Temporal lobe epilepsy after experimental prolonged febrile seizures: prospective analysis

Abstract

Experimental prolonged febrile seizures (FS) lead to structural and molecular changes that promote hippocampal hyperexcitability and reduce seizure threshold to further convulsants. However, whether these seizures provoke later-onset epilepsy, as has been suspected in humans, has remained unclear. Previously, intermittent EEGs with behavioural observations for motor seizures failed to demonstrate spontaneous seizures in adult rats subjected to experimental prolonged FS during infancy. Because limbic seizures may be behaviourally subtle, here we determined the presence of spontaneous limbic seizures using chronic video monitoring with concurrent hippocampal and cortical EEGs, in adult rats (starting around 3 months of age) that had sustained experimental FS on postnatal day 10. These subjects were compared with groups that had undergone hyperthermia but in whom seizures had been prevented (hyperthermic controls), as well as with normothermic controls. Only events that fulfilled both EEG and behavioural criteria, i.e. electro-clinical events, were considered spontaneous seizures. EEGs (over 400 recorded hours) were normal in all normothermic and hyperthermic control rats, and none of these animals developed spontaneous seizures. In contrast, prolonged early-life FS evoked spontaneous electro-clinical seizures in 6 out of 17 experimental rats (35.2%). These seizures consisted of sudden freezing (altered consciousness) and typical limbic automatisms that were coupled with polyspike/sharp-wave trains with increasing amplitude and slowing frequency on EEG. In addition, interictal epileptiform discharges were recorded in 15 (88.2%) of the experimental FS group and in none of the controls. The large majority of hippocampally-recorded seizures were heralded by diminished amplitude of cortical EEG, that commenced half a minute prior to the hippocampal ictus and persisted after seizure termination. This suggests a substantial perturbation of normal cortical neuronal activity by these limbic spontaneous seizures. In summary, prolonged experimental FS lead to later-onset limbic (temporal lobe) epilepsy in a significant proportion of rats, and to interictal epileptifom EEG abnormalities in most others, and thus represent a model that may be useful to study the relationship between FS and human temporal lobe epilepsy.

Fig 1

Study design.

Fig. 2

Epilepsy after experimental prolonged febrile seizures (FS): Quantitative analysis of the number of rats developing epileptic EEG and behavioural changes. The prolonged FS group (n = 17) sustained hyperthermia for 30 min on postnatal day (P)10, that elicited seizures lasting 24.1 ± 0.1 min. Chronic video-EEGs were recorded during P90–180 in this group and in normothermic and hyperthermic control rats (see Materials and methods for definitions). In both control groups, EEGs recorded over more than 400 h did not contain any epileptiform discharges. None of the controls developed spontaneous seizures. The FS evoked interictal events in 88.2% of rats, and spontaneous seizures, with both behavioural and electrographic manifestation, seizures in six (35.2%) rats.

Fig. 3

Typical spontaneous electrographic seizures recorded from hippocampal electrodes in adult rats that had sustained prolonged febrile seizures (FS) early in life. Seizures were defined as events meeting both EEG and behavioural criteria: electrographic parameters included presence of polyspikes or sharp-wave trains lasting over 6 s (see Materials and methods). (A) Hippocampal EEGs. Arrow points to onset and end of epileptiform discharges. The typical behaviour associated with these events was sudden cessation of activity accompanied by facial automatisms. Typically, behaviour changes slightly preceded the onset of hippocampal seizures, and lasted longer. Hippocampal EEG from a normothermic control rat shows low amplitude baseline trace. (B) Histogram of the duration of hippocampal EEG-recorded spontaneous seizures in adult rats that had endured prolonged FS early in life.

Fig. 4

Perturbation of cortical activity during spontaneous seizures evoked by experimental febrile seizures (FS) precedes and outlasts hippocampal discharges. (A) Example of reduced cortical EEG amplitude (indicated by the arrowheads) occurring before the onset of a hippocampally-evident spontaneous seizure and persisting during and beyond it, for a total duration of 92 s (arrows denote the onset and the end of hippocampal epileptiform discharges). (B) Histogram of the duration of these cortical perturbations during seizures in rats rendered epileptic by prolonged FS early in life. Note that reduced amplitude of cortical EEG lasts on average 49 ± 6.4 s, and in some cases up to 4 min.

Fig. 5

Postictal depression in hippocampal EEG follows spontaneous seizures evoked by early-life febrile seizures. (A) An example of postictal depression (arrowheads denote the onset and end of the depression) occurring immediately following the seizure. (B) Comparison of the mean durations of hippocampal and cortical postictal depressions, showing that hippocampal postictal depression averaged 3.2 ± 0.4 s, whereas cortical EEG postictal depression lasted 15.2 ± 3.1 s. *P < 0.05.

Fig. 6

Interictal epileptiform activity recorded from hippocampal electrodes in adult rats that had endured prolonged febrile seizures early in life. (A) Examples of interictal epileptiform activity (arrowheads), defined as polyspikes or sharp-waves shorter than 5 s, and without altered behaviour. (B) Histogram of the duration of the interictal discharges in the six epileptic rats. The distribution indicates that the majority of these events lasted between 2.5 and 4 s. (C) Histogram of the duration of the interictal epileptiform discharges analysed by rat. The distribution of the duration of these events seems to differ among individual animals. e.g. event durations in rat no. 3 were significantly lower than those in rats nos. 1, 4 or 5, P < 0.05.

Fig. 7

Raw energy spectra of individual nocturnal hippocampal EEGs from rats with febrile seizures (FS)-evoked spontaneous seizures (pink; n = 6) compared with FS-experiencing rats with epileptiform interictal discharges only (black; n = 9) and to normothermic control rats (blue; n = 8). A significant reduction of low-frequency energy (arrows) is apparent in rats with spontaneous seizures (epileptic), suggesting that these seizures may perturb global neuronal network activity. For each group, the solid line represents the mean of the raw energy; dashed line denotes the standard errors.

Fig. 8

Neuronal density analyses in hippocampal regions most affected in temporal lobe epilepsy do not indicate cell loss in epileptic rats. Analyses were performed on sections subjected to Nissl stain. This stain permits distinction between neurons (large nuclei, ample cytoplasm with abundant ‘Nissl material’ (ribosomes) and astrocytes (smaller nuclei, cant pale cytoplasm). (A) Illustration of areas selected for data sampling in hilus and CA1 (see Materials and methods). Neuronal densities in hilus (B) and CA1 (C) of rats that had become epileptic after early-life prolonged febrile seizures (pink bars) did not differ from densities in normothermic controls (blue bars). (D) Dorsal hippocampal (left y-axis) and hilar volume (right y-axis) were the same in epileptic and control rats, excluding volume changes as a potential confounder of these analyses.