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Simultaneous single neuron recording of O 2 consumption, [Ca 2+ ] i and mitochondrial membrane potential in glutamate toxicity
Author(s) -
Gleichmann Marc,
Collis Leon P.,
Smith Peter J. S.,
Mattson Mark P.
Publication year - 2009
Publication title -
journal of neurochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.75
H-Index - 229
eISSN - 1471-4159
pISSN - 0022-3042
DOI - 10.1111/j.1471-4159.2009.05997.x
Subject(s) - glutamate receptor , nmda receptor , membrane potential , biophysics , chemistry , cytosol , amperometry , receptor , biochemistry , biology , electrode , electrochemistry , enzyme
Abstract 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 (mΔψ) in single cortical neurons. Oxygen consumption was measured using an amperometric self‐referencing platinum electrode adjacent to neurons in which [Ca 2+ ] i and mΔψ 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 mΔψ occurred during this time period, and then, shortly before maximal O 2 consumption was reached, the mΔψ, as indicated by TMRE + fluorescence, dissipated. Maximal O 2 consumption lasted up to 5 min and then declined together with mΔψ and ATP levels, while [Ca 2+ ] i further increased. mΔψ 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 mΔψ. 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.