
chemical_list: D011983:Receptors, Purinergic; D019830:Adenosine-5'-(N-ethylcarboxamide); C027513:N(6)-cyclohexyladenosine; D000242:Cyclic AMP; D000262:Adenylyl Cyclases; D000241:Adenosine; D002118:Calcium