title: Functional expression of purinergic P2 receptors and transient receptor potential channels by the human urothelium.
abstract: In addition to its role as a physical barrier, the urothelium is considered to play an active role in mechanosensation. A key mechanism is the release of transient mediators that activate purinergic P2 receptors and transient receptor potential (TRP) channels to effect changes in intracellular Ca²⁺. Despite the implied importance of these receptors and channels in urothelial tissue homeostasis and dysfunctional bladder disease, little is known about their functional expression by the human urothelium. To evaluate the expression and function of P2X and P2Y receptors and TRP channels, the human ureter and bladder were used to separate urothelial and stromal tissues for RNA isolation and cell culture. RT-PCR using stringently designed primer sets was used to establish which P2 and TRP species were expressed at the transcript level, and selective agonists/antagonists were used to confirm functional expression by monitoring changes in intracellular Ca²⁺ and in a scratch repair assay. The results confirmed the functional expression of P2Y₄ receptors and excluded nonexpressed receptors/channels (P2X₁, P2X₃, P2X₆, P2Y₆, P2Y₁₁, TRPV5, and TRPM8), while a dearth of specific agonists confounded the functional validation of expressed P2X₂, P2X₄, P2Y₁, P2Y₂, TRPV2, TRPV3, TRPV6 and TRPM7 receptors/channels. Although a conventional response was elicited in control stromal-derived cells, the urothelial cell response to well-characterized TRPV1 and TRPV4 agonists/antagonists revealed unexpected anomalies. In addition, agonists that invoked an increase in intracellular Ca²⁺ promoted urothelial scratch repair, presumably through the release of ATP. The study raises important questions about the ligand selectivity of receptor/channel targets expressed by the urothelium. These pathways are important in urothelial tissue homeostasis, and this opens the possibility of selective drug targeting.
authors.affiliation: Jack Birch Unit of Molecular Carcinogenesis, Department of Biology, University of York, York, UK.
authors.forename: Saqib
authors.initials: S
authors.lastname: Shabir
authors.forename: William
authors.initials: W
authors.lastname: Cross
authors.forename: Lisa A
authors.initials: LA
authors.lastname: Kirkwood
authors.forename: Joanna F
authors.initials: JF
authors.lastname: Pearson
authors.forename: Peter A
authors.initials: PA
authors.lastname: Appleby
authors.forename: Dawn
authors.initials: D
authors.lastname: Walker
authors.forename: Ian
authors.initials: I
authors.lastname: Eardley
authors.forename: Jennifer
authors.initials: J
authors.lastname: Southgate
mesh_terms: D000255:Adenosine Triphosphate / Q000494:pharmacology; D000328:Adult; D000368:Aged; D002118:Calcium / Q000378:metabolism; D002211:Capsaicin / Q000494:pharmacology; D002478:Cells, Cultured; D017931:DNA Primers; D006706:Homeostasis / Q000502:physiology; D006801:Humans; D008875:Middle Aged; D058919:Purinergic P2 Receptor Antagonists / Q000494:pharmacology; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D018048:Receptors, Purinergic P2 / Q000096:biosynthesis*; D017154:Stromal Cells / Q000187:drug effects / Q000378:metabolism; D050051:Transient Receptor Potential Channels / Q000819:agonists / Q000037:antagonists & inhibitors / Q000096:biosynthesis*; D014513:Ureter / Q000473:pathology; D001743:Urinary Bladder / Q000473:pathology; D019459:Urothelium / Q000293:injuries / Q000378:metabolism*; D014945:Wound Healing / Q000502:physiology
chemical_list: D017931:DNA Primers; D058919:Purinergic P2 Receptor Antagonists; D012333:RNA, Messenger; D018048:Receptors, Purinergic P2; D050051:Transient Receptor Potential Channels; D000255:Adenosine Triphosphate; D002211:Capsaicin; D002118:Calcium
keywords: calcium; purinergic; transient receptor potential channel; urothelium