Animal Models of Acquired Epilepsies and Status Epilepticus

Chapter 36

Giuliano G. Avanzini

David M. Treiman

Jerome Engel Jr.

Introduction

In animals, epilepsies with focal seizures can be faithfully reproduced by various experimental procedures that can induce an initial status epilepticus (SE) followed after a latent period by a chronic seizure recurrence. Thus, the initial SE and the subsequent chronic epilepsy can be considered the biphasic expression of a common pathogenetic process, which is set in motion by the experimental manipulation.

This chapter reviews models for SE and for epilepsies with focal seizures obtained using experimental epileptogenic procedures in normal animals. Some procedures are relevant to both focal epilepsies and SE, whereas others are specific for either one; therefore, SE and focal epilepsy models are dealt with in two separate sections. Models of epileptogenic dysplasia are also discussed.

In principle, animals presenting with spontaneous seizures can be profitably studied to gain information on species-specific susceptibility and pathophysiologic mechanisms, but, in practice, their interest is limited by the difficulties of obtaining homogeneous populations and the obvious differences in the ability to study domesticated or laboratory species versus those in the wild.

In most domesticated species, spontaneously occurring SE has never been observed, whereas reports of spontaneous localization-related epilepsies are mostly anecdotal and do not provide a basis for defining any suitable model for systematic experimental studies. An exception is the dog, in which prevalence of epilepsy approaches that of humans.⁶⁴ Because there is evidence for genetically determined susceptibility to epilepsy and/or epileptogenic brain pathologies, canine epilepsies are not dealt with here. None of the other animal models of genetically determined epilepsies are included in this account because they also are models for generalized rather than focal epilepsies.

Reports of experimentally induced seizures in animals date back to the seventeenth century, when Robert Boyle²⁴ observed the occurrence of seizures in sparrows, larks, cats, and mice exposed to low air pressure in a decompression chamber. At the cusp of the nineteenth and twentieth centuries, pioneering studies by Fritsch and Hitzig,⁸¹ Openchowski,¹⁶³ Baglioni and Magnini,¹⁰ and later Kaufmann¹⁰⁹^,¹¹⁰ revealed that other physical and chemical means were effective in inducing epileptic manifestations, including cortical electrical stimulation, freezing, and the topical application of strychnine. Other agents later shown to be effective as topical convulsants include metallic compounds, which were studied for many years by Lenore and Nicholas Kopeloff after their first report,¹¹⁴ metabolic antagonists, and convulsant drugs (see Prince¹⁷² for review). The application of intracellular recording to experimentally induced cortical foci provided an early insight into the cellular mechanisms underlying epileptogenesis,¹³⁴^,¹³⁵ and since then, a number of studies have considerably deepened our understanding of focal epileptogenesis in experimental animals and established a number of pathogenetic concepts that can now be reliably applied to human epilepsies.⁸^,¹⁷³

Experimental models provide a unique means of testing the efficacy of antiepileptic drugs and antiepileptogenic strategies, although the results may not always be generally applicable. Species-specific or mechanism-specific effects may, in fact, prevent their reproducibility in different species or in other types of experimentally induced or naturally occurring epilepsies.

Chemically Induced Focal Epilepsies

General Characteristics

Focal seizures are defined as those seizures whose symptomatology indicates the initial activation of a system of neurons limited to a part of one cerebral hemisphere.⁶⁹ Epilepsy is defined as a chronic condition of the brain characterized by an enduring propensity to generate epileptic seizures,⁷⁵ a definition that implies the tendency of recurrence of ictal manifestations over time. Thus, only animals with recurrent focal seizures can be considered as models for focal epilepsies.

In several instances, SE or closely spaced seizures are the initial event of a process leading to epileptic disorders presenting with recurrent seizures that occur spontaneously and do not require any further exogenously delivered precipitating stimulus. These characteristics correspond to those of human focal epilepsies, which are further classified as idiopathic (i.e., not preceded or caused by a neurologic disorder other than a possible hereditary predisposition), symptomatic (i.e., secondary to a known disorder of the central nervous system), or cryptogenic (i.e., secondary to a disorder whose course is hidden or occult). By definition, experimentally induced animal epilepsies fall into the category of symptomatic epilepsies because the causative factor (i.e., the experimental epileptogenic procedure) is always known.

The models reviewed in this section reproduce one important characteristic of human epilepsy, that is, its chronic course. They can be obtained by experimental manipulations that lead to a gradual development of persistent focal epileptogenic activities or by procedures capable of inducing acute epileptic manifestations that are able to set in motion a progressive epileptogenic process leading to a permanent epileptic condition. A typical example of the first possibility is topical application of alumina hydroxide gel, which induces progressive neuropathologic changes affecting cortical excitability, whereas the second possibility is best exemplified by the pilocarpine model,

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in which the development of a chronic epilepsy is mediated by pilocarpine-induced SE. The role of the acutely induced limbic SE or closely spaced seizures is also important although not exclusive for kainic acid, whereas afterdischarges are relevant to epileptogenesis induced by focal electrical stimulation. Whether the epileptic manifestations occurring in the early phase contribute to the development of tetanus toxin–induced chronic epileptogenesis is not clear.

Animal models of acute seizures and in vitro models are dealt with in other chapters.

Interictal and Ictal Clinical Phenomenology

Ictal clinical phenomena depend on the complex of anatomic structures involved in the generation of ictal discharges, referred to as the epileptogenic zone. In principle, the main types of seizures observed in human epilepsies should be faithfully reproduced in experimental animals by creating epileptogenic zones in the appropriate cortical areas. However, the evaluation of ictal phenomenology in animals is necessarily limited to external observations of animal behavior and thus provide only an indirect assessment of the subjective experience, which is an important aspect of seizures originating in cortical areas involved in sensation, emotion, and higher cognitive function.

The evaluation of experimentally induced complex partial seizures presenting with variable sequences of typical symptoms (including loss of contact) is also difficult, as is the comparison of animal and human seizures because of species-related differences in central nervous system organization. Of course, the ictal involvement of some human-specific functions such as language can never be modeled in experimental animals.

Besides these intrinsic difficulties, further problems in assessing the reproducibility of human seizures in experimental animals derive from the scanty descriptions of ictal phenomenology in some experimental studies. For instance, it is insufficient to describe a seizure as focal with elementary motor symptomatology without a qualitative (clonic vs. tonic, tonic vs. postural, positive vs. suppressive, etc.) and topographic (massive vs. segmental, stationary vs. migrating, etc.) analysis of its motor manifestations.

Neurologic, psychic, and neuropsychological persisting signs during the interictal period have been reported in patients suffering from focal symptomatic epilepsies. Interictal neurologic and behavioral studies in animals made epileptic by different experimental procedures have been performed only occasionally (see later discussion).

Interictal and Ictal Electroencephalographic Features

In human localization-related epilepsies, interictal focal “epileptiform” electroencephalographic (EEG) activities are often seen as spikes, sharp and slow waves, fast rhythms, and so on.⁵³ The transition from an interictal to an ictal discharge may be represented by repetitive interictal discharges, localized fast activity (often with a recruiting character), or other rhythmic discharges not necessarily colocalized with the interictal focus (if present).⁵³ Similar EEG changes can be detected in animal models of partial epilepsies, although the internal frequency of interictal and ictal discharges may not be the same as that seen in humans. This may depend on both species-related differences and technical factors (the type of electrodes, interelectrode distances, etc.). In particular, the current use of bone screws or intracranial wire as recording electrodes may exclude or reduce the filtering effect of bone on fast activities. To make recordings from animals comparable with those from humans, standardized EEG methodology and terminology should be developed.⁷⁴

Natural History

Although human focal symptomatic epilepsies depend on various progressive, stationary, or remitting brain pathologies, their course is to some extent independent of the underlying disorder. In particular, the epilepsies secondary to acute brain insults (such as traumatic, vascular, and infectious epilepsies) display a typical biphasic course that includes an early phase with single or repeated “reactive”⁶⁸ seizures or SE, which may then subside to give rise to chronic epilepsies after a more or less prolonged interval. Some of the experimental models reviewed here have contributed significantly to clarifying the mechanisms underlying such a biphasic course.

Questions that can be Addressed by Animal Studies

Experimental studies of epilepsies try to answer various questions concerning brain physiology, pharmacology, and pathophysiology that are all basically related to the aim of gaining a better understanding of the mechanisms underlying epileptogenesis.

The characterization of a cellular hallmark of epilepsy in the “penicillin” focus of cat neocortex, the paroxysmal depolarization shift (PDS),¹³⁴^,¹³⁵ was seen as a major breakthrough that could directly lead to the unraveling of the “basic mechanism of epilepsy.” However, it was soon realized that the analysis of this characteristic burst-discharge seen in individual neurons only partially explains the biologic basis of epileptogenesis. Moreover, it became clear that similar shifts could result from a variety of experimental manipulations differentially affecting the excitable properties of cortical neurons, thus suggesting that a unitary explanation of epileptogenesis was unrealistic.¹⁷²

Selection of a model for study depends on the specific question being asked. However, whether designed to replicate a specific component or an overall process, the ideal model is one that most closely approximates the phenomenon or process of interest. Thus, experimental models should closely approximate behavior, electroencephalographic characteristics, and pharmacologic responses of the type of human epileptic manifestation being studied.

In considering the questions to be addressed by a given experimental model, it must be borne in mind that the answers an investigator may obtain are valid only for that specific model unless otherwise demonstrated. Therefore, the relevance that an experimentally demonstrated epileptogenic mechanism may have in relation to human epilepsies depends on the strength of the evidence of its involvement in human epilepsies. Ultimately, therefore, parallel studies in patients are required.

The following is a discussion of some of the models that have been in use for many years. Others not discussed in detail that are being developed include models of traumatic brain injury, hypoxia-induced epileptogenic encephalopathy, stroke, febrile seizures, intraparenchymal bleeding, and cerebral infection.¹⁶⁸

Alumina Model

The epileptogenic effect of the topical application of alumina hydroxide gel to monkey neocortex was discovered by Kopeloff et al.¹¹⁴ in the context of a study aimed at investigating

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immunologic factors in epilepsy. Among the many other metallic compounds that were later shown to induce epileptic foci, alumina cream is the most suitable for inducing chronic focal epilepsy.

Methods

Autoclaved aluminum hydroxide 4% gel is injected in a few adjacent sites of the exposed neocortex using a small syringe needle.²⁴⁷ Epileptogenic foci are optimally induced in primates but less consistently in dogs, cats, and guinea pigs. There are no reports of spontaneous seizures induced by intracortical alumina in lower forms. Alternatively, the systemic⁷⁶ and intracisternal¹¹¹ administration of aluminum salt or aluminum metallic powder²⁹ has been used to induce encephalopathy with multifocal seizures in susceptible animals (rabbits, cats, and ferrets, but not rats).

FIGURE 1. Sleep recording from a Macaca mulatta injected in left pre- and postcentral gyrus with aluminum hydroxide. The electrodes 1–8 and 10–13 are skull-screws. The electrode 9 is placed in temporalis muscle, and electrodes 14 and 15 are implanted in the mastoid process. Note the spike activity on the left hemisphere. (From Lockard JS. A primate model of clinical epilepsy: mechanism of action through quantification of therapeutic effects. In: Lockard JS, Ward AA. Epilepsy: A Window to Brain Mechanisms. New York: Raven Press, 1980;11–49; with permission.)

