Mepyramine, a Histamine H1 Receptor Inverse Agonist, Binds Preferentially to a G Protein-coupled Form of the Receptor and Sequesters G Protein

Accurate characterization of the molecular mechanisms of the action of ligands is an extremely important issue for their appropriate research, pharmacological, and therapeutic uses. In view of this fact, the aim of the present work was to investigate the mechanisms involved in the actions of mepyramine at the guinea pig H1 receptor stably expressed in Chinese hamster ovary cells. We found that mepyramine is able to decrease the basal constitutive activity of the guinea pig H1 receptor, to bind with high affinity to a Gq/11 protein-coupled form of the receptor and to promote a G protein-coupled inactive state of the H1 receptor that interferes with the Gq/11-mediated signaling of the endogenously expressed ATP receptor, as predicted by the Cubic Ternary Complex Model of receptor occupancy. The effect of mepyramine on ATP-induced signaling was specifically neutralized by Gα11 overexpression, indicating that mepyramine is able to reduce G protein availability for other non-related receptors associated with the same signaling pathway. Finally, we found a loss of mepyramine efficacy in decreasing basal levels of intracellular calcium at high Gα11 expression levels, which can be theoretically explained in terms of high H1 receptor constitutive activity. The whole of the present work sheds new light on H1 receptor pharmacology and the mechanisms H1 receptor inverse agonists could use to exert their observed negative efficacy. Accurate characterization of the molecular mechanisms of the action of ligands is an extremely important issue for their appropriate research, pharmacological, and therapeutic uses. In view of this fact, the aim of the present work was to investigate the mechanisms involved in the actions of mepyramine at the guinea pig H1 receptor stably expressed in Chinese hamster ovary cells. We found that mepyramine is able to decrease the basal constitutive activity of the guinea pig H1 receptor, to bind with high affinity to a Gq/11 protein-coupled form of the receptor and to promote a G protein-coupled inactive state of the H1 receptor that interferes with the Gq/11-mediated signaling of the endogenously expressed ATP receptor, as predicted by the Cubic Ternary Complex Model of receptor occupancy. The effect of mepyramine on ATP-induced signaling was specifically neutralized by Gα11 overexpression, indicating that mepyramine is able to reduce G protein availability for other non-related receptors associated with the same signaling pathway. Finally, we found a loss of mepyramine efficacy in decreasing basal levels of intracellular calcium at high Gα11 expression levels, which can be theoretically explained in terms of high H1 receptor constitutive activity. The whole of the present work sheds new light on H1 receptor pharmacology and the mechanisms H1 receptor inverse agonists could use to exert their observed negative efficacy. Traditionally, ligands acting at G protein-coupled receptors (GPCRs) 1The abbreviations used are: GPCR, guanine nucleotide binding regulatory protein (G protein)-coupled receptor; CTC, Cubic Ternary Complex Model; gp-H1r, guinea pig histamine H1 receptor; GTPγS, guanosine 5′-O-(3-thio)triphosphate; mepyramine, 2-((2-(dimethylamino)ethyl)(p-methoxybenzyl)amino)-pyridine; triprolidine, (E)-2-[1-(4-methylphenyl)-3-(1-pyrrolidinyl)-1-propenyl]pyridine; CHO, Chinese hamster ovary; DMEM, Dulbecco's modified Eagle's medium; PTX, pertussis toxin; InsP, inositol phosphate. were classified as agonists, accounting for ligands with observable positive efficacy, or as antagonists, for ligands with no observable efficacy. However, the identification of constitutively active receptors led to the identification of ligands with intrinsic negative efficacy, characterized by their ability to decrease basal constitutive receptor activity. In view of this property, those ligands were defined as inverse agonists (1Milligan G. Bond R.A. Lee M. Trends Pharmacol. Sci. 1995; 16: 10-13Abstract Full Text PDF PubMed Scopus (286) Google Scholar). Considering the importance accurate ligand classification has in terms of proper pharmacological use, the mechanism(s) by which these ligands exert their effects has to be extensively and carefully characterized (2Strange P.G. Trends Pharmacol. Sci. 2002; 23: 89-95Abstract Full Text Full Text PDF PubMed Scopus (116) Google Scholar). In an attempt to understand such mechanisms, several receptor occupancy models have been developed (3Leff P. Trends Pharmacol. Sci. 1995; 16: 89-97Abstract Full