1. The A1-adenosme receptor (A1AdoR) reserve for N6-cyclopentyladenosine (CPA) mediated inhibition of (-)isoprenaline stimulated cyclic AMP accumulation and stimulation of [35S]guanosine-5'-O-(thiotriphosphate) (GTPγS) binding, a measure of guanine nucleotide binding protein (G-protein) activation, was determined in DDT1MF-2 cells. 2. inactivation of the A1AdoRs 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 EC50value. In contrast, inactivation of A1AdoR's caused only a 1.7 fold rightward shift in the CPA concentration-response for stimulation of [35S]-GTPγS binding. 3. The A1AdoR 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% A1AdoR reserve. In contrast, the A1AdoR occupancy-response relationship for CPA mediated stimulation of [35S]-GTPγS binding was linear indicating little or no receptor reserve for G-protein activation. The relationship between CPA stimulation of [35S]-GTPγS binding and cyclic AMP inhibition was also hyperbolic with 44% G-protein activation sufficient to cause maximal inhibition. 4. The data suggest that the A1AdoR reserve for CPA mediated inhibition of cyclic AMP accumulation occurs at the level of G-protein interaction with adenylyl cyclase. However, each A1AdoR 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.
- 8-Cyclopentyl-3-[3-[[4(fluorosulphonyl)benzoyl]oxy]propyl]-1-propylxanthine (FSCPX)
- Cyclic AMP
- Irreversible antagonist
- Receptor reserve