Knocking down of the KCC2 in rat hippocampal neurons increases intracellular chloride concentration and compromises neuronal survival

J Physiol. 2011 May 15;589(Pt 10):2475-96. doi: 10.1113/jphysiol.2010.203703. Epub 2011 Mar 21.

Abstract

KCC2 is a neuron-specific potassium-chloride co-transporter controlling intracellular chloride homeostasis in mature and developing neurons. It is implicated in the regulation of neuronal migration, dendrites outgrowth and formation of the excitatory and inhibitory synaptic connections. The function of KCC2 is suppressed under several pathological conditions including neuronal trauma, different types of epilepsies, axotomy of motoneurons, neuronal inflammations and ischaemic insults. However, it remains unclear how down-regulation of the KCC2 contributes to neuronal survival during and after toxic stress. Here we show that in primary hippocampal neuronal cultures the suppression of the KCC2 function using two different shRNAs, dominant-negative KCC2 mutant C568A or DIOA inhibitor, increased the intracellular chloride concentration [Cl⁻]i and enhanced the toxicity induced by lipofectamine-dependent oxidative stress or activation of the NMDA receptors. The rescuing of the KCC2 activity using over-expression of the active form of the KCC2, but not its non-active mutant Y1087D, effectively restored [Cl⁻]i and enhanced neuronal resistance to excitotoxicity. The reparative effects of KCC2 were mimicked by over-expression of the KCC3, a homologue transporter. These data suggest an important role of KCC2-dependent potassium/chloride homeostasis under neurototoxic conditions and reveal a novel role of endogenous KCC2 as a neuroprotective molecule.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Cell Survival / drug effects
  • Cells, Cultured
  • Chlorides / metabolism*
  • Down-Regulation
  • Hippocampus / metabolism*
  • Lipids / adverse effects
  • Neurons / drug effects
  • Neurons / metabolism
  • Oxidative Stress / drug effects
  • Patch-Clamp Techniques
  • Rats
  • Rats, Wistar
  • Receptors, N-Methyl-D-Aspartate / agonists
  • Symporters / genetics
  • Symporters / metabolism*
  • gamma-Aminobutyric Acid / metabolism

Substances

  • Chlorides
  • Lipids
  • Lipofectamine
  • Receptors, N-Methyl-D-Aspartate
  • Symporters
  • potassium-chloride symporters
  • gamma-Aminobutyric Acid