Abstract

A failure in optimal bicarbonate supply, as seen in the airways of patients with cystic fibrosis, produces severe lung disease due to abnormal mucus maturation and acidic pH of airway surface liquid in the airways. The membrane proteins responsible for bicarbonate movement across airway epithelium include CFTR and TMEM16A channels in the apical membrane and a series of transporters of the SLC4 family in both apical and basolateral membranes. While most of these exchangers are electroneutral, the SCLA4A4 or NBCe1 Sodium/Bicarbonate cotransporter moves a ratio of 2-3 bicarbonate and 1 sodium ion into the cell. The aim of our study was to evaluate if the NBCe1 cotransporter is necessary for electrogenic bicarbonate transport in mouse airways. Ussing chamber experiments of freshly isolated mouse trachea showed that roughly half of the UTP-evoked anionic current is lost when bicarbonate is replaced with HEPES in the bath solution (-105±11 vs -47±3 µA*cm-2 respectively). To determine if UTP induces HCO3 - exit, acutely isolated airway cells from mouse trachea were loaded with BCECF to track pH changes. UTP induced rapid acidification of airway cells which was fully abolished when using a CO2 /HCO3 - free bath solution (ΔpH -0.21 ± 0.003 vs -0.016 ± 0.005, respectively). Using a specific NBCe1 inhibitor (S0859) we observed that both UTP-evoked short-circuit current and intracellular acidification were significantly decreased, supporting the involvement of NBCe1 in epithelial anionic secretion. Nevertheless, Ussing chamber experiments in the Slca4a4-/- mice could not be performed due to low tissue resistance (32.7±8.7Ωcm2 ). Preliminary observations of histological tissue samples obtained from 3 week old Slca4a4-/- mice showed signs of mucus accumulation in bronchi and lung emphysema. In summary, our electrophysiological and imaging data show that bicarbonate secretion occurs in mouse airways and appears to be dependent on NBCe1 cotransporter function. Genetic silencing of NBCe1 produces a lung phenotype with features often observed in cystic fibrosis human patients and could be due to restricted bicarbonate delivery and acidification of airways.