The incidence rate of status epilepticus is ~7% of all annual epilepsy cases (Lowenstein, 1999). Frequently the seizures are intractable to current anticonvulsants, and thus, there is a significant unmet clinical need for novel drugs to treat this disorder. This investigation was undertaken to develop and validate an in vitro model of status epilepticus using kainic acid (KA)-treated hippocampal organotypic cultures.

Hippocampal organotypic slice cultures were prepared from 5-7 day old male and female Wistar rats by the roller‑tube method (Gähwiler et al., 1997). At 14 days in vitro (DIV), KA (1 µM) was added for 24h. Following KA application, single-unit extracellular recordings of spontaneous discharge activity were recorded from CA1 neurones and analysed using NeuroEXplorer (Plexon Inc., USA); epileptiform‑like burst activity was defined as a3 spikes with an interspike interval <300ms and a minimum duration of 10ms. After 24h, the cultures were incubated with sodium valproate (100 and 300 µM) or levetiracetam (10 and 100 µM). Neuronal cell death was determined by propidium iodide (2 µM) uptake into the CA1 region 24h after KA treatment, visualized by confocal microscope and quantified by densitometry.

Prior to addition of KA, hippocampal cells exhibited spontaneous discharge activity (16.5±4.1 bursts/min, n=12 cells/12 cultures). A gradual increase in the burst-rate was observed which was significant 210min after KA application (control: 9.8±2.3 bursts/min, n=4 cells/culture; KA 42.2±9.4 bursts/min, n =8 cells/culture, P<0.05 one way ANOVA with post-hoc Bonferonni). The burst-rate remained significantly higher 24h following KA incubation (44.6±8.3 bursts/min, n=7 cells/7 cultures, P<0.05) and only returned to control levels 240min after removal of KA (control: 17.5±12.4 bursts/min, n=4 cells/4 cultures; KA: 29.09±4.9 bursts/min, n=7 cells/7 cultures, P>0.05). KA-induced burst firing at 15 DIV (80.8±3.6 bursts/min, n=3 cells/3 cultures) was concentration-dependently decreased by levetiracetam (10 µM: 40.9±4.1 bursts/min, P<0.05; 100 µM: 17.9±6.8 bursts/min, P<0.01) but sodium valproate was without effect (P>0.05) at either concentration. Exposure to KA (1 µM for 24hr) significantly increased (P<0.01) propidium iodide uptake into CA1 neurones (1.1±0.4%, n=9 cultures) compared with controls (0.1±0.06%, n=9 cultures).

These data demonstrate that KA (1 µM for 24hr) produces epileptiform-like burst activity and associated selective neuronal cell death in a manner similar to that observed clinically with status epilepticus. The contrasting effects of levetiracetam and sodium valproate on KA‑induced epileptiform‑like activity are consistent with their postulated clinical utility in status epilepticus. Thus, this in vitro system can be used to evaluate the potential anticonvulsant and neuroprotective effects of novel compounds for the treatment of status epilepticus.

Gähwiler BH, et al. (1997) Trends Neurosci. 20 471-477.
Lowenstein DH. (1999) Epilepsia 40 S3-S8.