FIGURE 2. Synaptic responses of two horseradish peroxidase–injected CA1 pyramidal neurons from control (A) and intracysternally AlCl₂-injected rabbits (B). Note in panel B the irregularities in dendritic size and the repetitive discharge superimposed on a prolonged depolarizing shift. (From Franceschetti S, Bugiani O, Panzica F, et al. Changes in excitability of CA1 pyramidal neurons in slices prepared from AlCl₃-treated rabbits. Epilepsy Res. 1990;6:39–48; with permission.)

Neuropathology

Dendritic distortion with decreased branching size irregularities and spine loss has been observed in pyramidal neurons within an alumina cortical focus.²⁵⁰ These changes are due to a toxic effect leading to neurofibrillary degeneration, which has been studied in aluminum encephalopathy induced by both systemic and intracisternal administration.²⁹^,¹¹¹ The further stages of toxic degeneration lead to neuronal loss with marked gliosis, which is particularly prominent in “mature” alumina cortical foci.¹⁰² A decrease in GABAergic nerve terminals at the sites of alumina foci has also been reported.¹⁷⁸ In most experiments, alumina has been topically applied to the sensorimotor cortex or the hippocampus; in both regions, pyramidal neurons have been found to be consistently affected after the intracisternal injection of aluminum powder.²⁹

Natural History and Clinical Phenomenology

Spontaneous clinical seizures appear between 2 weeks and 6 to 8 months after intracerebral injection of alumina cream, depending on the number of injections and the width of the involved area.²⁴⁷ Once established, the seizures spontaneously recur, probably throughout the life span of the animals; in monkeys, they have been observed for at least 7 years.²⁴⁷ With large foci, seizure frequency may increase to status epilepticus and require antiepileptic treatment to maintain a viable preparation. The seizure patterns depend on the location of the focus. According to Ward,²⁴⁷ in monkeys receiving alumina injections in the face and arm area of the somatomotor cortex, the seizures begin with contralateral facial or hand twitching, spread gradually to the entire contralateral side (mimicking a jacksonian march), and may eventually become generalized. During the seizures, muscular jerks occur at an increasing rate, which then fuse into a tonic contraction with superimposed strong generalized jerks, cyanosis, and sialorrhea. The tonic or tonic–clonic phase ceases abruptly, giving place to a postictal depression with hypotonia and unresponsiveness to external stimulation of variable duration. When a particularly intense epileptogenic focus has been obtained, focal motor status epilepticus can be observed, with continuous 1/10-second jerks of the contralateral face and hand, mimicking Kojeknikoff epilepsia partialis continua. Behavioral seizures reminiscent of human complex partial seizures of temporal origin can be observed after alumina injection in the anterior part of the temporal lobe and/or in the amygdala of monkeys and cats.⁸³^,²⁵¹ Soper et al.²⁰³ consistently obtained chronic temporal lobe epilepsy in monkeys only by means of bilateral alumina injections in the hippocampus; the clinical seizures consisted of head-turning, lip-smacking, mastication, and salivation.

Electroencephalogram

Serial recordings show the gradual development of interictal foci of slow and sharp waves, spikes, and delta activity in the region of the scalp corresponding to the underlying alumina focus (Fig. 1).⁹⁶ The transition to ictal discharge may be difficult to identify when interictal activity is sustained; otherwise, it is characterized by focal fast activity of increasing amplitude.²⁰² As the seizure progresses, spikes and sharp waves tend to recur rhythmically and spread to other regions. In animals with bilateral foci, ictal discharges invariably begin at the site of the older focus.²⁰²

Pharmacology

Antiepileptic drug efficacy on interictal EEG discharges or seizure frequency has been widely studied in correlation with pharmacokinetic parameters.¹²¹ Phenytoin (Dilantin), phenobarbital (Arco-Lase, Bellergal-S, Donnatal, Quadrinal Mudrane, Rexatol, Solfoton), primidone (Mysoline), and carbamazepine (Atretol, Tegretol) showed a significant effect on both seizure frequency and interictal EEG discharge. The efficacy was correlated with drug plasma levels. A less clear level/effect correlation was found for sodium valproate (which improved when the correlation was evaluated on an hour-by-hour basis).

Pathophysiology

Neurons belonging to alumina foci have a high probability of burst-firing, closely associated with surface epileptiform waves.¹⁷³ These putative epilepsy-related changes are not maintained in in vitro slices prepared from monkey alumina foci, which have been found to have physiologic properties that are not significantly different from those of control tissue.¹⁸⁶ On the contrary, Franceschetti et al.⁷⁷ observed significant changes in the excitable properties of hippocampal pyramidal neurons in slices prepared from intracisternally AlCl₂-injected rabbit. These changes were accounted for by a decrease in the efficacy of Ca²⁺-dependent K⁺ conductances and GABAergic transmission and by electrotonic shortening due to dendritic debranching (Fig. 2). They were clearly detectable in the very early phases of the aluminum-induced encephalopathy, even before the manifestation of neurofibrillary degeneration, cell loss, and gliosis. On the other hand, the hypothesized epileptogenic role of the gliotic scar (which is believed to impair K⁺ regulation and thus lead to K⁺ accumulation in the interstitial space) is controversial¹⁰² and could not be directly demonstrated by Heinemann and Dietzel,¹⁰⁰ who found that the spatial buffer capacity of gliotic tissue for K⁺ was not severely impaired in cat alumina cream foci.

FIGURE 3. Schematic representation of temporal evolution of behavioral and electroencephalographic changes induced by an intraperitoneal injection of pilocarpine (380 mg/kg) in rats. (Redrawn from Cavalheiro EA, Leite JP, Bortolotto ZA, et al. Long-term effects of pilocarpine in rats: structural damage of the brain triggers kindling and spontaneous recurrent seizures. Epilepsia. 1991;32:778–782; with permission.)

FIGURE 4. Skull-screw electrode electroencephalographic recording from frontoparietal region of a rat at different time intervals after intraperitoneal injection of pilocarpine (400 mg/kg). Continuous spike activity appeared 50 minutes after injection and lasted up to 8 hours. Isolated spikes persisted up to 24 hours, then disappeared during the latent period to show up again 12 days after injection. (From Liu Z, Nagao T, Desjardins GC, et al. Quantitative evaluation of neuronal loss in the dorsal hippocampus in rats with long-term pilocarpine seizures. Epilepsy Res. 1994;17:237–247; with permission.)

Pilocarpine Model

The first evidence that rats with brain damage induced by the cholinergic agent pilocarpine develop spontaneous recurrent seizures after a silent period of 14 to 15 days was provided by Turski et al.,²²⁶ who were investigating the acute effects of pilocarpine treatment. As was stressed later by the same authors,³⁴ this natural history is reminiscent of that of human temporal lobe epilepsy, which often begins with prolonged status epilepticus in infancy and develops with recurrent seizures in later life.⁸⁹

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Methods

Thirty minutes after subcutaneous pretreatment with scopolamine, 1 mg/kg (to minimize peripheral cholinergic effects), a single high dose of pilocarpine (300–380 mg/kg) is injected intraperitoneally in rats and mice.³³ The acute pilocarpine-induced status epilepticus must be continuously monitored by behavioral observation and EEG recording because of the high mortality rate (30%),³³ which can be partially prevented by repeated intraperitoneal injections of diazepam 10 mg/kg plus phenobarbital 30 mg/kg¹¹⁷ at 30 minutes 1, 2, and 6 hours after the beginning of the status epilepticus. Further behavioral-EEG monitoring is indicated during the silent period to detect the onset of chronic recurrent seizures.

Neuropathology

At the end of the acute phase, widespread bilateral morphologic changes appear that involve the hippocampus, amygdala, thalamus, pyriform and entorhinal cortices, neocortex, and substantia nigra.²²⁶ These consist of massive swelling of dendrites and neuronal cell bodies with relative sparing of axons and astroglial cell swelling.⁴⁰^,¹⁵⁹ During chronic stages, there is invariably cell loss in all of the structures mentioned, including the hilus of the dentate gyrus. Clear-cut evidence of supragranular sprouting of the mossy fibers, beginning 4 days after the episode of status epilepticus and reaching a plateau after 100 days, can be found using Timm staining.¹⁵² The severity of the chronic neuropathologic changes is proportional to the duration of the status episode¹¹⁷ but not to the number of chronic seizures.¹²⁰

Natural History and Clinical Phenomenology

The temporal evaluation of pilocarpine-induced behavioral and EEG changes is schematically illustrated in FIGURE 3. Immediately after the pilocarpine injection, the animal is hypoactive; subsequently, there is the appearance of facial automatisms, including chewing and eye blinking, followed by head bobbing and motor limbic seizures (forelimb clonus, salivation, and rearing on hind limbs). Generalized convulsions and limbic status usually occur 40 to 80 minutes after the injection, depending on the injected dose.³³^,¹²⁰^,²²⁶ After a silent period of 4 to 44 days, the spontaneous seizures that characterize the chronic period appear and recur with a variable frequency of 2 to 15 per month, with no evidence of spontaneous remission for at least 6 months.³³

Electroencephalogram

During the acute phase, the electroencephalographic changes evolve from early surface low-voltage fast activity correlated with a theta hippocampal rhythm to high-voltage fast activity associated with spiking in the hippocampus and, finally, surface spiking activity that correlates with head bobbing.¹²⁰ Limbic seizures are associated with high-frequency, high-voltage spike discharges that become continuous during limbic status. The EEG progressively normalizes at the end of the acute phase, although spontaneous spike discharges reappear after the silent period. The ictal electrographic discharges of the chronic period consistently originate in the hippocampus and subsequently spread to the cortical electrodes.³³ FIGURE 4 summarizes the evolution of EEG changes.

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Pharmacology

Diazepam (Dizac, Valium) and scopolamine, 10 mg/kg, provide effective protection from acute seizures and limbic status.³³ Phenobarbitone 40 mg/kg/d, phenytoin 100 mg/kg/d, and carbamazepine 120 mg/kg/d have been found to be effective against the spontaneous seizures occurring during the chronic phase. Sodium valproate has been found to be effective only at very high doses (600 mg/kg/d); ethosuximide (Zarontin) 400 mg/kg/d is totally ineffective.¹¹⁶

Pathophysiology

In vitro experiments¹⁵^,¹⁶ have shown that the epileptogenic effect of cholinergic agents depends on the facilitation of burst discharges in hippocampal pyramidal neurons by means of a block of the K⁺ transmembrane current I_M. This mechanism explains the massive activation of hippocampal neurons during pilocarpine-induced status epilepticus, which leads to cell death, axonal sprouting, and a synaptic reorganization of hippocampal circuitry that results in permanent epileptogenic changes.³³^,³⁵^,¹²⁰ The neuronal damage is not attributed to a direct toxic effect of pilocarpine, but to a seizure-related excitotoxic effect involving glutamate receptors and Ca²⁺ influx.

