Enhanced burst discharges in the CA1 area of the immature versus adult hippocampus: patterns and cellular mechanisms.

Burst discharges in the immature brain may contribute to its enhanced seizure susceptibility. The cellular mechanisms underlying burst discharges in the CA1 area of the immature versus adult hippocampus were investigated with simultaneous whole-cell and field-potential recordings. When GABAA receptors were blocked pharmacologically, bursts in CA1 were either graded or all-or-none (or mixed) as a function of electrical stimulation intensity. Most CA1 minislices from immature rats displayed all-or-none or mixed bursts, whereas the slices from adult rats predominantly elicited graded bursts. The frequency and amplitude of spontaneous excitatory postsynaptic currents (sEPSCs) were greater in CA1 pyramidal cells from the immature than the adult slices. The developmental differences in CA1 bursting were also detected in slices adjusted for maturational changes in brain volume (i.e., 350 µm thick for immature vs. 450 µm thick for adult rats). Neither N-methyl-d-aspartate (NMDA) nor group I metabotropic glutamate (mGlu1) receptor antagonists blocked the network-driven bursts in immature CA1, but an α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor blocker abolished them. Robust excitatory postsynaptic potentials (EPSPs) occurred after bursts in some immature CA1 slices (23%) but never in slices from the adult. The input-output (amount of current injected vs. number of action potentials generated) relationship was markedly greater in CA1 pyramidal cells in the immature compared with the adult hippocampus. These data suggest that the CA1 area of the immature brain is capable of generating network-driven bursts, which declines in adult rats. The increased propensity of burst generation in immature CA1 appears to involve a greater AMPA receptor-mediated synaptic network and an increased intrinsic spike-generating ability.NEW & NOTEWORTHY Burst discharges in the developing brain can provide valuable insights into epileptogenesis. We show that the immature hippocampal CA1 area is capable of generating all-or-none (i.e., network) bursts, which transitions to graded (i.e., nonnetwork) bursts in the mature brain via both synaptic and intrinsic mechanisms. Our results provide new clues to help understand possible mechanisms that may be shared in the immature and epileptic brain and how the normal brain becomes seizure prone (i.e., epileptogenesis).

Sensitivity of unilateral- versus bilateral-onset spike-wave discharges to ethosuximide and carbamazepine in the fluid percussion injury rat model of traumatic brain injury.

Unilateral-onset spike-wave discharges (SWDs) following fluid percussion injury (FPI) in rats have been used for nearly two decades as a model for complex partial seizures in human posttraumatic epilepsy (PTE). This study determined if SWDs with a unilateral versus bilateral cortical onset differed. In this experiment, 2-mo-old rats received severe FPI (3 atm) or sham surgery and were instrumented for chronic video-electrocorticography (ECoG) recording (up to 9 mo). The antiseizure drug, carbamazepine (CBZ), and the antiabsence drug, ethosuximide (ETX), were administered separately to determine if they selectively suppressed unilateral- versus bilateral-onset SWDs, respectively. SWDs did not significantly differ between FPI and sham rats on any measured parameter (wave-shape, frequency spectrum, duration, or age-related progression), including unilateral (∼17%) versus bilateral (∼83%) onsets. SWDs with a unilateral onset preferentially originated ipsilateral to the craniotomy in both FPI and sham rats, suggesting that the unilateral-onset SWDs were related to surgical injury and not specifically to FPI. ETX profoundly suppressed SWDs with either unilateral or bilateral onsets, and CBZ had no effect on either type of SWD. These results suggest that SWDs with either a unilateral or bilateral onset have a pharmacosensitivity similar to absence seizures and are very different from the complex partial seizures of PTE. Therefore, SWDs with a unilateral onset after FPI are not a model of the complex partial seizures that occur in PTE, and their use for finding new treatments for PTE could be counterproductive, particularly if their close similarity to normal brain oscillations is not acknowledged.NEW & NOTEWORTHY Unilateral-onset spike-wave discharges (SWDs) in rats have been used to model complex partial seizures in human posttraumatic epilepsy (PTE), compared to bilateral-onset SWDs thought to reflect human absence seizures. Here, we show that both unilateral- and bilateral-onset SWDs following traumatic brain injury are suppressed by the antiabsence drug ethosuximide and are unaffected by the antiseizure drug carbamazepine. We propose that unilateral-onset SWDs are not useful for studying mechanisms of, or treatments for, PTE.

