Abstract
1. We analysed the kinetics of coupling between cytoplasmic calcium ([Ca2+](i)) and ciliary beat frequency (CBF) using simultaneous single cilium recording and single cell [Ca2+](i) measurements from cultured ovine tracheal epithelial cells. 2. CBF and [Ca2+](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). 3. Fourier transform analysis of 3 s data segments of light intensity from phase-contrast microscopy showed no significant delay between changes in [Ca2+](i) and CBF during a 2 min exposure to ACh and subsequent washout. 4. CBF time resolution was improved by computing instantaneous beat frequency. This revealed that CBF lagged the rapid increase in [Ca2+](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. 5. 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 [Ca2+](i) and CBF. Similarly, phosphatase inhibitors (okadaic acid and cyclosporin A; each n = 5), changed neither baseline CBF nor the kinetic coupling between [Ca2+](i) and CBF. 6. These data suggest that the timing of Ca2+ action on CBF in ovine airway epithelial cells, is unlikely to be determined by phosphorylation reactions involving calmodulin or kinase/phosphatase reactions. 7. A simple model for Ca2+ stimulation of CBF is presented. Fits of the model to the data suggest four or more Ca2+ ions bind cooperatively to speed up CBF.
| Original language | English |
|---|---|
| Pages (from-to) | 851-865 |
| Number of pages | 15 |
| Journal | Journal of Physiology |
| Volume | 520 |
| Issue number | 3 |
| DOIs | |
| State | Published - Nov 1 1999 |
Fingerprint
ASJC Scopus subject areas
- Physiology
Cite this
Mode of Ca2+ action on ciliary beat frequency in single ovine airway epithelial cells. / Salathe, Matthias A; Bookman, Richard J.
In: Journal of Physiology, Vol. 520, No. 3, 01.11.1999, p. 851-865.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Mode of Ca2+ action on ciliary beat frequency in single ovine airway epithelial cells
AU - Salathe, Matthias A
AU - Bookman, Richard J.
PY - 1999/11/1
Y1 - 1999/11/1
N2 - 1. We analysed the kinetics of coupling between cytoplasmic calcium ([Ca2+](i)) and ciliary beat frequency (CBF) using simultaneous single cilium recording and single cell [Ca2+](i) measurements from cultured ovine tracheal epithelial cells. 2. CBF and [Ca2+](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). 3. Fourier transform analysis of 3 s data segments of light intensity from phase-contrast microscopy showed no significant delay between changes in [Ca2+](i) and CBF during a 2 min exposure to ACh and subsequent washout. 4. CBF time resolution was improved by computing instantaneous beat frequency. This revealed that CBF lagged the rapid increase in [Ca2+](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. 5. 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 [Ca2+](i) and CBF. Similarly, phosphatase inhibitors (okadaic acid and cyclosporin A; each n = 5), changed neither baseline CBF nor the kinetic coupling between [Ca2+](i) and CBF. 6. These data suggest that the timing of Ca2+ action on CBF in ovine airway epithelial cells, is unlikely to be determined by phosphorylation reactions involving calmodulin or kinase/phosphatase reactions. 7. A simple model for Ca2+ stimulation of CBF is presented. Fits of the model to the data suggest four or more Ca2+ ions bind cooperatively to speed up CBF.
AB - 1. We analysed the kinetics of coupling between cytoplasmic calcium ([Ca2+](i)) and ciliary beat frequency (CBF) using simultaneous single cilium recording and single cell [Ca2+](i) measurements from cultured ovine tracheal epithelial cells. 2. CBF and [Ca2+](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). 3. Fourier transform analysis of 3 s data segments of light intensity from phase-contrast microscopy showed no significant delay between changes in [Ca2+](i) and CBF during a 2 min exposure to ACh and subsequent washout. 4. CBF time resolution was improved by computing instantaneous beat frequency. This revealed that CBF lagged the rapid increase in [Ca2+](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. 5. 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 [Ca2+](i) and CBF. Similarly, phosphatase inhibitors (okadaic acid and cyclosporin A; each n = 5), changed neither baseline CBF nor the kinetic coupling between [Ca2+](i) and CBF. 6. These data suggest that the timing of Ca2+ action on CBF in ovine airway epithelial cells, is unlikely to be determined by phosphorylation reactions involving calmodulin or kinase/phosphatase reactions. 7. A simple model for Ca2+ stimulation of CBF is presented. Fits of the model to the data suggest four or more Ca2+ ions bind cooperatively to speed up CBF.
UR - http://www.scopus.com/inward/record.url?scp=0033231444&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=0033231444&partnerID=8YFLogxK
U2 - 10.1111/j.1469-7793.1999.00851.x
DO - 10.1111/j.1469-7793.1999.00851.x
M3 - Article
C2 - 10545149
AN - SCOPUS:0033231444
VL - 520
SP - 851
EP - 865
JO - Journal of Physiology
JF - Journal of Physiology
SN - 0022-3751
IS - 3
ER -