Effect of carbamazepine on spontaneous recurrent seizures recorded from the dentate gyrus in rats with kainate-induced epilepsy.

OBJECTIVE: Animal models of chronic epilepsy with spontaneous recurrent seizures (SRSs) may be useful in the discovery and mechanistic analyses of antiseizure drugs (ASDs). Carbamazepine (CBZ), a widely used ASD with a well-defined mechanism, was analyzed in this proof-of-principle study to determine how a traditional ASD affects the properties of SRSs. METHODS: The effects of CBZ on electrographic SRSs recorded from the dentate gyrus were studied in freely behaving rats using a repeated, low-dose kainate model of acquired epilepsy with a repeated-measures, crossover protocol. RESULTS: Almost all seizure durations were >20 seconds. Both seizure likelihood and duration appeared to be similar between 1 and 8 hours after individual CBZ injections. CBZ-induced decreases in seizure frequency were not significant at 10 mg/kg; however, at 30 mg/kg, seizure frequency was significantly reduced for convulsive but not nonconvulsive seizures. At 100 mg/kg, CBZ strongly suppressed both convulsive and nonconvulsive seizures. Although CBZ had a dose-dependent effect on seizure frequency, CBZ did not affect seizure duration at any dose. The preceding interictal interval did not affect seizure duration; however, at 30 mg/kg CBZ, nearly all seizures were nonconvulsive when the interictal interval was <30 minutes (ie, during clusters). SIGNIFICANCE: Increased doses of CBZ (10-100 mg/kg) suppressed the frequency but not the duration of convulsive and nonconvulsive seizures in the repeated, low-dose kainate model. The repeated-measures, crossover protocol, which requires relatively few animals and compensates for progressive increases in seizure frequency during epileptogenesis after status epilepticus, allowed quantitative analyses of clinically relevant and translatable properties of SRSs.

Maternal Stress Combined with Terbutaline Leads to Comorbid Autistic-Like Behavior and Epilepsy in a Rat Model.

Human autism is comorbid with epilepsy, yet, little is known about the causes or risk factors leading to this combined neurological syndrome. Although genetic predisposition can play a substantial role, our objective was to investigate whether maternal environmental factors alone could be sufficient. We examined the independent and combined effects of maternal stress and terbutaline (used to arrest preterm labor), autism risk factors in humans, on measures of both autistic-like behavior and epilepsy in Sprague-Dawley rats. Pregnant dams were exposed to mild stress (foot shocks at 1 week intervals) throughout pregnancy. Pups were injected with terbutaline on postnatal days 2-5. Either maternal stress or terbutaline resulted in autistic-like behaviors in offspring (stereotyped/repetitive behaviors and deficits in social interaction or communication), but neither resulted in epilepsy. However, their combination resulted in severe behavioral symptoms, as well as spontaneous recurrent convulsive seizures in 45% and epileptiform spikes in 100%, of the rats. Hippocampal gliosis (GFAP reactivity) was correlated with both abnormal behavior and spontaneous seizures. We conclude that prenatal insults alone can cause comorbid autism and epilepsy but it requires a combination of teratogens to achieve this; testing single teratogens independently and not examining combinatorial effects may fail to reveal key risk factors in humans. Moreover, astrogliosis may be common to both teratogens. This new animal model of combined autism and epilepsy permits the experimental investigation of both the cellular mechanisms and potential intervention strategies for this debilitating comorbid syndrome.

Seizure-related regulation of GABAA receptors in spontaneously epileptic rats.

In this study, we analyzed the impact that spontaneous seizures might have on the plasma membrane expression, composition and function of GABAA receptors (GABAARs). For this, the tissue of chronically epileptic rats was collected within 3h of seizure occurrence (≤3h group) or at least 24h after seizure occurrence (≥24h group). A retrospective analysis of seizure frequency revealed that selecting animals on the bases of seizure proximity also grouped animals in terms of overall seizure burden with a higher seizure burden observed in the ≤3h group. A biochemical analysis showed that although animals with more frequent/recent seizures (≤3h group) had similar levels of GABAAR at the plasma membrane they showed deficits in inhibitory neurotransmission. By contrast, the tissue obtained from animals experiencing infrequent seizures (≥24h group) had increased plasma membrane levels of GABAAR and showed no deficit in inhibitory function. Together, our findings offer an initial insight into the molecular changes that might help to explain how alterations in GABAAR function can be associated with differential seizure burden. Our findings also suggest that increased plasma membrane levels of GABAAR might act as a compensatory mechanism to more effectively maintain inhibitory function, repress hyperexcitability and reduce seizure burden. This study is an initial step towards a fuller characterization of the molecular events that trigger alterations in GABAergic neurotransmission during chronic epilepsy.

Effect of spontaneous seizures on GABAA receptor α4 subunit expression in an animal model of temporal lobe epilepsy.