The Kv7 Modulator, Retigabine, is an Efficacious Antiseizure Drug for Delayed Treatment of Organophosphate-induced Status Epilepticus.

Benzodiazepines are the primary treatment option for organophosphate (OP)-induced status epilepticus (SE), but these antiseizure drugs (ASDs) lose efficacy as treatment is delayed. In the event of a mass civilian or military exposure, significant treatment delays are likely. New ASDs that combat benzodiazepine-resistant, OP-induced SE are critically needed, particularly if they can be efficacious after a long treatment delay. This study evaluated the efficacy of the Kv7 channel modulator, retigabine, as a novel therapy for OP-induced SE. Adult, male rats were exposed to soman or diisopropyl fluorophosphate (DFP) to elicit SE and monitored by electroencephalogram (EEG) recording. Retigabine was administered alone or adjunctive to midazolam (MDZ) at delays of 20- or 40-min in the soman model, and 60-min in the DFP model. Following EEG recordings, rats were euthanized and brain tissue was collected for Fluoro-Jade B (FJB) staining to quantify neuronal death. In the DFP model, MDZ + 15 mg/kg retigabine suppressed seizure activity and was neuroprotective. In the soman model, MDZ + 30 mg/kg retigabine suppressed seizures at 20- and 40-min delays. Without MDZ, 15 mg/kg retigabine provided partial antiseizure and neuroprotectant efficacy in the DFP model, while 30 mg/kg without MDZ failed to attenuate soman-induced SE. At 60 mg/kg, retigabine without MDZ strongly reduced seizure activity and neuronal degeneration against soman-induce SE. This study demonstrates the antiseizure and neuroprotective efficacy of retigabine against OP-induced SE. Our data suggest retigabine could be a useful adjunct to standard-of-care and has potential for use in the absence of MDZ.

Delayed Adjunctive Treatment of Organophosphate-Induced Status Epilepticus in Rats with Phenobarbital, Memantine, or Dexmedetomidine.

Organophosphate (OP) exposure induces status epilepticus (SE), a medical emergency with high morbidity and mortality. Current standard medical countermeasures lose efficacy with time so that treatment delays, in the range of tens of minutes, result in increasingly poor outcomes. As part of the Countermeasures Against Chemical Threats Neurotherapeutics Screening Program, we previously developed a realistic model of delayed treatment of OP-induced SE using the OP diisopropyl fluorophosphate (DFP) to screen compounds for efficacy in the termination of SE and elimination of neuronal death. Male rats were implanted for electroencephalogram (EEG) recordings 7 days prior to experimentation. Rats were then exposed to DFP, and SE was induced for 60 minutes and then treated with midazolam (MDZ) plus one of three antiseizure drugs (ASDs)-phenobarbital (PHB), memantine (MEM), or dexmedetomidine (DMT)-in conjunction with antidotes. EEG was recorded for 24 hours, and brains were stained with Fluoro-Jade B for quantification of degenerating neurons. We found that PHB + MDZ induced a prolonged suppression of SE and reduced neuronal death. MEM + MDZ treatment exacerbated SE and increased mortality; however, surviving rats had fewer degenerating neurons. DMT + MDZ significantly suppressed SE with only a minimal reduction in neuronal death. These data demonstrate that delayed treatment of OP-induced SE with other ASDs, when added to MDZ, can achieve greater seizure suppression with additional reduction in degenerating neurons throughout the brain compared with MDZ alone. The effect of a drug on the severity of seizure activity did not necessarily determine the drug's effect on neuronal death under these conditions. SIGNIFICANCE STATEMENT: This study assesses the relative effectiveness of three different delayed-treatment regimens for the control of organophosphate-induced status epilepticus and reduction of subsequent neuronal death. The data demonstrate the potential for highly effective therapies despite significant treatment delay and a potential disconnect between seizure severity and neuronal death.

Screening for Efficacious Anticonvulsants and Neuroprotectants in Delayed Treatment Models of Organophosphate-induced Status Epilepticus.