Tetanus Toxin

The epileptogenic properties of intracerebral tetanus toxin have been known since the end of the nineteenth century¹⁸⁰ but have been exploited in experimental studies of epileptogenesis only more recently after studies published by Brooks and Asanuma²⁸ and Carrea and Lanari³² in cat and dog neocortex and by Mellanby et al.¹⁵² in rat hippocampus and neocortex.¹⁰⁶

Methods

Tetanus toxin is a protein with a molecular weight of 150 kDa released by Clostridium tetani and is now available in a purified form. It is quite stable and can be kept at 4°C for months. Tetanus toxin is extremely toxic and must be handled only by effectively immunized people wearing protective clothing (gloves and a mask). Tetanus toxin is active in all of the mammalian species tested¹⁸⁰ when very small amounts are injected in appropriate regions. The dose is usually expressed in toxicologic units that correspond to different quantities, depending on the degree of purification: 5 to 20 mouse half lethal dose (LD₅₀) has been used in rat hippocampus,¹⁰⁶ 2 to 30 minimum lethal dose in rat neocortex,²⁴ 10 to 10³ mouse lethal dose in cat neocortex,²⁵ and 28 to 83 LD₅₀ in dog neocortex.³² It is important to bear in mind that tetanus toxin is very efficiently transported along axons.

Neuropathology

Early tetanus toxin–induced histologic changes with minimum effective doses injected into the hippocampus are very mild and consist of a loss of neurons in the CA1 region. Much higher doses cause neuronal loss at the injection site. It is suspected that late changes in hippocampal circuitry occur, but these have not yet been systematically investigated.¹⁰⁶

Natural History and Clinical Phenomenology

Animals that are injected with low doses in either the neocortex or hippocampus develop seizures 3 to 7 days after the injection.²⁷^,¹⁰⁶ Shorter intervals are reported with higher doses.²⁸ In rats receiving hippocampal injections of the toxin, there is a reduction in the number of seizures after some weeks of intense epileptic activity; the seizures eventually disappear but may relapse in the longer term.¹⁰⁶ In neocortex-injected rats²⁷ and cats,¹²⁵ tetanus toxin–induced foci can be permanent. Hippocampal injections induce limbic seizures that start with a behavioral arrest, followed by vibrissal and facial twitching and forelimb myoclonus that may eventually develop into a clonic–tonic seizure involving the hindlimbs. Injections in rat cortex are followed by early focal motor seizures in the jaws and/or contralateral limbs, tending to progress to generalized convulsive seizures after 16 hours.²⁷

Electroencephalogram

Interictal biphasic sharp waves are recorded from the injected hemisphere 3 to 5 days after the injection and, a few days later, from the contralateral homotopic sites with synchronous or asynchronous expression (Fig. 5).²⁷ Ictal discharges of 3- to 20-Hz spikes and spikes-and-waves may be limited to one hemisphere or may spread rapidly to the contralateral side. The injected hemisphere leads in the early phases, but 5 to 13 days after the injection, the ictal discharge can be initiated in the contralateral hemi-sphere.²⁷

FIGURE 5. Skull-screw recording in a rat 22 days after injection of tetanus toxin. Interictal spikes occur in both injected (IPSI) and contralateral hemisphere (CONTRA) (A) either synchronously (B) or asynchronously (C). (From Brener K, Amitai Y, Jefferys JGR, et al. Chronic epileptic foci in neocortex: in vivo and in vitro effects of tetanus toxin. Eur J Neurosci. 1991;3:47–54; with permission.)

FIGURE 6. A, B: Simultaneous field (upper trace) and intracellular (lower trace) recordings made from A and B sites indicated in the schematic diagram (C) of a slice prepared from a rat injected with tetanus toxin in parietal neocortex 8 days before. Although the two recording points are equidistant from the injection track, site A generates epileptiform response, whereas site B appears relatively normal. (From Brener K, Amitai Y, Jefferys JGR, et al. Chronic epileptic foci in neocortex: in vivo and in vitro effects of tetanus toxin. Eur J Neurosci. 1991;3:47–54; with permission.)

Pharmacology

Drugs effective in human focal epilepsies, such as carbamaze-pine, have also been found to be quite effective in the tetanus toxin model but not the N-methyl-D-aspartate (NMDA)–receptor antagonist 2-amino-5-phosphonopentanoic acid (APV).¹⁰⁶

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Pathophysiology

The tetanus toxin–dependent epileptogenic process has various phases. At the very beginning, the toxin induces epileptiform discharges in the injection site as a result of a block of γ-aminobutyric acid (GABA) release,⁴⁶ leading to the impairment of the local inhibitory circuits established by GABA interneurons in both the hippocampus and the cerebral cortex. The epileptiform neuronal aggregate is contained in hippocampal and neocortical slices prepared from the injected areas, and this has allowed a detailed electrophysiologic analysis.⁶⁷^,¹⁰⁵ In a second phase, the toxin is transported through the axons to even remote sites, where it can move transsynaptically inside local GABAergic neurons. A transport velocity of as much as 200 mm/d has been estimated in peripheral nerves,¹⁸⁵ and, therefore, transport-related patches of epileptiform activity can be generated in a few hours through corticocortical and callosal connections (Fig. 6). This mechanism might account for the rapid generation of mirror foci.²⁷ In a third chronic phase, GABA-mediated inhibition recovers, due probably to an increased RNA expression of glutamic acid dehydrogenase (GAD). The epileptic activity persisting in this chronic phase is attributed to seizure-induced plastic changes in the hippocampal or neocortical circuits, leading to the functional disconnection of GABAergic neurons or the sprouting of new excitatory axons.¹⁰⁶

Kainic Acid

Kainic acid is a highly potent glutamate agonist that is obtained from the seaweed Digenea simplex and used as an ascaricide. Kainic acid was found to be excitatory when applied iontophoretically to rat cortex¹⁹⁵ and to induce seizures when injected intracranially¹⁹^,²⁰ or systemically.²¹^,⁴⁷^,¹²⁶ Olney et al.¹⁶⁰ highlighted its neurotoxic properties and reported its toxicity to be particularly prominent on the hippocampus, even when systemically injected. Kainic acid–induced cell injury is attributed to an excitotoxic mechanism triggered by the activation of excitatory amino acid (EAA) receptors. The same mechanism is also responsible for the hippocampal seizures occurring in animals injected, either intravenously¹²⁶ or in the hippocampus,³⁴ with doses less than those required to produce direct cell damage.

Methods

Most of the studies with kainic acid have been done in rats. Systemic doses of 4 mg/kg¹²⁶ and local injections of 0.4 mg⁸ are effective in inducing persistent seizures. Similar effects have been obtained by means of intracisternal ventricular injections in mice,²²⁷ for which a 50% convulsive dose of 0.3 nmol has been calculated. A single intra-amygdaloid injection of 1 μg in cats has been found to be effective in inducing focal status epilepticus.¹²⁶^,²¹⁴

Neuropathology

The systemic administration of kainic acid induces a variable pattern of cortical and subcortical damage involving the pyriform and entorhinal cortices, the hippocampus, the lateral septum, and several thalamic and amygdaloid nuclei.¹⁵⁹ In the hippocampus, degenerating neurons are found in the dentate hilus, CA1, and CA3.⁷⁴ A similar pattern of hippocampal degeneration has been found after local hippocampal injection during the acute phase,¹³³ with evidence of active cellular phagocytosis of the necrotic zone in the next active phase. The Timm²¹⁷ sulfide silver method for heavy metals, subsequently modified,⁵⁴ shows an initial sprouting of zinc-containing mossy fibers into the inner molecular layer. This sprouting progressively increases during the latent phase, reaching its maximum at the beginning of the chronic phase, when a thick band of Timm-positive fibers is permanently found in the inner molecular layer (Fig. 7). With unilateral injection, the changes are largely unilateral, resembling human hippocampal sclerosis.

FIGURE 7. Interictal recording from a rat bilaterally injected with kainic acid in the hippocampus 122 (A) and 130 (B) days before. Fr, single wire electrodes placed into the frontal cortex; HC1, 2, double-twisted wire electrodes inserted in the hippocampus; L, left; R, right. The independent epileptiform transient (arrows) are seen mostly in the left hippocampus, which showed evidence of mossy fiber sprouting, as shown by Fig. 8. (From Mathern GW, Cifuentes F, Leite JP, et al. Hippocampal EEG excitability and chronic spontaneous seizures are associated with aberrant synaptic reorganization in the rat intrahippocampal kainate model. Electroencephalogr Clin Neurophysiol. 1993;87:326–339; with permission.)

FIGURE 8. In the same animal whose electrographic recording is shown in Fig. 7, Timm-stained sections show a zinc-positive band (arrows) in the left (A) but not in the right (B) inner molecular layer (IML). Note that both hippocampi (C, D) show evidence of neuron loss in the stratum granulosum (SG) and hilus (H). (From Mathern GW, Cifuentes F, Leite JP, et al. Hippocampal EEG excitability and chronic spontaneous seizures are associated with aberrant synaptic reorganization in the rat intrahippocampal kainate model. Electroencephalogr Clin Neurophysiol. 1993;87:326–339; with permission.)

Natural History and Clinical Phenomenology

The evolution of the epileptic phenomenology and electrophysiologic correlates induced by low doses of kainic acid injected

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bilaterally in rat hippocampus has been longitudinally studied by Mathern et al.¹³³ and correlated with concomitant histologic changes. Similar asymmetric changes occur with unilateral hippocampal injections, which more accurately reproduce the predominantly unilateral condition of human mesial temporal lobe epilepsy.²⁵ On the basis of the clinical and electrophysiologic course, four phases have been recognized as sequentially occurring after kainic acid injection at the following approximate time intervals in days: acute (0–10), active (10–30), latent (30–90), and chronic. The earliest seizures characterizing the acute phase occur within 1 hour after hippocampal injection and consist of behavioral arrest; the association of sniffing and facial myoclonus is ascribed to the frontal spread of the discharge. After several hours, the seizures increase in duration and may eventually evolve into a partial status characterized by simple staring. The active phase is characterized by short-duration seizures (<1 minute) with generalized motor clonus and very rapid behavioral recovery. The seizures disappear during the latent period but relapse during the chronic phase, with characteristics similar to those of the acute stage: A motionless stare that may occasionally progress to a more complex phenomenology, including facial automatisms, forelimb clonus, and generalized clonic–tonic seizures. The chronic seizures do not tend to subside, but rather increase in frequency and duration and may generalize over time.¹³³

Electroencephalogram

Mathern et al.¹³³ and Bragin et al.²⁵ monitored the hippocampal activity recorded by stereotaxically implanted deep electrodes and the frontal activity derived from intracortical wires.

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In the acute phase, discharges of spikes and multispikes in the hippocampus are associated with behavioral arrest and spread of the discharges toward the frontal cortex with sniffing and facial myoclonus. Postictal diffuse slowing of background activity is regularly observed and is sometimes associated with periodic lateralized epileptiform discharges (PLEDs). The active phase is characterized by interictal fast activity and polyspikes limited to the hippocampus, the transition to the ictal discharge being marked by a pronounced spike activity that rapidly generalizes. During the latent phase, there is a reduction in the interictal spikes, and background activity is almost normalized; in the late part of the latent phase, however, spikes and sharp waves are increasingly recorded, and there is a marked tendency to asynchronous expression in both hippocampi. The asynchrony of interictal spikes and sharp waves becomes more and more evident during the chronic phase, when ictal discharges limited to one hippocampus reappear in association with motionless staring episodes. A bilateral spread of discharges is observed during generalized seizures (Fig. 8). In addition to the clinical seizures, more frequent electrographic seizures can also be recorded, usually during sleep.²⁵ Whereas the EEG patterns during clinical seizures resemble the low-voltage fast ictal onset of human mesial temporal lobe epilepsy, the electrographic events resemble the hypersynchronous onsets usually associated with auras in patients.

