title: Species and response dependent differences in the effects of MAPK inhibitors on P2X(7) receptor function.
abstract: BACKGROUND
  Recent studies have implicated the mitogen activated protein kinase (MAPK) in cellular permeability changes following P2X(7) receptor activation in native tissues. In this study we have further studied the effect of MAPK inhibitors on recombinant and native P2X(7) receptors.
  
  
  METHODS
  The MAPK inhibitors SB-203580, SB-202190 and SB-242235 were examined in HEK293 cells expressing recombinant P2X(7) receptors and in THP-1 cells expressing native human P2X(7) receptors using a range of experimental approaches.
  
  
  RESULTS
  At human recombinant P2X(7) receptors, SB-203580 and SB-202190 were weak, non-competitive inhibitors (pIC(50)= 4.8 - 6.4) of ethidium accumulation stimulated by 2'- & 3'-O-(4benzoylbenzoyl)-ATP (BzATP) but SB-242235 (0.1-10 microM) had no effect. SB-203580 and SB-202190 had no effect on rat or mouse recombinant P2X(7) receptors and studies with chimeric P2X(7) receptors suggested that SB-203580 was only effective in chimeras containing the N-terminal 255aa of the human P2X(7) receptor. SB-203580 did not consistently affect BzATP-mediated increases in cell calcium levels and, in electrophysiological studies, it slightly decreased responses to 30 microM BzATP but potentiated responses to 100 microM BzATP. In THP1 cells, SB-203580 modestly inhibited BzATP-stimulated ethidium accumulation (pIC(50) 5.7 - < 5) but SB-202190 had no effect. Finally, SB-203580 did not block BzATP-stimulated interleukin-1beta release in THP-1 cells.
  
  
  CONCLUSIONS
  This study confirms that high concentrations of SB-203580 and SB-202190 can block human P2X(7) receptor-mediated increases in cellular ethidium accumulation but suggest this is not related to MAPK inhibition. Overall, the data cast doubt on a general role of MAPK in mediating P2X(7) receptor mediated changes in cellular permeability.
authors.affiliation: Neurology and GI Centre of Excellence for Drug Discovery, GlaxoSmithKline Research and Development Limited, New Frontiers Science Park, Harlow, Essex, UK. anton.d.michel@gsk.com
authors.forename: A D
authors.initials: AD
authors.lastname: Michel
authors.forename: K M
authors.initials: KM
authors.lastname: Thompson
authors.forename: J
authors.initials: J
authors.lastname: Simon
authors.forename: I
authors.initials: I
authors.lastname: Boyfield
authors.forename: E
authors.initials: E
authors.lastname: Fonfria
authors.forename: P P A
authors.initials: PP
authors.lastname: Humphrey
mesh_terms: D000255:Adenosine Triphosphate / Q000031:analogs & derivatives / Q000494:pharmacology; D000818:Animals; D020013:Calcium Signaling / Q000187:drug effects; D002460:Cell Line; D002463:Cell Membrane Permeability / Q000187:drug effects*; D004305:Dose-Response Relationship, Drug; D004996:Ethidium; D006801:Humans; D007093:Imidazoles / Q000494:pharmacology; D007202:Indicators and Reagents; D053583:Interleukin-1beta / Q000378:metabolism; D008564:Membrane Potentials / Q000187:drug effects; D051379:Mice; D020928:Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors*; D009000:Monocytes / Q000187:drug effects / Q000378:metabolism; D047428:Protein Kinase Inhibitors / Q000494:pharmacology*; D011725:Pyridines / Q000494:pharmacology; D051381:Rats; D018048:Receptors, Purinergic P2 / Q000187:drug effects* / Q000235:genetics; D058486:Receptors, Purinergic P2X7; D011993:Recombinant Fusion Proteins / Q000187:drug effects; D013045:Species Specificity; D014162:Transfection
chemical_list: D007093:Imidazoles; D007202:Indicators and Reagents; D053583:Interleukin-1beta; C546468:P2RX7 protein, human; C546469:P2rx7 protein, mouse; C546470:P2rx7 protein, rat; D047428:Protein Kinase Inhibitors; D011725:Pyridines; D018048:Receptors, Purinergic P2; D058486:Receptors, Purinergic P2X7; D011993:Recombinant Fusion Proteins; C403289:SB 242235; C004282:periodate-oxidized adenosine 5'-triphosphate; C033901:3'-O-(4-benzoyl)benzoyladenosine 5'-triphosphate; D000255:Adenosine Triphosphate; D020928:Mitogen-Activated Protein Kinases; D004996:Ethidium; C093642:SB 203580; C090942:4-(4-fluorophenyl)-2-(4-hydroxyphenyl)-5-(4-pyridyl)imidazole