Organophosphorus (OP) compounds are deadly chemicals that exert their intoxicating effects through the irreversible inhibition of acetylcholinesterase (AChE). In addition to an excess of peripheral ailments, OP intoxication induces status epilepticus (SE) which if left untreated may lead to permanent brain damage or death. Benzodiazepines are typically the primary therapies for OP-induced SE, but these drugs lose efficacy as treatment time is delayed. The CounterACT Neurotherapeutic Screening (CNS) Program was therefore established by the National Institutes of Health (NIH) to discover novel treatments that may be administered adjunctively with the currently approved medical countermeasures for OP-induced SE in a delayed treatment scenario. The CNS program utilizes in vivo EEG recordings and Fluoro-JadeB (FJB) histopathology in two established rat models of OP-induced SE, soman (GD) and diisopropylfluorophosphate (DFP), to evaluate the anticonvulsant and neuroprotectant efficacy of novel adjunct therapies when administered at 20 or 60 min after the induction of OP-induced SE. Here we report the results of multiple compounds that have previously shown anticonvulsant or neuroprotectant efficacy in other models of epilepsy or trauma. Drugs tested were ganaxolone, diazoxide, bumetanide, propylparaben, citicoline, MDL-28170, and chloroquine. EEG analysis revealed that ganaxolone demonstrated the most robust anticonvulsant activity, whereas all other drugs failed to attenuate ictal activity in both models of OP-induced SE. FJB staining demonstrated that none of the tested drugs had widespread neuroprotective abilities. Overall these data suggest that neurosteroids may represent the most promising anticonvulsant option for OP-induced SE out of the seven unique mechanisms tested here. Additionally, these results suggest that drugs that provide significant neuroprotection from OP-induced SE without some degree of anticonvulsant activity are elusive, which further highlights the necessity to continue screening novel adjunct treatments through the CNS program.

Enaminone Modulators of Extrasynaptic α4ß3δ γ-Aminobutyric AcidA Receptors Reverse Electrographic Status Epilepticus in the Rat After Acute Organophosphorus Poisoning.

Seizures induced by organophosphorus nerve agent exposure become refractory to treatment with benzodiazepines because these drugs engage synaptic γ-aminobutyric acid-A receptors (GABAARs) that rapidly internalize during status epilepticus (SE). Extrasynaptic GABAARs, such as those containing α4ß3δ subunits, are a putative pharmacological target to comprehensively manage nerve agent-induced seizures since they do not internalize during SE and are continuously available for activation. Neurosteroids related to allopregnanolone have been tested as a possible replacement for benzodiazepines because they target both synaptic and extrasynaptic GABAARs receptors. A longer effective treatment window, extended treatment efficacy, and enhanced neuroprotection represent significant advantages of neurosteroids over benzodiazepines. However, neurosteroid use is limited by poor physicochemical properties arising from the intrinsic requirement of the pregnane steroid core structure for efficacy rendering drug formulation problematic. We tested a non-steroidal enaminone GABAAR modulator that interacts with both synaptic and extrasynaptic GABAARs on a binding site distinct from neurosteroids or benzodiazepines for efficacy to control electrographic SE induced by diisopropyl fluorophosphate or soman intoxication in rats. Animals were treated with standard antidotes, and experimental therapeutic treatment was given following 1 h (diisopropyl fluorophosphate model) or 20 min (soman model) after SE onset. We found that the enaminone 2-261 had an extended duration of seizure termination (>10 h) in the diisopropyl fluorophosphate intoxication model in the presence or absence of midazolam (MDZ). 2-261 also moderately potentiated MDZ in the soman-induced seizure model but had limited efficacy as a stand-alone anticonvulsant treatment due to slow onset of action. 2-261 significantly reduced neuronal death in brain areas associated with either diisopropyl fluorophosphate- or soman-induced SE. 2-261 represents an alternate chemical template from neurosteroids for enhancing extrasynaptic α4ß3δ GABAAR activity to reverse SE from organophosphorous intoxication.

Spontaneous Recurrent Absence Seizure-like Events in Wild-Caught Rats.

