Functional coupling of the human dopamine D 2 receptor with Gα i1 , Gα i2 , Gα i3 and Gα o G proteins: evidence for agonist regulation of G protein selectivity

The human dopamine D 2long (D 2L ) receptor was expressed with four different G proteins in Sf9 cells using the baculovirus expression system. When co‐expressed with G i /G o G proteins (G i1 α, G i2 α, G i3 α, or G o α, plus Gβ 1 and Gγ 2 ), the receptor displayed a high‐affinity binding site for the agonists (dopamine and NPA), which was sensitive to GTP (100 μ M ), demonstrating interaction between the receptor and the different G proteins. The receptor to G protein ratio (R : G ratio) was evaluated using [ 3 H]‐spiperone saturation binding (R) and [ 35 S]‐GTPγS saturation binding (G). R : G ratios of 1 : 12, 1 : 3, 1 : 14 and 1 : 5 were found for G i1 , G i2 , G i3 , and G o preparations, respectively. However, when R : G ratios of 1 : 2 and 1 : 12 were compared for G i2 and G o , no difference was found for the stimulation of [ 35 S]‐GTPγS binding. Several agonists were tested for their ability to stimulate [ 35 S]‐GTPγS binding to membranes co‐expressing the receptor and various G proteins. All the compounds tested showed agonist activity in preparations expressing G i3 and G o . However, for G i2 and G i1 preparations, compounds such as S ‐(−)‐3‐PPP and p ‐tyramine were unable to stimulate [ 35 S]‐GTPγS binding. Most of the compounds showed higher relative efficacies (compared to dopamine) and higher potencies in the preparation expressing G o . Comparison of the effects of different agonists in the different preparations showed that each agonist differentially activates the four G proteins. We conclude that the degree of selectivity of G protein activation by the D 2L receptor can depend on the conformation of the receptor stabilised by an agonist.British Journal of Pharmacology (2003) 138 , 775–786. doi: 10.1038/sj.bjp.0705116