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1 Department of Medicine-Laboratory of Epithelial Cell Biology, University of Pittsburgh, Pittsburgh, PA, USA; Department of Pharmacology, University of Pittsburgh, Pittsburgh, PA, USA
2 Department of Medicine-Laboratory of Epithelial Cell Biology, University of Pittsburgh, Pittsburgh, PA, USA
3 Department of Pharmacology, University of Pittsburgh, Pittsburgh, PA, USA
4 Clinical Veterinary Sciences, Ohio State University, Columbus, OH, USA
* To whom correspondence should be addressed. E-mail: lbirder{at}pitt.edu.
ATP can be released from a variety of cell types by mechanical stimulation, however the mechanism for this release and the influence of pathology is not well understood. The present study examined intracellular signaling mechanisms involved in swelling-evoked (exposure to a hypotonic solution) release of ATP in urothelial cells from normal cats and cats diagnosed with interstitial cystitis (feline interstitial cystitis, FIC). Using the luciferin-luciferase bioluminescent assay we demonstrate that swelling-evoked ATP release is significantly elevated in FIC cells. In both normal and FIC cells, ATP release was significantly decreased (mean 70% decrease) by application of blockers of stretch activated channels (amiloride or gadolinium), as well as brefeldin A and monensin (mean 90% decrease), suggesting that ATP release occurs when ATP containing vesicles fuse with the plasma membrane. Swelling-evoked release was reduced following removal of external calcium (65%), and release was blocked by incubation with BAPTA-AM, or agents that interfere with internal calcium stores (caffeine, ryanodine, heparin or 2-APB). In addition, agents known to act through IP3 receptors (thapsigargin; acetylcholine) release significantly more ATP in FIC as compared to normal urothelium. Taken together, these results suggest that FIC results in a novel hypersensitivity to mechanical stimuli that may involve alterations in IP3-sensitive pathways.
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