Simultaneous single neuron recording of O2 consumption, [Ca2+]i and mitochondrial membrane potential in glutamate toxicity

J Neurochem. 2009 Apr;109(2):644-55. doi: 10.1111/j.1471-4159.2009.05997.x. Epub 2009 Feb 16.


In order to determine the sequence of cellular processes in glutamate toxicity, we simultaneously recorded O(2) consumption, cytosolic Ca(2+) concentration ([Ca(2+)](i)), and mitochondrial membrane potential (mDeltapsi) in single cortical neurons. Oxygen consumption was measured using an amperometric self-referencing platinum electrode adjacent to neurons in which [Ca(2+)](i) and mDeltapsi were monitored with Fluo-4 and TMRE(+), respectively, using a spinning disk laser confocal microscope. Excitotoxic doses of glutamate caused an elevation of [Ca(2+)](i) followed seconds afterwards by an increase in O(2) consumption which reached a maximum level within 1-5 min. A modest increase in mDeltapsi occurred during this time period, and then, shortly before maximal O(2) consumption was reached, the mDeltapsi, as indicated by TMRE(+) fluorescence, dissipated. Maximal O(2) consumption lasted up to 5 min and then declined together with mDeltapsi and ATP levels, while [Ca(2+)](i) further increased. mDeltapsi and [Ca(2+)](i) returned to baseline levels when neurons were treated with an NMDA receptor antagonist shortly after the [Ca(2+)](i) increased. Our unprecedented spatial and time resolution revealed that this sequence of events is identical in all neurons, albeit with considerable variability in magnitude and kinetics of changes in O(2) consumption, [Ca(2+)](i), and mDeltapsi. The data obtained using this new method are consistent with a model where Ca(2+) influx causes ATP depletion, despite maximal mitochondrial respiration, minutes after glutamate receptor activation.

Publication types

  • Comparative Study
  • Research Support, N.I.H., Extramural
  • Research Support, N.I.H., Intramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Calcium / metabolism*
  • Cations, Divalent / metabolism
  • Cell Survival / drug effects
  • Cell Survival / physiology
  • Cells, Cultured
  • Cerebral Cortex / cytology
  • Cerebral Cortex / drug effects
  • Cerebral Cortex / metabolism
  • Cytosol / metabolism*
  • Glutamic Acid / toxicity*
  • Membrane Potential, Mitochondrial / drug effects
  • Membrane Potential, Mitochondrial / physiology*
  • Mitochondrial Membranes / drug effects
  • Mitochondrial Membranes / metabolism*
  • Neurons / drug effects
  • Neurons / metabolism*
  • Oxygen Consumption / drug effects
  • Oxygen Consumption / physiology*
  • Rats
  • Rats, Sprague-Dawley


  • Cations, Divalent
  • Glutamic Acid
  • Calcium