Excitotoxic mitochondrial depolarisation requires both calcium and nitric oxide in rat hippocampal neurons

J Physiol. 1999 Nov 1;520 Pt 3(Pt 3):797-813. doi: 10.1111/j.1469-7793.1999.00797.x.


1. Glutamate neurotoxicity has been attributed to cellular Ca2+ overload. As mitochondrial depolarisation may represent a pivotal step in the progression to cell death, we have used digital imaging techniques to examine the relationship between cytosolic Ca2+ concentration ([Ca2+]c) and mitochondrial potential (DeltaPsim) during glutamate toxicity, and to define the mechanisms underlying mitochondrial dysfunction. 2. In cells of > 11 days in vitro (DIV), exposure to 50 mM potassium or 100 microM glutamate had different consequences for DeltaPsim. KCl caused a small transient loss of DeltaPsim but in response to glutamate there was a profound loss of DeltaPsim. In cells of 7-10 DIV, glutamate caused only a modest and reversible drop in DeltaPsim. 3. Using fura-2 to measure [Ca2+]c, responses to KCl and glutamate did not appear significantly different. However, use of the low affinity indicator fura-2FF revealed a difference in the [Ca2+]c responses to KCl and glutamate, which clearly correlated with the loss of DeltaPsim. Neurons exhibiting a profound mitochondrial depolarisation also showed a large secondary increase in the fura-2FF ratio. 4. The glutamate-induced loss of DeltaPsim was dependent on Ca2+ influx. However, inhibition of nitric oxide synthase (NOS) by L-NAME significantly attenuated the loss of DeltaPsim. Furthermore, photolysis of caged NO at levels that had no effect alone promoted a profound mitochondrial depolarisation when combined with high [Ca2+]c, either in response to KCl or to glutamate in cultures at 7-10 DIV. 5. In cells that showed only modest mitochondrial responses to glutamate, induction of a mitochondrial depolarisation by the addition of NO was followed by a secondary rise in [Ca2+]c. These data suggest that [Ca2+]c and nitric oxide act synergistically to cause mitochondrial dysfunction and impaired [Ca2+]c homeostasis during glutamate toxicity.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Calcium / metabolism
  • Calcium / physiology*
  • Culture Techniques
  • Cytosol / metabolism
  • Electrophysiology
  • Fluorescent Dyes
  • Fura-2
  • Glutamic Acid / poisoning*
  • Hippocampus / cytology
  • Hippocampus / physiology*
  • Mitochondria / drug effects
  • Mitochondria / physiology*
  • Neurons / physiology*
  • Nitric Oxide / physiology*
  • Osmolar Concentration
  • Rats
  • Rhodamine 123


  • Fluorescent Dyes
  • Rhodamine 123
  • Nitric Oxide
  • Glutamic Acid
  • Calcium
  • Fura-2