Mode of Ca2+ action on ciliary beat frequency in single ovine airway epithelial cells

• Published in: The Journal of physiology Vol. 520; no. 3; pp. 851 - 865
• Main Authors: Salathe, Matthias, Bookman, Richard J.
• Format: Journal Article
• Language: English
• Published: Oxford, UK The Physiological Society 01.11.1999; Blackwell Science Ltd; Blackwell Science Inc
• Subjects: Animals; Calcium - physiology; Cells, Cultured; Cilia - physiology; Cytoplasm - metabolism; Epithelial Cells - physiology; Models, Biological; Original; Osmolar Concentration; Sheep; Trachea - cytology; Trachea - physiology
• ISSN: 0022-3751; 1469-7793
• DOI: 10.1111/j.1469-7793.1999.00851.x

We analysed the kinetics of coupling between cytoplasmic calcium ([Ca 2+ ] i ) and ciliary beat frequency (CBF) using simultaneous single cilium recording and single cell [Ca 2+ ] i measurements from cultured ovine tracheal epithelial cells. CBF and [Ca 2+ ] i (indicated by fura-2) were measured at rest and in response to activation of the G-protein coupled M3 muscarinic receptor by 10 μM acetylcholine (ACh). Fourier transform analysis of 3 s data segments of light intensity from phase-contrast microscopy showed no significant delay between changes in [Ca 2+ ] i and CBF during a 2 min exposure to ACh and subsequent washout. CBF time resolution was improved by computing instantaneous beat frequency. This revealed that CBF lagged the rapid increase in [Ca 2+ ] i in response to ACh with a delay of less than 1 beat cycle (143 ms at 7 Hz). When CBF was estimated by an improved Fourier method, this delay was observed to be 70 ± 30 ms (mean ± s.e.m .; n = 20 cilia). During the slower return to baseline, a lag of 8 ± 3.2 s was observed, indicative of hysteresis. While calmodulin inhibitors (calmidazolium and W-7; each n = 5) decreased baseline CBF by an average of 1.1 ± 0.1 Hz, they did not alter the kinetic relationship between [Ca 2+ ] i and CBF. Similarly, phosphatase inhibitors (okadaic acid and cyclosporin A; each n = 5), changed neither baseline CBF nor the kinetic coupling between [Ca 2+ ] i and CBF. These data suggest that the timing of Ca 2+ action on CBF in ovine airway epithelial cells, is unlikely to be determined by phosphorylation reactions involving calmodulin or kinase/phosphatase reactions. A simple model for Ca 2+ stimulation of CBF is presented. Fits of the model to the data suggest four or more Ca 2+ ions bind cooperatively to speed up CBF.