History of the G Protein–Coupled Receptor (GPCR) Assays From Traditional to a State-of-the-Art Biosensor Assay

Abstract

Abstract. The G protein-coupled receptors (GPCRs) form the largest and the most versatile superfamily that share a seven-transmembrane-spanning architecture. GPCR-signaling is involved in vision, taste, olfaction, sympathetic/parasympathetic nervous functions, metabolism, and immune regulation, indicating that GPCRs are extremely important therapeutic targets for various diseases. Cellular dielectric spectroscopy (CDS) is a novel technology that employs a label-free, real-time and cell-based assay approach for the comprehensive pharmacological evaluation of cells that exogenously or endogenously express GPCRs. Among the biosensors that use CDS technology, the CellKey TM system not only detects the activation of GPCRs but also distinguishes between signals through different subtypes of the Ga protein (Gs, Gi/o, and Gq). In this review, we discuss the traditional assays and then introduce the principles by which the CellKey TM system evaluates GPCR activation, followed by a perspective on the advantages and future prospects of this system.

Tricyclic Antidepressant Amitriptyline-induced Glial Cell Line-derived Neurotrophic Factor Production Involves Pertussis Toxin-sensitive Gαi/o Activation in Astroglial Cells

“…This study demonstrated that the PTX-sensitive G i/o subunits, especially G o and G i3 , play important roles in the activation of ERK evoked by amitriptyline. The CellKey TM assay confirmed the pharmacological and siRNA findings, in that the pattern of the change in impedance induced by amitriptyline was similar to that of activation of G i/o -coupled receptors, and not of G s -or G q -coupled receptors (13,14,17,18). Although the C6 cells is an astrocyte model cell line derived from rat glioma, activation of the MMP/FGFR/FRS2␣/ERK cascade occurs in both normal human and rat astrocytes as well (9,11), in a PTX-sensitive manner.…”

“…The G ␣ family of G proteins is composed of the G s , G i/o , and G q subfamilies (14). Among the G protein inhibitors used in this study, only PTX inhibited amitriptyline-induced GDNF production and the amitriptyline-mediated response of the cascade (FRS2␣ and ERK), indicating that PTX-sensitive G i/o plays a critical role in the intracellular response to amitriptyline.…”

“…The effect of amitriptyline on G protein activation in living C6 cells and rat astrocytes was analyzed using a novel biosensor technology, the CellKey TM assay (MDS Sciex, Ontario, Canada). The CellKey TM assay utilizes cellular dielectric spectroscopy, which is specifically tailored to G protein-coupled receptor (GPCR) detection and can distinguish signals between the G␣ s (G s ), G␣ i/o (G i/o ), and G␣ q (G q ) subfamilies (13,14).…”

“…To show typical different ⌬Z values caused by each ligand to specific types of GPCRs, HEK293 cells expressing -opioid receptors, coupled to G i/o , and C6 cells endogenously expressing ␤-adrenergic receptors, coupled to G s , were used (14,35). In addition, HEK293 cells, which endogenously express the muscarinic (M 3 ) acetylcholine receptor, were used to show the activation of G q -coupled receptors (36,37).…”

Mechanisms involved in the regulation of neuropeptide-mediated neurite outgrowth: a minireview

“…With regard to the neuropeptides, it is important to distinguish G protein subunits with different functional consequences. Based on the amino acid sequence homologies of Gα subunit, G proteins are classifi ed into Gαs (Gs), Gαi/o (Gi/o), Gαq (Gq) (Miyano et al 2014). Gs protein activates adenylate cyclase (AC) that induces an increase in the intracellular concentration of cyclic adenosine monophosphate (cAMP) and activation of protein kinase A (PKA).…”