Demonstration of Ri-type adenosine receptors in bovine myocardium by radioligand binding.

Adenosine has been shown to have negative inotropic, chronotropic and dromotropic effects on the heart. The pharmacological profiles of these effects suggest that they are mediated via Ri (A1) adenosine receptors, but a direct demonstration of these receptors is still missing. In the present study we report direct labelling of these receptors with (-)N6-[125I]-p-hydroxyphenylisopropyladenosine [( 125I]HPIA)1. The radioligand bound in a saturable and reversible manner to a crude membrane preparation, the Bmax-value was 30.5 fmol/mg protein and the KD-value 1.1 nmol/l. A similar affinity of the ligand was obtained in kinetic and competition experiments. Competition experiments with a variety of adenosine analogues gave a pharmacological profile characteristic of Ri adenosine receptors with high affinities of N6-substituted derivatives and a marked stereospecificity for N6-phenylisopropyladenosine (PIA). Purification of the membrane preparation by density gradient centrifugation resulted in a 30-fold increase in the number of binding sites which was paralleled by a similar increase in the number of binding sites for [3H]ouabain. Guanine nucleotides decreased binding of [125I]HPIA in a dose-dependent manner, but the IC50-values were considerably higher than those reported in other tissues. Finally, binding of [125I]HPIA appeared to be entropy-driven which has been shown to be characteristic of agonist binding to Ri adenosine receptors. These results suggest the presence of Ri adenosine receptors in ventricular myocardium which may be responsible for the mediation of the effects of adenosine and its analogues.