Species Differences in Ligand Affinity at Central A₃-Adenosine Receptors

[Table: see text] Binding affinities of purine derivatives at A₃ adenosine receptors in different species were compared. Binding was carried out using the novel high affinity agonist ligand [¹²⁵I]AB-MECA (3-iodo-4-aminobenzyladenosine-5'-N-methyluronamide) in the presence of 1.0 microM XAC (8-[4-[[[[(2-aminoethyl)amino]carbonyl]methyl]oxy]phenyl]-1,3-dipropylxanthine), an A₁- and A_2a-adenosine antagonist. XAC was added to eliminate binding to non-A₃ receptors. In rat brain membranes [¹²⁵I]AB-MECA exhibited saturable, specific binding with a K_d of 2.28 nM and a B_max of 43 fmol/mg protein. The affinity of [¹²⁵I]AB-MECA at the gerbil and rabbit brain A₃-receptors was similar to the rat, suggesting that the affinity of this agonist is not highly species dependent. The affinity of various xanthine derivatives was measured in [¹²⁵I]AB-MECA competition binding assays. Gerbil and rabbit brain A₃-receptors were similar in the affinity of antagonists whose potency order in both species was: BWA522 ≥ CPX > XCC, XAC, SPX, BWA1433 > theophylline. The affinities of 8-arylxanthines at the rat, rabbit, and gerbil brain A₃ receptors were considerably less than the previously reported affinities at cloned sheep and human A₃ receptors. Species differences in agonist affinity were assessed by comparing K_i values at cloned rat brain A₃ receptors expressed in CHO cells with cloned sheep and human A₃ receptors. Human and rat brain A₃ receptors were highly similar in the relative affinities of agonists, and sheep brain A₃ receptors were unlike either human or rat A₃ receptors in agonist affinity.