Intra-Hippocampal Kainate Model (mouse)

The intra-hippocampal kainate mouse model of mesial temporal lobe epilepsy (MTLE) recapitulates many of the characteristics observed in human patients with temporal lobe epilepsy (TLE). The MTLE mouse is characterized by an initial neurotoxic event, a unilateral intrahippocampal injection of kainic acid (KA) into the dorsal hippocampus, which induces a non-convulsive status epilepticus (SE) lasting several hours. This initial event is followed by a period of two to three weeks during which epileptogenesis takes place with the appearance of spikes, bursts, and discharges. Spontaneous, recurrent, electrographic hippocampal paroxysmal discharges (HPD) are afterwards stably observed in the injected hippocampus and stereotyped for the whole life of the animal (Bouilleret et al., 1999; Riban et al., 2002; Duveau et al., 2016, Duveau and Roucard, 2017; Twele et al., 2017). These HPDs occur spontaneously about 30-60 times per hour when the animals are in a state of quiet wakefulness, generally last 15-20 sec, and are associated with behavioral arrest and/or mild motor automatisms. To reliably record HPDs, the animals need to be implanted with an intra-hippocampal depth electrode.

Experiments are performed on male C57BL/6J mice of at least 10 weeks of age using a protocol similar to that described by Duveau et al., 2016. Briefly, animals are anesthetized (2-3% isoflurane in oxygen) and placed in a stereotactic frame in a flat skull position and 1 nmol KA solution (Sigma) in 0.9% sterile saline is injected into the right dorsal hippocampus. After KA injection, all mice are implanted with a bipolar electrode into the injected hippocampus. Before and between 6 and 8h after surgery, animals receive an injection of buprenorphine (0.05mg/kg). After surgery, mice are housed in individual cages with food and water ad libitum and kept under a 12/12h light-dark controlled cycle. All experiments are performed during the light hours of the cycle. The animals are allowed to recover for four weeks (to allow the development of HPDs) prior to evaluation in the screening protocol.

To detect and quantify electrographic HPDs, the electroencephalogram (EEG) is recorded in freely moving animals placed in individual plexiglass cages using SystemPlus Evolution (Micromed). Offline analysis is performed by an expert to detect HPDs, defined as rhythmic high-amplitude sharp waves occurring with a frequency of 5-10 Hz and lasting at least 5 seconds with a minimum interevent interval of 1 second. Using a cohort of validated MTLE mice, the compounds are typically first tested using the effective dose identified in prior mouse screening tests (e.g., 6 Hz 44 mA model). If a compound demonstrates efficacy at this dose, the dose can then be increased depending on other relevant factors, i.e., the known TD50, to vary the protection observed. Animals are used as their own controls. EEG recordings are performed on freely moving animals for 20 minutes pre-injection (reference period) and up to 240 minutes post-injection depending on the compound’s time of peak effect. Data are analyzed for the period of 10 minutes before and 10 minutes after peak time of effect of the investigational compound, as determined from prior mouse screening tests. The effects of the injected compound are compared to the reference period. Data are presented as the raw number of HPDs during the analyzed 20 min period (10 minutes before and 10 minutes after peak time of effect of compound) for each MTLE mouse, group mean number of HPDs (n = 4-8 MTLE mice), and effect on suppression of epileptic discharges as represented by percent of baseline HPD values. Any accompanying observed effect on animal behavior is noted.