Abstracts

Rationale: Epilepsy is a common neurological disorder in which the random nature of seizures poses difficult research challenges. We recently developed a novel in vivo model of ictogenesis, allowing experimental modulation of the risk of temporal lobe seizures. In the present study, we used this model to search for biochemical changes associated with increased seizure risk. Methods: Male Sprague-Dawley rats with intraperitoneal pilocarpine were prepared as described in Luna-Munguia et al. (2017) to generate Epileptic (n=15) and Control (n=15) animals. A cannula implanted into nucleus reuniens was used for local KCl or PBS injection while another cannula was implanted into left hippocampus for microdialysis experiments. During intracerebral microdialysis experiments, KCl or PBS were injected (120 mM or 1X, respectively; 0.1 µl/min over 5 min) into the nucleus reuniens of freely moving rats, a process recently shown to increase the risk of seizures up to three-fold. This injection was a total of 9 times, with 15 min between injections, comprising 180 min. Dialysates were collected before (6 collections), during (27 collections) and after (4 collections) KCl or PBS injections, derivatized immediately, and analyzed by liquid chromatography-mass spectrometry (LC-MS) to assess the extracellular concentrations of 24 different neurotransmitters. Results: The majority of Epileptic animals had seizures during the KCl injections. The LC-MS analysis revealed that pilocarpine produced significant differences in baseline levels of several neurotransmitters within the hippocampus, the most significant of which were decreased levels of adenosine (79%), homovanillic acid (55%), and serotonin (86%) and increased choline (286%). During the KCl injection into nucleus reuniens, which increased the risk of seizures, there were additional significant changes in several neurotransmitters, most significantly increased 5-HIAA and decreased serine and aspartate. Both the difference in baseline concentrations (between animals with and without pilocarpine-induced seizures) and the difference during the KCl injection showed complex interactions between multiple neurotransmitters, an effect that we quantified using stepwise logistic regression.  There was no difference in measurements taken less than one minute prior to a seizure.  Conclusions: Our results are the first to show how the pilocarpine model alters the basal hippocampal extracellular concentrations of 24 neurotransmitters simultaneously. In addition, we have identified several neurotransmitters that are altered during a time period of increased seizure risk. These neurotransmitters, and the interaction between them, are candidates for future experiments investigating the basic mechanisms of ictogenesis, search for biomarkers associated with that risk, and potentially develop and optimize more effective antiseizure therapies. Funding: NIH 1K08NS0697831R01NS094399The Michigan Brain Initiative Working Group1K01ES026839