α2-Adrenoceptor regulation of adenylyl cyclase in CHO cells: Dependence on receptor density, receptor subtype and current activity of adenylyl cyclase

Abstract

Chinese hamster ovary (CHO) cells stably transfected to express different densities of the human α 2A-, α 2B- and α 2C-adrenoceptor subtypes, were used to characterize the regulation of adenylyl cyclase activity by α 2-adrenoceptor agonists. In isolated cell membranes, activation of α 2A- and α 2C-adrenoceptors did not affect basal enzyme activity, but activation of α 2B-adrenoceptors stimulated adenylyl cyclase activity. The extent of stimulation was dependent on the receptor density and was insensitive to pertussis toxin treatment. In the presence of 5 μM forskolin all three receptor subtypes mediated inhibition of adenylyl cyclase activity in a pertussis toxin-sensitive manner. In experiments performed with intact cells the same pattern could be seen: the basal production of cAMP was not affected when α 2C-adrenoceptors were activated, but activated α 2B-adrenoceptors mediated stimulation of cAMP production. In the presence of forskolin, both receptor subtypes mediated inhibition of cAMP production. Our results suggest that α 2B-adrenoceptors are coupled to both G i and G s proteins. The signal transduction pathway to which the receptor is coupled is not dependent on receptor density, but its effect on adenylyl cyclase regulation is dependent on the current activity of adenylyl cyclase. The results also suggest that the α 2A- and α 2C-subtypes are preferentially coupled to G i and transduce only inhibition of adenylyl cyclase activity in transfected CHO cells. At low densities of α 2C-adrenoceptors, clonidine was a partial agonist, but in clones expressing high levels of α 2C-adrenoceptors, clonidine acted as a full agonist by inhibiting cAMP accumulation with the same efficacy as (−)-noradrenaline. This demonstrates that receptor reserve can mask partial agonist activity.