Allosteric modulators affect the internalization of human adenosine A1 receptors.

ABSTRACT

To study the effect of allosteric modulators on the internalization of human adenosine A(1) receptors, the receptor was equipped with a C-terminal yellow fluorescent protein tag. The introduction of this tag did not affect the radioligand binding properties of the receptor. CHO cells stably expressing this receptor were subjected during 16 h to varying concentrations of the agonist N(6)-cyclopentyladenosine (CPA) in the absence or presence of 10 microM of the allosteric enhancer PD 81,723 ((2-amino-4,5-dimethyl-3-thienyl)-[3-(trifluoromethyl)phenyl]methanone) or the allosteric inhibitor SCH-202676 (N-(2,3-diphenyl-1,2,4-thiadiazol-5(2H)-ylidene)methanamine). CPA itself was able to internalize 25% and 40% of the receptors at a concentration of 400 nM or 4 muM, respectively. Addition of either PD 81,723 or SCH-202676 alone had no effect on internalization. However, with PD 81,723 a slight amount of internalization was obtained already at 40 nM of CPA and at 400 nM CPA 59% of the receptors internalized. SCH-202676 on the other hand effectively prevented CPA-induced internalization of the receptor.