Text PDF PubMed Scopus (477) Google Scholar, 4Samama P. Cotecchia S. Costa T. Lefkowitz R.J. J. Biol. Chem. 1993; 268: 4625-4636Abstract Full Text PDF PubMed Google Scholar, 5Weiss J.M. Morgan P. Lutz M. Kenakin T.P. J. Theor. Biol. 1996; 178: 151-167Crossref Scopus (178) Google Scholar, 6Weiss J.M. Morgan P. Lutz M. Kenakin T.P. J. Theor. Biol. 1996; 178: 169-182Crossref Scopus (97) Google Scholar, 7Weiss J.M. Morgan P.H. Lutz M.W. Kenakin T.P. J. Theor. Biol. 1996; 181: 381-397Crossref PubMed Scopus (165) Google Scholar). For all of the currently accepted models, basal activity is proportional to the population of receptors active and coupled to G protein. The simplest explanation for the negative efficacy observed for inverse agonists is, then, the selective affinity of the ligand for an inactive state of the receptor. Binding of the ligand would then deplete the levels of active receptors coupled to G protein, resulting in a decrease in receptor constitutive activity. Alternatively, ligands with observed negative efficacy could promote uncoupling from G proteins, leading to G protein-free receptor species, which are unable to evoke response. However, the Cubic Ternary Complex Model (CTC) predicts a third mechanism accounting for negative efficacy (5Weiss J.M. Morgan P. Lutz M. Kenakin T.P. J. Theor. Biol. 1996; 178: 151-167Crossref Scopus (178) Google Scholar, 6Weiss J.M. Morgan P. Lutz M. Kenakin T.P. J. Theor. Biol. 1996; 178: 169-182Crossref Scopus (97) Google Scholar, 7Weiss J.M. Morgan P.H. Lutz M.W. Kenakin T.P. J. Theor. Biol. 1996; 181: 381-397Crossref PubMed Scopus (165) Google Scholar). According to the CTC, a ligand may bind with high affinity to a G protein-coupled but inactive form of the receptor, resulting in a decrease of spontaneous receptor activity. The distinctive feature of this proposed mechanism is that, by promoting receptor-G protein coupling, binding of the ligand would result in a reduction of the G protein availability to other receptors associated with the same pathway, thus interfering with the signaling of those GPCRs (8Monczor F. Fernandez N. Legnazzi B.L. Riveiro M.E. Baldi A. Shayo C. Davio C. Mol. Pharmacol. 2003; 64: 512-520Crossref PubMed Scopus (37) Google Scholar). Importantly, experimental evidence supporting all the three aforementioned mechanisms has been reported for different inverse agonists. Mepyramine, traditionally classified as a histamine H1 receptor antagonist, is the reference ligand for studying H1 receptor pharmacology (9van der Goot H. Timmerman H. Eur. J. Med. Chem. 2000; 35: 5-20Crossref PubMed Scopus (106) Google Scholar). Together with other formerly named H1 antagonists, it has been recently reclassified as an inverse agonist, in view of its negative efficacy at the constitutively active human H1 receptor observed in transiently transfected COS-7 cells (10Bakker R.A. Wieland K. Timmerman H. Leurs R. Eur. J. Pharmacol. 2000; 387: 5-7Crossref PubMed Scopus (153) Google Scholar). Taking into account the clinically widespread use of histamine H1 receptor antihistamines in the treatment of human diseases, such as allergic rhinitis, and proper classification of these ligands and accurate characterization of their mechanism(s) of action is of The aim of the present work was to the mechanisms involved in mepyramine actions at the guinea pig H1 receptor stably expressed in Chinese hamster ovary cells. We found that mepyramine as an inverse decreasing the basal spontaneous activity of the mepyramine inositol and intracellular calcium with a decrease in the to histamine at high mepyramine effect was observed with triprolidine, H1 receptor inverse of mepyramine with the Gq/11-mediated signaling of the endogenously expressed ATP receptor was that mepyramine at in exert its action by promoting a G protein-coupled inactive state of the H1 receptor as predicted by the the effect of mepyramine was neutralized by of Gα11 protein, indicating that mepyramine is able to reduce the G protein availability for other non-related receptors associated with the same signaling pathway. Finally, we found a decrease in the efficacy of mepyramine to the basal levels of intracellular calcium at high Gα11 expression G efficacy observed for mepyramine can be explained in terms of the high H1 receptor constitutive activity. The whole of the present work sheds new light into the of H1 receptor pharmacology and the mechanisms inverse agonists could to exert their observed negative efficacy. and were from mepyramine pertussis and were from was from form and were from DMEM, and were from and all the were from and were from other were of and of the H1 pig