Article date: March 1999
By: Laurent Daeffler, Fabien Schmidlin, Jean‐Pierre Gies, Yves Landry, in Volume 126, Issue 5, pages 1246-1252
The intrinsic properties of muscarinic ligands were studied through their binding properties and their abilities to modulate the GTPase activity of G proteins coupled to muscarinic M2 receptors in pig atrial sarcolemma.
Competition binding experiments were performed with [3H]‐oxotremorine‐M to assess the affinity of receptors coupled to G proteins (R*), with [3H]‐N‐methylscopolamine ([3H]‐NMS) to estimate the affinities of coupled and uncoupled receptors (R*+R) and with [3H]‐NMS in the presence of GppNHp to assess the affinity of uncoupled receptors (R).
The ranking of Ki values for the agonist carbachol was R*<<R*+R<<R (0.95, 124 and 1017 nM). Ki values for atropine and AF‐DX 116 were similar for the three binding conditions (0.34, 0.42, 0.41 and 19, 22, 32 nM). The ranking of Ki values for pirenzepine was R*>R*+R>R (174, 155, 115 nM), suggesting inverse agonism.
The Vmax of the basal high affinity GTPase activity of pig atrial sarcolemma was increased by mastoparan and decreased by GPAnt‐2 indicating the relevance of this activity to G proteins coupled to receptors (R*). The KM value (0.26–0.33 μM) was not modified by mastoparan or GPAnt‐2.
Carbachol increased the Vmax of GTP hydrolysis (EC50 8.1±0.3 μM), whereas atropine and AF‐DX 116, up to 1 mM, did not modify it. Pirenzepine decreased the Vmax of GTP hydrolysis (EC50 77.5±10.3 μM). This effect was enhanced when KCl was substituted for NaCl (EC50 11.0±0.8 μM) and was antagonized by atropine and AF‐DX 116 (IC50 0.91±0.71 and 197±85 nM).
Pirenzepine is proposed as an inverse agonist and atropine and AF‐DX 116 as neutral antagonists at the muscarinic M2 receptor.
The intrinsic properties of muscarinic ligands were studied through their binding properties and their abilities to modulate the GTPase activity of G proteins coupled to muscarinic M2 receptors in pig atrial sarcolemma.
Competition binding experiments were performed with [3H]‐oxotremorine‐M to assess the affinity of receptors coupled to G proteins (R*), with [3H]‐N‐methylscopolamine ([3H]‐NMS) to estimate the affinities of coupled and uncoupled receptors (R*+R) and with [3H]‐NMS in the presence of GppNHp to assess the affinity of uncoupled receptors (R).
The ranking of Ki values for the agonist carbachol was R*<<R*+R<<R (0.95, 124 and 1017 nM). Ki values for atropine and AF‐DX 116 were similar for the three binding conditions (0.34, 0.42, 0.41 and 19, 22, 32 nM). The ranking of Ki values for pirenzepine was R*>R*+R>R (174, 155, 115 nM), suggesting inverse agonism.
The Vmax of the basal high affinity GTPase activity of pig atrial sarcolemma was increased by mastoparan and decreased by GPAnt‐2 indicating the relevance of this activity to G proteins coupled to receptors (R*). The KM value (0.26–0.33 μM) was not modified by mastoparan or GPAnt‐2.
Carbachol increased the Vmax of GTP hydrolysis (EC50 8.1±0.3 μM), whereas atropine and AF‐DX 116, up to 1 mM, did not modify it. Pirenzepine decreased the Vmax of GTP hydrolysis (EC50 77.5±10.3 μM). This effect was enhanced when KCl was substituted for NaCl (EC50 11.0±0.8 μM) and was antagonized by atropine and AF‐DX 116 (IC50 0.91±0.71 and 197±85 nM).
Pirenzepine is proposed as an inverse agonist and atropine and AF‐DX 116 as neutral antagonists at the muscarinic M2 receptor.
British Journal of Pharmacology (1999) 126, 1246–1252; doi:10.1038/sj.bjp.0702407
DOI: 10.1038/sj.bjp.0702407
View this article