ALPHA(2)-ADRENOCEPTOR REGULATION OF ADENYLYL-CYCLASE IN CHO CELLS - DEPENDENCE ON RECEPTOR DENSITY, RECEPTOR SUBTYPE AND CURRENT ACTIVITY OF ADENYLYL-CYCLASE

Chinese hamster ovary (CHO) cells stably transfected to express differ ent densities of the human alpha(2A)-, alpha(2B)- and alpha(2C)-adreno ceptor subtypes, were used to characterize the regulation of adenylyl cyclase activity by alpha(2)-adrenoceptor agonists. In isolated cell m embranes, activation of alpha(2A)- and alpha(2C)-adrenoceptors did not affect basal enzyme activity, but activation of alpha(2B)-adrenocepto rs stimulated adenylyl cyclase activity, The extent of stimulation was dependent on the receptor density and was insensitive to pertussis to xin treatment. In the presence of 10 mu M forskolin all three receptor subtypes mediated inhibition of adenylyl cyclase activity in a pertus sis toxin-sensitive manner. In experiments performed with intact cells the same pattern could be seen: the basal production of cAMP was not affected when alpha(2C)-adrenoceptors were activated, but activated al pha(2B)-adrenoceptors mediated stimulation of cAMP production. In the presence of forskolin, both receptor subtypes mediated inhibition of c AMP production. Our results suggest that alpha(2B)-adrenoceptors are c oupled 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 cu rrent activity of adenylyl cyclase. The results also suggest that the alpha(2A)- and alpha(2C)-subtypes are preferentially coupled to G(i) a nd transduce only inhibition of adenylyl cyclase activity in transfect ed CHO cells. At low densities of alpha(2C)-adrenoceptors, clonidine w as a partial agonist, but in clones expressing high levels of alpha(2C )-adrenoceptors, clonidine acted as a full agonist by inhibiting cAMP accumulation with the same efficacy as (-)-noradrenaline. This demonst rates that receptor reserve can mask partial agonist activity. (C) 199 7 Elsevier Science B.V.