histamine H1 receptor was recently C. Fernandez N. Legnazzi B.L. F. A. Baldi A. Davio C. Mol. Pharmacol. PubMed Scopus Google with from the of the the Leurs R. J.M. J. J. M. J. PubMed Scopus Google were used to a from guinea pig from guinea pig the a and the in a and the a an was and the resulting were and into the The was for the and the was found to be to the reported For the was by with and into the of the expression at the to the for The and of the was by The resulting was used to and a with high histamine H1 receptor expression was by the was the same and were with the cells were at in a and cells were in and cells were in the same and H1 Binding from and cells were and binding were as (8Monczor F. Fernandez N. Legnazzi B.L. Riveiro M.E. Baldi A. Shayo C. Davio C. Mol. Pharmacol. 2003; 64: 512-520Crossref PubMed Scopus (37) Google Scholar). cells were in for and from the a in of binding The cells were then by in binding and at for to and cells. The was for at and the was in an of binding was by the and the was in binding at a protein of and at For binding of were in with different of from to in a of for For binding were with in in the of different of in a of for For of were in a to of several of at a of were with for by the of mepyramine H. C. S. S. M. Eur. J. Pharmacol. 2002; PubMed Scopus Google Scholar). In all were at in the effect of on were at for in in the of of binding was in the of that was of and for the were and with binding were in of and was in a inositol was as C. N. H. Pharmacol. 2002; PubMed Scopus Google Scholar). cells were in and for in were then and the was for the of and for the was and with and for were then for with histamine in from to in a of in the or of H1 in the in The was by the of of and were by the of of and of The was then at for and the inositol was by of the was a were expressed as the was to from the of the in intracellular were by the Mol. Pharmacol. PubMed Scopus Google Scholar). cells were in for cells were with of of and and in and The cells were then in the of for at The cells were and for at to the of the to the was by of in of was by a of in of was in a with the to with The was at and was at of to basal levels, agonists were in and the of intracellular was for the of the by was to by to was by the by an of cells. these intracellular were as by G. M. J. Biol. Chem. Full Text PDF PubMed Scopus Google Scholar). were for were expressed as the of the of the in the of levels in the were in all of the was as N. F. C. Baldi A. Davio C. Shayo C. Mol. Pharmacol. 2002; PubMed Scopus Google Scholar). cells were in and and to were for and were in and to The binding were with in and were with or in were with the same was developed an or to and to the and were for binding was by of binding from of with was Binding and at the H1 to the binding and signaling of mepyramine at the expressed in cells. we a named In these mepyramine and binding binding for mepyramine with affinity and of and a with affinity and of is with from several other M. Pharmacol. PubMed Scopus Google Scholar, S. R. G. PubMed Scopus Google Scholar, M. M. Pharmacol. PubMed Scopus Google Scholar). of the ligands classified as H1 were recently reclassified as inverse agonists in view of the reported constitutive activity of the human histamine H1 receptor (10Bakker R.A. Wieland K. Timmerman H. Leurs R. Eur. J. Pharmacol. 2000; 387: 5-7Crossref PubMed Scopus (153) Google Scholar). this in COS-7 we the effects of mepyramine on levels in cells. mepyramine was able to decrease basal levels this we the effect of triprolidine, formerly H1 antagonist, on basal levels in cells. observed with mepyramine, was able to decrease the basal in cells Importantly, the of inverse agonists was could of the basal levels of InsP, indicating that is a ligand mepyramine be classified as a inverse agonist, mepyramine may be classified as inverse at the expressed in cells. the constitutive activity of the gp-H1r, the negative efficacy for inverse agonists, in with with the human H1 receptor R.A. Timmerman H. Leurs R. Mol. Pharmacol. PubMed Scopus Google Scholar). of on Binding at the the of by the P. Cotecchia S. Costa T. Lefkowitz R.J. J. Biol. Chem. 1993; 268: 4625-4636Abstract Full Text PDF PubMed Google the negative efficacy observed for inverse agonists is explained by a selective affinity of the ligand for a state of the receptor (2Strange P.G. Trends Pharmacol. Sci. 2002; 23: 89-95Abstract Full Text Full Text PDF PubMed Scopus (116) Google Scholar). this the affinity of an inverse be by uncoupling the receptor from the G protein P. Cotecchia S. Costa T. Lefkowitz R.J. J. Biol. Chem. 1993; 268: 