Constitutive activity of GPR40/FFA1 intrinsic or assay dependent?

Free fatty acid receptor 1 (FFA1; previously designated GPR40) is a potential therapeutic target for the treatment of diabetes and related metabolic disorders. Agonist-independent or constitutive activity is a feature associated with essentially all G protein-coupled receptors but the extent of this varies substantially between family members. In many situations, detection of such activity can be both assay- and context-dependent and may reflect the presence in the assay of an endogenous agonist. In studies on FFA1, experiments employing cell membrane preparations and the binding of [(35)S]guanosine 5'-O-[γ-thio]triphosphate to G proteins produce data consistent with a high-level constitutive activity of this receptor. Herein, we detail these assays and discuss approaches to determine if this is a measure of intrinsic receptor constitutive activity or if such results reflect the presence of an endogenous agonist. FFA1 is coupled predominantly to G proteins of the Gα(q) subfamily. Activation of the receptor results, therefore, in the transient elevation of intracellular [Ca(2+)]. We also detail assays to measure such signals and consider whether they are appropriate to detect receptor constitutive activity