Activation of an apical Cl- conductance by Ca2+ ionophores in cystic fibrosis airway epithelia

Abstract

Cystic fibrosis (CF) airway epithelia express a defect in adenosine 3',5'-cyclic monophosphate (cAMP)-dependent regulation of apical membrane Cl- channels. Recent patch-clamp studies have raised the possibility that Ca2+ -dependent mechanisms for the activation of Cl- secretion may be preserved in CF airway epithelia. To determine 1) whether intact normal (N1) and CF airway epithelia exhibit a Ca2+ -dependent mechanism for activation of Cl- secretion and 2) whether Ca2+ -dependent mechanism for activation of Cl- secretion and 2) whether Ca2+ -dependent mechanisms initiate Cl- secretion via activation of an apical membrane Cl- conductance (GCl-), nasal epithelia from N1 and CF subjects were cultured on collagen membranes, and responses to isoproterenol or Ca2- ionophores [A23187 10(-6) M; ionomycin (10(-5)M)] were measured with transepithelial and intracellular techniques. Isoproterenol induced activation of an apical membrane GCl- in N1 cultures but was ineffective in CF. In contrast, in both N1 and CF amiloride-pretreated cultures, A23187 induced an increase in the equivalent short-circuit current that was associated with an activation of an apical membrane Gc1- and was bumetanide inhibitable. A23187 addition during superfusion of the lumen with a low Cl- (3 mM) solution reduced intracellular Cl- activity of CF cells. A Ca2+ ionophore of different selectivity properties, ionomycin, was also an effective Cl- secretagogue in both N1 and CF cultures. We conclude that 1) the A23187 induced Cl- secretion via activation of an apical GCl- in N1 human nasal epithelium, and 2) in contrast to an isoproterenol-dependent path, a Ca2+ -dependent path for GCl- activation is preserved in CF epithelia.

Second‐messenger regulation of sodium transport in mammalian airway epithelia.

“…However, ionomycin produces very similar effects (Willumsen & Boucher, 1989) and has been shown to directly elevate intracellular calcium in human nasal epithelium (Murphy, Cheng, Yankaskas, Stutts & Boucher, 1988). No stimulation of cyclic AMP levels by A23187 was seen in our tissues.…”

Defective regulatory volume decrease in human cystic fibrosis tracheal cells because of altered regulation of intermediate conductance Ca 2+ -dependent potassium channels

“…Vectorial ion transport is greatly affected in many CF epithelia (1). CF cells, unlike normal epithelial cells, do not elicit a net Cl Ϫ secretion when challenged with cAMPelevating agents (20), whereas Cl Ϫ secretion in response to agonists that elevate intracellular Ca 2ϩ is preserved (20)(21)(22)(23). To check the phenotypic characteristics of the original ⌬F508 cells (CFT1) and the cells permanently transfected with ␤gal (CFT1-LC3) or wild-type CFTR (CFT1-LCFSN), we studied the secretory responses of these cells.…”

Cystic Fibrosis Airway Epithelial Ca2+ Signaling

“…ATP/UTP) are greater in CF patients than in normal subjects (9,10). In vitro studies also detected a larger apical Ca 2ϩ i -dependent Cl Ϫ conductance in CF airway epithelia (9,(11)(12)(13). In addition, we recently reported that apical P2Y 2 -R activation promotes greater Ca 2ϩ i mobilization in CF compared with normal human nasal epithelia (4).…”