title: Contribution of pannexin 1 and connexin 43 hemichannels to extracellular calcium-dependent transport dynamics in human blood-brain barrier endothelial cells.
abstract: 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.
authors: Division of Membrane Transport and Drug Targeting, Graduate School of Pharmaceutical Sciences, Tohoku University, Sendai, Japan (Y.K., M.T., R.A., Y.U., T.T.); Department of Applied Chemistry and Biochemistry, Faculty of Engineering, Kyushu Sangyo University, Fukuoka, Japan (K.F., M.I.); INSERM, U1016, Institut Cochin and CNRS, UMR8104, and Université Paris Descartes, Paris, France (P.-O.C.); Department of Pharmaceutical Microbiology, Faculty of Life Sciences, Kumamoto University, Kumamoto, Japan (S.O.); and Department of Pharmaceutics, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, Toyama, Japan (K.H.).
authors: Yosuke
authors: Y
authors: Kaneko
authors: Division of Membrane Transport and Drug Targeting, Graduate School of Pharmaceutical Sciences, Tohoku University, Sendai, Japan (Y.K., M.T., R.A., Y.U., T.T.); Department of Applied Chemistry and Biochemistry, Faculty of Engineering, Kyushu Sangyo University, Fukuoka, Japan (K.F., M.I.); INSERM, U1016, Institut Cochin and CNRS, UMR8104, and Université Paris Descartes, Paris, France (P.-O.C.); Department of Pharmaceutical Microbiology, Faculty of Life Sciences, Kumamoto University, Kumamoto, Japan (S.O.); and Department of Pharmaceutics, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, Toyama, Japan (K.H.) tachik-dds@umin.ac.jp.
authors: Masanori
authors: M
authors: Tachikawa
authors: Division of Membrane Transport and Drug Targeting, Graduate School of Pharmaceutical Sciences, Tohoku University, Sendai, Japan (Y.K., M.T., R.A., Y.U., T.T.); Department of Applied Chemistry and Biochemistry, Faculty of Engineering, Kyushu Sangyo University, Fukuoka, Japan (K.F., M.I.); INSERM, U1016, Institut Cochin and CNRS, UMR8104, and Université Paris Descartes, Paris, France (P.-O.C.); Department of Pharmaceutical Microbiology, Faculty of Life Sciences, Kumamoto University, Kumamoto, Japan (S.O.); and Department of Pharmaceutics, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, Toyama, Japan (K.H.).
authors: Ryo
authors: R
authors: Akaogi
authors: Division of Membrane Transport and Drug Targeting, Graduate School of Pharmaceutical Sciences, Tohoku University, Sendai, Japan (Y.K., M.T., R.A., Y.U., T.T.); Department of Applied Chemistry and Biochemistry, Faculty of Engineering, Kyushu Sangyo University, Fukuoka, Japan (K.F., M.I.); INSERM, U1016, Institut Cochin and CNRS, UMR8104, and Université Paris Descartes, Paris, France (P.-O.C.); Department of Pharmaceutical Microbiology, Faculty of Life Sciences, Kumamoto University, Kumamoto, Japan (S.O.); and Department of Pharmaceutics, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, Toyama, Japan (K.H.).
authors: Kazuhisa
authors: K
authors: Fujimoto
authors: Division of Membrane Transport and Drug Targeting, Graduate School of Pharmaceutical Sciences, Tohoku University, Sendai, Japan (Y.K., M.T., R.A., Y.U., T.T.); Department of Applied Chemistry and Biochemistry, Faculty of Engineering, Kyushu Sangyo University, Fukuoka, Japan (K.F., M.I.); INSERM, U1016, Institut Cochin and CNRS, UMR8104, and Université Paris Descartes, Paris, France (P.-O.C.); Department of Pharmaceutical Microbiology, Faculty of Life Sciences, Kumamoto University, Kumamoto, Japan (S.O.); and Department of Pharmaceutics, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, Toyama, Japan (K.H.).
authors: Megumi
authors: M
authors: Ishibashi
authors: Division of Membrane Transport and Drug Targeting, Graduate School of Pharmaceutical Sciences, Tohoku University, Sendai, Japan (Y.K., M.T., R.A., Y.U., T.T.); Department of Applied Chemistry and Biochemistry, Faculty of Engineering, Kyushu Sangyo University, Fukuoka, Japan (K.F., M.I.); INSERM, U1016, Institut Cochin and CNRS, UMR8104, and Université Paris Descartes, Paris, France (P.-O.C.); Department of Pharmaceutical Microbiology, Faculty of Life Sciences, Kumamoto University, Kumamoto, Japan (S.O.); and Department of Pharmaceutics, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, Toyama, Japan (K.H.).
