Muscarinic-induced mucin secretion and intracellular signaling by hamster tracheal goblet cells.
Hamster tracheal epithelial cell cultures were used to investigate muscarinic regulation of high-molecular-weight glycoconjugate (HMWG) secretion by airway goblet cells. HMWG were radiolabeled with N-acetyl-D-[1-(3)H]glucosamine, precipitated with trichloroacetic acid and phosphotungstic acid, and counted by liquid scintillation. Carbachol (100 microM) increased HMWG secretion (166.6 +/- 18.7%, P < 0.001, n = 20), and this response was blocked by the muscarinic receptor antagonist atropine. Ca2+ may not be essential for carbachol response since 1) carbachol-activated secretion was not inhibited by chelating extracellular Ca2+ with 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA) or by reducing both extracellular and intracellular Ca2+ with BAPTA-acetoxymethyl ester in low-Ca2+ medium; 2) the carbachol response was only partially blocked in low-Ca2+ medium; and 3) calcium ionophore did not stimulate HMWG secretion. However, carbachol-stimulated secretion was abolished by pertussis toxin (PTX), indicating the involvement of a PTX-sensitive guanine nucleotide-binding regulatory protein (G protein), and by the protein kinase C (PKC) inhibitor chelerythrine chloride. Furthermore, carbachol-stimulated secretion was not inhibited by overnight incubation with phorbol 12-myristate 13-acetate. In conclusion, carbachol-stimulated secretion of HMWG appears to be coupled to a PTX-sensitive G protein and requires the activation of a phorbol ester-insensitive PKC isoform.
Steel
D M
DM
Department of Physiology, Respiratory Health Network of Centres of Excellence, McGill University, Montreal, Quebec, Canada.
Hanrahan
J W
JW
D000470:Alkaloids; D000818:Animals; D053119:Benzophenanthridines; D002217:Carbachol / Q000494:pharmacology; D002478:Cells, Cultured; D006224:Cricetinae; D004533:Egtazic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006001:Glycoconjugates / Q000737:chemistry / Q000378:metabolism; D007425:Intracellular Membranes / Q000502:physiology*; D008297:Male; D008647:Mesocricetus; D008970:Molecular Weight; D009077:Mucins / Q000378:metabolism*; D009116:Muscarine / Q000378:metabolism*; D018721:Muscarinic Agonists / Q000494:pharmacology; D019331:NG-Nitroarginine Methyl Ester / Q000494:pharmacology; D037342:Pertussis Toxin; D010617:Phenanthridines / Q000494:pharmacology; D015398:Signal Transduction; D014132:Trachea / Q000166:cytology / Q000378:metabolism* / Q000502:physiology*; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
D000470:Alkaloids; D053119:Benzophenanthridines; D004791:Enzyme Inhibitors; D006001:Glycoconjugates; D009077:Mucins; D018721:Muscarinic Agonists; D010617:Phenanthridines; D010566:Virulence Factors, Bordetella; C070379:1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid acetoxymethyl ester; D004533:Egtazic Acid; D009116:Muscarine; D002217:Carbachol; C016299:chelerythrine; D037342:Pertussis Toxin; C025603:1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid; D019331:NG-Nitroarginine Methyl Ester