title: ATP release from freshly isolated guinea-pig bladder urothelial cells: a quantification and study of the mechanisms involved.
abstract: OBJECTIVE
  To quantify the amount of ATP released from freshly isolated bladder urothelial cells, study its control by intracellular and extracellular calcium and identify the pathways responsible for its release.
  
  
  METHODS
  Urothelial cells were isolated from male guinea-pig urinary bladders and stimulated to release ATP by imposition of drag forces by repeated pipetting. ATP was measured using a luciferin-luciferase assay and the effects of modifying internal and external calcium concentration and blockers of potential release pathways studied.
  
  
  RESULTS
  Freshly isolated guinea-pig urothelial cells released ATP at a mean (sem) rate of 1.9 (0.1) pmoles/mm(2) cell membrane, corresponding to about 700 pmoles/g of tissue, and about half [49 (6)%, n = 9) of the available cell ATP. This release was reduced to a mean (sem) of 0.46 (0.08) pmoles/mm(2) (160 pmoles/g) with 1.8 mm external calcium, and was increased about two-fold by increasing intracellular calcium. The release from umbrella cells was not significantly different from a mixed intermediate and basal cell population, suggesting that all three groups of cells release a similar amount of ATP per unit area. ATP release was reduced by ≈ 50% by agents that block pannexin and connexin hemichannels. It is suggested that the remainder may involve vesicular release.
  
  
  CONCLUSIONS
  A significant fraction of cellular ATP is released from isolated urothelial cells by imposing drag forces that cause minimal loss of cell viability. This release involves multiple release pathways, including hemichannels and vesicular release.
authors.affiliation: Department of Biochemistry and Physiology, FHMS, University of Surrey, Guildford, UK.
authors.forename: Linda M
authors.initials: LM
authors.lastname: McLatchie
authors.affiliation: Department of Physiology and Pharmacology, University of Bristol, Bristol, UK.
authors.forename: Christopher H
authors.initials: CH
authors.lastname: Fry
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism*; D000818:Animals; D020126:Brefeldin A / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism / Q000494:pharmacology; D002478:Cells, Cultured; D017630:Connexins / Q000378:metabolism; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism*; D005033:Ethylmaleimide / Q000494:pharmacology; D006168:Guinea Pigs; D008297:Male; D019459:Urothelium / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*
chemical_list: D017630:Connexins; D020126:Brefeldin A; D000255:Adenosine Triphosphate; D005033:Ethylmaleimide; D002118:Calcium
keywords: ATP; bladder; calcium; hemichannel; urothelial cells; vesicular release