Absence epilepsy is a heritable human neurological disorder characterized by brief nonconvulsive seizures with behavioral arrest, moderate-to-severe loss of consciousness (absence), and distinct spike-wave discharges (SWDs) in the EEG and electrocorticogram (ECoG). Genetic models of this disorder have been created by selectively inbreeding rats for absence seizure-like events with similar electrical and behavioral characteristics. However, these events are also common in outbred laboratory rats, raising concerns about whether SWD/immobility accurately reflects absence epilepsy as opposed to "normal" rodent behavior. We hypothesized that, if SWD/immobility models absence seizures, it would not exist in wild-caught rats due to the pressures of natural selection. To test this hypothesis, we compared chronic video/electrocorticogram recordings from male and female wild-caught (Brown-Norway [BN]) rats to recordings from laboratory outbred BN, outbred Long-Evans, and inbred WAG/Rij rats (i.e., a model of absence epilepsy). Wild-caught BN rats displayed absence-like SWD/immobility events that were highly similar to outbred BN rats in terms of spike-wave morphology, frequency, diurnal rhythmicity, associated immobility, and sensitivity to the anti-absence drug, ethosuximide; however, SWD bursts were less frequent and of shorter duration in wild-caught and outbred BN rats than the outbred Long-Evans and inbred WAG/Rij strains. We conclude that SWD/immobility in rats does not represent absence seizures, although they appear to have many similarities. In wild rats, SWD/immobility appears to represent normal brain activity that does not reduce survival in natural environments, a conclusion that logically extends to outbred laboratory rats and possibly to those that have been inbred to model absence epilepsy.SIGNIFICANCE STATEMENT Spike-wave discharges (SWDs), behavioral arrest, and diminished consciousness are cardinal signs of seizures in human absence epilepsy and are used to model this disorder in inbred rats. These characteristics, however, are routinely found in outbred laboratory rats, leading to debate on whether SWD/immobility is a valid model of absence seizures. The SWD/immobility events in wild-caught rats appear equivalent to those found in outbred and inbred rat strains, except for lower incidence and shorter durations. Our results indicate that the electrophysiological and behavioral characteristics of events underlying hypothetical absence epilepsy in rodent models are found in wild rats captured in their natural environment. Other criteria beyond observation of SWDs and associated immobility are required to objectively establish absence epilepsy in rat models.

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.

Methodologic recommendations and possible interpretations of video-EEG recordings in immature rodents used as experimental controls: A TASK1-WG2 report of the ILAE/AES Joint Translational Task Force.

The use of immature rodents to study physiologic aspects of cortical development requires high-quality recordings electroencephalography (EEG) with simultaneous video recording (vEEG) of behavior. Normative developmental vEEG data in control animals are fundamental for the study of abnormal background activity in animal models of seizures or other neurologic disorders. Electrical recordings from immature, freely behaving rodents can be particularly difficult because of the small size of immature rodents, their thin and soft skull, interference with the recording apparatus by the dam, and other technical challenges. In this report of the TASK1 Working Group 2 (WG2) of the International League Against Epilepsy/American Epilepsy Society (ILAE/AES) Joint Translational Task Force, we provide suggestions that aim to optimize future vEEG recordings from immature rodents, as well as their interpretation. We focus on recordings from immature rodents younger than 30 days old used as experimental controls, because the quality and correct interpretation of such recordings is important when interpreting the vEEG results of animals serving as models of neurologic disorders. We discuss the technical aspects of such recordings and compare tethered versus wireless approaches. We also summarize the appearance of common artifacts and various patterns of electrical activity seen in young rodents used as controls as a function of behavioral state, age, and (where known) sex and strain. The information herein will hopefully help improve the methodology of vEEG recordings from immature rodents and may lead to results and interpretations that are more consistent across studies from different laboratories.

Age-dependent behaviors, seizure severity and neuronal damage in response to nerve agents or the organophosphate DFP in immature and adult rats.

Exposure to nerve agents (NAs) and other organophosphates (OPs) can initiate seizures that rapidly progress to status epilepticus (SE). While the electrographic and neuropathological sequelae of SE evoked by NAs and OPs have been characterized in adult rodents, they have not been adequately investigated in immature animals. In this study postnatal day (PND) 14, 21 and 28 rat pups, along with PND70 animals as adult controls, were exposed to NAs (sarin, VX) or another OP (diisopropylfluorophosphate, DFP). We then evaluated behavioral and electrographic (EEG) correlates of seizure activity, and performed neuropathology using Fluoro-Jade B. Although all immature rats exhibited behaviors that are often characterized as seizures, the incidence, duration, and severity of the electrographic seizure activity were age-dependent. No (sarin and VX) or brief (DFP) EEG seizure activity was evoked in PND14 rats, while SE progressively increased in severity as a function of age in PND21, 28 and 70 animals. Fluoro-Jade B staining was observed in multiple brain regions of animals that exhibited prolonged seizure activity. Neuronal injury in PND14 animals treated with DFP was lower than in older animals and absent in rats exposed to sarin or VX. In conclusion, we found that NAs and an OP provoked robust SE and neuronal injury similar to adults in PND21 and PND28, but not in PND14, rat pups. Convulsive behaviors were often present independent of EEG seizures and were unaccompanied by neuronal damage. These differential responses should be considered when investigating medical countermeasures for NA and OP exposure in pediatric populations.

