Emerging Concepts in Ion Channel Biophysics

Emerging Concepts in Ion Channel Biophysics

Poster Abstracts

24-POS Board 24 Hemichannel Closure Triggered by Extracellular ATP at Millimolar Concentration Impairs Airways Ciliary Activity Karla Droguett , Mariana Rios, Llilian Arzola, Manuel Villalón, Nelson P. Barrera. Pontificia Universidad Católica de Chile, Santiago, Chile. Mucociliary clearance (MCC) in the airway epithelium protects the lungs against contaminants and microorganisms present in the inspired air. However, in inflammatory chronic respiratory diseases the MCC is impaired, which in turn contributes to airway inflammation and respiratory disease progression. Extracellular ATP is a signalling molecule in the epithelium regulating both basal ciliary beat frequency (CBF) and increased CBF, associated to [Ca 2+ ] i , after mechanical stimulation. In addition, the ATP effect on CBF is dependent on pannexin and connexin hemichannels (Panx/Cx HCs) activity, through an autocrine purinergic mechanism involving ATP release and Ca 2+ entry. Under pathophysiological conditions of the airways, such as asthma, high levels of ATP had been measured in broncho alveolar fluid, however it is unclear how these ATP concentrations affect ciliary activity and MCC. Our goal was to determine the cellular effect of 1 mM ATP on primary cultures of mouse tracheal epithelium. A series of experimental methods were carried out, hemichannel functionality through the uptake of ethidium bromide (EtBr), CBF and ciliary beating forces using atomic force microscopy, and [Ca 2+ ] i measurements by fluorimetric assays. The ATP addition reduced the uptake rate of EtBr, effect that was also observed in the presence of carbenoxolone (100 μM), a Panx/Cx HCs inhibitor. The Ca 2+ ionophore ionomycin (10 μM) increases the uptake rate of EtBr, which was blocked by ATP. Furthermore, ATP lowered CBF, ciliary forces and [Ca 2+ ] i . These results show that high levels of extracellular ATP, measured in chronic respiratory diseases, trigger epithelial hemichannel closure that impairs airway ciliary activity and MCC. Funded by CONICYT- FONDECYT 3150652 (KD), CONICYT 21160416 (LlA) and CONICYT-DPI20140080.

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