Enhancement of airway ciliary beating mediated via voltage-gated Ca2+ channels/α7-nicotinic receptors in mice

Acetylcholine (ACh), which activates muscarinic ACh receptors (mAChRs) and nicotinic ACh receptors (nAChRs), enhances airway ciliary beating by increasing the intracellular Ca2+ concentration ([Ca2+]i). The mechanisms enhancing airway ciliary beating by nAChRs have remained largely unknown, although those by mAChRs are well understood. In this study, we focused on the effects of α7-nAChRs and voltage-gated Ca2+ channels (CaVs) on the airway ciliary beating. The activities of ciliary beating were assessed by frequency (CBF, ciliary beat frequency) and amplitude (CBD, ciliary bend distance) measured by high-speed video microscopy. ACh enhanced CBF and CBD by 25% mediated by an [Ca2+]i increase stimulated by mAChRs and α7-nAChRs (a subunit of nAChR) in airway ciliary cells of mice. Experiments using PNU282987 (an agonist of α7-nAChR) and MLA (an inhibitor of α7-nAChR) revealed that CBF and CBD enhanced by α7-nAChR are approximately 50% of those enhanced by ACh. CBF, CBD, and [Ca2+]i enhanced by α7-nAChRs were inhibited by nifedipine, suggesting activation of CaVs by α7-nAChRs. Experiments using a high K+ solution with/without nifedipine (155.5 mM K+) showed that the activation of CaVs enhances CBF and CBD via an [Ca2+]i increase. Immunofluorescence and immunoblotting studies demonstrated that Cav1.2 and α7-nAChR are expressed in airway cilia. Moreover, IL-13 stimulated MLA-sensitive increases in CBF and CBD in airway ciliary cells, suggesting an autocrine regulation of ciliary beating by CaV1.2/α7-nAChR/ACh. In conclusion, a novel Ca2+ signalling pathway in airway cilia, CaV1.2/α7-nAChR, enhances CBF and CBD and activates mucociliary clearance maintaining healthy airways.