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Investigation of ion channel activity in alveolar epithelial cells. The development of lung slice in-vitro model.

Bourke, Steven 2010. Investigation of ion channel activity in alveolar epithelial cells. The development of lung slice in-vitro model. MPhil Thesis, Cardiff University.
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Abstract

The alveolar epithelium maintains lung fluid homeostasis by active ion transport. The molecular machinery required to create an osmotic gradient across the alveolar epithelium, which in turn moves water out of the airspaces, has been identified by number of experimental techniques. The alveolar epithelium, which covers 95% of the surface area of the lung, is composed of two morphologically distinct epithelial cell types: alveolar epithelial type one and alveolar epithelial type two cells. The much studied alveolar epithelial type two cell, which cover <5% of the surface, contain the myriad of ion channel and transports required to undertake fluid transport. The alveolar epithelial type one cell, which covers >95% of the surface area, is less well characterised. Direct functional electrophysiological measurements from the alveolar epithelial type one cell has proved difficult to obtain. In order to obtain novel recordings the unfilled lung slice experimental model was developed. This experimental model demonstrated excellent cellular viability. VIIIB2 mouse monoclonal antibody, a specific cell marker for alveolar epithelial type one cells, was used to obtain live cell immunohistochemistry, the first such reported in peripheral tissue. The results presented in this thesis represent the first published ion channel recordings from identified alveolar epithelial type one cells within a rat lung slice (Bourke, S. et al 2005). Cell-attached, patch-clamp recordings demonstrated the presence of a voltage-dependant, K+ selective ion channel with a measured unitary conductance of 21 pS. The findings contained within this thesis present a novel experimental technique to investigate the functional physiological role of the alveolar epithelial cells. The alveolar epithelium is reliant upon Ca2+ signalling mechanisms to undertake a number of normal physiological activities. The release of surfactant from the alveolar epithelial type two cell lamellar bodies relies on a Ca2+ dependant exocytosis step. To investigate further the function of Ca2+ in the alveolar cells, experiments were undertaken using Ca2+ fluorescent indicator dye Fluo-3. Slices demonstrated spontaneous Ca2+ oscillations. Application of a number of physiological and pharmacological substances failed to modulate the observed activity.

Item Type: Thesis (MPhil)
Status: Unpublished
Schools: Biosciences
Subjects: Q Science > QP Physiology
Date of First Compliant Deposit: 30 March 2016
Last Modified: 19 Mar 2016 23:02
URI: http://orca-mwe.cf.ac.uk/id/eprint/36121

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