Abstract

[125I]N6-(p-aminobenzyl)adenosine and [125I]N6-(p-azidobenzyl)adenosine, which are potent agonists at A1 (Ri) adenosine receptors, have been used to characterize the adenosine receptor in membranes prepared from newborn chick heart. Scatchard analyses of [125I]N6-(p-aminobenzyl)adenosine binding to cardiac membranes revealed that the ligand bound to two affinity states of the receptor with Kd values of 0.7 and 9.9 nM. The corresponding maximum binding (Bmax) values were 25 and 86 fmol/mg of protein, respectively. In the presence of 0.1 mM 5'-guanylyl imidodiphosphate, a single affinity state was detected with a Kd of 9.4 nM and a Bmax of 96 fmol/mg of protein. Direct and indirect ligand binding studies with several adenosine receptor agonists and antagonists were used to compare the characteristics of the cardiac receptor with those of the A1 receptor in the cerebellum. The binding properties of the receptors in the two tissues were very similar although marked differences were observed in the binding kinetics of [125I]N6-(p-azidobenzyl)adenosine. Photo-affinity labeling experiments followed by sodium dodecyl sulfate-gel electrophoresis showed that the cardiac receptor had a apparent molecular weight of 37,600, which was slightly but significantly higher than that of the cerebellar receptor (35,500). The present results show that the cardiac receptor has ligand binding properties and a minimal subunit molecular weight similar to the more thoroughly studied A1 receptor in neural tissue.