4625-4636Abstract Full Text PDF PubMed Google Scholar). the effect of receptor uncoupling from G protein, from cells were with of these mepyramine binding with a of and a effect of at We then to the effect of G protein uncoupling on mepyramine binding a of of with this a decrease in the affinity of mepyramine for the gp-H1r, the of mepyramine from a of in the of to a of in the of are in with with the expressed in M. S. M. J. Pharmacol. PubMed Scopus Google Scholar). of Gα11 the a in the to a of transfected the same experimental the affinity of for the was with observed of in the of in the of and in the Gα11 that mepyramine and intrinsic negative efficacy at the gp-H1r, may different receptor binding was to uncoupling of the receptor from G protein. In mepyramine affinity for the was by with and Gα11 protein was indicating a affinity for receptor coupled to G protein. We have recently reported with the histamine receptor inverse (8Monczor F. Fernandez N. Legnazzi B.L. Riveiro M.E. Baldi A. Shayo C. Davio C. Mol. Pharmacol. 2003; 64: 512-520Crossref PubMed Scopus (37) Google Scholar). of from cells with the affinity of for the human receptor, indicating that binding of the inverse could the a G protein-coupled but inactive receptor predicted by the of receptor occupancy J.M. Morgan P.H. Lutz M.W. Kenakin T.P. J. Theor. Biol. 1996; 181: 381-397Crossref PubMed Scopus (165) Google Scholar). binding of mepyramine could a receptor state of the H1 receptor, we to the effects of mepyramine on histamine intrinsic efficacy at the H1 receptor expressed in cells. of on at the the effect of mepyramine on histamine intrinsic efficacy at gp-H1r, different were used to of the experimental on the intrinsic efficacy was by with of mepyramine and of the histamine with of histamine in used to its effects on histamine response. effects of mepyramine on histamine have been recently a Mol. Pharmacol. PubMed Scopus Google Scholar). the of the reduction in the to histamine observed with that was the reduction we that is to be from a on a T. Pharmacol. 2002; PubMed Scopus Google Scholar). In view of the observed in binding we to the effects of on histamine intrinsic efficacy at the in and a of histamine response. that ligands intrinsic negative efficacy at the gp-H1r, the mechanisms that mepyramine and use to this effect are effects on have been recently reported for other formerly H1 J.M. J. PubMed Scopus Google Scholar). were explained in by a of of histamine and its at the H1 receptor the used for intracellular calcium resulting in an this we to the of mepyramine from In could from the the of mepyramine with a of is in with the that mepyramine is a ligand at the histamine H1 receptor, by several other M. S. M. J. Pharmacol. PubMed Scopus Google Scholar). the for mepyramine binding at the could that in the of calcium histamine may mepyramine from a binding the with the ligands were for that that be explained in terms of a of mepyramine at the gp-H1r, that other H1 may a G protein-coupled inactive state of the H1 receptor. the of such a receptor state with the signaling of other non-related GPCRs (8Monczor F. Fernandez N. Legnazzi B.L. Riveiro M.E. Baldi A. Shayo C. Davio C. Mol. Pharmacol. 2003; 64: 512-520Crossref PubMed Scopus (37) Google we to the effects of mepyramine on the intrinsic efficacy of ATP at the receptor endogenously expressed in cells J.M. J. Pharmacol. PubMed Scopus Google Scholar). of on ATP at the in ATP as an at the receptor endogenously expressed in of the receptor and the signaling in and intracellular calcium J.M. J. Pharmacol. PubMed Scopus Google Scholar). the H1 receptor has been associated with the same Timmerman H. J.M. R. Pharmacol. Google Scholar). Considering these we to the effects of of mepyramine on the by in of mepyramine the to ATP in cells. the that mepyramine is of a G protein-coupled receptor state unable to evoke of no effect on ATP indicating that may use different mechanism to the spontaneous activity of the that histamine mepyramine or in with histamine effect on basal levels in mepyramine with the observed in cells indicating that the effects of mepyramine are specifically by the expressed and the that mepyramine may have effects on the ATP receptor or on G proteins, endogenously expressed in cells. the effects of ATP in cells were to that the signaling of the ATP receptor is by Gq/11 as reported J. Pharmacol. 