Abstracts

Rationale: Temporal lobe epilepsy (TLE) is the most common form of epilepsy with focal seizures. TLE is also frequently associated with resistance to currently available antiepileptic drugs (AEDs). Current AEDs work primarily by targeting neurons through enhancement of GABAergic neurotransmission or attenuation of glutamatergic neurotransmission. Astrocytes play a key role in the modulation of neurotransmission and glutamate homeostasis through glutamate transporters glutamate transporter-1 (GLT-1) and glutamate aspartate transporter (GLAST). Previously, GLT-1 has been shown to be reduced at epileptic foci in patients with TLE. Here, we aim to determine the role of glutamate transporters in glutamate modulation during epileptogenesis in the intrahippocampal kainic acid model of epilepsy. Methods: 8- to 10-week-old male CD1 wild-type (WT) mice were used in these experiments. At day 0, animals were injected with either 64 nL of saline or 20 mM kainic acid into the dorsal hippocampus. Following injection, IHKA-injected mice experienced status epilepticus for at least 3 hours. Mice were sacrificed at 1, 3, 7 and 30 days post-IHKA and used for synaptosomal fraction and Western blot analysis. A subset of mice had an electrode and biosensor cannula implanted in the ipsilateral dorsal hippocampus immediately following kainic acid administration. At 14 days post-IHKA, the dummy cannula was replaced with an enzymatic glutamate biosensor. The biosensor and electrode were connected to an acquisition device to record extracellular glutamate and electrical activity in the hippocampus for 24 hours. Results: Following crude synaptosomal fractionation, Western blot analysis showed significant downregulation in synaptosomal GLT-1 protein levels in the ipsilateral dorsal hippocampus 7 days post-IHKA (p = 0.0064) that returned to near baseline levels by 30 days post-IHKA (p > 0.05). There were no changes observed in synaptosomal GLT-1 expression in the contralateral dorsal hippocampus during epileptogenesis (p > 0.05). Western blot analysis showed a significant upregulation in synaptosomal GLAST expression at 30 days post-IHKA compared with 7 days post-IHKA in the ipsilateral dorsal hippocampus (p = 0.033) but not contralaterally (p > 0.05). At 14 days post-IHKA induced status-epilepticus, mice had an average of 14 spontaneous seizures per hour. Based on our results, we categorized glutamate spike events associated with spontaneous seizures into four separate categories: pre-seizure glutamate spike events, intra-seizure glutamate spike events, post-seizure glutamate spike events, and interictal glutamate spike events. Pre-seizure glutamate spike events ranged from 0.835 μM - 8.559 μM glutamate increase from baseline. Post-seizure spike events ranged from 0.501 μM- 6.818 μM glutamate increase from baseline.  Conclusions: We found that synaptosomal GLT-1 protein levels were significantly downregulated 7 days post-IHKA in the ipsilateral hippocampus. GLAST synaptosomal protein levels were not significantly different from saline-injected animals (control). We also characterized glutamate spike events at 14 days post-IHKA induced status epilepticus. Overall, our data support the hypothesis that modulation of glutamate transport may be an important antiepileptogenic strategy. Funding: No funding