In situ measurement of airway surface liquid [K+] using a ratioable K+-sensitive fluorescent dye

Wan Namkung, Yuanlin Song, Aaron D. Mills, Prashant Padmawar, Walter E. Finkbeiner, Alan S. Verkman

Research output: Contribution to journalArticlepeer-review

33 Citations (Scopus)

Abstract

The airway surface liquid (ASL) is the thin fluid layer lining airway surface epithelial cells, whose volume and composition are tightly regulated and may be abnormal in cystic fibrosis (CF). We synthesized a two-color fluorescent dextran to measure ASL [K+], TAC-Lime-dextran-TMR, consisting of a green-fluorescing triazacryptand K+ ionophore-Bodipy conjugate, coupled to dextran, together with a red fluorescing tetramethylrhodamine reference chromophore. TAC-Lime-dextran-TMR fluorescence was K+-selective, increasing >4-fold with increasing [K+] from 0 to 40 mM. In well differentiated human airway epithelial cells, ASL [K+] was 20.8 ± 0.3 mM and decreased by inhibition of the Na+/K+ pump (ouabain), ENaC (amiloride), CF transmembrane conductance regulator (CFTRinh-172), or K+ channels (TEA or XE991). ASL [K+] was increased by forskolin but not affected by Na+/K+/2Cl-cotransporter inhibition (bumetanide). Functional and expression studies indicated the involvement of [K+] channels KCNQ1, KCNQ3, and KCNQ5 as determinants of ASL [K+]. [K+] in CF cultures was similar to that in non-CF cultures, suggesting that abnormal ASL [K+] is not a factor in CF lung disease. In intact airways, ASL [K+] was also well above extracellular [K+]: 22 ± 1 mM in pig trachea ex vivo and 16 ± 1 mM in mouse trachea in vivo. Our results provide the first noninvasive measurements of [K+] in the ASL and indicate the involvement of apical and basolateral membrane ion transporters in maintaining a high ASL [K+].

Original languageEnglish
Pages (from-to)15916-15926
Number of pages11
JournalJournal of Biological Chemistry
Volume284
Issue number23
DOIs
Publication statusPublished - 2009 Jun 5

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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