Allosteric modulation of the rat adenosine A(1) receptor: Differential effects on agonist and antagonist binding

The interaction of 2-amino-3-benzoyl-thiophene derivative, PD81,723, as wel l as other G protein-coupled receptor (GPCR)-modulating agents such as sura min (SUR), N-ethylmaleimide (NEM), GTP, and NaCl with the rat adenosine A(1 ) receptor was investigated using kinetic, saturation, as well as displacem ent experiments of various agonists, partial agonists or antagonists. PD81, 723 enhanced agonist (CPA, R-PIA, NECA) binding similar to2-fold, while its effect on CPA binding was increased (4-11-fold) with other modulators pres ent. In contrast, binding of antagonists (DPCPX, CPT, N-0840) was inhibited , while binding of partial agonists (8BCPA, MeSCPA) remained uninfluenced. The effect of PD81,723 is consistent with shifting Al receptors to an "acti ve" (R*) state with high affinity for agonists and low for antagonists. Fur ther, all described allosteric modulators influenced both agonist and antag onist binding. The IC50 values observed for the agonist CPA, ranging from 4 .7 nM in the presence of PD81,723 to a high value of 2.9 muM in the combine d presence of NEM, NaCl, and GTP, represented a greater than 600-fold affin ity shift. We suggest that the latter micromolar IC50 value may approximate CPA's "true" affinity (K-A) for the rat adenosine Al receptor. (C) 2001 Wi ley-Liss.