More recently, microelectrode recordings from unilateral intrahippocampal kainic acid–treated rats have led to the discovery of 150- to 500-Hz “Fast Ripple” oscillations identical to those recorded from human epileptic hippocampus.²⁵ Fast Ripples can also be recorded shortly after kainate injection and predict which rats will develop spontaneous seizures.²⁶ The significance of this novel finding is discussed in Chapter 13.

Pharmacology

Drug responsiveness of the kainate model is essentially the same as that for the pilocarpine model.

Pathophysiology

FIGURE 9. Diagrammatic representation of dentate network circuitry and the hypothesized translamellar mossy cell innervation of inhibitory basket cells. BC, basket cells; G, granule cells. (From Sloviter RS. The functional organization of the hippocampal dentate gyrus and its relevance to the pathogenesis of temporal lobe epilepsy. Ann Neurol. 1994;35:640–654; with permission.)

FIGURE 10. Different susceptibility to electrically induced afterdischarge in the cortex of monkey. Stimulation thresholds in volts (V) in different areas are indicated by the three coded patterns. (Originally from French et al.⁸⁰ From Ajmone Marsan C. Focal electrical stimulation. In: Purpura DP, Penry JK, Tower D, et al., eds. Experimental Models of Epilepsy. A Manual for the Laboratory Worker. New York: Raven Press; 1972:147–172; with permission.)

The early seizures occurring during the acute phase are attributed to the direct effect of the excitatory amino acid receptor agonist kainic acid, which is especially prominent in the hippocampus due to the particularly high concentration of kainic acid receptors in this region. The permanent changes in hippocampal excitability underlying chronic seizures is ascribed to neuronal damage of the vulnerable hilar mossy cells due to kainic acid excitotoxicity or the ensuing sprouting of mossy fibers. The first effect is thought to lead to a selective denervation of the neurons that mediate granule cell inhibition (Fig. 9), thus increasing their excitability.¹⁹⁹ On the other hand, the newly formed recurrent mossy fiber collaterals could re-excite the dentate granule cells through newly formed synapses.²¹⁵ The role of these two putative epileptogenic mechanisms will be further discussed later.

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Electrical-Stimulation–Induced Focal Epilepsies

Self-Sustained Status Epilepticus

The possibility of eliciting epileptic manifestations by means of the repeated electrical stimulation of discrete regions of the central nervous system has been known since the nineteenth century following the experimental studies of Fritsch and Hitzig,⁸¹ Ferrier,⁷³ and Luciani.¹²⁸ The analysis of the electrographic correlates of stimulation-induced seizures showed that the associated discharge may outlast the end of the stimulation train.³ Although self-sustained, this electrically evoked afterdischarge is still a stimulation-dependent acute epileptic phenomenon. Later, Alonso-De Florida and Delgado⁵ discovered that appropriate paradigms of repeated stimulation may induce permanent changes in excitability and lead to the recurrence of spontaneous seizures. Electrogenic models of SE are discussed later in the SE section.

Methods

The optimal stimuli for the production of afterdischarges are 4- to 6-second trains of repeated (25–60 Hz) 2.5- to 8.0-mA, 2- to 5-msec diphasic square pulses.⁴ After a first effective train, a second afterdischarge can be obtained only after a delay of at least 15 seconds. Using more prolonged and/or intense stimulating trains, Lothman et al.¹²⁶ and Handforth and Ackermann⁹³ overcame this refractory period and reduced the intertrain interval to 0.5 second by using a protocol currently defined as “continuous” hippocampal stimulation. Focal electrical stimulation has been found to be effective in all studied species, including humans. The greatest susceptibility is in the hippocampus and amygdala. All of the neocortical areas can generate afterdischarges, the most and least prone regions being the precentral motor area and the temporal gyri (Fig. 10).⁴

Neuropathology

By definition, afterdischarges are evoked in normal tissue, which should not be directly damaged by the stimulation procedure, provided appropriate parameters are used. However, Sloviter and Damiano²⁰¹ and Sloviter¹⁹⁸ found that indirect damage to hilar mossy cells can be induced by the stimulation of a perforant path capable of inducing repetitive discharges in their target granule cells, which innervate the hilar mossy cells themselves. In addition, there is evidence that, during repeated stimulation, use-dependent structural rearrangements of the involved neuronal network occur (e.g., mossy fiber sprouting) that are similar to those observed in other chronic models of hippocampal epilepsy.¹⁷

FIGURE 11. Electrocorticographic recording of a typical, relatively well-localized afterdischarge in an epileptic patient with surgical exposure of the occipital region, as indicated in brain outline with electrode position. The stimulation was delivered in the region indicated by the bent arrow. (From Ajmone Marsan C. Focal electrical stimulation. In: Purpura DP, Penry JK, Tower D, et al., eds. Experimental Models of Epilepsy. A Manual for the Laboratory Worker. New York: Raven Press; 1972:147–172; with permission.)

Natural History and Clinical Phenomenology

The stimulation-induced afterdischarge can be purely electrical or associated with clinical manifestations appropriate to the functional properties of the involved areas. Continuous hippocampal¹²⁴ or amygdala stimulation⁹³ leads to self-sustained epileptic states defined as “immobile, exploratory, minor convulsive, and clonic” according to a behavioral-electrographic hierarchy of severity.⁹⁴ After a latent period, as

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with pilocarpine and kainic acid, animals exhibit spontaneous seizures.

Electroencephalogram

The electrographic activity characterizing typical afterdischarges becomes evident 1 to 2 seconds after the end of the stimulating train. It begins as 15- to 30-Hz low-voltage oscillations, which progressively increase in amplitude and decrease in frequency, leading to large voltage oscillations (Fig. 11). The episode lasts from 10 to 90 seconds and ends abruptly, giving way to a postictal depression lasting from 2 to 15 seconds.⁴

Pharmacology

The effect of antiepileptic drugs can be tested on the threshold or duration of the afterdischarge. Carbamazepine, phenobarbital, and diazepam, but not sodium valproate, ethosuximide, or phenytoin, have been found to be effective on the maximal dentate activation obtained by means of contralateral region stimulation.²⁰⁹

Pathophysiology

The self-sustained character of the afterdischarge has always been attributed to a failure in inhibitory mechanisms,⁴ a view that was subsequently supported by the perforant path stimulation experiments of Sloviter and Damiano²⁰¹ and Sloviter.¹⁹⁸ The hyperexcitable state, leading to the facilitation of afterdischarges, electrogenic status epilepticus,⁹³^,¹²⁶ and, eventually, the epileptogenic process, is the result of complex changes involving cell excitability and circuitry rearrangements that are discussed for other models (see also next section on kindling).

Kindling

Although physiologic psychologists carrying out electrical stimulation studies in rats had noted for some time that a few eventually developed seizures, this was considered a nuisance because it disrupted their research paradigms. Goddard, however, recognized this to be an interesting phenomenon in itself and developed the concept of kindling.⁹⁰^,⁹¹ Electrical kindling refers to the process of brief subthreshold brain stimulation that, when repeated, gradually results in ictal behaviors. Stimulation-induced seizures then persist indefinitely after the kindling process is discontinued. Electrical kindling, therefore, was an ideal mechanism for bringing the process of epileptogenesis under laboratory control, and it rapidly became the most commonly used animal model for studying basic mechanisms of epilepsy. Kindling as usually practiced, however, is not a model for chronic epilepsy because seizures do not occur spontaneously but need to be provoked by electrical stimulation. Although animals kindled for prolonged periods of time after maximal seizures occur eventually do develop spontaneous seizures,¹⁶⁵ this model of chronic epilepsy is rarely used because of the time required and the difficulty in maintaining viable animals. Consequently, although kindling remains a useful model for the study of epileptic phenomena, particularly with respect to the limbic system,⁴⁹^,⁵⁰^,²³⁰^,²³¹^,²³²^,²³³^,²³⁴ it has now been largely replaced by the excitotoxic and stimulation-induced self-sustained status epilepticus models that more faithfully

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reproduce human mesial temporal lobe epilepsy with hippocampal sclerosis.

“Chemical kindling” refers to the process of repeated treatments with chemoconvulsants such as metrazol or carbacol, which, like electrical kindling, eventually results in epileptic seizures in response to the chemical treatment.⁸⁷ Although chemical kindling presumably produces many of the same epileptogenic plastic changes induced by electrical kindling, the former models have been less well studied.

Methods

The classical kindling model involves electrical stimulation of the amygdala, although stimulation of hippocampus, piriform cortex, and other limbic structures results in the same epileptogenic phenomenology.¹⁴⁵ The time course of kindling differs depending on the site of stimulation, stimulus parameters, interstimulation intervals, and species. Kindling can also be achieved with stimulation of nonlimbic brain areas such as neocortex, thalamus, and caudate, but this is associated with a different behavioral evolution and less consistent progression than limbic kindling. Many other structures, such as brainstem and cerebellum, however, do not support kindling.

Typically, limbic kindling begins with a stimulus intensity that produces afterdischarge without behavioral effects. With repeated stimulation, the afterdischarge duration increases and ictal behaviors begin with arrest, ipsilateral eye blinking, and chewing (stage 1), followed over time by head bobbing (stage 2), then forelimb clonus (stage 3), rearing (stage 4), and falling with generalized tonic–clonic movements (stage 5).¹⁷⁶ It is also possible to begin kindling with subthreshold stimulation intensity (below that necessary to induce afterdischarges), in which case repeated stimulation eventually produces afterdischarges, which then reiterate the electrographic and behavioral epileptogenic progression.¹⁷⁵ The usual stimulation parameters are 60 to 100 Hz delivered for 1 second, although slower frequencies and different durations are also effective. The time required for kindling to stage 5 depends on the area stimulated and the interval between stimulations. A typical paradigm involves stimulation for 1 second once a day, which requires approximately 2 weeks for amygdala kindling and 4 weeks for hippocampal kindling.

