N-methyl-D-aspartate receptor activation in human cerebral endothelium promotes intracellular oxidant stress

Christopher D. Sharp, J. Houghton, J. W. Elrod, A. Warren, T. H. Jackson IV, A. Jawahar, A. Nanda, A. Minagar, J. S. Alexander

Research output: Contribution to journalArticlepeer-review

62 Scopus citations

Abstract

Cerebral endothelial cells in the rat, pig, and, most recently, human have been shown to express several types of receptors specific for glutamate. High levels of glutamate disrupt the cerebral endothelial barrier via activation of N-methyl-D-aspartate (NMDA) receptors. We have previously suggested that this glutamate-induced barrier dysfunction was oxidant dependent. Here, we provide evidence that human cerebral endothelial cells respond to glutamate by generating an intracellular oxidant stress via NMDA receptor activation. Cerebral endothelial cells loaded with the oxidant-sensitive probe dihydrorhodamine were used to measure intracellular reactive oxygen species (ROS) formation in response to glutamate receptor agonists, antagonists, and second message blockers. Glutamate (1 mM) significantly increased ROS formation compared with sham controls (30 min). This ROS response was significantly reduced by 1) MK-801, a noncompetitive NMDA receptor antagonist; 2) 8-(N,N-diethylamino)-n-octyl-3,4,5-trimethoxybenzoate, an intracellular Ca 2+ antagonist; 3) LaCl3, an extracellular Ca2+ channel blocker; 4) diphenyleiodonium, a hemeferryl-containing protein inhibitor; 5) itraconazole, a cytochrome P-450 3A4 inhibitor; and 6) cyclosporine A, which prevents mitochondrial membrane pore transition required for mitochondrial-dependent ROS generation. Our results suggest that the cerebral endothelial barrier dysfunction seen in response to glutamate is Ca2+ dependent and may require several intracellular signaling events mediated by oxidants derived from reduced nicotinamide adenine dinucleotide oxidase, cytochrome P-450, and the mitochondria.

Original languageEnglish (US)
Pages (from-to)H1893-H1899
JournalAmerican Journal of Physiology - Heart and Circulatory Physiology
Volume288
Issue number4 57-4
DOIs
StatePublished - Apr 1 2005
Externally publishedYes

All Science Journal Classification (ASJC) codes

  • Physiology
  • Cardiology and Cardiovascular Medicine
  • Physiology (medical)

Keywords

  • Arachidonic acid
  • Brain
  • Human
  • Mitochondria
  • Reactive oxygen species
  • Reduced nicotinamide adenine dinucleotide oxidase

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