1993; PubMed Scopus Google Scholar). and calcium were in cells of and mepyramine on intracellular in cells. cells were with ATP ATP ATP mepyramine histamine histamine mepyramine and mepyramine or were transiently transfected with of Gα11 levels were as were as the of of G on the of the are explained by receptor occupancy models, we to investigate the of mepyramine with a receptor state to G of the Gq/11 resulting in G protein that is the of G of this would the effect of mepyramine on ATP in cells. Importantly, the histamine H1 receptor has been to to Gα11 R.A. Timmerman H. Leurs R. Mol. Pharmacol. PubMed Scopus Google Scholar, J. G. J. Biol. Chem. 2003; Full Text Full Text PDF PubMed Scopus Google Scholar). in the effects by mepyramine on ATP were by of cells and in the and in the of with of Gα11 the basal levels were to an that ATP was unable to those basal is with constitutive activity as for other constitutively active receptors G. J. S. C. K. Kenakin T. Mol. Pharmacol. 2000; PubMed Scopus Google Scholar). these experimental the effect of mepyramine was to decrease basal levels of intracellular the to to of with a to the in the were were to the same effects of or were no of the to ATP was that of Gq/11 could the negative effects of mepyramine on ATP intrinsic efficacy. the high basal levels of intracellular calcium by the of Gα11 were by mepyramine, indicating that were by the constitutive activity of the gp-H1r, the expression of Gα11 in the cells no effect on basal levels that mepyramine a G protein-coupled receptor state that is unable to evoke a response. this is predicted by the CTC, at this could the of constitutively active of G and on of cells with of Gα11 in of Gα11 protein expression and in basal levels of intracellular by the of receptors coupled to with the observed constitutive activity of the receptor. In all of the experimental mepyramine intracellular levels by the spontaneous activity of the the basal levels are by the constitutive activity of the in this a in the negative efficacy of mepyramine could be However, mepyramine a efficacy could be with the of the Considering and as the able to and into account the by the its and for that the of basal levels and by the ligand can be as a of G protein and are the of G protein in the or in the of of The of the G protein is a of the ligand present in the and be for ligand F. Fernandez N. Legnazzi B.L. Riveiro M.E. Baldi A. Shayo C. Davio C. Mol. Pharmacol. 2003; 64: 512-520Crossref PubMed Scopus (37) Google Scholar). The of the the ability of a ligand to basal receptor its efficacy, as by in the efficacy of an inverse with high affinity for inactive G protein-coupled of the receptor, as mepyramine is, to the receptor a efficacy could be predicted for high is the for receptor from to efficacy could be for receptors with high constitutive activity. of the to the reported in this the that the is constitutively active expressed in cells and that mepyramine acting as an inverse at the is able to a G protein-coupled receptor state that is unable to evoke are the to be of the ability of mepyramine to with the ATP receptor signaling but to the observed constitutive activity of the In the present work we stably expressed the in cells to the pharmacological of mepyramine and its mechanism of action as an inverse could be from these the intrinsic negative efficacy of mepyramine is on the of a G protein-coupled form of the H1 receptor, which is unable to evoke a response. as a of such a mechanism mepyramine interferes with the signaling of at GPCR, the ATP receptor, by the availability of Gq/11 this was by of Gα11 protein. issue has the therapeutic of inverse agonists. Finally, by the effects of mepyramine at high G protein a high of constitutive activity could be predicted for the with the of the a high of constitutive activity has been reported for the human H1 receptor (10Bakker R.A. Wieland K. Timmerman H. Leurs R. Eur. J. Pharmacol. 2000; 387: 5-7Crossref PubMed Scopus (153) Google indicating that constitutive activity could be a of the H1 on these we the importance of proper classification of ligands of constitutively active receptors but the importance of the molecular mechanisms by which these ligands exert their actions in terms of their and pharmacological In the ligands formerly as H1 have been reclassified as H1 inverse agonists in view of the observed constitutive activity of the human H1 receptor (10Bakker R.A. Wieland K. Timmerman H. Leurs R. Eur. J. Pharmacol. 2000; 387: 5-7Crossref PubMed Scopus (153) Google Scholar). The H1 antihistamines are the and the in the of their widespread use to is a to use as to the treatment of of diseases, allergic and allergic the of or effects of importance 2002; PubMed Scopus Google Scholar). in the of their mechanism of action is of importance to their and several pharmacological the human histamine H1 receptor and the guinea pig histamine receptor have been recently reported R. K. T. S. A. S. J. Pharmacol. 