Neuropathology

Although limbic kindling is associated with mossy fiber sprouting, resembling this pathologic finding in human hippocampal sclerosis and chronic animal models of this disorder,²¹⁰ cell death, particularly in the dentate hilus, is minimal. Neurogenesis and astrocytic proliferations have also been reported, but there are no neuropathologic findings specific to kindling.²¹¹

Natural History and Clinical Phenomenology

Kindling is not a single phenomenon, but a series of phenomena that underlie a progressive epileptogenic process. With subthreshold kindling, for instance, there are changes that occur only at the tips of the stimulating electrodes that eventually result in the appearance of afterdischarge. As stimulation continues, or with classical kindling, the increase in duration of afterdischarge also involves epileptogenic plastic changes at the electrode tips, but most likely in surrounding tissue as well. The appearance of clinical signs reflects propagation of epileptiform afterdischarge to distant structures, requiring transsynaptic alterations. This process evolves to recruit primary motor cortex and other brain areas, eventually engaging brainstem systems responsible for tonic–clonic seizures. The progressive transsynaptic recruitment is not merely a reflection of more intense afterdischarge at the electrode tips, but true synaptic plasticity, because stage 5 seizures, once achieved, almost always occur with any stimulation at the kindling site capable of inducing afterdischarge. Many widespread areas of the brain, therefore, develop an enduring epileptogenic potential with focal kindling. If kindling proceeds for months, spontaneous seizures eventually appear, and these typically do not originate at the site of kindling stimulation but from their efferent projection areas.¹⁶⁵

Other enduring distant effects of kindling also occur. For instance, “transfer” refers to the fact that whereas it may take 2 weeks of daily stimulation to reach a stage 5 seizure with unilateral amygdala kindling, stage 5 seizures can then be provoked with only a few stimulations of the contralateral amygdale.¹⁴⁴ On the other hand, contralateral kindling has an interhemispheric seizure-suppressing effect, in that it might take several additional days of stimulation to reestablish a stage 5 seizure with stimulation of the primary site.¹⁴⁴ Ipsilateral seizure facilitating and seizure-suppressing effects can also be demonstrated with electrical kindling. Other evidence of kindling-related seizure suppression also exists during the postictal period. There is a refractory period after kindled seizures during which time another stimulation will not generate a seizure. Animals that have been subjected to frequent seizures during a relatively short period of time develop postictal seizure refractoriness that can last weeks.¹⁵⁵ Investigations into this phenomenon could provide insights into natural homeostatic mechanisms that protect against recurrent seizures.

Neocortical, thalamic, and caudate kindling produces an entirely different epileptogenic progression. Rats do not go through the initial stages associated with limbic behaviors but eventually exhibit focal motor and generalized tonic–clonic seizures. Whereas limbic kindling progresses consistently, once a stage is achieved, rarely does the animal backslide to a lower-stage seizure on the subsequent stimulation; backsliding is a common occurrence with neocortical, thalamic, and caudate kindling. Kindling-like phenomena can also be produced by stimulation of other subcortical structures such as nucleus accumbans, but these may not be epileptic behaviors.²⁰⁷ Electrical kindling has been carried out in a variety of species, from frogs to primates. In general, the higher the species is on the phylogenetic scale, the longer it takes to kindle. Genetic factors also appear to influence kindling rate; for instance, whereas it can take hundreds of stimulations to complete amygdala kindling in a rhesus monkey, it takes only about 70 in the Papio papio with genetic photosensitive epilepsy.²³⁴

The relationship of the various mechanisms of kindling to those of human epileptogenesis and seizure generation are unclear, but it is likely that kindling mechanisms (a) are involved in permitting subclinical electrical discharges eventually to manifest as behavioral seizures, (b) contribute to the progression of an epileptogenic abnormality, resulting in more frequent, more severe, and more pharmacoresistant seizures, (c) recruit distant structures in some forms of epilepsy, leading to secondary epileptogenesis (the development of new epileptogenic regions), and (d) induce enduring neuronal dysfunction, which could contribute to the appearance of interictal behavioral disturbances.

Electroencephalogram

As with human epilepsy, limbic kindling is associated with interictal EEG spikes that appear not only in the kindled limbic area, but also independently in other ipsilateral and contralateral limbic structures.¹⁰⁸ These interictal spikes appear early in the course of kindling and persist after stage 5 seizures occur. Interictal spikes are more common immediately following seizures, and there is some evidence that at least some types of postictal interictal spikes reflect mechanisms of seizure suppression.⁷⁰ Propagation of stimulation-induced ictal EEG discharges progressively recruits structures responsible for

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behavioral manifestations, but there is no strict correlation between the duration of afterdischarge and the severity of the behavioral seizure. It is interesting that the very high frequency (200–600 Hz) oscillations associated with interictal spikes, termed Fast Ripples (FR), which in chronic animal models of mesial temporal lobe epilepsy and in the human disorder are believed to indicate tissue capable of generating spontaneous seizures, do not occur in kindled animals, perhaps because these models do not generate spontaneous seizures.²⁵

Pharmacology

A number of studies over the years have been carried out to test the effectiveness of standard antiepileptic drugs as well as other compounds on kindling. It is of particular interest that drugs that are effective in preventing the development of kindling are not necessarily effective in preventing stimulation-induced seizures, whereas not all drugs effective in preventing stimulation-induced seizures disrupt the kindling process. This model, therefore, is capable of dissecting out antiepileptogenic versus anti-ictogenic properties of pharmacologic agents. Although kindling provides an effective model for screening potential antiepileptic compounds for antiepileptogenic and anti-ictogenic effects, it is much more labor intensive and expensive than the standard screening models of subcutaneous metrazol and maximal electroshock, and therefore it is rarely used. Recently, however, levetiracetam, which failed these two standard screening procedures, was tested and found to be effective against kindled seizures.¹²³ As a result, levetiracetam is now available as a highly effective agent against partial onset seizures.

An interesting phenomenon of contingent tolerance has been described whereby pretreatment with an effective anticonvulsant prior to, but not after, daily amygdala-kindled seizures results in a loss of efficacy.¹⁶⁹ Similarly, contingent inefficacy refers to the fact that pretreatment of an effective antiepileptic drug prior to daily stimulation during kindling results in the loss of effectiveness after kindling is achieved.¹⁷⁰ The mechanisms of contingent tolerance and inefficacy are unknown; however, they provide opportunities for studying neuronal processes underlying the development of pharmacoresistance and suggest that certain dosing schedules in patients may be counterproductive.

Chemical kindling can be achieved with a wide variety of chemical agents administered either intracerebrally or systemically.⁸⁷ Behavioral and electrographic features for most chemoconvulsants are similar to those of electrical limbic kindling, and once kindling is completed, the ability of subthreshold doses of chemoconvulsant to induce a stage 5 seizure is persistent. Reports of neuropathologic changes are variable. Crossovers among various chemoconvulsants as kindling agents, as well as between chemical and electrical kindling, suggest common mechanisms, although chemical kindling appears to involve much larger areas of the brain than localized electrical stimulation.

Pathophysiology

Although many investigations have been carried out to elucidate pathophysiologic mechanisms underlying kindling, no single underlying epileptogenic neuronal process has been revealed. This is undoubtedly due to the fact that kindling is not a unitary phenomenon but involves a great variety of alterations in cellular excitability, synaptic plasticity, neuronal loss, neurogenesis, glial proliferation, and synaptic reorganization, not only at the point of electrical stimulation for electrical kindling, but also transsynaptically in local and distant structures ultimately responsible for behavioral ictal manifestations, as well as related phenomena such as transference, the development of homeostatic seizure-protective mechanisms, and the generation of spontaneous seizures if kindling is continued. Specific fundamental neuronal mechanisms by which such changes occur are covered in more detail in other chapters in this section, and there is no indication that there are any unique to either electrical or chemical kindling.

Epileptogenic Dysplasia

The word “dysplasia” defines a tissue that has failed to develop perfectly during embryonic or fetal life as a result of genetic determinants⁵⁹^,⁶⁶ or pathogenic factors impairing the ordered sequence of maturational events, and it is known that dysplasias giving rise to macroscopic structural malformations that are clearly visible in neuroimaging studies are often associated with severe epilepsies.¹³^,⁶⁰^,¹⁶³

Genetically determined developmental brain abnormalities have been found in spontaneous mutant rodents such as dreher¹⁸⁹ and reeler mice,³⁶^,⁷¹ and local⁶ and bilateral subcortical neuronal heterotopias¹¹⁵ have also been detected in some strains of genetically epilepsy-prone rodents. Animal models of cerebral dysplasia can also be obtained by means of embryonic exposure to physical (i.e., X-rays, freezing) or chemical teratogenic agents (e.g., ethanol, methylazoxymethanol) capable of killing neuroblasts and/or disarranging neuron–glial relationships. Finally, targeted genetic manipulations can lead to a selective impairment of cortical development that may be associated with spontaneous seizures, as has been demonstrated by Acampora et al.² in a mouse model lacking the Otx1 gene.

However, it should be noted that most of the genetic and experimentally induced animal models of dysplasia do not present with obvious spontaneous epileptic seizures, which can only be induced by means of proconvulsant manipulations that are not effective (or significantly less effective) in control animals.

In this section, we concentrate on the models obtained by means of methylazoxymethanol (MAM), freezing, or the deletion of the orthodenticle gene (Otx1 1^-/-) and on TISH (telencephalic internal structural heterotopia) rats with laminar heterotopia.

Methylazoxymethanol Model

MAM is an alkylating agent extracted from the King Sago Palm (Cycas revoluta) that was found to induce cerebral dysplasia by Singh¹⁹⁷ and Johnston and Coyle.¹⁰⁷ It kills neuroblasts in mitotic phase, thus leading to a narrower and disarranged neocortical mantle. It also induces structural abnormalities of the radial glia that lead to the obstructed and misdirected migration of neocortical precursors that is responsible for subcortical heterotopias.

Methods

A single intraperitoneal injection of MAM 15 mg/kg administered to pregnant rats on embryonic day 15 (E15) regularly induces microcephaly, hippocampal heterotopias, and altered neocortical lamination in the offspring.¹¹^,¹²^,³⁸^,⁴⁵^,⁵⁷^,⁸⁶^,¹⁰⁷^,¹⁹⁶ In addition, the double transplacental administration of MAM (15 mg/kg in the morning and, 12 hours later, the same dose injected intraperitoneally on E15) induces large heterotopic aggregates surrounding the ventricular floor (in continuity with hippocampal heterotopia) and large irregular neuronal clusters in the sensorimotor cortical area.¹³^,⁴²

Neuropathology

Morphologic analyses of MAM-treated rat neocortex show disrupted layering with evidence of a thick subpial band of heterotopic neurons in layer I. As observed in human

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cortical dysplasia (see earlier discussion), the lateral somatosensory and auditory cortices contain clusters of enlarged cortical neurons positive for specific anti-SMI311 and anti-MAP2 antibody neuronal markers surrounded by a dense terminal network that is immunoreactive to antibodies against calbindin, a calcium-binding protein expressed by cortical interneurons.

The subcortical heterotopic nodules consist of normally differentiated cortical neurons randomly oriented within the nodule core with their major axis parallel to the edge of the nodule in the marginal zone. The nodule borders are characterized by a dense, GABA-positive network. In some animals, the periventricular nodules were in continuity with the overlying cortex, which was also characterized by a similar nodular structure or extended into the hippocampus and disrupted CA1 and CA2 layering.

Natural History and Clinical Phenomenology

The animals born of MAM-treated mothers show microcephaly, but postnatal developmental milestones (including eye opening, fur growth, sucking, ambulating, and grooming) are reached at the same age as controls. Spontaneous seizures and EEG changes indicating epileptic activities have never been reported, but there is evidence of increased susceptibility to epileptogenic agents or procedures such as fluorothyl,¹¹ hyperthermia,⁸⁶ kainic acid,⁸⁴ and kindling.⁸⁵

Electroencephalogram

EEG monitoring reveals an increased delta frequency that positively correlates with the severity of the cortical disarrangement but no evidence of epileptiform activity.⁸⁵ Intracellular recordings from neurons located 350 to 550 μm below the surface of brain slices of dysplastic sensorimotor cortex show a slightly higher percentage of intrinsically bursting (IB) neurons than that found in layer V of untreated rat sensorimotor cortex.¹⁸² Baraban and Schwartzkroin¹¹ reported a similar finding in the CA2 hippocampal region, with a definitely higher percentage of IB neurons in MAM-treated rats than in controls.

