Glutamate toxicity in single cortical neurons

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

Neurons are electrically excitable cells which produce relatively large amounts of ATP to drive the ion pumps that restore intracellular Na⁺ and Ca²⁺ levels after synaptic activity and action potentials. Glutamate, the major excitatory neurotransmitter in mammals, is responsible for fast synaptic transmission. Excessive activation of glutamate receptors can result in irreversible disruption of ion homeostasis and cell death, a process called excitotoxicity. Many of the physiological effects of glutamate on neurons are mediated by Ca²⁺ influx and excessive Ca²⁺ influx is believed to play a major role in excitotoxicity. However, the exact mechanisms are still a matter of debate.

In order to determine the sequence of cellular processes in glutamate toxicity, Marc Gleichmann et al. simultaneously recorded O₂ consumption, cytosolic Ca²⁺ concentration ([Ca²⁺]_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 [Ca²⁺]_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 [Ca²⁺]_i followed seconds afterwards by an increase in O₂ 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 O₂ consumption was reached, the mΔψ, as indicated by TMRE⁺ fluorescence, dissipated. Maximal O₂ consumption lasted up to 5 min and then declined together with mΔψ and ATP levels, while [Ca²⁺]_i further increased. mΔψ and [Ca²⁺]_i returned to baseline levels when neurons were treated with an NMDA receptor antagonist shortly after the [Ca²⁺]_i increased.

Their 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 O₂ consumption, [Ca²⁺]_i, and mΔψ. Their method will also allow the measurement of O₂ consumption in cases where cell numbers are limited, such as preparations of neurons from adult animal brain or human biopsies.

Web-Link: http://onlinelibrary.wiley.com/doi/10.1111/j.1471-4159.2009.05997.x/full.

Keywords: excitotoxicity; glutamate; oxygen consumption

Reference: Gleichmann, M., Collis, L. P., Smith, P. J. S. and Mattson, M. P. (2009), Simultaneous single neuron recording of O₂ consumption, [Ca²⁺]_i and mitochondrial membrane potential in glutamate toxicity. Journal of Neurochemistry, 109: 644-655.