Hormonal regulation of Cl transport in polar airway epithelia measured by a fluorescent indicator.
Cl transport mechanisms in polarized cultures of canine tracheal epithelium were examined using an Ussing-type chamber with independent mucosal and serosal perfusion. Cl activity was monitored continuously from fluorescence of entrapped 6-methoxy-N-(3-sulfopropyl)quinolinium (SPQ). When added to the serosal (but not mucosal) solution, isoproterenol increased Cl fluxes across the apical membrane Cl more than fourfold. Apical Cl transport was sensitive to diphenylamine-2-carboxylate (DPC) but not to furosemide, whereas basolateral membrane Cl transport was sensitive to furosemide but not to DPC. Based on a mathematical model of Cl transport, we developed a sensitive protocol to measure hormone-sensitive Cl transport. In Cl-loaded cells in which basolateral Cl transport was partially inhibited by furosemide, mucosal Cl removal caused no Cl efflux before but rapid efflux (0.25 mM/s) after addition of isoproterenol or chlorophenylthio-cAMP. In the presence of indomethacin to block prostaglandin production, elevation of intracellular Ca by bradykinin or 4-bromo-A23187 did not cause Cl efflux, nor did Ca buffering with 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid affect stimulation by the cAMP pathway. Phorbol 12-myristate 13-acetate increased Cl efflux submaximally (0.09 mM/s) but did not affect maximal stimulation by cAMP agonists. Methoxamine did not alter apical or basolateral membrane Cl transport.(ABSTRACT TRUNCATED AT 250 WORDS)
Verkman
A S
AS
Department of Medicine, University of California, San Francisco 94143-0532.
Chao
A C
AC
Hartmann
T
T
D000818:Animals; D001692:Biological Transport; D002118:Calcium / Q000502:physiology; D002478:Cells, Cultured; D002712:Chlorides / Q000378:metabolism*; D000242:Cyclic AMP / Q000502:physiology; D004847:Epithelial Cells; D004848:Epithelium / Q000378:metabolism; D005456:Fluorescent Dyes; D006728:Hormones / Q000502:physiology*; D007545:Isoproterenol / Q000494:pharmacology; D008729:Methoxamine / Q000494:pharmacology; D011493:Protein Kinase C / Q000502:physiology; D011806:Quinolinium Compounds; D015398:Signal Transduction; D014132:Trachea / Q000166:cytology / Q000378:metabolism*; D062367:ortho-Aminobenzoates / Q000494:pharmacology
D002712:Chlorides; D005456:Fluorescent Dyes; D006728:Hormones; D011806:Quinolinium Compounds; D062367:ortho-Aminobenzoates; C052335:6-methoxy-N-(3-sulfopropyl)quinolinium; C001209:fenamic acid; D000242:Cyclic AMP; D011493:Protein Kinase C; D008729:Methoxamine; D007545:Isoproterenol; D002118:Calcium