Aberrant firing patterns of repetitive bursts of action potentials, which gradually increase in duration and eventually merge in a long-lasting discharge, can be induced by both depolarizing current pulses¹⁸² and an increase in extracellular K⁺ concentrations.¹¹^,¹²

Pharmacology

There are no data concerning the effect of drugs on the seizures (because there are no spontaneous seizures) or susceptibility to epileptogenic agents.

Pathophysiology

The neurotoxic effect of MAM on dividing neuroblasts, which is due to the methylation of nucleic acids, affects the migrating neuronal progenitors and the radial glia along which they migrate. The results are cortical disarrangement and the formation of subcortical nodules consisting of heterotopic neurons that were originally committed to the neocortex but do not find a normally permissive environment for correct migration to their final destination.

FIGURE 12. Pyramidal neurons recorded in dysplastic neocortex (A) and in periventricular heterotopia (PVH) (B) of MAM-treated rats. In panel A, the microphotograph of a thionin-counterstained slice shows abnormal neocortical layering and irregular neuronal clusters. The pyramidal neuron drawn on the left responded to both synaptic and direct activation with short burst firing. Panel B shows intracellular recordings from two PVH neurons. The left neuron shows normal responses to both synaptic and direct stimulations (uppermost traces), and the right neuron responds with a short burst to the synaptic activation and with an excessive bursting behavior to the direct stimulation with injection of a depolarizing pulse. (Modified from Sancini G, Franceschetti S, Battaglia G, et al. Dysplastic neocortex and subcortical heterotopias in methylazoxymethanol-treated rats: an intracelular study of identified pyramidal neurons. Neurosci Lett. 1998;246:181–185.)

Incoming inputs can affect the neurons in dysplastic neocortical areas, as well as those located in periventricular and parahippocampal heterotopias (Fig. 12). There is also evidence of reciprocal synaptic connections establishing aberrant cortico-subcortical circuitry³⁸^,⁴²^,⁷⁸^,¹⁸² that is excessively susceptible to epileptogenic agents. The factors responsible for this greater susceptibility are the larger proportion of IB neurons and changes in intrinsic K⁺ current-related properties,¹¹^,¹⁸² and the possible influence of clustered GABAergic hyperinnervation should also be taken into account. The association of a smaller number of GABA-reactive neurons with a dense network of parvalbumin (PV)-immunoreactive terminals suggests that axonal sprouting of the PV-positive subpopulation of GABAergic neurons may take place during the development of MAM-treated rats. The expected functional consequence is a pacing effect due to the occurrence of highly synchronized inhibitory postsynaptic potentials in a large population of pyramidal neurons, which are then simultaneously released from inhibition and thus become prone to synchronous discharges.

Freezing-Induced Layered Microgyria in Rats

The word polymicrogyria indicates an excessive number of small and prominent convolutions separated by shallow and enlarged sulci, which give the cortical surface a lumpy appearance. Two types of polymicrogyria have been recognized: (a) unlayered polymicrogyria due to exogenous insults occurring between gestational weeks 13 and 18 or to genetic factors and (b) four-layered polymicrogyria due to a perfusion failure between gestational weeks 20 and 24. The perfusion failure causes laminar necrosis of the intermediate layers with a consequent late migration disorder and the postmigratory overturning of cortical organization. Human polymicrogyria is quite often associated with epileptic manifestations.

The application of a freeze probe to the skull of newborn rats generates a focal region of necrosis with a loss of deep layers whose basic structure is quite similar to that observed in four-layered human polymicrogyria.⁶²^,⁶³ The results of stimulation and lesion experiments suggest that aberrant development in the zone adjacent to the microgyrus underlies epileptogenesis.¹⁷⁴

Methods

A freeze lesion is induced in rat pups 3 to 30 hours after birth by means of a freeze probe with a diameter of 1 mm to a few millimeters that is cooled to between -40°C and -70°C and applied to the skull for 3 to 8 seconds.¹⁰⁴^,¹²⁸ Neocortical slices for electrophysiologic recordings containing the lesion and the surrounding neocortex have been prepared from rats at postnatal ages ranging from 9 to 118 days.¹⁰⁴^,¹²⁸

Neuropathology

A freeze lesion consists of an infolded cortex that creates a more or less deepening microsulcus and a surrounding mic-rogyrus that typically contains four distinguishable layers: The first and second correspond to the molecular and II/III layers of the adjacent neocortex, the third layer contains some glia and a few neurons, and the fourth layer partially corresponds to the VI layer of the adjacent neocortex (VIb).¹⁰⁴ The parvalbumin immunoreactive neurons normally concentrated in layers IV and Vb are completely absent in the microgyrus during the first 13 postnatal days but normalize after postnatal day 21.¹⁷⁹

Natural History and Clinical Phenomenology

Scantlebury et al.¹⁸⁵ demonstrated increased susceptibility to hyperthermia-induced seizures in freeze-lesioned immature rats. Generalized, convulsive hyperthermic seizures were evoked by significantly lower temperatures in lesioned pups than in controls.

Electroencephalogram

No in vivo demonstration of spontaneous epileptic EEG abnormalities has been provided in rats with microgyrus, but the

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ictal EEG correlates of hyperthermic seizures induced in freeze-lesioned rats show a topographic correlation between spike activity and the microgyrus.¹⁸⁵ In slice experiments, interictal-like epileptiform activity can be evoked from freeze-lesioned rats after postnatal day 12 and have been found until postnatal day 118, with a decreased incidence being observed after postnatal day 40 in rats lesioned on postnatal day 0 but not in those lesioned on postnatal day 1. Jacobs et al.¹⁰⁴ obtained evidence of decreased synaptic inhibition leading to local multiphasic epileptiform activity in neocortical areas closely surrounding the microgyri, and Luhmann and Raabe,¹²⁹ Luhmann et al.,¹³⁰ and Prince and Jacobs¹⁷⁵ reported similar results.

Pharmacology

The multiphasic discharges are reversibly blocked by NMDA-receptor antagonists in slices from both mature and immature freeze-lesioned rats, which suggest that they are attributable to disinhibited excitatory postsynaptic potentials mediated by the excitatory glutamate or aspartate amino acids.

Pathophysiology

Freezing interferes with cortical development through mechanisms similar to those responsible for human polymicrogyria because neuronal migration in rodents continues after birth until postnatal days 2 to 3.¹⁸⁷ It has been suggested that the basic mechanism supporting the increased seizure susceptibility is an imbalance between synaptically driven excitation and inhibition.¹³⁰ A number of observations suggest that aberrant synaptic connectivity develops in the rat cortex surrounding the microgyrus and causes a focal epileptogenic zone whose capacity to generate epileptiform activities does not depend on connections with the malformation itself because the evoked epileptiform activities in the paramicrogyral cortex remain unaltered if this zone is separated from the microgyrus by means of a transcortical cut in adjacent area. It has been hypothesized

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that afferents originating from cortical and extracortical sites lose their targets in the region of the malformation but make appropriate laminar contacts in the cortex adjacent to the malformation and thus create an overabundance of excitatory inputs to this cortical zone. The resulting imbalance between the excitatory and inhibitory synaptic systems may be further aggravated by the loss of the parvalbumin-immunoreactive GABAergic neurons found in early developmental stages.

Otx1^-/- Model

Mutations leading to the replication of body segments (a process called homeosis) were first observed in insects, and it has since been established that they affect a special category of genes that contain a DNA motif (or homeobox) coding for a 61–amino acid domain called the homeodomain. The proteins containing homeodomains act as activators or repressors of downstream target genes, thus controlling the development of body segments. The role of the Otx1 homeobox gene in mouse corticogenesis was investigated by Acampora et al.,² who confirmed that Otx1 is required for the development of the entire dorsal telencephalic cortex, has a more pronounced effect in the temporal and perirhinal areas, and may affect the mechanisms specifying neuronal identity. Moreover, Otx1-null mice exhibit epileptic seizures.

Methods

Otx1-knockout mice were generated by replacing the Otx1 gene with the Escherichia coli lacZ gene.² Heterozygous Otx1^+/- mice are healthy and fertile, and their cross-breeding generated the homozygous mice Otx1^-/-, which are smaller and show epileptic manifestations; 30% die during the first postnatal month. Electrographic recordings from Otx1^-/- mice have been made using screw electrodes over the occipital cortex and deep electrodes inserted in the hippocampus.² Intracellular recordings have been obtained from pyramidal neurons in neocortical slices prepared from Otx1^-/- and control mice.¹⁸³

Neuropathology

Gross examination of the brain reveals that the adult Otx1^-/- mouse brain is about 25% lighter than that of heterozygous and wild-type mice. The overall thickness of the neocortex is reduced in the homozygous mutant, especially in the temporal and perirhinal areas, where the reduction may be as much as 40%. The sulcus rhinalis is displaced dorsally and the hippocampus is shrunken with a divarication of the dentate gyrus, whereas the volume of the colliculi is increased, and there is an additional protuberance between the superior and inferior colliculi. The cerebellar abnormalities include the presence of an additional lobule and a duplication of the rostral end of the declivus.² Morphologic analyses of Otx1^-/- neocortex have shown that, despite their overall reduction in thickness, all of the cortical layers are recognizable; however, the cells appear to be more tightly packed in the mutant IV layer, whereas their density is clearly reduced in the V layer and particularly in the VA sublayer.¹⁸³ Outside of the central nervous system, Otx1^-/- mice have no ciliary process in the eye, no lachrymal or Harderian glands, and no lateral semicircular duct in the inner ear.

Natural History and Clinical Phenomenology

Epileptic symptoms occur in all Otx1^-/- mice but have never been observed in heterozygous Otx1^+/- mice. The seizures are of two main types: (a) short, 30-second episodes of head bobbing and teeth chattering that may subside or evolve into (b) generalized seizures characterized by upper extremity clonus, rearing and falling, and convulsions that last for approximately 60 seconds with complete recovery or, occasionally, evolve into status epilepticus and exitus. The frequency of the seizures tends to be less in older Otx^-/- mice, but they never disappear. In addition, Otx1^-/- mice show nonepileptic turning behavior.

FIGURE 13. Morphologic (A, B), histologic (C, D), and electroencephalographic (EEG) (E) comparison of wild-type (wt) and Otx^-/- mice. Note the reduction of temporal (Te) perirhinal (PR) cortices and hippocampus (Hi) in Otx^-/- mice. Representative EEG recordings from neocortex (Cx) and Hi during a convulsive seizure of Otx^-/- mice is shown in panel E, where the EEG recording from a wild-type (wt) animal is shown for comparison. (From Acampora D, Barone P, Simeone A. Otx genes in corticogenesis and brain development. Cereb Cortex. 1999;9:533–542; with permission.) (See color insert.)

