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Method for identifying ligands activating either excitatory or inhibitory G-protein-coupled receptors by functional coexpression in Xenopus oocytes.

Xenopus oocytes devoid of their follicular enclosure provide a frequently used expression system for investigating receptors that transduce through activation of adenylyl cyclase following injection of the appropriate mRNA. However, due to a low basal activity of the cyclase they cannot be utilized to investigate receptor-mediated reductions in endogenous cAMP levels. In order to overcome this limitation, a model was designed in which test clones for such inhibitory receptors were co-expressed with a beta 2-adrenoceptor, which elevated cAMP upon exposure to isoproterenol. Following injection of mRNA to express the alpha 2 test receptor in the oocytes, marked reduction in cAMP could be measured after exposure to clonidine. Attenuation of cAMP levels was also seen following co-expression of the dopamine D2 receptor along with dopamine administration. Thus, after inducing a receptor-mediated tone in adenylyl cyclase activity, Xenopus oocytes can be conveniently used to study also ligands that bind to inhibitory G-protein coupled receptors.