Progression of convulsive and nonconvulsive seizures during epileptogenesis after pilocarpine-induced status epilepticus.

Although convulsive seizures occurring during pilocarpine-induced epileptogenesis have received considerable attention, nonconvulsive seizures have not been closely examined, even though they may reflect the earliest signs of epileptogenesis and potentially guide research on antiepileptogenic interventions. The definition of nonconvulsive seizures based on brain electrical activity alone has been controversial. Here we define and quantify electrographic properties of convulsive and nonconvulsive seizures in the context of the acquired epileptogenesis that occurs after pilocarpine-induced status epilepticus (SE). Lithium-pilocarpine was used to induce the prolonged repetitive seizures characteristic of SE; when SE was terminated with paraldehyde, seizures returned during the 2-day period after pilocarpine treatment. A distinct latent period ranging from several days to >2 wk was then measured with continuous, long-term video-EEG. Nonconvulsive seizures dominated the onset of epileptogenesis and consistently preceded the first convulsive seizures but were still present later. Convulsive and nonconvulsive seizures had similar durations. Postictal depression (background suppression of the EEG) lasted for >100 s after both convulsive and nonconvulsive seizures. Principal component analysis was used to quantify the spectral evolution of electrical activity that characterized both types of spontaneous recurrent seizures. These studies demonstrate that spontaneous nonconvulsive seizures have electrographic properties similar to convulsive seizures and confirm that nonconvulsive seizures link the latent period and the onset of convulsive seizures during post-SE epileptogenesis in an animal model. Nonconvulsive seizures may also reflect the earliest signs of epileptogenesis in human acquired epilepsy, when intervention could be most effective. NEW & NOTEWORTHY Nonconvulsive seizures usually represent the first bona fide seizure following a latent period, dominate the early stages of epileptogenesis, and change in severity in a manner consistent with the progressive nature of epileptogenesis. This analysis demonstrates that nonconvulsive and convulsive seizures have different behavioral outcomes but similar electrographic signatures. Alternatively, epileptiform spike-wave discharges fail to recapitulate several key seizure features and represent a category of electrical activity separate from nonconvulsive seizures in this model.

The antihelminthic moxidectin enhances tonic GABA currents in rodent hippocampal pyramidal neurons.

Macrocyclic lactones (MLs) are commonly used treatments for parasitic worm and insect infections in humans, livestock, and companion animals. MLs target the invertebrate glutamate-activated chloride channel that is not present in vertebrates. MLs are not entirely inert in vertebrates, though; they have been reported to have activity in heterologous expression systems consisting of ligand-gated ion channels that are present in the mammalian central nervous system (CNS). However, these compounds are typically not able to reach significant concentrations in the CNS because of the activity of the blood-brain barrier P-glycoprotein extrusion system. Despite this, these compounds are able to reach low levels in the CNS that may be useful in the design of novel "designer" ligand-receptor systems that can be used to directly investigate neuronal control of behavior in mammals and have potential for use in treating human neurological diseases. To determine whether MLs might affect neurons in intact brains, we investigated the activity of the ML moxidectin (MOX) at native GABA receptors. Specifically, we recorded tonic and phasic miniature inhibitory postsynaptic currents (mIPSCs) in ex vivo brain slices. Our data show that MOX potentiated tonic GABA currents in a dose-dependent manner but had no concomitant effects on phasic GABA currents (i.e., MOX had no effect on the amplitude, frequency, or decay kinetics of mIPSCs). These studies indicate that behavioral experiments that implement a ML-based novel ligand-receptor system should take care to control for potential effects of the ML on native tonic GABA receptors. NEW & NOTEWORTHY We have identified a novel mechanism of action in the mammalian central nervous system for the antihelminthic moxidectin, commonly prescribed to animals worldwide and currently being evaluated for use in humans. Specifically, moxidectin applied to rodent brain slices selectively enhanced the tonic GABA conductance of hippocampal pyramidal neurons.

