The chloride transporter Na(+)-K(+)-Cl- cotransporter isoform-1 contributes to intracellular chloride increases after in vitro ischemia.

Journal Article

Ischemic episodes in the CNS cause significant disturbances in neuronal ionic homeostasis. To directly measure changes in intracellular Cl- concentration ([Cl-]i) during and after ischemia, we used Clomeleon, a novel ratiometric optical indicator for Cl-. Hippocampal slices from adult transgenic mice expressing Clomeleon in hippocampal neurons were subjected to 8 min of oxygen-glucose deprivation (OGD) (an in vitro model for ischemia) and reoxygenated in the presence of glucose. This produced mild neuronal damage 3 h later that was prevented when the extracellular [Cl-] was maintained at 10 mm during reoxygenation. OGD induced a transient decrease in fluorescence resonance energy transfer within Clomeleon, indicating an increase in [Cl-]i. During reoxygenation, there was a partial recovery in [Cl-]i, but [Cl-]i rose again 45 min later. To investigate sources of Cl- accumulation, we examined the effects of Cl- transport inhibitors on the rises in [Cl-]i during and after OGD. Bumetanide and furosemide, which inhibit Cl- influx through the Na(+)-K(+)-Cl- cotransporter isoform-1 (NKCC-1) and efflux through the K(+)-Cl- cotransporter isoform-2, were unable to inhibit the first rise in [Cl-]i, yet entirely prevented the secondary rise in [Cl-]i during reoxygenation. In contrast, picrotoxin, which blocks the GABA-gated Cl- channel, did not inhibit the secondary rise in [Cl-]i after OGD. [Cl-]i increases during reoxygenation were accompanied by an increase in phosphorylation of NKCC-1, an indication of increased NKCC-1 activity after OGD. We conclude that NKCC-1 plays an important role in OGD-induced Cl- accumulation and subsequent neuronal damage.

Full Text

Duke Authors

Cited Authors

  • Pond, BB; Berglund, K; Kuner, T; Feng, G; Augustine, GJ; Schwartz-Bloom, RD

Published Date

  • February 1, 2006

Published In

Volume / Issue

  • 26 / 5

Start / End Page

  • 1396 - 1406

PubMed ID

  • 16452663

Electronic International Standard Serial Number (EISSN)

  • 1529-2401

Digital Object Identifier (DOI)

  • 10.1523/JNEUROSCI.1421-05.2006

Language

  • eng

Conference Location

  • United States