Differential A 1 ‐adenosine receptor reserve for inhibition of cyclic AMP accumulation and G‐protein activation in DDT 1 MF‐2 cells

The A 1 ‐adenosine receptor (A 1 AdoR) reserve for N 6 ‐cyclopentyladenosine (CPA) mediated inhibition of (−)isoprenaline stimulated cyclic AMP accumulation and stimulation of [ 35 S]‐guanosine‐5′‐O‐(thiotriphosphate) (GTPγS) binding, a measure of guanine nucleotide binding protein (G‐protein) activation, was determined in DDT 1 MF‐2 cells. Inactivation of the A 1 AdoRs with the chemoreactive ligand 8‐cyclopentyl‐3‐[3‐[[4‐(fluorosulphonyl)benzoyl]oxy]propyl]‐1‐propylxanthine (FSCPX) caused a progressive rightward shift of the concentration‐response curves for CPA to inhibit cyclic AMP accumulation, with a maximum of 10 fold increase in the EC 50 value. In contrast, inactivation of A 1 AdoR's caused only a 1.7 fold rightward shift in the CPA concentration‐response for stimulation of [ 35 S]‐GTPγS binding. The A 1 AdoR occupancy‐response relationship for CPA inhibition of cyclic AMP accumulation was hyperbolic with 43% receptor occupancy required to elicit the maximal response, i.e. a 57% A 1 AdoR reserve. In contrast, the A 1 AdoR occupancy‐response relationship for CPA mediated stimulation of [ 35 S]‐GTPγS binding was linear indicating little or no receptor reserve for G‐protein activation. The relationship between CPA stimulation of [ 35 S]‐GTPγS binding and cyclic AMP inhibition was also hyperbolic with 44% G‐protein activation sufficient to cause maximal inhibition. The data suggest that the A 1 AdoR reserve for CPA mediated inhibition of cyclic AMP accumulation occurs at the level of G‐protein interaction with adenylyl cyclase. However, each A 1 AdoR appears to activate a constant fraction of the total G‐protein population suggesting signal amplification at the receptor‐G‐protein level which may also contribute to the receptor reserve for CPA.British Journal of Pharmacology (2000) 130 , 1156–1164; doi: 10.1038/sj.bjp.0703405