Targeted Interneuron Ablation in the Mouse Hippocampus Can Cause Spontaneous Recurrent Seizures.

The death of GABAergic interneurons has long been hypothesized to contribute to acquired epilepsy. These experiments tested the hypothesis that focal interneuron lesions cause acute seizures [i.e., status epilepticus (SE)] and/or chronic epilepsy [i.e., persistent spontaneous recurrent seizures (SRSs)]. To selectively ablate interneurons, Gad2-ires-Cre mice were injected unilaterally in the CA1 area of the dorsal hippocampus with an adeno-associated virus containing the diphtheria toxin receptor (DTR). Simultaneously, an electrode, connected to a miniature telemetry device, was positioned at the injection site for chronic recordings of local field potentials (LFPs). Two weeks after virus transfection, intraperitoneal injection of DT consistently caused focal, specific, and extensive ablation of interneurons. Long-term, continuous monitoring revealed that all mice with DT-induced interneuron lesions had SRSs. Seizures lasted tens of seconds and interseizure intervals were several hours (or days); therefore, these interneuron lesions did not induce SE. The SRSs occurred 3-5 d after DT treatment, which is the estimated time required for DT-induced cell death; therefore, induction of SRSs occurred without the latent period typical of acquired epilepsy. In five of six DT-treated mice, SRSs stopped within days, suggesting that the DT-induced interneuron lesions did not usually cause epilepsy. In one mouse, however, SRSs occurred for ≥34 d after interneuron ablation, similar to epilepsy after experimental SE. Sham control mice had no detectable seizures, confirming that the SRSs were due to ablation of interneurons. These data show that selective interneuron ablation consistently caused SRSs but not SE; and, at least under the conditions used here, interneuron lesions rarely led to persistent SRSs (i.e., epilepsy).

Voluntary Control of Epileptiform Spike-Wave Discharges in Awake Rats.

Genetically inherited absence epilepsy in humans is typically characterized by brief (seconds) spontaneous seizures, which involve spike-wave discharges (SWDs) in the EEG and interruption of consciousness and ongoing behavior. Genetic (inbred) models of this disorder in rats have been used to examine mechanisms, comorbidities, and antiabsence drugs. SWDs have also been proposed as models of complex partial seizures (CPSs) following traumatic brain injury (post-traumatic epilepsy). However, the ictal characteristics of these rat models, including SWDs and associated immobility, are also prevalent in healthy outbred laboratory rats. We therefore hypothesized that SWDs are not always associated with classically defined absence seizures or CPSs. To test this hypothesis, we used operant conditioning in male rats to determine whether outbred strains, Sprague Dawley and Long-Evans, and/or the inbred WAG/Rij strain (a rat model of heritable human absence epilepsy) could exercise voluntary control over these epileptiform events. We discovered that both inbred and outbred rats could shorten the duration of SWDs to obtain a reward. These results indicate that SWD and associated immobility in rats may not reflect the obvious cognitive/behavioral interruption classically associated with absence seizures or CPSs in humans. One interpretation of these results is that human absence seizures and perhaps CPSs could permit a far greater degree of cognitive capacity than often assumed and might be brought under voluntary control in some cases. However, these results also suggest that SWDs and associated immobility may be nonepileptic in healthy outbred rats and reflect instead voluntary rodent behavior unrelated to genetic manipulation or to brain trauma.SIGNIFICANCE STATEMENT Our evidence that inbred and outbred rats learn to control the duration of spike-wave discharges (SWDs) suggests a voluntary behavior with maintenance of consciousness. If SWDs model mild absence seizures and/or complex partial seizures in humans, then an opportunity may exist for operant control complementing or in some cases replacing medication. Their equal occurrence in outbred rats also implies a major potential confound for behavioral neuroscience experiments, at least in adult rats where SWDs are prevalent. Alternatively, the presence and voluntary control of SWDs in healthy outbred rats could indicate that these phenomena do not always model heritable absence epilepsy or post-traumatic epilepsy in humans, and may instead reflect typical rodent behavior.

Staged anticonvulsant screening for chronic epilepsy.

