Dynamic Changes in Cytosolic ATP Levels in Cultured Glutamatergic Neurons During NMDA-Induced Synaptic Activity...

We have previously shown that synaptic transmission fails in cultured neurons in the presence of lactate as the sole substrate. Thus, to test the hypothesis that the failure of synaptic transmission is a consequence of insufficient energy supply, ATP levels were monitored employing the ATP biosensor Ateam1.03YEMK. While inducing synaptic activity by subjecting cultured neurons to two 30 s pulses of NMDA (30 µM) with a 4 min interval, changes in relative ATP levels were measured in the presence of lactate (1 mM), glucose (2.5 mM) or the combination of the two. ATP levels reversibly declined following NMDA-induced neurotransmission activity, as indicated by a reversible 10-20 % decrease in the response of the biosensor. The responses were absent when the NMDA receptor antagonist memantine was present. In the presence of lactate alone, the ATP response dropped significantly more than in the presence of glucose following the 2nd pulse of NMDA (approx. 10 vs. 20 %). Further, cytosolic Ca(2+) homeostasis during NMDA-induced synaptic transmission is partially inhibited by verapamil indicating that voltage-gated Ca(2+) channels are activated. Lastly, we showed that cytosolic Ca(2+) homeostasis is supported equally well by both glucose and lactate, and that a pulse of NMDA causes accumulation of Ca(2+) in the mitochondrial matrix. In summary, we have shown that ATP homeostasis during neurotransmission activity in cultured neurons is supported by both glucose and lactate. However, ATP homeostasis seems to be negatively affected by the presence of lactate alone, suggesting that glucose is needed to support neuronal energy metabolism during activation.