Small inhaled particles, which are entrapped by the mucous layer that is maintained by mucous secretion via mucin exocytosis and fluid secretion, are removed from the nasal cavity by beating cilia. The functional activities of beating cilia are assessed by their frequency and the amplitude. Nasal ciliary beating is controlled by intracellular ions (Ca²⁺, H⁺ and Cl⁻), and is enhanced by a decreased concentration of intracellular Cl⁻ ([Cl⁻]_i) in ciliated human nasal epithelial cells (cHNECs) in primary culture, which increases the ciliary beat amplitude. A novel method to measure both ciliary beat frequency (CBF) and ciliary beat distance (CBD, an index of ciliary beat amplitude) in cHNECs has been developed using high-speed video microscopy, which revealed that a decrease in [Cl⁻]_i increased CBD, but not CBF, and an increase in [Cl⁻]_i decreased both CBD and CBF. Thus, [Cl⁻]_i inhibits ciliary beating in cHNECs, suggesting that axonemal structures controlling CBD and CBF may have Cl⁻ sensors and be regulated by [Cl⁻]_i. These observations indicate that the activation of Cl⁻ secretion stimulates ciliary beating (increased CBD) mediated via a decrease in [Cl⁻]_i in cHNECs. Thus, [Cl⁻]_i is critical for controlling ciliary beating in cHNECs. This review introduces the concept of Cl⁻ regulation of ciliary beating in cHNECs.

Sei-hai-to (TJ-90, Qing Fei Tang), a Chinese traditional medicine, increases ciliary beat frequency (CBF) and ciliary bend angle (CBA) mediated via cAMP (3′,5′-cyclic adenosine monophosphate) accumulation modulated by Ca²⁺-activated phosphodiesterase 1 (PDE1A). A high concentration of TJ-90 (≥40 μg/mL) induced two types of CBF increases, a transient increase (an initial increase, followed by a decrease) and a sustained increase without any decline, while it only sustained the CBA increase. Upon inhibiting increases in intracellular Ca²⁺ concentration ([Ca²⁺]_i) by 10 μM BAPTA-AM (Ca²⁺-chelator, 1,2-Bis(2-aminophenoxy)ethane-N,N,N′,N′-tetraacetic acid tetrakis(acetoxymethyl ester) or Ca²⁺/calmodulin-dependent PDE1 by 8MmIBMX (a selective PDE1 inhibitor), TJ-90 (400 μg/mL) induced only the sustained CBF increase without any transient CBF increase. The two types of the CBF increase (the transient increase and the sustained increase) induced by TJ-90 (≥40 μg/mL) were mimicked by the stimulation with both procaterol (100 pM) and ionomycin (500 nM). Thus, TJ-90 stimulates small increases in the intracellular cAMP concentration ([cAMP]_i) and [Ca²⁺]_i in airway ciliary cells of mice. These small increases in [cAMP]_i and [Ca²⁺]_i cause inducing a transient CBF increase or a sustained CBF increase in an airway ciliary cells, depending on the dominant signal, Ca²⁺-signal, or cAMP-signal.

Mucociliary clearance, which is conducted by beating cilia cooperating with the surface mucous layer, is a major host defense mechanism of the airway epithelium. Ezrin, a crosslinker between membrane proteins and the actin cytoskeleton, is located in microvilli and around the basal bodies in airway ciliary cells. It is also likely that ezrin plays an important role in apical localization of β2 adrenergic receptor (β2AR) in airway ciliary cells. Here, we studied the physiological roles of ezrin by using trachea and airway epithelial cells prepared from ezrin-knockdown (Vil2kd/kd) mice. The trachea and airway ciliary cells of Vil2kd/kd mice presented a normal morphology and basal body orientation, suggesting that ezrin is not directly involved in development and planar cell polarity of cilia. Procaterol stimulates ciliary beating (frequency and amplitude) via β2AR in the airway ciliary cells. In the Vil2kd/kd mice, airway ciliary beating stimulated with procaterol was partly inhibited due to the impairment of cell surface expression of β2AR. These results suggest that ezrin regulates the beating of airway ciliary cells by promoting the apical surface localization of β2AR. This article has an associated First Person interview with the first author of the paper.

MQAE is a ‘non-ratiometric’ chloride ion (Cl⁻)-quenched fluorescent indicator that is used to determine intracellular Cl⁻ concentration ([Cl⁻]_i). MQAE-based two-photon microscopy is reported to be a useful method to measure [Cl⁻]_i, but it is still controversial because a change in cell volume may alter the MQAE concentration, leading to a change in the fluorescence intensity without any change in [Cl⁻]_i. In an attempt to elucidate the effect or lack of effect of cell volume on MQAE concentration, we studied the effects of changes in cell volume, achieved by applying different levels of osmotic stress, on the intensity of MQAE fluorescence in airway ciliary cells. To study solely the effect of changes in cell volume on MQAE fluorescence intensity, i.e., excluding the effect of any change in [Cl⁻]_i, we first conducted the experiments in a Cl⁻-free nitrate (NO₃⁻) solution to substitute NO₃⁻ (non-quenching anion for MQAE fluorescence) for Cl⁻ in the intracellular fluid. Hypo- (− 30 mM NaNO₃) or hyper-osmotic stress (+ 30 mM NaNO₃) effected changes in cell volume, but the stress did not result in any significant change in MQAE fluorescence intensity. The experiments were also carried out in Cl⁻-containing solution. Hypo-osmotic stress (− 30 mM NaCl) increased both MQAE fluorescence intensity and cell volume, while hyper-osmotic stress (+ 30 mM NaCl) decreased both of these properties. These results suggest that the osmotic stress-induced change in MQAE fluorescence intensity was caused by the change in [Cl⁻]_i and not by the MQAE concentration. Moreover, the intracellular distribution of MQAEs was heterogeneous and not affected by the changes in osmotic stress-induced cell volume, suggesting that MQAEs are bound to un-identified subcellular structures. These bound MQAEs appear to have enabled the measurement of [Cl⁻]_i in airway ciliary cells, even under conditions of cell volume change.