OBJECTIVE: Current anticonvulsant screening programs are based on seizures evoked in normal animals. One-third of epileptic patients do not respond to the anticonvulsants discovered with these models. We evaluated a tiered program based on chronic epilepsy and spontaneous seizures, with compounds advancing from high-throughput in vitro models to low-throughput in vivo models. METHODS: Epileptogenesis in organotypic hippocampal slice cultures was quantified by lactate production and lactate dehydrogenase release into culture media as rapid assays for seizure-like activity and cell death, respectively. Compounds that reduced these biochemical measures were retested with in vitro electrophysiological confirmation (i.e., second stage). The third stage involved crossover testing in the kainate model of chronic epilepsy, with blinded analysis of spontaneous seizures after continuous electrographic recordings. RESULTS: We screened 407 compound-concentration combinations. The cyclooxygenase inhibitor, celecoxib, had no effect on seizures evoked in normal brain tissue but demonstrated robust antiseizure activity in all tested models of chronic epilepsy. INTERPRETATION: The use of organotypic hippocampal cultures, where epileptogenesis occurs on a compressed time scale, and where seizure-like activity and seizure-induced cell death can be easily quantified with biomarker assays, allowed us to circumvent the throughput limitations of in vivo chronic epilepsy models. Ability to rapidly screen compounds in a chronic model of epilepsy allowed us to find an anticonvulsant that would be missed by screening in acute models.

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.

Epileptogenesis.

Epileptogenesis is a chronic process that can be triggered by genetic or acquired factors, and that can continue long after epilepsy diagnosis. In 2015, epileptogenesis is not a treatment indication, and there are no therapies available in clinic to treat individuals at risk of epileptogenesis. However, thanks to active research, a large number of animal models have become available for search of molecular mechanisms of epileptogenesis. The first glimpses of treatment targets and biomarkers that could be developed to become useful in clinic are in sight. However, the heterogeneity of the epilepsy condition, and the dynamics of molecular changes over the course of epileptogenesis remain as challenges to overcome.

Long-term Continuous EEG Monitoring in Small Rodent Models of Human Disease Using the Epoch Wireless Transmitter System.

Many progressive neurologic diseases in humans, such as epilepsy, require pre-clinical animal models that slowly develop the disease in order to test interventions at various stages of the disease process. These animal models are particularly difficult to implement in immature rodents, a classic model organism for laboratory study of these disorders. Recording continuous EEG in young animal models of seizures and other neurological disorders presents a technical challenge due to the small physical size of young rodents and their dependence on the dam prior to weaning. Therefore, there is not only a clear need for improving pre-clinical research that will better identify those therapies suitable for translation to the clinic but also a need for new devices capable of recording continuous EEG in immature rodents. Here, we describe the technology behind and demonstrate the use of a novel miniature telemetry system, specifically engineered for use in immature rats or mice, which is also effective for use in adult animals.

Progressive, Seizure-Like, Spike-Wave Discharges Are Common in Both Injured and Uninjured Sprague-Dawley Rats: Implications for the Fluid Percussion Injury Model of Post-Traumatic Epilepsy.

Variable-duration oscillations and repetitive, high-voltage spikes have been recorded in the electrocorticogram (ECoG) of rats weeks and months after fluid percussion injury (FPI), a model of traumatic brain injury. These ECoG events, which have many similarities to spike-wave-discharges (SWDs) and absence seizures, have been proposed to represent nonconvulsive seizures characteristic of post-traumatic epilepsy (PTE). The present study quantified features of SWD episodes in rats at different time points after moderate to severe FPI, and compared them with age-matched control rats. Control and FPI-injured rats at 1 year of age displayed large-amplitude and frequent SWD events at frontal and parietal recording sites. At 3-6 months, SWDs were shorter in duration and less frequent; extremely brief SWDs (i.e., "larval") were detected as early as 1 month. The onset of the SWDs was nearly always synchronous across electrodes and of larger amplitude in frontal regions. A sensory stimulus, such as a click, immediately and consistently stopped the occurrence of the SWDs. SWDs were consistently accompanied by behavioral arrest. All features of SWDs in control and experimental (FPI) rats were indistinguishable. None of the FPI-treated rats developed nonconvulsive or convulsive seizures that could be distinguished electrographically or behaviorally from SWDs. Because SWDs have features similar to genetic absence seizures, these results challenge the hypothesis that SWDs after FPI reflect PTE.