title: Estimation of agonist activity at G protein-coupled receptors: analysis of M2 muscarinic receptor signaling through Gi/o,Gs, and G15.
abstract: We developed novel methods for analyzing the concentration-response curve of an agonist to estimate the product of observed affinity and intrinsic efficacy, expressed relative to that of a standard agonist. This parameter, termed intrinsic relative activity (RA(i)), is most applicable for the analysis of responses at G protein-coupled receptors. RA(i) is equivalent to the potency ratios that agonists would exhibit in a hypothetical, highly sensitive assay in which all agonists behave as full agonists, even those with little intrinsic efficacy. We investigated muscarinic responses at the M(2) receptor, including stimulation of phosphoinositide hydrolysis through G(alpha15) in HEK 293T cells, inhibition of cAMP accumulation through G(i) in Chinese hamster ovary (CHO) cells, and stimulation of cAMP accumulation through G(s) in CHO cells treated with pertussis toxin. The RA(i) values of carbachol, oxotremorine-M, and the enantiomers of aceclidine were approximately the same in the three assay systems. In contrast, the activity of 4-[[N-[3-chlorophenyl]carbamoy]oxy-2-butynyl]trimethylammonium chloride (McN-A-343) was approximately 10-fold greater at M(2) receptors coupled to G(alpha15) in HEK 293T cells compared with M(2) receptors coupled to G(i) in the same cells or in CHO cells. Our results show that the RA(i) estimate is a useful measure for quantifying agonist activity across different assay systems and for detecting agonist directed signaling.
authors: Department of Physical Sciences, Chapman University, Orange, California, USA.
authors: Michael T
authors: MT
authors: Griffin
authors: Katherine W
authors: KW
authors: Figueroa
authors: Sarah
authors: S
authors: Liller
authors: Frederick J
authors: FJ
authors: Ehlert
mesh_terms: D008455:(4-(m-Chlorophenylcarbamoyloxy)-2-butynyl)trimethylammonium Chloride / Q000494:pharmacology; D000465:Algorithms; D000818:Animals; D016466:CHO Cells; D002217:Carbachol / Q000494:pharmacology; D002460:Cell Line; D005576:Colforsin / Q000494:pharmacology; D006224:Cricetinae; D003412:Cricetulus; D000242:Cyclic AMP / Q000378:metabolism; D004158:Diphenylacetic Acids / Q000494:pharmacology; D019206:GTP-Binding Protein alpha Subunits, Gi-Go / Q000037:antagonists & inhibitors / Q000378:metabolism*; D043802:GTP-Binding Protein alpha Subunits, Gq-G11 / Q000235:genetics / Q000378:metabolism*; D019205:GTP-Binding Protein alpha Subunits, Gs / Q000378:metabolism*; D020962:Heterotrimeric GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D007545:Isoproterenol / Q000494:pharmacology; D010095:Oxotremorine / Q000031:analogs & derivatives / Q000494:pharmacology; D037342:Pertussis Toxin / Q000494:pharmacology; D010716:Phosphatidylinositols / Q000378:metabolism; D010880:Piperidines / Q000494:pharmacology; D010890:Pirenzepine / Q000031:analogs & derivatives / Q000494:pharmacology; D011812:Quinuclidines / Q000494:pharmacology; D043585:Receptor, Muscarinic M2 / Q000819:agonists* / Q000037:antagonists & inhibitors / Q000235:genetics; D015398:Signal Transduction / Q000187:drug effects*; D014162:Transfection
chemical_list: D004158:Diphenylacetic Acids; D010716:Phosphatidylinositols; D010880:Piperidines; D011812:Quinuclidines; D043585:Receptor, Muscarinic M2; C084650:aceclidine; C082230:N-(2-bromoethyl)-4-piperidinyl diphenylacetate; D005576:Colforsin; D010890:Pirenzepine; D008455:(4-(m-Chlorophenylcarbamoyloxy)-2-butynyl)trimethylammonium Chloride; D010095:Oxotremorine; C042743:oxotremorine M; D002217:Carbachol; D000242:Cyclic AMP; D037342:Pertussis Toxin; C112631:G protein alpha 16; D019206:GTP-Binding Protein alpha Subunits, Gi-Go; D043802:GTP-Binding Protein alpha Subunits, Gq-G11; D019205:GTP-Binding Protein alpha Subunits, Gs; D020962:Heterotrimeric GTP-Binding Proteins; D007545:Isoproterenol; C048593:otenzepad