Interictal Spikes Precede Ictal Discharges in an Organotypic Hippocampal Slice Culture Model of Epileptogenesis

Abstract

In organotypic hippocampal slice cultures, principal neurons form aberrant excitatory connections with other principal cells in response to slicing-induced deafferentation, similar to mechanisms underlying epileptogenesis in post-traumatic epilepsy. To investigate the consequences of this synaptogenesis, we recorded field potential activity from area CA3 during perfusion with the complete growth medium used during incubation. At 7 days in vitro (DIV), slice cultures only displayed multi-unit activity. At 14 DIV the majority displayed population bursts reminiscent of interictal-like spikes, but sustained synchronous activity was rare. Bandpass filtering of interictal discharges revealed fast ripple-like complexes, similar to in vivo recordings. Spontaneous ictal-like activity became progressively more prevalent with age: at 21 DIV 50% of organotypic hippocampal slice cultures displayed long lasting, ictal-like discharges that could be suppressed by phenytoin, whereas interictal-activity was not suppressed. The fraction of cultures displaying ictal events continually increased with incubation time. Quantification of population spike activity throughout epileptogenesis using automatic detection and clustering algorithms confirmed the appearance of interictal-like activity before ictal-like discharges, and also revealed high frequency pathological multi-unit activity in slice cultures at 14–17 DIV. These experiments indicate that interictal-like spikes precede the appearance of ictal-like activity in a reduced in vitro preparation. Epileptiform activity in cultures resembled in vivo epilepsy, including sensitivity to anticonvulsants and steadily increasing seizure incidence over time, although seizure frequency and rate of epileptogenesis were higher in vitro. Organotypic hippocampal slice cultures comprise a useful model system for investigating mechanisms of epileptogenesis as well as developing anti-epileptic and anti-epileptogenic drugs.

Epileptogenesis

“…Early epileptiform activity in electrographic recordings is a promising predictor of epilepsy after brain injury induced by kainic acid . Similar results were obtained in vitro in the organotypic hippocampal slice culture model of epileptogenesis (Dyhrfjeld-Johnsen et al 2010). These studies undertook a simple but important step beyond prior EEG biomarker studies by greatly increasing the duration of the EEG sample from the standard clinical EEG of tens of minutes up to 24-h recording epochs (Jennett and Van De Sande 1975).…”

Adenosine Release during Seizures Attenuates GABAAReceptor-Mediated Depolarization

“…The pH was adjusted to be between 7.35 and 7.40 using NaOH. Previous studies have shown that hippocampal organotypic slices can be used as a post-traumatic model of epileptogenesis (Dyhrfjeld-Johnsen et al, 2010;Berdichevsky et al, 2012). By 7 d in vitro the tissue has recovered from the acute trauma associated with the slicing process and the neurons exhibit hyperexcitability, probably due to increased connectivity (Berdichevsky et al, 2012).…”

Staged anticonvulsant screening for chronic epilepsy

“…The use of organotypic hippocampal cultures, where epileptogenesis occurs on a compressed time scale,23, 24 and where seizures and seizure‐induced cell death can be easily quantified with biomarker assays,30, 31 allowed us to circumvent the throughput limitations of in vivo chronic epilepsy models 4. Thus, we were able to screen 140 compounds and combinations of compounds, in over 400 separate drug‐concentration experiments, and identify over 40 hits.…”

“…In light of this abundant reciprocal excitation, it is not surprising that organotypic slice cultures are hyperexcitable19 and even spontaneously epileptic20, 21 (see Heinemann and Staley22 for review). We have characterized epileptogenesis in this preparation 23, 24. Increases in extracellular lactate accompanied seizures in human hippocampus,25, 26 and we found that lactate production was correlated with seizure‐like activity in organotypic cultures.…”