Contribution of pannexin 1 and connexin 43 hemichannels to extracellular calcium-dependent transport dynamics in human blood-brain barrier endothelial cells

Dysregulation of blood-brain barrier (BBB) transport function is thought to exacerbate neuronal damage in acute ischemic stroke. The purpose of this study was to clarify the characteristics of pannexin (Px) and/or connexin (Cx) hemichannel(s)-mediated transport of organic anions and cations in human...

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Veröffentlicht in:The Journal of pharmacology and experimental therapeutics 2015-04, Vol.353 (1), p.192-200
Hauptverfasser: Kaneko, Yosuke, Tachikawa, Masanori, Akaogi, Ryo, Fujimoto, Kazuhisa, Ishibashi, Megumi, Uchida, Yasuo, Couraud, Pierre-Olivier, Ohtsuki, Sumio, Hosoya, Ken-ichi, Terasaki, Tetsuya
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container_title The Journal of pharmacology and experimental therapeutics
container_volume 353
creator Kaneko, Yosuke
Tachikawa, Masanori
Akaogi, Ryo
Fujimoto, Kazuhisa
Ishibashi, Megumi
Uchida, Yasuo
Couraud, Pierre-Olivier
Ohtsuki, Sumio
Hosoya, Ken-ichi
Terasaki, Tetsuya
description Dysregulation of blood-brain barrier (BBB) transport function is thought to exacerbate neuronal damage in acute ischemic stroke. The purpose of this study was to clarify the characteristics of pannexin (Px) and/or connexin (Cx) hemichannel(s)-mediated transport of organic anions and cations in human BBB endothelial cell line hCMEC/D3 and to identify inhibitors of hemichannel opening in hCMEC/D3 cells in the absence of extracellular Ca(2+), a condition mimicking acute ischemic stroke. In the absence of extracellular Ca(2+), the cells showed increased uptake and efflux transport of organic ionic fluorescent dyes. Classic hemichannel inhibitors markedly inhibited the enhanced uptake and efflux. Quantitative targeted absolute proteomics confirmed Px1 and Cx43 protein expression in plasma membrane of hCMEC/D3 cells. Knockdown of Px1 and Cx43 with the small interfering RNAs significantly inhibited the enhanced uptake and efflux of organic anionic and cationic fluorescent dyes. Clinically used cilnidipine and progesterone, which have neuroprotective effects in animal ischemia models, were identified as inhibitors of hemichannel opening. These findings suggest that altered transport dynamics at the human BBB in the absence of extracellular Ca(2+) is at least partly attributable to opening of Px1 and Cx43 hemichannels. Therefore, we speculate that Px1 and Cx43 may be potential drug targets to ameliorate BBB transport dysregulation during acute ischemia.
doi_str_mv 10.1124/jpet.114.220210
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The purpose of this study was to clarify the characteristics of pannexin (Px) and/or connexin (Cx) hemichannel(s)-mediated transport of organic anions and cations in human BBB endothelial cell line hCMEC/D3 and to identify inhibitors of hemichannel opening in hCMEC/D3 cells in the absence of extracellular Ca(2+), a condition mimicking acute ischemic stroke. In the absence of extracellular Ca(2+), the cells showed increased uptake and efflux transport of organic ionic fluorescent dyes. Classic hemichannel inhibitors markedly inhibited the enhanced uptake and efflux. Quantitative targeted absolute proteomics confirmed Px1 and Cx43 protein expression in plasma membrane of hCMEC/D3 cells. Knockdown of Px1 and Cx43 with the small interfering RNAs significantly inhibited the enhanced uptake and efflux of organic anionic and cationic fluorescent dyes. Clinically used cilnidipine and progesterone, which have neuroprotective effects in animal ischemia models, were identified as inhibitors of hemichannel opening. These findings suggest that altered transport dynamics at the human BBB in the absence of extracellular Ca(2+) is at least partly attributable to opening of Px1 and Cx43 hemichannels. 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Clinically used cilnidipine and progesterone, which have neuroprotective effects in animal ischemia models, were identified as inhibitors of hemichannel opening. These findings suggest that altered transport dynamics at the human BBB in the absence of extracellular Ca(2+) is at least partly attributable to opening of Px1 and Cx43 hemichannels. 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Tachikawa, Masanori ; Akaogi, Ryo ; Fujimoto, Kazuhisa ; Ishibashi, Megumi ; Uchida, Yasuo ; Couraud, Pierre-Olivier ; Ohtsuki, Sumio ; Hosoya, Ken-ichi ; Terasaki, Tetsuya</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c404t-a950a5f6b4b17d0991d2b90498b246aab40ad21fe29a7433ca4e25e0358b471e3</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2015</creationdate><topic>Anions</topic><topic>Biological Transport - drug effects</topic><topic>Blood-Brain Barrier - cytology</topic><topic>Blood-Brain Barrier - drug effects</topic><topic>Blood-Brain Barrier - metabolism</topic><topic>Calcium - metabolism</topic><topic>Cations</topic><topic>Cell Line</topic><topic>Cell Membrane - metabolism</topic><topic>Connexin 43 - antagonists &amp; inhibitors</topic><topic>Connexin 43 - metabolism</topic><topic>Connexins - antagonists &amp; inhibitors</topic><topic>Connexins - metabolism</topic><topic>Endothelial Cells - drug effects</topic><topic>Endothelial Cells - metabolism</topic><topic>Extracellular Space - metabolism</topic><topic>Fluorescent Dyes - metabolism</topic><topic>Humans</topic><topic>Ion Channel Gating</topic><topic>Microvessels - cytology</topic><topic>Microvessels - metabolism</topic><topic>Nerve Tissue Proteins - antagonists &amp; 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Clinically used cilnidipine and progesterone, which have neuroprotective effects in animal ischemia models, were identified as inhibitors of hemichannel opening. These findings suggest that altered transport dynamics at the human BBB in the absence of extracellular Ca(2+) is at least partly attributable to opening of Px1 and Cx43 hemichannels. Therefore, we speculate that Px1 and Cx43 may be potential drug targets to ameliorate BBB transport dysregulation during acute ischemia.</abstract><cop>United States</cop><pmid>25670633</pmid><doi>10.1124/jpet.114.220210</doi><tpages>9</tpages><oa>free_for_read</oa></addata></record>
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subjects Anions
Biological Transport - drug effects
Blood-Brain Barrier - cytology
Blood-Brain Barrier - drug effects
Blood-Brain Barrier - metabolism
Calcium - metabolism
Cations
Cell Line
Cell Membrane - metabolism
Connexin 43 - antagonists & inhibitors
Connexin 43 - metabolism
Connexins - antagonists & inhibitors
Connexins - metabolism
Endothelial Cells - drug effects
Endothelial Cells - metabolism
Extracellular Space - metabolism
Fluorescent Dyes - metabolism
Humans
Ion Channel Gating
Microvessels - cytology
Microvessels - metabolism
Nerve Tissue Proteins - antagonists & inhibitors
Nerve Tissue Proteins - metabolism
title Contribution of pannexin 1 and connexin 43 hemichannels to extracellular calcium-dependent transport dynamics in human blood-brain barrier endothelial cells
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