Heterogeneity of adenosine A₁ receptor binding in brain tissue

A possible heterogeneity of adenosine A₁ receptors in brain was evaluated by comparing drug specificity, regional distribution, and temperature influences on ³H-labeled ligand binding in several mammalian species. Adenosine receptors were labeled with the agonists [³H] N⁶-cyclohexyladenosine ([³H]CHA) and [³H]N⁶-phenylisopropyladenosine ([³H]PIA) or the antagonist [³H]1,3-diethyl-8-phenylxanthine ([³H]DPX). The binding properties of adenosine analogues are similar in the species studied. In all cases, the N⁶-substituted adenosine analogues have nanomolar affinity for [³H]CHA binding sites with a marked stereoselectivity for the isomers of PIA. The xanthine derivative DPX manifests considerable differences in affinity for [³H]CHA-labeled A₁ receptors of different species. DPX shows highest affinity for A₁ receptors in calf brain and lowest affinity for A₁ receptors in guinea pig and human brain. [³H]DPX labels adenosine A₁ receptors in calf, rat, and rabbit cerebral cortical brain membranes. By contrast, very little specific [³H]DPX binding is detected in human and guinea pig brain, and the limited level of binding is inhibited poorly by PIA or xanthines. The failure of [³H]DPX to label adenosine A₁ receptors in human and guinea pig cortex may be related to the very low affinity of DPX for A₁ receptors in these species as labeled by [³H]CHA. Temperature variations affect agonist and antagonist binding differentially. Adenosine derivatives have higher affinities for A₁ receptors at higher temperatures, whereas xanthine analogues have higher affinities at lower temperatures. Similar temperature effects on A₁ receptor binding occur in all species examined. Temperature effects can account for the differences in drug affinities at [³H]agonist- and [³H]antagonist-labeled sites. Temperature effects cannot account for the failure of [³H]DPX to label A₁ receptors in guinea pig and human brain. Our thermodynamic analysis of temperature influences on A₁ receptor binding indicates that agonist binding is entropy-driven.