Abstracts

Rationale: Approximately 1% of the general population has epilepsy, and up to 30% of individuals with this disease cannot control their seizures with antiepileptic drugs. One of the most devastating features of epilepsy is the uncertainty of when a seizure will occur. Therefore, the discovery of new approaches aimed at predicting and treating epileptic seizures is particularly important. We hypothesize that unique alterations of the extracellular brain chemistry precede by several hours the occurrence of spontaneous seizures in patients with drug-resistant focal epilepsies. We postulate that a better understanding of these alterations could facilitate the development of predictive biomarkers and more efficacious treatments for epileptic seizures.Methods: Using chemical profiling (metabolomics) by mass spectrometry, we explored seizure-associated changes in the brain extracellular fluid (ECF) of nine patients with drug-resistant focal epilepsies. These patient underwent EEG-electrode implantation to localize their seizure focus. Brain ECF samples were collected via microdialysis catheters located within the shaft of the depth electrodes. On average, there were 2 catheters per patient. Three groups of samples were analyzed: (Pre-sz), collected 5-24 hrs before a seizure, (Sz), collected during a seizure, and (Post-sz), collected 1-96 hrs after a seizure. The samples were analyzed using ultra-performance liquid chromatography followed by Q-TOF mass spectrometry. Principal component analysis and paired t-tests were used to assess for group differences. The resultant accurate masses of parent and fragment ions were run against three databases for preliminary identification: KEGG, METLIN and The Human Metabolome Database. Final identification was performed by comparing parent and fragment ions from patient samples with that of analytical grade standards of the candidate chemicals.Results: More than 1,000 different chemicals were detected in the samples. Unsupervised principal component analysis of the chemical changes revealed, with a few exceptions, clear clustering into the 3 comparison groups. Between 10 and 20 chemicals were significantly changed in concentration among the 3 comparison groups. Some of these chemicals are involved in the metabolic pathways of glutamate and GABA.Conclusions: This is the first high-throughput chemical profiling study of the brain extracellular fluid in patients with drug-resistant focal epilepsies. We have demonstrated the feasibility of this approach by detecting more than 1,000 different chemicals in dilute, low volume (5-10 uL) brain microdialysis samples. We are presently identifying as many of the significantly changed chemicals as possible. Future studies are aimed at defining the exact time course and biological significance of the chemical changes by supplementary studies in human subjects and relevant laboratory models. The information obtained from these studies is likely to improve our understanding of the mechanism of epileptic seizures, and thereby facilitate the development of predictive biomarkers and more efficacious therapeutics against drug-resistant seizures.