title: Differential Rho-kinase dependency of full and partial muscarinic receptor agonists in airway smooth muscle contraction.
abstract: In airway smooth muscle (ASM), full and partial muscarinic receptor agonists have been described to have large differences in their ability to induce signal transduction, including Ca2+-mobilization. Despite these differences, partial agonists are capable of inducing a submaximal to maximal ASM contraction. To further elucidate transductional differences between full and partial muscarinic receptor agonists, we investigated the contribution of Rho-kinase (an important regulator of Ca2+-sensitization) to methacholine-, pilocarpine- and McN-A-343-induced bovine tracheal smooth muscle (BTSM) contraction, using the selective Rho-kinase inhibitor Y-27632. In addition, we measured Ca2+-mobilization and -influx in BTSM cells in response to these agonists in the absence and presence of Y-27632. Whereas treatment with Y-27632 (1 microM) significantly decreased potency (pEC50) for all agonists, maximal contraction (Emax) was reduced by 23.4+/-2.8 and 50.4+/-7.9% for the partial agonists pilocarpine and McN-A-343, respectively, but was unaffected for the full agonist methacholine. However, Emax of methacholine became Rho-kinase dependent after taking away its receptor reserve using the irreversible muscarinic receptor antagonist propylbenzilylcholine mustard. Pilocarpine and McN-A-343 induced a very small Ca2+-mobilization and -influx as compared to methacholine. In addition, an inverse relationship of these two parameters with the Rho-kinase dependency was observed. Interestingly, no inhibitory effects of Y-27632 were observed on Ca2+-mobilization and-influx for all three agonists, indicating that the effects of Y-27632 on contraction are most likely on the level of Ca2+-sensitization. In conclusion, in contrast to the full agonist methacholine, the partial muscarinic receptor agonists pilocarpine and McN-A-343 are dependent on Rho-kinase for their maximal contractile effects, presumably as a consequence of differences in transductional reserve, indicating an agonist-dependent role for Rho-kinase in ASM contraction. Moreover, an inverse relationship exists between Rho-kinase dependency and both Ca2+-mobilization and Ca2+-influx for these agonists.
authors.affiliation: Department of Molecular Pharmacology, University of Groningen, A. Deusinglaan 1, 9713 AV Groningen, The Netherlands. D.Schaafsma@rug.nl
authors.forename: Dedmer
authors.initials: D
authors.lastname: Schaafsma
authors.forename: Mark
authors.initials: M
authors.lastname: Boterman
authors.forename: Anne-Margreet
authors.initials: AM
authors.lastname: de Jong
authors.forename: Iris
authors.initials: I
authors.lastname: Hovens
authors.forename: Jelte-Maarten
authors.initials: JM
authors.lastname: Penninks
authors.forename: S Adriaan
authors.initials: SA
authors.lastname: Nelemans
authors.forename: Herman
authors.initials: H
authors.lastname: Meurs
authors.forename: Johan
authors.initials: J
authors.lastname: Zaagsma
mesh_terms: D008455:(4-(m-Chlorophenylcarbamoyloxy)-2-butynyl)trimethylammonium Chloride / Q000494:pharmacology; D000478:Alkylation; D000577:Amides / Q000494:pharmacology; D000818:Animals; D016085:Bronchoconstrictor Agents / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism; D002417:Cattle; D004791:Enzyme Inhibitors / Q000494:pharmacology; D047908:Intracellular Signaling Peptides and Proteins; D007537:Isometric Contraction / Q000187:drug effects; D016210:Methacholine Chloride / Q000494:pharmacology; D018721:Muscarinic Agonists / Q000494:pharmacology*; D009119:Muscle Contraction / Q000187:drug effects; D009130:Muscle, Smooth / Q000187:drug effects*; D009924:Organ Culture Techniques; D010862:Pilocarpine / Q000494:pharmacology; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000502:physiology*; D011725:Pyridines / Q000494:pharmacology; D011976:Receptors, Muscarinic / Q000187:drug effects*; D014132:Trachea / Q000187:drug effects; D054460:rho-Associated Kinases
chemical_list: D000577:Amides; D016085:Bronchoconstrictor Agents; D004791:Enzyme Inhibitors; D047908:Intracellular Signaling Peptides and Proteins; D018721:Muscarinic Agonists; D011725:Pyridines; D011976:Receptors, Muscarinic; D010862:Pilocarpine; D016210:Methacholine Chloride; C108830:Y 27632; D008455:(4-(m-Chlorophenylcarbamoyloxy)-2-butynyl)trimethylammonium Chloride; D017346:Protein Serine-Threonine Kinases; D054460:rho-Associated Kinases; D002118:Calcium