title: Purinergic autocrine regulation of mechanosensitivity and serotonin release in a human EC model: ATP-gated P2X3 channels in EC are downregulated in ulcerative colitis.
abstract: BACKGROUND
  Alterations in 5-hydroxytryptamine (HT) signaling in inflamed gut may contribute to pathogenesis of inflammatory bowel diseases. Adenosine 5'-triphosphate (ATP) regulates mucosal-mechanosensory reflexes and ATP receptors are sensitive to mucosal inflammation. Yet, it remains unknown whether ATP can modulate 5-HT signaling in enterochromaffin cells (EC). We tested the novel purinergic hypothesis that ATP is a critical autocrine regulator of EC mechanosensitivity and whether EC expression of ATP-gated P2X3-ion channels is altered in inflammatory bowel diseases.
  
  
  METHODS
  Laser confocal (fluo-4) Ca imaging was performed in 1947 BON cells. Chemical stimulation or mechanical stimulation (MS) was used to study 5-HT or ATP release in human BON or surgical mucosal specimens, and purine receptors by reverse transcription-polymerase chain reaction, Western Blot, or P2X3-immunoreactivity in BON or 5-HT human EC (hEC) in 11 control and 10 severely inflamed ulcerative colitis (UC) cases.
  
  
  RESULTS
  ATP or MS triggered Ca-transients or 5-HT release in BON. ATP or adenosine diphosphate increased 5-HT release 5-fold. MS caused ATP release, detected after 5'ecto-ATPase inhibition by ARL67156. ARL67156 augmented and apyrase blocked Ca/5-HT mechanosensitive responses. 2-Methyl-thio-adenosine diphosphate 5'-monophosphate-evoked (P2Y1,12) or mechanically-evoked responses were blocked or augmented by a P2Y1,12 antagonist, MRS2179, in different cells or inhibited by U73122. A P2Y12 antagonist, 2MeSAMP, augmented responses. A P2X1,3 agonist, α,β-MeATP, triggered Ca responses, whereas a P2X1,2/3,3 antagonist, 2',3'-O-(2,4,6-trinitrophenyl)-ATP, blocked mechanical responses or cell-surface 5'ATP- labeling. In hEC, α,β-MeATP stimulated 5-HT release. In UC, P2X3-immunoreactivity decreased from 15% to 0.2% of 5-HThECs. Human mucosa and BON expressed P2X1, P2X3, P2X4, P2X5, P2Y1, P2Y2, P2Y4, P2Y6, P2Y11, and P2Y12R-messenger RNA transcripts.
  
  
  CONCLUSIONS
  ATP is a critical determinant of mechanosensation and 5-HT release via autocrine activation of slow P2Y1-phospholipase C/inositol-1,4,5-triphosphate-Ca or inhibitory P2Y12-purinergic pathways, and fast ATP-gated P2X3-channels. UC downregulation of P2X3-channels (or A2B) is postulated to mediate abnormal 5-HT signaling.
authors.affiliation: Departments of *Anesthesiology and †Pathology, The Wexner Medical Center at The Ohio State University, Columbus, Ohio; ‡Center for Perinatal Research, §Division of Pediatric Gastroenterology, The Research Institute at Nationwide Children's Hospital, Columbus, Ohio; and ‖Departments of Surgery and **Neuroscience, The Wexner Medical Center at The Ohio State University, Columbus, Ohio.
authors.forename: Andrómeda
authors.initials: A
authors.lastname: Liñán-Rico
authors.forename: Jacqueline E
authors.initials: JE
authors.lastname: Wunderlich
authors.forename: Iveta S
authors.initials: IS
authors.lastname: Grants
authors.forename: Wendy L
authors.initials: WL
authors.lastname: Frankel
authors.forename: Jianjing
authors.initials: J
authors.lastname: Xue
authors.forename: Kent C
authors.initials: KC
authors.lastname: Williams
authors.forename: Alan E
authors.initials: AE
authors.lastname: Harzman
authors.forename: Joshua T
authors.initials: JT
authors.lastname: Enneking
authors.forename: Helen J
authors.initials: HJ
authors.lastname: Cooke
authors.forename: Fievos L
authors.initials: FL
authors.lastname: Christofi
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism*; D019898:Autocrine Communication; D002478:Cells, Cultured; D003093:Colitis, Ulcerative / Q000188:drug therapy / Q000378:metabolism* / Q000601:surgery; D004759:Enterochromaffin Cells / Q000187:drug effects / Q000378:metabolism*; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D007413:Intestinal Mucosa / Q000187:drug effects / Q000378:metabolism*; D040542:Mechanotransduction, Cellular / Q000187:drug effects*; D058477:Receptors, Purinergic P2X3 / Q000737:chemistry / Q000378:metabolism*; D012701:Serotonin / Q000378:metabolism*
chemical_list: D058477:Receptors, Purinergic P2X3; D012701:Serotonin; D000255:Adenosine Triphosphate
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