A 2a/D 2 receptor interactions are not observed in COS-7 cells transiently transfected with dopamine D 2 and adenosine A 2a receptor cDNA

Abstract

The rat D 2 receptor and the dog A 2a receptor subcloned into the pXM vector were transiently transfected into COS-7 cells using the DEAE-dextran method. The transfected cells expressed approx. 200 fmol D 2 receptors/mg protein and approx. 5 pmol/mg protein of the A 2a receptor as judged by binding experiments with [ 3H]raclopride [ or[ 3 H]-N- propyl-apomorphine (NPA)] and [ 3H]-CGS 21680, respectively. The high affinity K D values were 0.43 and 19 nM for D 2 and A 2a receptors, respectively, in agreement with results obtained from other cells and tissues. The non-selective adenosine receptor agonist NECA stimulated cAMP accumulation both in non-transfected and transfected COS-7 cells with only a slight difference in potency, suggesting that most of the stimulation is due to activation of A 2b receptors known to be present on virtually every cell. The two A 2a selective agonists CGS 21680 and CV-1808 were essentially inactive in transfected COS-7 cells, but were very active in PC-12 cells known to possess functional A 2a receptors. Dopamine did not decrease cAMP accumulation in the transfected COS-7 cells. CGS 21680 (30 nM) did not affect the binding characteristics of D 2 receptors in the co-transfected COS-7 cells in contrast to the increased K H , K L and R H values found previously in rat striatal membranes after CGS 21680 treatment. The present findings indicate that transiently transfected A 2a and D 2 receptors in COS-7 cells have normal binding properties, but couple poorly to adenylyl cyclase, despite the presence of G s protein and adenylyl cyclase in these cells. Our results also demonstrate that the previously reported interactions between A 2a receptors and D 2 receptors do not occur when only the receptor proteins are expressed in COS-7 cells, suggesting that the two receptor molecules do not interact directly to influence binding characteristics.