OBJECTIVE: Temporal lobe epilepsy (TLE) is frequently medically intractable and often progressive. Compromised inhibitory neurotransmission due to altered γ-aminobutyric acid (GABA)A receptor α4 subunit (GABAA Rα4) expression has been emphasized as a potential contributor to the initial development of epilepsy following a brain insult (primary epileptogenesis), but the regulation of GABAA Rα4 during chronic epilepsy, specifically, how expression is altered following spontaneous seizures, is less well understood. METHODS: Continuous video-electroencephalography (EEG) recordings from rats with pilocarpine-induced TLE were used to capture epileptic animals within 3 h of a spontaneous seizure (SS), or >24 h after the last SS, to determine whether recent occurrence of a seizure was associated with altered levels of GABAA Rα4 expression. We further evaluated whether this GABAA Rα4 plasticity is regulated by signaling mechanisms active in primary epileptogenesis, specifically, increases in brain-derived neurotrophic factor (BDNF) and early growth response factor 3 (Egr3). RESULTS: Elevated levels of GABAA Rα4 messenger RNA (mRNA) and protein were observed following spontaneous seizures, and were associated with higher levels of BDNF and Egr3 mRNA. SIGNIFICANCE: These data suggest that spontaneous, recurrent seizures that define chronic epilepsy may influence changes in GABAA Rα4 expression, and that signaling pathways known to regulate GABAA Rα4 expression after status epilepticus may also be activated after spontaneous seizures in chronically epileptic animals.

Molecular pathways controlling inhibitory receptor expression.

Epilepsy is a disease of complex etiology, and multiple molecular mechanisms contribute to its development. Temporal lobe epilepsy (TLE) may result from an initial precipitating event such as hypoxia, head injury, or prolonged seizure (i.e., status epilepticus [SE]), that is followed by a latent period of months to years before spontaneous seizures occur. γ-Aminobutyric acid (GABA)(A) receptor (GABA(A) R) subunit changes occur during this latent period and may persist following the onset of spontaneous seizures. Research into the molecular mechanisms regulating these changes and potential targets for intervention to reverse GABA(A) R subunit alterations have uncovered seizure-induced pathways that contribute to epileptogenesis. Several growth or transcription factors are known to be activated by SE, including (but not limited to): brain-derived neurotrophic factor (BDNF), cAMP response element binding protein (CREB), inducible cAMP early repressor (ICER), and early growth response factors (Egrs). Results of multiple studies suggest that these factors transcriptionally regulate GABA(A) R subunit gene expression in a way that is pertinent to the development of epilepsy. This article focuses on these signaling elements and describes their possible roles in gene regulatory pathways that may be critical in the development of chronic epilepsy.

Time-frequency analysis using damped-oscillator pseudo-wavelets: Application to electrophysiological recordings.

The damped-oscillator pseudo-wavelet is presented as a method of time-frequency analysis along with a new spectral density measure, the data power. An instantaneous phase can be defined for this pseudo-wavelet, and it is easily inverted. The data power measure is tested on both computer generated data and in vivo intrahippocampal electrophysiological recordings from a rat. The data power spectral density is found to give better time and frequency resolution than the more conventional total energy measure, and additionally shows intricate time-frequency structure in the rat that is altered in association with the emergence of epilepsy. With epileptogenesis, the baseline theta oscillation is severely degraded and is absorbed into a broader gamma band. There are also broad 600 Hz and 2000 Hz bands which localize to hippocampal layers that are distinct from those of the theta and gamma bands. The 600 Hz band decreases in prominence with epileptogenesis while the 2000 Hz band increases in prominence. The origins of these high frequency bands await further study. In general, we find that the damped-oscillator pseudo-wavelet is easy to use and is particularly suitable for problems where a wide range of oscillator frequencies is expected.

Narp regulates homeostatic scaling of excitatory synapses on parvalbumin-expressing interneurons.

Homeostatic synaptic scaling alters the strength of synapses to compensate for prolonged changes in network activity and involves both excitatory and inhibitory neurons. The immediate-early gene Narp (neuronal activity-regulated pentraxin) encodes a secreted synaptic protein that can bind to and induce clustering of AMPA receptors (AMPARs). We found that Narp prominently accumulated at excitatory synapses on parvalbumin-expressing interneurons (PV-INs). Increasing network activity resulted in a homeostatic increase of excitatory synaptic strength onto PV-INs that increased inhibitory drive and this response was absent in neurons cultured from Narp-/- mice. Activity-dependent changes in the strength of excitatory inputs on PV-INs in acute hippocampal slices were also dependent on Narp and Narp-/- mice had increased sensitivity to kindling-induced seizures. We propose that Narp recruits AMPARs at excitatory synapses onto PV-INs to rebalance network excitation/inhibition dynamics following episodes of increased circuit activity.

