CFTR in Calu-3 human airway cells: Channel properties and role in cAMP- activated Cl- conductance

C. Haws, W. E. Finkbeiner, Jonathan Widdicombe, J. J. Wine

Research output: Contribution to journalArticlepeer-review

140 Scopus citations

Abstract

Calu-3, a cell line derived from a lung adenocarcinoma, forms tight junctions, expresses cystic fibrosis transmembrane conductance regulator (CFTR), and secretes Cl- in response to adenosine 3',5'-cyclic monophosphate (cAMP)-elevating agents. Anion conductance of Calu-3 cells was assessed with isotopic flux and patch-clamp methods at 22°C. Iodide efflux was increased by cAMP-elevating agents and brief trypsin treatment. A 7.1 ± 0.4-pS voltage-independent Cl- channel with linear current-voltage relation was the most common channel observed in cell-attached recordings and was identified as CFTR on the basis of shared features with recombinant CFTR. In unstimulated cells, the mean minimum number of active CFTR channels per patch was 1 ± 1 (n = 12), increasing to 6 ± 8 (n = 40) after stimulation with cAMP-elevating agents or after brief trypsin treatment. Channel closure after excision was biexponential with τ1 ≃4 s and τ2 ≃79 s; typically channels were open continuously until closing permanently. In 11 of 12 excised patches, channels were reactivated by exposure to cAMP-dependent protein kinase (PKA) plus ATP. Efficacy of reactivation was inversely related to the duration from excision to addition of PKA. Channels were blocked by 20-40 μM 5-nitro-2-(3-phenylpropylamino)benzoate on cytosolic but not external side. Active CFTR channels were recorded in 83% of total patches. Other types of Cl- channels were observed in 5 of 52 (10%) cell-attached patches and in 17 of 34 (50%) excised patches, including an outwardly rectifying channel in 2 patches. CFTR channels are the predominant pathway for cAMP-stimulated Cl- conductance in Calu-3 cells; the long open times in the absence of ATP are not explained by present models of CFTR activation.

Original languageEnglish (US)
JournalAmerican Journal of Physiology - Lung Cellular and Molecular Physiology
Volume266
Issue number5 10-5
StatePublished - 1994
Externally publishedYes

Keywords

  • cell culture
  • cystic fibrosis
  • epithelia
  • patch clamp
  • submucosal gland

ASJC Scopus subject areas

  • Pulmonary and Respiratory Medicine
  • Cell Biology
  • Physiology

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