Characterization of cAMP dependent CFTR‐chloride channels in human trache gland cells

Abstract

Human tracheal gland cells are believed to be a major site at the origin of cystic fibrosis. Since this disease is due to mutations in a protein called CFTR, we looked for the activity of CFTR in human tracheal gland cells in culture. We have identified CFTR‐like chloride‐selective channels as having a linear current‐voltage relationship and unitary conductance of 7 pS in these cells. In cell‐attached patches, theophylline (1 mM), IBMX (1 mM), or a cocktail of dibutyryl cAMP (1 mM) and IBMX (0.1 mM) promoted the opening of channels. The unitary current had a reversal potential close to the cell resting potential. Replacement of choline by K+ or Na+ in the pipette solution was without effect on the current‐voltage relationship, the reversal potential or the unitary conductance, which is consistent with the chloride selectivity of the channel. Channels were always found clustered and their opening probability was not noticeably dependent on membrane potential. This work therefore represents the first observation of a CFTR‐like channel activity in submucosal gland cells.

Absent Secretion to Vasoactive Intestinal Peptide in Cystic Fibrosis Airway Glands

“…However, ion and fluid secretion is reduced in cultures of CF gland cells (18,19) and in glands pharmacologically treated with CFTR inhibitors (20). This is consistent with patch clamp studies of primary cultures of serous cells (21) and the Calu-3 serous cell model (22), which indicate that CFTR is the only physiologically relevant apical anion channel in such cells. Ussing chamber studies with permeabilized cell sheets of Calu-3 cells confirm those studies and further indicate that functional CFTR is required for secretion to calcium-elevating agonists (23,24).…”

Acid in the airways. Focus on “Hyperacidity of secreted fluid from submucosal glands in early cystic fibrosis”

“…Some of the results were surprising. In the absence of HCO 3 CFTR-dependent mechanism is at work here in the absence of bath HCO 3 Ϫ or CO 2 ? Song et al (23a) also showed that if the droplets initially had a high pH, the surface airway acidified them, and this process was unaffected by CFTR inhibitors or by amiloride.…”

“…In the model by Devor et al (6), forskolin stimulates Calu-3 cells to secrete but does not activate Na ϩ -K ϩ -2Cl, the main transport pathway by which Cl Ϫ is moved into the cell across the basolateral membrane, leaving HCO 3 Ϫ as the major anion mediating fluid secretion; this model was supported by direct measures of Calu-3 fluid secreted in response to forskolin or VIP, which had average pH Ͼ8 and calculated HCO 3 Ϫ concentrations of 70 -90 mM (11,17). In addition, there is good evidence that CF surface epithelia, at least in culture, produce a fluid that is more acidic than normal (4).…”

“…After the surface was cleaned with a cotton swab and coated with oil, 300-nl fluid droplets were placed under the oil in such a way that they contacted the surface yet retained their spherical shapes. These fluid droplets contained BCECF-dextran and were buffered either with HCO 3 Ϫ or HEPES to match the bath solutions, and were then gassed appropriately. Both solutions and bath also contained various inhibitors or activators and were initially set to either pH 7.6 or 6.4.…”

Intracellular free Ca2+ dynamic changes to histamine are reduced in cystic fibrosis human tracheal gland cells

“…A decreased CI-permeability [9] and a constitutive hypersecretion [10] have been reported in cultured CF tracheobronchial gland cells. Furthermore, a high expression of CFTR mRNA and protein [11][12][13] and of CFTR-Iike C1 channels [14] were demonstrated in normal human tracheobronchial glands. We recently showed [12] that CFTR was specifically associated with the membrane of serous cell secretory granules.…”