Cerebral microvascular endothelial cell Na/H exchange: Evidence for the presence of NHE1 and NHE2 isoforms and regulation by arginine vasopressin

Tina I. Lam, Phyllis M. Wise, Martha E O'Donnell

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Abstract

Blood-brain barrier (BBB) Na transporters are essential for brain water and electrolyte homeostasis. However, they also contribute to edema formation during the early hours of ischemic stroke by increased transport of Na from blood into brain across an intact BBB. We previously showed that a luminal BBB Na-K-Cl cotransporter is stimulated by hypoxia, aglycemia, and AVP and that inhibition of the cotransporter by intravenous bumetanide significantly reduces edema and infarct in the rat middle cerebral artery occlusion (MCAO) model of stroke. More recently, we found evidence that intravenous cariporide (HOE-642), a highly potent Na/H exchange inhibitor, also reduces brain edema after MCAO. The present study was conducted to investigate which Na/H exchange protein isoforms are present in BBB endothelial cells and to evaluate the effects of ischemic factors on BBB Na/H exchange activity. Western blot analysis of bovine cerebral microvascular endothelial cells (CMEC) and immunoelectron microscopy of perfusion-fixed rat brain revealed that Na/H exchanger isoforms 1 and 2 (NHE1 and NHE2) are present in BBB endothelial cells. Using microspectrofluorometry and the pH-sensitive dye BCECF, we found that hypoxia (2% O2, 30 min), aglycemia (30 min), and AVP (1-200 nM, 5 min) significantly increased CMEC Na/H exchange activity, assessed as Na-dependent, HOE-642-sensitive H + flux. We found that AVP stimulation of CMEC Na/H exchange activity is dependent on intracellular Ca concentration and is blocked by V1, but not V2, vasopressin receptor antagonists. Our findings support the hypothesis that a BBB Na/H exchanger, possibly NHE1 and/or NHE2, is stimulated during ischemia to participate in cerebral edema formation.

Original languageEnglish (US)
JournalAmerican Journal of Physiology - Cell Physiology
Volume297
Issue number2
DOIs
StatePublished - Aug 2009

Fingerprint

Arginine Vasopressin
Blood-Brain Barrier
Protein Isoforms
Endothelial Cells
Sodium-Hydrogen Antiporter
Middle Cerebral Artery Infarction
Brain Edema
Edema
Brain
Sodium-Potassium-Chloride Symporters
Stroke
Bumetanide
Immunoelectron Microscopy
Electrolytes
Homeostasis
Coloring Agents
Ischemia
Perfusion
Western Blotting
Water

Keywords

  • Blood-brain barrier
  • Brain edema
  • Cariporide
  • Cerebral ischemia
  • Stroke

ASJC Scopus subject areas

  • Cell Biology
  • Physiology

Cite this

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title = "Cerebral microvascular endothelial cell Na/H exchange: Evidence for the presence of NHE1 and NHE2 isoforms and regulation by arginine vasopressin",
abstract = "Blood-brain barrier (BBB) Na transporters are essential for brain water and electrolyte homeostasis. However, they also contribute to edema formation during the early hours of ischemic stroke by increased transport of Na from blood into brain across an intact BBB. We previously showed that a luminal BBB Na-K-Cl cotransporter is stimulated by hypoxia, aglycemia, and AVP and that inhibition of the cotransporter by intravenous bumetanide significantly reduces edema and infarct in the rat middle cerebral artery occlusion (MCAO) model of stroke. More recently, we found evidence that intravenous cariporide (HOE-642), a highly potent Na/H exchange inhibitor, also reduces brain edema after MCAO. The present study was conducted to investigate which Na/H exchange protein isoforms are present in BBB endothelial cells and to evaluate the effects of ischemic factors on BBB Na/H exchange activity. Western blot analysis of bovine cerebral microvascular endothelial cells (CMEC) and immunoelectron microscopy of perfusion-fixed rat brain revealed that Na/H exchanger isoforms 1 and 2 (NHE1 and NHE2) are present in BBB endothelial cells. Using microspectrofluorometry and the pH-sensitive dye BCECF, we found that hypoxia (2{\%} O2, 30 min), aglycemia (30 min), and AVP (1-200 nM, 5 min) significantly increased CMEC Na/H exchange activity, assessed as Na-dependent, HOE-642-sensitive H + flux. We found that AVP stimulation of CMEC Na/H exchange activity is dependent on intracellular Ca concentration and is blocked by V1, but not V2, vasopressin receptor antagonists. Our findings support the hypothesis that a BBB Na/H exchanger, possibly NHE1 and/or NHE2, is stimulated during ischemia to participate in cerebral edema formation.",
keywords = "Blood-brain barrier, Brain edema, Cariporide, Cerebral ischemia, Stroke",
author = "Lam, {Tina I.} and Wise, {Phyllis M.} and O'Donnell, {Martha E}",
year = "2009",
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journal = "American Journal of Physiology - Renal Fluid and Electrolyte Physiology",
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TY - JOUR

T1 - Cerebral microvascular endothelial cell Na/H exchange

T2 - Evidence for the presence of NHE1 and NHE2 isoforms and regulation by arginine vasopressin

AU - Lam, Tina I.

