Agonist-independent activation of Gz by the 5-hydroxytryptamine1A receptor co-expressed in Spodoptera frugiperda cells. Distinguishing inverse agonists from neutral antagonists.
The human 5-hydroxytryptamine1A receptor, when expressed in Spodoptera frugiperda (Sf9) cells, facilitates the binding of [35S]GTPgammaS to a co-expressed GTP-binding regulatory protein, Gz, consistent with constitutive activity. The antagonists 4-(2'-methoxyphenyl)-1-[2'(n-2"-pyridinyl)-p-iodobenzamido]ethyl-p ipe razine (p-MPPI) and the related fluorobenzamido analogue p-MPPF had little (p-MPPI) or no (p-MPPF) effect on this activity. In contrast, a third antagonist, the neuroleptic spiperone, produced an almost complete suppression. Thus, using G protein activation as an index of receptor activity, p-MPPF was classified as a neutral antagonist, p-MPPI as a partial inverse agonist, and spiperone as essentially a full inverse agonist. As predicted, spiperone displayed properties consistent with a special form of noncompetitive antagonism when used to displace the agonist [125I]R-(+)-trans-8-hydroxy-2-[N-n-propyl-N-(3'-iodo-2'-propenyl)amin o]tetralin. Our data profile Sf9 cells as a unique vehicle for the characterization of inverse agonists, as these cells support a systematic pairing of mammalian receptors and G proteins, quantitative assays of G protein activation, and unambiguously labeled populations of coupled and uncoupled receptors.
Barr
A J
AJ
Department of Pharmacology, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania 19104-6084, USA.
Manning
D R
DR
D000631:Aminopyridines / Q000378:metabolism; D000818:Animals; D004791:Enzyme Inhibitors / Q000378:metabolism; D044385:GTP-Binding Protein alpha Subunits; D019204:GTP-Binding Proteins / Q000378:metabolism*; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000378:metabolism; D020962:Heterotrimeric GTP-Binding Proteins; D006801:Humans; D007700:Kinetics; D008024:Ligands; D010879:Piperazines / Q000378:metabolism; D011985:Receptors, Serotonin / Q000378:metabolism*; D044263:Receptors, Serotonin, 5-HT1; D012702:Serotonin Antagonists / Q000378:metabolism*; D013134:Spiperone / Q000378:metabolism; D018411:Spodoptera; D013764:Tetrahydronaphthalenes / Q000378:metabolism
D000631:Aminopyridines; D004791:Enzyme Inhibitors; C099387:GNAZ protein, human; D044385:GTP-Binding Protein alpha Subunits; D008024:Ligands; D010879:Piperazines; D011985:Receptors, Serotonin; D044263:Receptors, Serotonin, 5-HT1; D012702:Serotonin Antagonists; D013764:Tetrahydronaphthalenes; C087205:4-(2'-methoxyphenyl)-1-(2'-(N-(2''-pyridinyl)-4-iodobenzamido)ethyl)piperazine; C083828:8-hydroxy-2-(N-n-propyl-N-(3'-iodo-2'-propenyl)amino)tetralin; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D013134:Spiperone; C099463:4-(2' methoxyphenyl)-1-(2'-(N-(2''-pyridinyl)-4-fluorobenzamido)ethyl)piperazine; D019204:GTP-Binding Proteins; D020962:Heterotrimeric GTP-Binding Proteins