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1 Neuroscience and Cell Biology, University of Texas Medial Branch, Galveston, Texas, United States
* To whom correspondence should be addressed. E-mail: slewis{at}utmb.edu.
Recent reports have proposed that the urothelium can sense mechanical stretch and communicate this information to sensory afferent neurons by the release of ATP into the vicinity of P2X containing neurons. This report investigates the bidirectional release of ATP by in vitro rabbit urothelium. ATP was measured using the luciferin-luciferase assay. Immediately after washing both sides of the epithelium, there was a linear increase in ATP content in the mucosal compartment with a rate of 23±6.5 fmoles/min/cm2 (n=18). Serosal ATP content increased as a saturating exponential function, suggesting a constant rate of release and degradation of ATP by ectonucleotidases/exonucleotidases. The presence of a serosal ectonucleotidase/exonucleotidases was demonstrated by the time dependent decrease of exogenously added ATP. The maximum rate of hydrolysis was 11 pmoles/min/cm2 with a km of 0.49 µM. The time course of serosal ATP release was modeled as a constant rate of release (d: moles/min/cm2 ) and rate constant of hydrolysis (kh : min-). In control conditions d was 18 fmoles/min/cm2 and kh of 0.056±0.01 min- (n=18). Steady state serosal chamber content is 370±90 fmoles/cm2 and concentration is 50±1.2 x10-12 M. Stretching the tissue resulted in a transient 5-fold increase in the rate of mucosal ATP release and a transient 6-fold increase in serosal ATP release. 1/2 osmotic strength solutions increased mucosal release by 10-fold and serosal release by 5-fold. Tissue damage resulted in a step increase in mucosal chamber ATP content by 6.6±1 pmoles/cm2 and serosal chamber ATP by 0.1±0.06 pmoles/cm2 (n=5).
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