AU - Wise, Phyllis M.

AU - O'Donnell, Martha E

PY - 2009/8

Y1 - 2009/8

N2 - Blood-brain barrier (BBB) Na transporters are essential for brain water and electrolyte homeostasis. However, they also contribute to edema formation during the early hours of ischemic stroke by increased transport of Na from blood into brain across an intact BBB. We previously showed that a luminal BBB Na-K-Cl cotransporter is stimulated by hypoxia, aglycemia, and AVP and that inhibition of the cotransporter by intravenous bumetanide significantly reduces edema and infarct in the rat middle cerebral artery occlusion (MCAO) model of stroke. More recently, we found evidence that intravenous cariporide (HOE-642), a highly potent Na/H exchange inhibitor, also reduces brain edema after MCAO. The present study was conducted to investigate which Na/H exchange protein isoforms are present in BBB endothelial cells and to evaluate the effects of ischemic factors on BBB Na/H exchange activity. Western blot analysis of bovine cerebral microvascular endothelial cells (CMEC) and immunoelectron microscopy of perfusion-fixed rat brain revealed that Na/H exchanger isoforms 1 and 2 (NHE1 and NHE2) are present in BBB endothelial cells. Using microspectrofluorometry and the pH-sensitive dye BCECF, we found that hypoxia (2% O2, 30 min), aglycemia (30 min), and AVP (1-200 nM, 5 min) significantly increased CMEC Na/H exchange activity, assessed as Na-dependent, HOE-642-sensitive H + flux. We found that AVP stimulation of CMEC Na/H exchange activity is dependent on intracellular Ca concentration and is blocked by V1, but not V2, vasopressin receptor antagonists. Our findings support the hypothesis that a BBB Na/H exchanger, possibly NHE1 and/or NHE2, is stimulated during ischemia to participate in cerebral edema formation.

AB - Blood-brain barrier (BBB) Na transporters are essential for brain water and electrolyte homeostasis. However, they also contribute to edema formation during the early hours of ischemic stroke by increased transport of Na from blood into brain across an intact BBB. We previously showed that a luminal BBB Na-K-Cl cotransporter is stimulated by hypoxia, aglycemia, and AVP and that inhibition of the cotransporter by intravenous bumetanide significantly reduces edema and infarct in the rat middle cerebral artery occlusion (MCAO) model of stroke. More recently, we found evidence that intravenous cariporide (HOE-642), a highly potent Na/H exchange inhibitor, also reduces brain edema after MCAO. The present study was conducted to investigate which Na/H exchange protein isoforms are present in BBB endothelial cells and to evaluate the effects of ischemic factors on BBB Na/H exchange activity. Western blot analysis of bovine cerebral microvascular endothelial cells (CMEC) and immunoelectron microscopy of perfusion-fixed rat brain revealed that Na/H exchanger isoforms 1 and 2 (NHE1 and NHE2) are present in BBB endothelial cells. Using microspectrofluorometry and the pH-sensitive dye BCECF, we found that hypoxia (2% O2, 30 min), aglycemia (30 min), and AVP (1-200 nM, 5 min) significantly increased CMEC Na/H exchange activity, assessed as Na-dependent, HOE-642-sensitive H + flux. We found that AVP stimulation of CMEC Na/H exchange activity is dependent on intracellular Ca concentration and is blocked by V1, but not V2, vasopressin receptor antagonists. Our findings support the hypothesis that a BBB Na/H exchanger, possibly NHE1 and/or NHE2, is stimulated during ischemia to participate in cerebral edema formation.

KW - Blood-brain barrier

KW - Brain edema

KW - Cariporide

KW - Cerebral ischemia

KW - Stroke

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U2 - 10.1152/ajpcell.00093.2009

DO - 10.1152/ajpcell.00093.2009

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JO - American Journal of Physiology - Renal Fluid and Electrolyte Physiology

JF - American Journal of Physiology - Renal Fluid and Electrolyte Physiology

SN - 1931-857X

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