authors: Yasuo
authors: Y
authors: Uchida
authors: Division of Membrane Transport and Drug Targeting, Graduate School of Pharmaceutical Sciences, Tohoku University, Sendai, Japan (Y.K., M.T., R.A., Y.U., T.T.); Department of Applied Chemistry and Biochemistry, Faculty of Engineering, Kyushu Sangyo University, Fukuoka, Japan (K.F., M.I.); INSERM, U1016, Institut Cochin and CNRS, UMR8104, and Université Paris Descartes, Paris, France (P.-O.C.); Department of Pharmaceutical Microbiology, Faculty of Life Sciences, Kumamoto University, Kumamoto, Japan (S.O.); and Department of Pharmaceutics, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, Toyama, Japan (K.H.).
authors: Pierre-Olivier
authors: PO
authors: Couraud
authors: Division of Membrane Transport and Drug Targeting, Graduate School of Pharmaceutical Sciences, Tohoku University, Sendai, Japan (Y.K., M.T., R.A., Y.U., T.T.); Department of Applied Chemistry and Biochemistry, Faculty of Engineering, Kyushu Sangyo University, Fukuoka, Japan (K.F., M.I.); INSERM, U1016, Institut Cochin and CNRS, UMR8104, and Université Paris Descartes, Paris, France (P.-O.C.); Department of Pharmaceutical Microbiology, Faculty of Life Sciences, Kumamoto University, Kumamoto, Japan (S.O.); and Department of Pharmaceutics, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, Toyama, Japan (K.H.).
authors: Sumio
authors: S
authors: Ohtsuki
authors: Division of Membrane Transport and Drug Targeting, Graduate School of Pharmaceutical Sciences, Tohoku University, Sendai, Japan (Y.K., M.T., R.A., Y.U., T.T.); Department of Applied Chemistry and Biochemistry, Faculty of Engineering, Kyushu Sangyo University, Fukuoka, Japan (K.F., M.I.); INSERM, U1016, Institut Cochin and CNRS, UMR8104, and Université Paris Descartes, Paris, France (P.-O.C.); Department of Pharmaceutical Microbiology, Faculty of Life Sciences, Kumamoto University, Kumamoto, Japan (S.O.); and Department of Pharmaceutics, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, Toyama, Japan (K.H.).
authors: Ken-ichi
authors: K
authors: Hosoya
authors: Division of Membrane Transport and Drug Targeting, Graduate School of Pharmaceutical Sciences, Tohoku University, Sendai, Japan (Y.K., M.T., R.A., Y.U., T.T.); Department of Applied Chemistry and Biochemistry, Faculty of Engineering, Kyushu Sangyo University, Fukuoka, Japan (K.F., M.I.); INSERM, U1016, Institut Cochin and CNRS, UMR8104, and Université Paris Descartes, Paris, France (P.-O.C.); Department of Pharmaceutical Microbiology, Faculty of Life Sciences, Kumamoto University, Kumamoto, Japan (S.O.); and Department of Pharmaceutics, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, Toyama, Japan (K.H.).
authors: Tetsuya
authors: T
authors: Terasaki
mesh_terms: D000838:Anions; D001692:Biological Transport / Q000187:drug effects; D001812:Blood-Brain Barrier / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism*; D002412:Cations; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D018031:Connexin 43 / Q000037:antagonists & inhibitors / Q000378:metabolism*; D017630:Connexins / Q000037:antagonists & inhibitors / Q000378:metabolism*; D042783:Endothelial Cells / Q000187:drug effects / Q000378:metabolism*; D005110:Extracellular Space / Q000378:metabolism; D005456:Fluorescent Dyes / Q000378:metabolism; D006801:Humans; D015640:Ion Channel Gating; D055806:Microvessels / Q000166:cytology / Q000378:metabolism; D009419:Nerve Tissue Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism*
chemical_list: D000838:Anions; D002412:Cations; D018031:Connexin 43; D017630:Connexins; D005456:Fluorescent Dyes; C500216:GJA1 protein, human; D009419:Nerve Tissue Proteins; C492764:PANX1 protein, human; D002118:Calcium