2003; PubMed Scopus Google the guinea pig has been used to and of the currently used H1 antihistamines Timmerman H. J.M. R. Pharmacol. Google and as of the reference models to H1 antihistamines in M. T. Pharmacol. PubMed Scopus Google and M. T. K. K. K. H. J. Pharmacol. PubMed Scopus Google Scholar). The of the molecular mechanisms by H1 inverse agonists may to effects of According to the currently accepted models, an inverse may the constitutive activity of a by at three different of it may by binding to the inactive state of the receptor (3Leff P. Trends Pharmacol. Sci. 1995; 16: 89-97Abstract Full Text PDF PubMed Scopus (477) Google Scholar). it could bind to a G state of the receptor P. Cotecchia S. Costa T. Lefkowitz R.J. J. Biol. Chem. 1993; 268: 4625-4636Abstract Full Text PDF PubMed Google Scholar). third is that the observed intrinsic negative efficacy is by the receptor to an G protein-coupled (2Strange P.G. Trends Pharmacol. Sci. 2002; 23: 89-95Abstract Full Text Full Text PDF PubMed Scopus (116) Google Scholar). the mechanism is predicted by the it the of an inactive form of the receptor coupled to G protein J.M. Morgan P.H. Lutz M.W. Kenakin T.P. J. Theor. Biol. 1996; 181: 381-397Crossref PubMed Scopus (165) Google it is that experimental evidence to all of the three mechanisms P. Cotecchia S. Costa T. Lefkowitz R.J. J. Biol. Chem. 1993; 268: 4625-4636Abstract Full Text PDF PubMed Google Scholar, K. Mol. Pharmacol. PubMed Scopus Google Scholar, P.G. J. 2000; PubMed Scopus Google Scholar, Mol. Pharmacol. Google Scholar, M.W. J. Pharmacol. Google Scholar, A. Mol. Pharmacol. Google Scholar, J. Pharmacol. Google Scholar, M. S. M. M. F. F. J. G. P. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). According to the third a ligand its action by promoting a G protein-coupled inactive of the receptor may the activity of the receptor but may with the signaling of other GPCRs by G protein the availability of the G protein to those GPCRs (8Monczor F. Fernandez N. Legnazzi B.L. Riveiro M.E. Baldi A. Shayo C. Davio C. Mol. Pharmacol. 2003; 64: 512-520Crossref PubMed Scopus (37) Google Scholar). may result in observed with other inverse agonists that promote the same G protein-coupled In this and mepyramine as inverse agonists at the gp-H1r, the binding to and Gα11 protein a in the mechanism used to exert their intrinsic negative efficacy. mepyramine but a decrease on response. the explanation to this of may the of several other models have been developed to ligand G. F. P. Trends Pharmacol. Sci. Full Text Full Text PDF PubMed Scopus Google Scholar, J. Pharmacol. PubMed Scopus Google Scholar, P. Trends Pharmacol. Sci. Full Text PDF PubMed Scopus Google Scholar, A. R.A. C. Eur. J. Pharmacol. 2000; PubMed Scopus Google Scholar). In the of the could be an inactive form of the receptor that with high resulting in the effects of mepyramine and on could be explained by in the mechanisms of action of ligands at the ligands different effects on the ATP-induced signaling at the endogenously expressed receptor as with the signaling by the ATP receptor to by the availability of Gq/11 to the receptor. no effect on ATP-induced indicating a different mechanism of In with of the has that in of histamine and affinity for mepyramine, binding of is K. R. Timmerman H. Leurs R. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google that mepyramine and may the same receptor to was for mepyramine at the gp-H1r, with of the receptor and with of the receptor that those ligands may promote G protein-coupled inactive of the receptors as However, in the present work we theoretically and that the by mepyramine on the signaling of the ATP receptor expressed in cells on the the of the signaling pathway, this effect was by of Gα11 protein. in the signaling distinctive of experimental result in of the Importantly, the of the different of the signaling is to be for the constitutive activity of GPCRs T. Pharmacol. 2002; PubMed Scopus Google Scholar). with the that the of the G protein the efficacy of a ligand such as For efficacy a for high efficacy a to G protein The of of Gα11 to that the efficacy of mepyramine as an inverse of the a as predicted for a receptor with constitutive activity. In on we that the of the efficacy of a ligand that a G protein-coupled inactive receptor such as mepyramine, in with of G protein is a to constitutive activity of the receptor. that the molecular of the mechanisms of action of a ligand are is an important to be into account accurate and of the pharmacological is We are to S. for on the