Probenecid blocks human P2X7 receptor-induced dye uptake via a pannexin-1 independent mechanism

Archana Bhaskaracharya; Phuong Dao-Ung; Iman Jalilian; Mari Spildrejorde; Kristen K. Skarratt; Stephen J. Fuller; Ronald Sluyter; Leanne Stokes

doi:10.1371/journal.pone.0093058

Probenecid blocks human P2X7 receptor-induced dye uptake via a pannexin-1 independent mechanism

Archana Bhaskaracharya, Phuong Dao-Ung, Iman Jalilian, Mari Spildrejorde, Kristen K. Skarratt, Stephen J. Fuller, Ronald Sluyter, Leanne Stokes

Molecular and Tissue Pharmacology

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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.

Original language	English
Article number	e93058
Journal	PLoS One
Volume	9
Issue number	3
DOIs	https://doi.org/10.1371/journal.pone.0093058
Publication status	Published - 26 Mar 2014

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10.1371/journal.pone.0093058Licence: CC BY

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Copyright 2014 Bhaskaracharya et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Final published version, 1.33 MBLicence: CC BY

Leanne Stokes
- L.Stokesuea.acuk
- School of Chemistry, Pharmacy and Pharmacology - Associate Professor in Pharmacology
- Molecular and Tissue Pharmacology - Member
Person: Research Group Member, Academic, Teaching & Research

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@article{08143f9e571e41a180cebeaf3b0000a7,

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.",

author = "Archana Bhaskaracharya and Phuong Dao-Ung and Iman Jalilian and Mari Spildrejorde and Skarratt, {Kristen K.} and Fuller, {Stephen J.} and Ronald Sluyter and Leanne Stokes",

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doi = "10.1371/journal.pone.0093058",

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T1 - Probenecid blocks human P2X7 receptor-induced dye uptake via a pannexin-1 independent mechanism

AU - Bhaskaracharya, Archana

AU - Dao-Ung, Phuong

AU - Jalilian, Iman

AU - Spildrejorde, Mari

AU - Skarratt, Kristen K.

AU - Fuller, Stephen J.

AU - Sluyter, Ronald

AU - Stokes, Leanne

PY - 2014/3/26

Y1 - 2014/3/26

N2 - 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.

AB - 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.

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DO - 10.1371/journal.pone.0093058

M3 - Article

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VL - 9

JO - PLoS One

JF - PLoS One

SN - 1932-6203

IS - 3

M1 - e93058

ER -

Probenecid blocks human P2X7 receptor-induced dye uptake via a pannexin-1 independent mechanism

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Leanne Stokes

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