Abstracts

Rationale: The characteristics of the epileptogenic human hippocampus include: above normal interictal extracellular glutamate levels and enhanced glutamate release during spontaneous seizures with abnormally slow post-ictal glutamate clearance. Glutamine synthase expression and activity are below normal in patients with mesial temporal sclerosis as is the rate of the glutamate-glutamine cycle measured during surgery. Based on these data we hypothesize that elevated extracellular glutamate is a consequence of impaired glial function due to both decreased rates of glial uptake and/or metabolism. A limitation of our previous results is that the rate of glutamine synthesis was not measured directly.Methods: We measured the interictal rate of glutamine synthesis from multiple sites with 16 microdialysis probes of six awake patients with refractory localization-related epilepsy during long-term intracranial EEG monitoring. Yale University Human Investigations Committee approved these experiments. Patients were infused intravenously with 3.75 grams of sodium-1-13C-acetate over 2.5 hours on day 4 after depth electrode implantation. The patients were given 10 grams of 1-13C-glucose over 2.5 hours on the next day. Microdialysate was collected at a rate of 20 microliter every 10 minutes and analyzed by liquid chromatography tandem mass spectrometry for glutamate and glutamine levels and their isotopic enrichments.Results: After the 13C-acetate infusion, the isotopic enrichment of extracellular glutamine was significantly less (p < 0.006) in the probes with high extracellular glutamate levels than the probes with normal glutamate levels. Log-linear regression analysis showed a highly significant (R 0.746, n 16, p < 0.001) association between the ECF glutamate levels and ECF %13C-glutamine. After the 13C-glucose infusion, the %13C-glutamine was significantly less (p < 0.001) in the probes with the high ECF glutamate levels than the probes with normal glutamate levels. Log-linear regression analysis showed a highly significant (R 0.804, n 16, p < 0.001) association between the ECF glutamate levels and ECF %13C-glutamine.Conclusions: The isotopic enrichment of microdialysis glutamine was higher for probes with nearly normal glutamate after infusion of labeled substrates, which suggests the rate of glutamine synthesis is lower in those areas of the brain with above normal extracellular glutamate. The data obtained with an infusion of 13C-glucose or 13C-acetate on different days in the same subjects were the same, which increases our confidence in the findings. The variation among multiple probes in the same patient reflects regional variation in glutamine synthesis and extracellular glutamate. Glutamine synthesis is limited to glia; therefore, our data suggest glial dysfunction in regions with above normal extracellular glutamate concentrations. Funding: NIH-NINDS RO1-NS-054038