FIGURE 14. Recordings from an Otx1^+/± pyramidal neuron. A1: Responses to two different stimulus intensities under control conditions. Note the postinhibitory depolarizing event reaching the threshold for action potential generation (arrow) that is abolished by the N-methyl-D-aspartate (NMDA) antagonist D-2-amino-5-phosphonovalerate (AP5) (A2). A3: The addition of both AP5 and the α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) led to an almost complete block of excitatory synaptic activity that was reversed after 20 minutes of washout (A4). B: Magnification of the foremost segment of the same traces as in panels A1 to A3, showing more clearly the effect of AP5 and CNQX (the stimulus is marked by open triangles). C: Characterization of the tested neuron by means of low-amplitude depolarizing and hyperpolarizing current pulse injections. (From Sancini G, Franceschetti S, Lavazza T, et al. Potentially epileptogenic dysfunction of cortical NMDA- and GABA-mediated neurotransmission in Otx1^-/- mice. Eur J Neurosci. 2001;14:1065–1074; with permission.)

Electroencephalogram

The “minor” episodes are associated with high-voltage spikes in the hippocampus and some fast activity in the neocortex, thus suggesting that the epileptogenic discharges have a localized origin; the convulsive seizures correlate with high-voltage synchronized activity involving both the hippocampus and neocortex (Figs. 13 and 14). Electrophysiologic experiments on neocortical slices have revealed some important differences in synaptic activities, which are characterized by pronounced multisynaptic excitatory postsynaptic potentials often leading to late action potential generation and strong GABA_A- and GABA_B-mediated inhibitory postsynaptic potentials that have a pacing effect on pyramidal firing.⁹ Both late excitatory postsynaptic potentials and postinhibitory excitation are selectively suppressed by NMDA-receptor antagonists but not by α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) antagonists.¹⁸³

Pharmacology

The effect of antiepileptic drugs on the seizures occurring in Otx1^-/- animals has never been systematically tested.

Pathophysiology

The fact that the layer V neurons so far reconstructed from the mutants seem to be smaller than those of control animals suggests that neurons originally committed to the upper layers may not migrate properly but be stopped in the fifth layer, where they induce the reorganization of GABAergic circuitry.

On the basis of the foregoing data, it can be concluded that the abnormalities in Otx1^-/- mice neocortex due to the selective loss of large projecting neurons lead to a complex rearrangement of local circuitry that is characterized by an excessive NMDA polysynaptic excitation that is counteracted by GABA-mediated inhibition in only a limited range of stimulus intensity. Prominent postsynaptic inhibitory potentials may also act as a further proepileptogenic event by synchronizing abnormal excitatory potentials.

Mutant Rats With Telencephalic Internal Structural Heterotopia

TISH rats are mutants presenting subcortical band heterotopia. They were identified by means of postmortem anatomic analyses during the course of unrelated experiments on Sprague Dawley rats by Lee et al.¹¹⁶: Spontaneous recurrent partial seizures with variable secondary generalization were present in some TISH rats.

Methods

A breeding colony was established by identifying the living relatives of deceased TISH individuals and screening them by means of magnetic resonance imaging.¹¹⁶ Crosses between two affected animals produced 100% TISH progeny, whereas crosses between one affected and one unaffected animal produced no affected offspring, but their intercrossing produced 29% of affected offspring. The overall incidence of affected males and females was respectively 47% and 53%, which is

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consistent with an autosomal-recessive pattern of inheritance of a single-gene defect.

Neuropathology

The brain of these mutants has a large region of heterotopic gray matter located bilaterally beneath the neocortex and extending from the frontal to the occipital lobe (Fig. 15). It is particularly prominent in the frontal and parietal cortices but is usually absent from the temporal cortex. The normotopic neocortex overlying the heterotopia is usually organized into six laminae (but thinner than in normal animals), whereas the heterotopia lacks precise lamination. Pyramidal neurons are present in both normo- and heterotopic cortices, but the apical dendrites in the latter are not consistently radially oriented and may even be inverted, and the dendrites near the edge of the heterotopic region often bend to follow the contour of the band. The heterotopia also contains nonpyramidal neurons (fusiform and stellate cells) lacking a regular tectonic organization. The normotopic cortex shows a normal laminar organization of the different cell types, but there are significantly fewer parvalbumin-positive interneurons (43% of those found in control neocortex), and the intensity of the parvalbumin plexus is likewise reduced in the neuropil of layer V.²⁴ Other laminar structures, such as the hippocampus and cerebellum, show a normal laminar pattern and do not contain heterotopic neurons.

Natural History and Clinical Phenomenology

Seizure activity was observed in several of the animals in the colony established in Charlottesville, Virginia, by Lee et al.¹¹⁶ Chen et al.³⁷ made prolonged video-electrographic recordings lasting 4 to 6 months in a sample of animals and found that seizure frequency ranged from 1.5 to 15.1 events per week. The seizure phenomenology consists of twitching of the face and paws and turning to one side, occasionally followed by falling and convulsive activity; seizure duration is between 1 and 2 minutes. No seizures have been observed in animals younger than postnatal day 30.

Electroencephalogram

Electrographic recordings made using depth electrodes positioned bilaterally on both normotopic and heterotopic areas show that ictal spiking activity arises almost simultaneously in the normotopic and heteroptopic areas of one hemisphere and spreads rapidly to the homologous contralateral regions. Field potential recordings from in vitro slices bathed with epileptogenic agents show synchronous spiking activity in normo- and heterotopic areas. Making a cut between the two bands significantly decreases the threshold for epileptiform spiking in the

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normotopic area but significantly increases the threshold in the heterotopia. Patch-clamp recordings from identified pyramidal neurons show a consistent reduction in inhibitory postsynaptic current (IPSC) amplitude and frequency in both normo- and heterotopic cortical areas.²²⁴

Pharmacology

No data are available concerning the drug sensitivity of the seizures observed in TISH rats.

Pathophysiology

Injecting 5-bromo-2′-deoxyuridine (BrdU) to label cells in the S-phase in pregnant dams has demonstrated that the heterotopic neurons are generated during the normal period of cortical neurogenesis and that the inside-out pattern of neurogenesis is intact in the normotopic neocortex of TISH mutants.¹¹⁵ Alterations in cell proliferation and migration are considered to be responsible for the TISH malformation.¹¹⁵

In relation to the mechanisms underlying epileptogenesis, electrophysiologic studies have shown that both normo- and heterotopic areas are involved in seizure activities, which are initiated by the normotopic cortex (see earlier discussion). Moreover, there is evidence that normotopic neurons have an excitatory synaptic influence on heterotopic neurons, which, conversely, have a net inhibitory effect on normotopic cortical cells. The results of Trotter et al.²²⁴ discussed earlier provide evidence of a reduced synaptic inhibition of pyramidal neurons that may account for an epileptogenic increase in excitability.

Status Epilepticus

General Characteristics

A variety of chemical convulsants have been administered systemically to induce various forms of experimental status epilepticus (SE). Chemical convulsants used to induce experimental SE do so by either increasing neuronal excitation or decreasing neuronal inhibition. Experimental models based on systemic administration of chemical convulsants have the advantage of simplicity—SE can be induced simply by parenteral administration of the convulsant agent. The disadvantage of such agents is their continuing presence once SE has been induced. Results may be confounded by the continuing presence of the inducing agent or by potential drug interactions between the inducing agent and an experimental therapeutic agent. Electrogenic models have the advantage that the status-inducing stimulus is eliminated once the stimulation has stopped, so that

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subsequent pathophysiologic changes or consequences of status epilepticus can be construed as being due to the seizure activity without the possible confounding effect of the initial stimulus. However, electrogenic models tend to be labor intensive, and thus do not lend themselves to large-scale studies.

FIGURE 15. Top: Three-dimensional reconstruction of the telencephalic internal structural heterotopia (TISH) shown in red at the cut surface of the brain and in pink where it is viewed through the overlying cortex. Bottom: Electroencephalographic (EEG) recordings of a convulsive seizure in a TISH rat. The four lines show continuous EEG recording from a single electrode positioned in the normotopic neocortex (arrow indicates seizure onset). Seizure activity can be observed as changes in the frequency and amplitude of the EEG. (Modified from Lee KS, Schottler F, Collins JL, et al. A genetic animal model of human neocortical heterotopia associated with seizures. J Neurosci. 1997;17:6236–6242). (See color insert.)

FIGURE 16. Examples of the five electroencephalographic (EEG) stages observed in human patients during generalized convulsive status epilepticus and in all animal models of experimental status epilepticus in which EEG changes over time have been systematically studied. The time required for initial appearance of epileptiform activity and for progression through the five stages varies with the model, as do details of the morphology of the ictal discharges. Nonetheless, the fundamental progression from discrete electrographic seizures (stage I), through the waxing and waning pattern (stage II), the continuous pattern (stage III), continuous ictal discharges punctuated by brief periods of relative flattening of the EEG (stage IV), to periodic epileptiform discharges on a relatively flat background (stage V) is seen in all models. See Table 1 for technical details. Co/Homo, cobalt/homocysteine; Li/Pilo, LiCl/pilocarpine.

Ictal Phenomenology

Most epileptic seizures last only from a few seconds to a few minutes. This is because seizure-terminating mechanisms operate during an isolated single seizure. A refractory period follows during which it is difficult to elicit a subsequent seizure. However, under some circumstances, the mechanisms responsible for seizure termination and the postictal refractory period fail, so that epileptic seizures recur before there has been complete recovery from the neurochemical and pathophysiologic consequences of the preceding seizure or persist beyond the usual, quite short, duration of individual seizures. This, in pathophysiologic terms, is the operational definition of SE,²¹⁸^,²¹⁹ whereas in clinical terms SE can be defined as “a condition in which epileptic seizures recur before there has been complete recovery from the consequences of the preceding seizure.”⁴⁸^,⁷⁵

Natural History

Status epilepticus is a surprisingly common⁵⁸^,⁹⁸ and potentially life-threatening medical emergency. There is increasing recognition that status epilepticus is a dynamic condition, with an evolution of clinical phenomenology, EEG changes, response to treatment, histopathologic changes, and behavioral consequences if it is untreated or inadequately treated.²¹⁸ Nonetheless, much remains to be learned about this disorder. The mechanisms of the transition from a single seizure to SE are not known. Our understanding of the pathophysiologic changes that cause SE to be progressively resistant to treatment is incomplete. We have only begun to understand the metabolic and pathologic consequences of sustained seizure activity; the time when permanent neuronal damage occurs during the course of SE is still not well worked out. Treatment of clinical SE is not always successful, and there remains a need for the development of more effective and less toxic drugs for this purpose. The potential role for neuroprotective agents in the clinical management of SE is unresolved, as is the cause of the differential susceptibility of young and adult brains to SE-induced neuronal damage.

