title: Probenecid blocks human P2X7 receptor-induced dye uptake via a pannexin-1 independent mechanism.
abstract: P2X7 is a ligand-gated ion channel which is activated by ATP and displays secondary permeability characteristics. The mechanism of development of the secondary permeability pathway is currently unclear, although a role for the hemichannel protein pannexin-1 has been suggested. In this study we investigated the role of pannexin-1 in P2X7-induced dye uptake and ATP-induced IL-1β secretion from human monocytes. We found no pharmacological evidence for involvement of pannexin-1 in P2X7-mediated dye uptake in transfected HEK-293 cells with no inhibition seen for carbenoxolone and the pannexin-1 mimetic inhibitory peptide, 10Panx1. However, we found that probenecid inhibited P2X7-induced cationic and anionic dye uptake in stably transfected human P2X7 HEK-293 cells. An IC50 value of 203 μM was calculated for blockade of ATP-induced responses at human P2X7. Probenecid also reduced dye uptake and IL-1β secretion from human CD14+ monocytes whereas carbenoxolone and 10Panx1 showed no inhibitory effect. Patch clamp and calcium indicator experiments revealed that probenecid directly blocks the human P2X7 receptor.
authors.affiliation: Sydney Medical School Nepean, University of Sydney, Nepean Hospital, Penrith, New South Wales, Australia.
authors.forename: Archana
authors.initials: A
authors.lastname: Bhaskaracharya
authors.affiliation: Sydney Medical School Nepean, University of Sydney, Nepean Hospital, Penrith, New South Wales, Australia.
authors.forename: Phuong
authors.initials: P
authors.lastname: Dao-Ung
authors.affiliation: School of Biological Sciences, Illawarra Health and Medical Research Institute, University of Wollongong, Wollongong, New South Wales, Australia.
authors.forename: Iman
authors.initials: I
authors.lastname: Jalilian
authors.affiliation: School of Biological Sciences, Illawarra Health and Medical Research Institute, University of Wollongong, Wollongong, New South Wales, Australia.
authors.forename: Mari
authors.initials: M
authors.lastname: Spildrejorde
authors.affiliation: Sydney Medical School Nepean, University of Sydney, Nepean Hospital, Penrith, New South Wales, Australia.
authors.forename: Kristen K
authors.initials: KK
authors.lastname: Skarratt
authors.affiliation: Sydney Medical School Nepean, University of Sydney, Nepean Hospital, Penrith, New South Wales, Australia.
authors.forename: Stephen J
authors.initials: SJ
authors.lastname: Fuller
authors.affiliation: School of Biological Sciences, Illawarra Health and Medical Research Institute, University of Wollongong, Wollongong, New South Wales, Australia.
authors.forename: Ronald
authors.initials: R
authors.lastname: Sluyter
authors.affiliation: Sydney Medical School Nepean, University of Sydney, Nepean Hospital, Penrith, New South Wales, Australia; Health Innovations Research Institute, School of Medical Sciences, RMIT University, Bundoora, Melbourne, Victoria, Australia.
authors.forename: Leanne
authors.initials: L
authors.lastname: Stokes
mesh_terms: D000255:Adenosine Triphosphate / Q000502:physiology; D001693:Biological Transport, Active; D020013:Calcium Signaling; D017630:Connexins / Q000378:metabolism*; D004996:Ethidium / Q000378:metabolism; D005456:Fluorescent Dyes / Q000378:metabolism; D057809:HEK293 Cells; D006801:Humans; D020128:Inhibitory Concentration 50; D053583:Interleukin-1beta / Q000378:metabolism; D007546:Isoquinolines / Q000378:metabolism; D008070:Lipopolysaccharides / Q000502:physiology; D009000:Monocytes / Q000276:immunology / Q000378:metabolism; D009419:Nerve Tissue Proteins / Q000378:metabolism*; D011339:Probenecid / Q000494:pharmacology*; D058920:Purinergic P2X Receptor Antagonists / Q000494:pharmacology*; D058486:Receptors, Purinergic P2X7 / Q000378:metabolism*
chemical_list: D017630:Connexins; D005456:Fluorescent Dyes; D053583:Interleukin-1beta; D007546:Isoquinolines; D008070:Lipopolysaccharides; D009419:Nerve Tissue Proteins; C492764:PANX1 protein, human; D058920:Purinergic P2X Receptor Antagonists; D058486:Receptors, Purinergic P2X7; D000255:Adenosine Triphosphate; C017475:lucifer yellow; D004996:Ethidium; D011339:Probenecid