NMDA-evoked consumption and recovery of mitochondrially targeted aequorin suggests increased Ca²⁺ uptake by a subset of mitochondria in hippocampal neurons

Activation of NMDA receptors produces large increases in cytosolic Ca ²⁺ that are taken up into mitochondria. We used recombinant aequorin targeted to mitochondria to report changes in matrix Ca²⁺ in rat hippocampal neurons in culture. Upon binding Ca²⁺, aequorin emits a photon in a one-shot reaction that consumes the indicator. Here we show that stimulation with NMDA produced a mitochondrial Ca²⁺ response that rapidly inactivated. However, following a 30-min recovery period the response was restored, suggesting the presence of a pool of indicator that was not exposed to high Ca²⁺ during the initial stimulus. We speculate that aequorin distant from the Ca²⁺ source was protected from microdomains of high Ca²⁺ near the plasmalemma and that this aequorin moved, either by movement of individual mitochondria or via the mitochondrial tubular network, to replenish consumed indicator during the recovery time. A large Ca²⁺ increase in a subset of mitochondria could produce local changes in energy metabolism, regional Ca²⁺ buffering, and foci that initiate neurotoxic processes.