Differential Rho-kinase dependency of full and partial muscarinic receptor agonists in airway smooth muscle contraction.
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.
Schaafsma
Dedmer
D
Department of Molecular Pharmacology, University of Groningen, A. Deusinglaan 1, 9713 AV Groningen, The Netherlands. D.Schaafsma@rug.nl
Boterman
Mark
M
de Jong
Anne-Margreet
AM
Hovens
Iris
I
Penninks
Jelte-Maarten
JM
Nelemans
S Adriaan
SA
Meurs
Herman
H
Zaagsma
Johan
J
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
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