title: Cholinergic regulation of epithelial sodium channels in rat alveolar type 2 epithelial cells.
abstract: We and others have shown that epithelial Na(+) channels (ENaC) in alveolar type 2 (AT2) cells are activated by β2 agonists, steroid hormones, elevated oxygen tension, and by dopamine. Although acetylcholine receptors (AChRs) have been previously described in the lung, there are few reports of whether cholinergic agonists alter sodium transport in the alveolar epithelium. Therefore, we investigated how cholinergic receptors regulate ENaC activity in primary cultures of rat AT2 cells using cell-attached patch-clamp recordings to assess ENaC activity. We found that the muscarinic agonists, carbachol (CCh) and oxotremorine, activated ENaC in a dose-dependent manner but that nicotine did not. CCh-induced activation of ENaC was blocked by atropine. Western blotting and immunohistochemistry suggested that muscarinic M2 and M3 receptors (mAChRs) but not nicotinic receptors were present in AT2 cells. Endogenous RhoA and GTP-RhoA increased in response to CCh and the increase was reduced by pretreatment with atropine. We showed that Y-27632, an inhibitor of Rho-associated protein kinase (ROCK), abolished endogenous ENaC activity and inhibited the activation of ENaC by CCh. We also showed that ROCK signaling was necessary for ENaC stability in 2F3 cells, a model for AT2 cells. Our results showed that muscarinic agonists activated ENaC in rat AT2 cells through M2 and/or M3 mAChRs probably via a RhoA/ROCK signaling pathway.
authors.affiliation: Department of Physiology, Emory University School of Medicine, Atlanta, GA 30322, USA.
authors.forename: Yoshizumi
authors.initials: Y
authors.lastname: Takemura
authors.forename: My N
authors.initials: MN
authors.lastname: Helms
authors.forename: Amity F
authors.initials: AF
authors.lastname: Eaton
authors.forename: Julie
authors.initials: J
authors.lastname: Self
authors.forename: Semra
authors.initials: S
authors.lastname: Ramosevac
authors.forename: Lucky
authors.initials: L
authors.lastname: Jain
authors.forename: Hui-Fang
authors.initials: HF
authors.lastname: Bao
authors.forename: Douglas C
authors.initials: DC
authors.lastname: Eaton
mesh_terms: D056809:Alveolar Epithelial Cells / Q000187:drug effects / Q000378:metabolism* / Q000502:physiology; D000577:Amides / Q000494:pharmacology; D000818:Animals; D001285:Atropine / Q000494:pharmacology; D002217:Carbachol / Q000494:pharmacology; D002478:Cells, Cultured; D004789:Enzyme Activation; D062708:Epithelial Sodium Channel Agonists / Q000494:pharmacology; D053503:Epithelial Sodium Channels / Q000378:metabolism*; D008564:Membrane Potentials / Q000187:drug effects; D051379:Mice; D008810:Mice, Inbred C57BL; D018721:Muscarinic Agonists / Q000494:pharmacology; D018727:Muscarinic Antagonists / Q000494:pharmacology; D009538:Nicotine / Q000494:pharmacology; D018722:Nicotinic Agonists / Q000494:pharmacology; D010095:Oxotremorine / Q000494:pharmacology; D018408:Patch-Clamp Techniques; D011725:Pyridines / Q000494:pharmacology; D051381:Rats; D017207:Rats, Sprague-Dawley; D043585:Receptor, Muscarinic M2 / Q000378:metabolism / Q000502:physiology; D043587:Receptor, Muscarinic M3 / Q000378:metabolism / Q000502:physiology; D054460:rho-Associated Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D020742:rhoA GTP-Binding Protein / Q000378:metabolism
chemical_list: D000577:Amides; D062708:Epithelial Sodium Channel Agonists; D053503:Epithelial Sodium Channels; D018721:Muscarinic Agonists; D018727:Muscarinic Antagonists; D018722:Nicotinic Agonists; D011725:Pyridines; D043585:Receptor, Muscarinic M2; D043587:Receptor, Muscarinic M3; C108830:Y 27632; D010095:Oxotremorine; D009538:Nicotine; D001285:Atropine; D002217:Carbachol; D054460:rho-Associated Kinases; D020742:rhoA GTP-Binding Protein