Allosteric modulation of the rat adenosine A1 receptor:: Differential effects on agonist and antagonist binding

The interaction of 2-amino-3-benzoyl-thiophene derivative, PD81,723, as well as other G protein-coupled receptor (GPCR)-modulating agents such as suramin (SUR), N-ethylmaleimide (NEM), GTP, and NaCl with the rat adenosine A(1) receptor was investigated using kinetic, saturation, as well as displacement 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 present. 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 "active" (R*) state with high affinity for agonists and low for antagonists. Further, all described allosteric modulators influenced both agonist and antagonist 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 combined presence of NEM, NaCl, and GTP, represented a greater than 600-fold affinity 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 Wiley-Liss.