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An autocrine ATP release mechanism regulates basal ciliary activity in airway epithelium.
J Physiol. 2017 07 15; 595(14):4755-4767.JP

Abstract

KEY POINTS

Extracellular ATP, in association with [Ca2+ ]i regulation, is required to maintain basal ciliary beat frequency. Increasing extracellular ATP levels increases ciliary beating in airway epithelial cells, maintaining a sustained response by inducing the release of additional ATP. Extracellular ATP levels in the millimolar range, previously associated with pathophysiological conditions of the airway epithelium, produce a transient arrest of ciliary activity. The regulation of ciliary beat frequency is dependent on ATP release by hemichannels (connexin/pannexin) and P2X receptor activation, the blockage of which may even stop ciliary movement. The force exerted by cilia, measured by atomic force microscopy, is reduced following extracellular ATP hydrolysis. This result complements the current understanding of the ciliary beating regulatory mechanism, with special relevance to inflammatory diseases of the airway epithelium that affect mucociliary clearance.

ABSTRACT

Extracellular nucleotides, including ATP, are locally released by the airway epithelium and stimulate ciliary activity in a [Ca2+ ]i -dependent manner after mechanical stimulation of ciliated cells. However, it is unclear whether the ATP released is involved in regulating basal ciliary activity and mediating changes in ciliary activity in response to chemical stimulation. In the present study, we evaluated ciliary beat frequency (CBF) and ciliary beating forces in primary cultures from mouse tracheal epithelium, using videomicroscopy and atomic force microscopy (AFM), respectively. Extracellular ATP levels and [Ca2+ ]i were measured by luminometric and fluorimetric assays, respectively. Uptake of ethidium bromide was measured to evaluate hemichannel functionality. We show that hydrolysis of constitutive extracellular ATP levels with apyrase (50 U ml-1) reduced basal CBF by 45% and ciliary force by 67%. The apyrase effect on CBF was potentiated by carbenoxolone, a hemichannel inhibitor, and oxidized ATP, an antagonist used to block P2X7 receptors, which reduced basal CBF by 85%. Additionally, increasing extracellular ATP levels (0.1-100 μm) increased CBF, maintaining a sustained response that was suppressed in the presence of carbenoxolone. We also show that high levels of ATP (1 mm), associated with inflammatory conditions, lowered basal CBF by reducing [Ca2+ ]i and hemichannel functionality. In summary, we provide evidence indicating that airway epithelium ATP release is the molecular autocrine mechanism regulating basal ciliary activity and is also the mediator of the ciliary response to chemical stimulation.

Authors+Show Affiliations

Department of Physiology, Faculty of Biological Sciences, Pontificia Universidad Católica de Chile, Santiago, Chile.Department of Physiology, Faculty of Biological Sciences, Pontificia Universidad Católica de Chile, Santiago, Chile.Department of Physiology, Faculty of Biological Sciences, Pontificia Universidad Católica de Chile, Santiago, Chile.Department of Physiology, Faculty of Biological Sciences, Pontificia Universidad Católica de Chile, Santiago, Chile.Department of Physiology, Faculty of Biological Sciences, Pontificia Universidad Católica de Chile, Santiago, Chile.Department of Physiology, Faculty of Biological Sciences, Pontificia Universidad Católica de Chile, Santiago, Chile.Department of Physiology, Faculty of Biological Sciences, Pontificia Universidad Católica de Chile, Santiago, Chile.

