title: ATP released via pannexin-1 hemichannels mediates bladder overactivity triggered by urothelial P2Y6 receptors.
abstract: In contrast to the well-known signaling role of urothelial ATP to control bladder function, the hypothesis that uracil nucleotides (UTP and/or UDP) also exert autocrine/paracrine actions only recently gained experimental support. Urothelial cells express UDP-sensitive P2Y6 receptors, yet their role in the control of bladder activity has been mostly neglected. This study was designed to investigate the ability of PSB0474, a stable UDP analogue which exhibits selectivity for P2Y6 receptors, to modulate urodynamic responses in the anaesthetized rat in vivo. Instillation of PSB0474 into the bladder increased the voiding frequency (VF) without affecting the amplitude (A) and the duration (Δt) of bladder contractions. PSB0474-induced bladder overactivity was prevented by the selective P2Y6 antagonist, MRS2578. The increase in the VF produced by PSB0474 was also blocked by inhibitors of pannexin-1 hemichannels, (10)Panx or carbenoxolone, when these drugs were applied inside the bladder lumen but not when they were administered intravenously. Reduction of hemichannels pore permeability with H1152 also prevented PSB0474-induced bladder overactivity, but the exocytosis inhibitor, Exo-1, was inactive. PSB0474 increased by 3-fold the urinary ATP content. Implication of hemichannels permeability on PSB0474-induced ATP release was demonstrated by real-time fluorescence video-microscopy measuring the uptake of propidium iodide by intact urothelial cells in the absence and in the presence of MRS2578 or carbenoxolone. Confocal microscopy studies confirmed the co-localization of pannexin-1 and P2Y6 receptors in the rat urothelium. Data indicate that activation of P2Y6 receptors causes bladder overactivity in the anaesthetized rat indirectly by releasing ATP from the urothelium via pannexin-1 hemichannels.
authors.affiliation: Laboratório de Farmacologia e Neurobiologia/UMIB, Instituto de Ciências Biomédicas Abel Salazar-Universidade do Porto (ICBAS-UP), Porto, Portugal.
authors.forename: M Alexandrina
authors.initials: MA
authors.lastname: Timóteo
authors.affiliation: Laboratório de Farmacologia e Neurobiologia/UMIB, Instituto de Ciências Biomédicas Abel Salazar-Universidade do Porto (ICBAS-UP), Porto, Portugal.
authors.forename: Inês
authors.initials: I
authors.lastname: Carneiro
authors.affiliation: Laboratório de Farmacologia e Neurobiologia/UMIB, Instituto de Ciências Biomédicas Abel Salazar-Universidade do Porto (ICBAS-UP), Porto, Portugal.
authors.forename: Isabel
authors.initials: I
authors.lastname: Silva
authors.affiliation: Laboratório de Farmacologia e Neurobiologia/UMIB, Instituto de Ciências Biomédicas Abel Salazar-Universidade do Porto (ICBAS-UP), Porto, Portugal.
authors.forename: José Bernardo
authors.initials: JB
authors.lastname: Noronha-Matos
authors.affiliation: Laboratório de Farmacologia e Neurobiologia/UMIB, Instituto de Ciências Biomédicas Abel Salazar-Universidade do Porto (ICBAS-UP), Porto, Portugal.
authors.forename: Fátima
authors.initials: F
authors.lastname: Ferreirinha
authors.affiliation: Laboratório de Farmacologia e Neurobiologia/UMIB, Instituto de Ciências Biomédicas Abel Salazar-Universidade do Porto (ICBAS-UP), Porto, Portugal; Serviço de Urologia, Centro Hospitalar do Porto (CHP), Porto, Portugal.
authors.forename: Miguel
authors.initials: M
authors.lastname: Silva-Ramos
authors.affiliation: Laboratório de Farmacologia e Neurobiologia/UMIB, Instituto de Ciências Biomédicas Abel Salazar-Universidade do Porto (ICBAS-UP), Porto, Portugal. Electronic address: farmacol@icbas.up.pt.
authors.forename: Paulo
authors.initials: P
authors.lastname: Correia-de-Sá
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism* / Q000502:physiology*; D000818:Animals; D002478:Cells, Cultured; D017630:Connexins / Q000378:metabolism*; D008297:Male; D009419:Nerve Tissue Proteins / Q000378:metabolism*; D009924:Organ Culture Techniques; D051381:Rats; D017208:Rats, Wistar; D018048:Receptors, Purinergic P2 / Q000378:metabolism*; D001743:Urinary Bladder / Q000378:metabolism*; D019459:Urothelium / Q000378:metabolism*
chemical_list: D017630:Connexins; D009419:Nerve Tissue Proteins; D018048:Receptors, Purinergic P2; C479936:pannexin 1, rat; C099845:purinoceptor P2Y6; D000255:Adenosine Triphosphate
keywords: ABC; ATP release; ATP-binding cassette transporters; Anaesthetized rat; Cx43; E-NTPDases; LDH; Micturition reflex; P2Y(6) receptor; Pannexin hemichannels; Panx1; TTX; UDP; UTP; Urothelium; connexin 43; ecto-nucleoside triphosphate diphosphohydrolases; lactate dehydrogenase; mRNA; messenger ribonucleic acid; pannexin-1; tetrodotoxin; uridine diphosphate; uridine triphosphate