Ambroxol-enhanced frequency and amplitude of beating cilia mediated by pHi and [Cl-]i in lung airway epithelial cells of mice

Abstract

Abstract Ambroxol (ABX), a frequently prescribed secretolytic agent that stimulates Ca2+ release from acidic stores and Ca2+ entry through CaV1.2, enhanced the ciliary beat frequency (CBF) and ciliary bend angle (CBA, an index of amplitude) by 30% in ciliated lung airway epithelial cells (c-LAECs) of mice. An increase in the intracellular Ca2+ concentration ([Ca2+]i) stimulated by ABX triggers two signals in c-LAECs; an increase in pHi (pH pathway) and a decrease in [Cl-]i (Cl- pathway). The pH pathway, which was activated by the HCO3- entry through Na+-HCO3- cotransporter (NBC) and inhibited by 4,4'-diisothiocyano-2,2'-stilbenedisulfonic acid (DIDS), enhanced the CBF (by 30%) and CBA (by 15-20%) and the Cl- pathway, which was activated by Cl- secretion through anoctamine 1 (ANO1) and inhibited by Cl- channel blockers (NPPB and T16Ainh), enhanced the CBA (by 10-15%). The enhancement of the CBF and CBA stimulated by ABX was decreased to 50% by a Ca2+-free solution or nifedipine (an inhibitor of L-type Ca2+ channels) and was abolished by BAPTA-AM in c-LAECs, indicating that an increase in [Ca2+]i stimulated by ABX is essential for activating the pH pathway and the Cl- pathway. The enhancement of CBF and CBA enhanced by ABX was mimicked by applying a CO2/HCO3--free Cl--free solution, which increased the pHi and decreased the [Cl-]i. In conclusion, ABX increased the CBF and CBA by 30% via the pH pathway and the Cl- pathway triggered by the [Ca2+]i increase in c-LAECs of mice.