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Mechanical Stimulation-induced Calcium Wave Propagation in Cell Monolayers: The Example of Bovine Corneal Endothelial Cells
Published on: July 16, 2013
Regulation of gap junction coupling in bovine ciliary epithelium
Zhao Wang1, Chi Wai Do, Virginijus Valiunas
1Department of Physiology, University of Pennsylvania School of Medicine, Philadelphia, PA, USA.
American Journal of Physiology. Cell Physiology
|January 22, 2010
Summary
Connexin 43 (Cx43) is key for communication between bovine ciliary epithelial cells, influencing aqueous humor production. Cyclic AMP (cAMP) can inhibit this communication via protein kinase A (PKA).
Area of Science:
- Ophthalmology
- Cell Biology
- Molecular Biology
Background:
- Aqueous humor formation involves fluid transfer across pigmented (PE) and nonpigmented (NPE) ciliary epithelial cells.
- Gap junctions, formed by connexins (Cx), facilitate intercellular communication between PE and NPE cells.
- The specific connexins involved in bovine PE-NPE gap junctions are not fully elucidated.
Purpose of the Study:
- To determine if small interfering RNA against Cx43 (siCx43) affects bovine PE-NPE communication.
- To investigate the role of cyclic AMP (cAMP) in modulating bovine PE-NPE communication.
Main Methods:
- Quantitative reverse transcription PCR (qRT-PCR) and Western immunoblotting to assess connexin expression.
- Dual-cell patch clamping and Lucifer Yellow (LY) dye transfer to measure cell-to-cell communication.
- Treatment with siCx43, dibutyryl-cAMP, and PKA inhibitor H-89.
Main Results:
- Cx43 demonstrated significantly higher expression than Cx40 in bovine ciliary epithelial cells.
- siCx43 reduced Cx43 protein expression and significantly decreased LY transfer and junctional currents.
- Dibutyryl-cAMP also reduced LY transfer and junctional currents, an effect prevented by the PKA inhibitor H-89.
Conclusions:
- Cx43 is a major functional component of bovine PE-NPE gap junctions.
- cAMP, acting through PKA, can inhibit bovine PE-NPE gap junctional communication under specific conditions.
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