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

28422293

Citation

Droguett, Karla, et al. "An Autocrine ATP Release Mechanism Regulates Basal Ciliary Activity in Airway Epithelium." The Journal of Physiology, vol. 595, no. 14, 2017, pp. 4755-4767.
Droguett K, Rios M, Carreño DV, et al. An autocrine ATP release mechanism regulates basal ciliary activity in airway epithelium. J Physiol (Lond). 2017;595(14):4755-4767.
Droguett, K., Rios, M., Carreño, D. V., Navarrete, C., Fuentes, C., Villalón, M., & Barrera, N. P. (2017). An autocrine ATP release mechanism regulates basal ciliary activity in airway epithelium. The Journal of Physiology, 595(14), 4755-4767. https://doi.org/10.1113/JP273996
Droguett K, et al. An Autocrine ATP Release Mechanism Regulates Basal Ciliary Activity in Airway Epithelium. J Physiol (Lond). 2017 07 15;595(14):4755-4767. PubMed PMID: 28422293.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - An autocrine ATP release mechanism regulates basal ciliary activity in airway epithelium. AU - Droguett,Karla, AU - Rios,Mariana, AU - Carreño,Daniela V, AU - Navarrete,Camilo, AU - Fuentes,Christian, AU - Villalón,Manuel, AU - Barrera,Nelson P, Y1 - 2017/06/15/ PY - 2016/12/31/received PY - 2017/04/10/accepted PY - 2017/4/20/pubmed PY - 2018/4/18/medline PY - 2017/4/20/entrez KW - ATP KW - airway epithelium KW - ciliary beat frequency KW - ciliary force SP - 4755 EP - 4767 JF - The Journal of physiology JO - J. Physiol. (Lond.) VL - 595 IS - 14 N2 - KEY POINTS: Extracellular ATP, in association with [Ca2+ ]i regulation, is required to maintain basal ciliary beat frequency. Increasing extracellular ATP levels increases ciliary beating in airway epithelial cells, maintaining a sustained response by inducing the release of additional ATP. Extracellular ATP levels in the millimolar range, previously associated with pathophysiological conditions of the airway epithelium, produce a transient arrest of ciliary activity. The regulation of ciliary beat frequency is dependent on ATP release by hemichannels (connexin/pannexin) and P2X receptor activation, the blockage of which may even stop ciliary movement. The force exerted by cilia, measured by atomic force microscopy, is reduced following extracellular ATP hydrolysis. This result complements the current understanding of the ciliary beating regulatory mechanism, with special relevance to inflammatory diseases of the airway epithelium that affect mucociliary clearance. ABSTRACT: Extracellular nucleotides, including ATP, are locally released by the airway epithelium and stimulate ciliary activity in a [Ca2+ ]i -dependent manner after mechanical stimulation of ciliated cells. However, it is unclear whether the ATP released is involved in regulating basal ciliary activity and mediating changes in ciliary activity in response to chemical stimulation. In the present study, we evaluated ciliary beat frequency (CBF) and ciliary beating forces in primary cultures from mouse tracheal epithelium, using videomicroscopy and atomic force microscopy (AFM), respectively. Extracellular ATP levels and [Ca2+ ]i were measured by luminometric and fluorimetric assays, respectively. Uptake of ethidium bromide was measured to evaluate hemichannel functionality. We show that hydrolysis of constitutive extracellular ATP levels with apyrase (50 U ml-1) reduced basal CBF by 45% and ciliary force by 67%. The apyrase effect on CBF was potentiated by carbenoxolone, a hemichannel inhibitor, and oxidized ATP, an antagonist used to block P2X7 receptors, which reduced basal CBF by 85%. Additionally, increasing extracellular ATP levels (0.1-100 μm) increased CBF, maintaining a sustained response that was suppressed in the presence of carbenoxolone. We also show that high levels of ATP (1 mm), associated with inflammatory conditions, lowered basal CBF by reducing [Ca2+ ]i and hemichannel functionality. In summary, we provide evidence indicating that airway epithelium ATP release is the molecular autocrine mechanism regulating basal ciliary activity and is also the mediator of the ciliary response to chemical stimulation. SN - 1469-7793 UR - https://www.unboundmedicine.com/medline/citation/28422293/An_autocrine_ATP_release_mechanism_regulates_basal_ciliary_activity_in_airway_epithelium_ L2 - https://doi.org/10.1113/JP273996 DB - PRIME DP - Unbound Medicine ER -