A new potential AED, carisbamate, substantially reduces spontaneous motor seizures in rats with kainate-induced epilepsy.

PURPOSE: Animal models with spontaneous epileptic seizures may be useful in the discovery of new antiepileptic drugs (AEDs). The purpose of the present study was to evaluate the efficacy of carisbamate on spontaneous motor seizures in rats with kainate-induced epilepsy. METHODS: Repeated, low-dose (5 mg/kg), intraperitoneal injections of kainate were administered every hour until each male Sprague-Dawley rat had experienced convulsive status epilepticus for at least 3 h. Five 1-month trials (n = 8-10 rats) assessed the effects of 0.3, 1, 3, 10, and 30 mg/kg carisbamate on spontaneous seizures. Each trial involved six AED-versus-vehicle tests comprised of carisbamate or 10% solutol-HS-15 treatments administered as intraperitoneal injections on alternate days with a recovery day between each treatment day. RESULTS: Carisbamate significantly reduced motor seizure frequency at doses of 10 and 30 mg/kg, and caused complete seizure cessation during the 6-h postdrug epoch in seven of the eight animals at 30 mg/kg. The effects of carisbamate (0.3-30 mg/kg) on spontaneous motor seizures appeared dose dependent. CONCLUSIONS: These data support the hypothesis that a repeated-measures, crossover protocol in animal models with spontaneous seizures is an effective method for testing AEDs. Carisbamate reduced the frequency of spontaneous motor seizures in a dose-dependent manner, and was more effective than topiramate at reducing seizures in rats with kainate-induced epilepsy.

Anticonvulsant effects of carbamazepine on spontaneous seizures in rats with kainate-induced epilepsy: comparison of intraperitoneal injections with drug-in-food protocols.

PURPOSE: The present study evaluated the effectiveness of intraperitoneal (IP) injections and oral administration of carbamazepine (CBZ) in food on the frequency of spontaneous motor seizures in rats with kainate-induced epilepsy. The purpose was to develop a convenient drug-in-food approach for continuous, long-term administration of potential antiepileptic drugs (AEDs). METHODS: Single IP injections of CBZ (10-100 mg/kg) were compared to vehicle injections via six AED-versus-vehicle tests using a repeated-measures, crossover protocol. Similar protocols were used with CBZ-containing or control food pellets. RESULTS: CBZ significantly reduced motor seizure frequency at 30 and 100 mg/kg after single IP injections, and these doses completely blocked motor seizures during a 6-h postdrug epoch in 25% and 70% of the animals, respectively. Single administrations of 30 mg/kg and 100 mg/kg CBZ in food also significantly reduced motor seizures, and blocked seizures in 33% and 89% of the rats, respectively. CBZ administered in food three times per day (100 mg/kg x3 CBZ in food) continuously blocked nearly all motor seizures over a 5-day period, and completely suppressed motor seizures in 50% of the animals tested. CONCLUSIONS: CBZ strongly suppresses spontaneous motor seizures, and single doses of CBZ in food are as effective as IP injections in rats with kainate-induced epilepsy. CBZ administered regularly in food continuously blocks nearly all motor seizures, and may provide a relatively simple method to test AEDs in chronic models of epilepsy.

Use of chronic epilepsy models in antiepileptic drug discovery: the effect of topiramate on spontaneous motor seizures in rats with kainate-induced epilepsy.

PURPOSE: Potential antiepileptic drugs (AEDs) are typically screened on acute seizures in normal animals, such as those induced in the maximal electroshock and pentylenetet-razole models. As a proof-of-principle test, the present experiments used spontaneous epileptic seizures in kainate-treated rats to examine the efficacy of topiramate (TPM) with a repeated-measures, crossover protocol. METHODS: Kainic acid was administered in repeated low doses (5 mg/kg) every hour until each Sprague-Dawley rat experienced convulsive status epilepticus for >3 h. Six 1-month trials (n = 6-10 rats) assessed the effects of 0.3-100 mg/kg TPM on spontaneous seizures. Each trial involved six pairs of TPM and saline-control treatments administered as intraperitoneal injections on alternate days with a recovery day between each treatment day. Data analysis included a log transformation to compensate for the asymmetric distribution of values and the heterogeneous variances, which appeared to arise from clustering of seizures. RESULTS: A significant effect of TPM was observed for 12 h (i.e., two 6-h periods) after a 30-mg/kg injection, and full recovery from the drug effect was complete within 43 h. TPM exerted a significant effect at doses of 10, 30, and 100 mg/kg, and the effects of TPM (0.3-100 mg/kg) were dose dependent. CONCLUSIONS: These data suggest that animal models with spontaneous seizures, such as kainate- and pilocarpine-treated rats, can be used efficiently for rapid testing of AEDs with a repeated-measures, crossover protocol. Furthermore, the results indicate that this design allows both dose-effect and time-course-of-recovery studies.