Electroencephalogram

One of the major advances in the use of animal models of status epilepticus has been the recognition of a predictable sequence of electrographic changes that is common to patients experiencing generalized convulsive status epilepticus²²¹ and all experimental models of SE that have been carefully studied, including lithium/pilocarpine,²²¹^,²³⁶ kainic acid in adult²²¹ and juvenile¹⁵³ rats, cobalt-homocysteine,²²¹^,²³⁵ high-dose pilocarpine in adult¹¹¹ and juvenile²²² rats, three different electrogenic models,⁹³^,⁹⁵^,¹²⁶ soman in rats¹¹⁵^,¹⁴⁰ and rhesus monkeys (McDonough, personal communication), and in an extended hippocampal slice model.¹⁷⁸ FIGURE 16 illustrates the sequence in a number of different models. Initially discrete electrographic seizures are seen, separated by interictal generalized slowing (stage I; Table 1). However, if the episode of SE is untreated or undertreated, the discrete seizures begin to merge together to produce a waxing and waning of amplitude, frequency, and distribution of the ictal discharges (stage II). Subsequently, the ictal discharges become continuous (stage III). This stage is usually prolonged, but eventually the continuous ictal discharges begin to be punctuated by periods of relative flattening (stage IV), which lengthen as the rhythmic ictal discharges shorten,

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until finally the record exhibits only periodic epileptiform discharges on a relatively flat background (stage V). The fundamental sequence of patterns is always the same, although different models may exhibit differences in morphology within a specific stage (e.g., triphasic-like spike-wave patterns vs. rapid spiking in stage III, variable frequencies and morphologies of periodic epileptiform discharge [PEDs] in stage V), and different models may progress through the five patterns at different rates, just as has been observed in human generalized convulsive status epilepticus (GCSE) depending on the etiology of the episode of SE.

Table 1 Technical Details for the Five Electroencephalographic (EEG) Stages Illustrated in Figure 16

Model/EEG stage	Stage I	Stage II	Stage III	Stage IV	Stage V
Human²²¹	39-yr-old male	64-yr-old male	68-yr-old male	68-yr-old male	64-yr-old male
Kainic acid (KA)²²¹	26 m after KA	75 m after KA	103 m after KA	148 m after KA	5 hr 46 m after KA
Cobalt/homocysteine (HCT)²²¹	30 m after HCT	37 m after HCT	48 m after HCT	75 m after HCT	2 hr 12 m after HCT
LiCl/Pilocarpine²²¹	21 m after pilocarpine	24 m after pilocarpine	28 m after pilocarpine	109 m after pilocarpine	2 hr 19 m after pilocarpine
Soman¹¹⁵	Onset 3.1 ± 0.5 m after soman	Onset 5.1 ± 0.7 m after soman	Onset 19.3 ± 3.3 m after soman	Onset 33.9 ± 5.6 m after soman	Onset 231.4 ± 9.5 m after soman
Kainate in rat pups¹⁵⁴	P35 rat; mean onset 15 m (P15)/21 m (P35) after KA	P15 rat; mean onset 35 m (P15 and P35) after KA	P35 rat; mean onset 126 m (P15)/50 m (P35) after KA	P15 rat; mean onset 374 m (P15)/9 hr (P35) after kA	P35 rat; mean onset 10.25 hr (P15)/14.4 hr (P35) after KA
Pilocarpine in rat pups²²²	Onset 11.79 ± 11.85 m after pilocarpine	Onset 22.78 ± 22.11 m after pilocarpine	Onset 34.90 ± 13.15 m after pilocarpine	Onset 61.64 ± 21.76 m after pilocarpine	Onset 160.67 m after pilocarpine

Questions That Can Be Addressed by Animal Studies of Status Epilepticus

Experimental models have the potential of answering many of the questions posed earlier. Specifically, experimental models can be used to test drugs for the management of status epilepticus before they are tested in human clinical trials. Other questions that can be addressed using status epilepticus models include the reasons for other dynamic changes during status epilepticus, such as (a) the EEG changes discussed previously, (b) the progressive loss of motor activity as overt convulsive status epilepticus progresses to subtle and eventually nonconvulsive status, and (c) the progressively severe consequences of prolonged status epilepticus, including learning and memory deficits, neuronal damage at histology, and the development of chronic epilepsy. Models are now being used to understand the mechanisms that underlie the increasing refractoriness to treatment seen in human and experimental status epilepticus. Details of these studies are discussed in relation to the specific models described in what follows and in the chapter on generalized convulsive status epilepticus.

A number of experimental models of SE have been developed, including models in intact animals based on systemic and focal administration of chemical convulsants and focal and generalized electrical stimulation of the brain. More recently, some investigators have begun to study recurrent or sustained seizure activity in isolated neuronal circuits using slice preparations. The most useful and commonly used models of status epilepticus are discussed in what follows. Many of these models are also used for the induction of chronic epilepsy and are also discussed in the first section of this chapter. The discussion of the following models is focused on their use to further understand phenomenology and mechanisms of status epilepticus.

Bicuculline Model

Bicuculline is a highly potent alkaloid inhibitor of GABA-mediated neuronal inhibition⁵¹^,⁵² that has been used to induce experimental status epilepticus, initially by Meldrum and Horton in baboons.¹⁵⁰

Methods

Bicuculline must be dissolved in a weak acid (0.1 N HCl) and then titrated with a weak base (0.1 N NaOH) to pH 5.6.⁵⁶ Bicuculline can be administered intraperitoneally (2–4 mg/kg for developing rats, 6–8 mg/kg for juveniles or adults)²²⁸ or intravenously (2 mg/kg).²⁵³

Neuropathology

Meldrum and Brierley¹⁴⁹ described SE-induced ischemic-like neuronal damage in neocortex, cerebellum, and hippocampus in baboons. Similar but less severe changes were also seen in neocortex, thalamus, and hippocampus in paralyzed and mechanically ventilated baboons,¹⁵¹ thus demonstrating that SE-induced neuronal damage is primarily due to ongoing seizure activity rather than to motor convulsions.

Natural History and Clinical Phenomenology

Meldrum and Horton¹⁵⁰ induced severe, generalized status epilepticus lasting up to 5 hours in adolescent baboons by injecting 0.4 to 1.4 mg/kg bicuculline intravenously. Seizures began with generalized myoclonic jerks that evolved within seconds to generalized flexor spasms. In rat pups, myoclonic seizures evolve to clonic and then clonic–tonic seizures. After day 18, spike-wave discharges are seen, associated with motionless “freezing” of behavior.²⁵³

Electroencephalogram

A sequence of EEG changes, similar to those described by Treiman et al.²²¹ in generalized convulsive status epilepticus, can be observed in one figure in Meldrum and Horton’s report, although the progressive nature of the EEG changes is not characterized in detail.

Pharmacology

Bicuculline-induced seizures respond readily to antiepileptic drugs, especially the benzodiazepines,⁵⁵ but the bicuculline model has not been used systematically to study potential anti-SE drugs. Peterson et al.¹⁶⁵ induced SE in rats pretreated with LiCl by focal injection of bicuculline methiodide into the deep prepiriform cortex. This procedure resulted in a model of SE that could be stopped with 5 mg/kg of diazepam, in contrast to the lithium-pilocarpine model.

Pathophysiology

Bicuculline induces seizures by competitive antagonism at the GABA_A receptor. More recently, blockade of K⁺ channels and prolongation of Ca²⁺ action potentials has also been suggested as a possible mechanism of seizure induction by bicuculline.¹⁹⁰

Cobalt-Homocysteine Model

Walton and Treiman²³⁵ developed a model of secondarily generalized convulsive SE that closely approximates human GCSE in the natural history and characteristics of the induced seizures, the EEG changes, and the response to antistatus drugs. The model was specifically designed to test new agents for the treatment of GCSE, and a number of antiepileptic drugs have been studied in the model (see later discussion).

Methods

An epileptic lesion is created over the left motor cortex in adult male Sprague Dawley rats by placing 25 mg of powdered cobalt onto the dura when epidural screw electrodes are implanted. It is important to disrupt the dura and to pack the powdered cobalt into the screw well to reliably induce status epilepticus with the administration of the chemical convulsant, homocysteine thiolactone. When the lesion becomes electrographically active with brief focal ictal discharges and the rat is exhibiting intermittent focal motor seizures, usually about 7 days after surgery, SE is induced by intraperitoneal administration of 5.5 mmol/kg D,L-homocysteine thiolactone.

Neuropathology

The necrotic cobalt lesion can been seen on gross inspection of the intact brain, and a profound area of encephalomalacia is

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observable on histologic sections. However, the pathologic consequences of cobalt-homocysteine–induced SE have not been reported.

Table 2 A comparison of the serum concentration produced by the median effective dose (ED₅₀) for control of generalized tonic–clonic seizures (GTCS) in the cobalt/homocysteine secondarily generalized convulsive status epilepticus model with serum concentrations of the same drugs that have been reported to be clinically effective

Drug	Serum concentration produced by ED₅₀ vs. GTCS	Serum concentration reported to be clinically effective
Phenytoin	26.2 mg/mL	23.8 μg/mL
Diazepam	168 ng/mL	30–80 ng/mL 30–200 ng/mL
Phenobarbital	12.8 mg/mL	8.4 μg/mL 18.3 μg/mL
Lorazepam	196 ng/mL	30–160 ng/mL 70–330 ng/mL
Modified from Walton and Treiman.²⁴⁴

Natural History and Clinical Phenomenology

Status epilepticus in this model resembles human secondarily generalized tonic–clonic SE behaviorally,²³⁵ electrically,²²¹^,²³⁵ and pharmacologically²³⁵^,²⁴⁴ (Table 2), and thus fulfills criteria proposed by Walton and Treiman²³⁵ for an ideal model of SE: (a) induced seizures should be similar in appearance to those seen in human SE, (b) electrographic patterns seen should also be like those seen in human SE, and (c) the induced SE should be responsive to the same drugs used in treating human SE. In the initial description of this model, a mean of 18.3 convulsions occurred over a mean time of 103.8 minutes after injection of homocysteine. Initially, the seizures are true, focal-onset, secondarily generalized tonic–clonic seizures. However, as SE continues over many convulsions, the seizures alter in appearance and may become prominent on only one side of the body or exhibit only subtle manifestations, as has been described in humans.²¹⁸

Electroencephalogram

SE is induced when brief focal runs of spikes with phase reversal around the left frontal electrode (the cobalt site) are evident on the EEG. Ten to fifteen minutes after homocysteine injection, these epileptiform discharges increase in amplitude, frequency, and distribution until a generalized convulsion occurs about 20 to 30 minutes after injection. Over the next 90 minutes, the EEG progresses through the five EEG stages described by Treiman et al.²²¹ and illustrated in FIGURE 16, although not all animals exhibit the full progression and some die before reaching stage V.

Pharmacology

Walton and Treiman²⁴⁴ validated this model’s ability to predict clinical effectiveness of a putative anti-SE drug by demonstrating that serum concentrations of diazepam,²³⁵ lorazepam,²³⁹ phenytoin,²⁴⁶ and phenobarbital²³⁷ effective at stopping generalized convulsions in this model closely approximate serum concentrations of these drugs reported to be effective in human GCSE. Subsequently, they used this model to evaluate the potential usefulness for the treatment of GCSE of other marketed and experimental antiepileptic drugs, including valproic acid,²⁴¹ tiagabine,²⁴⁵ fosphenytoin,²³⁸ NPC-17742,²⁴² lamotrigine,²⁴⁶ and remacemide.²⁴³

Pathophysiology

For at least three decades, cobalt has been known to induce focal onset seizures.⁴³ However, the mechanism of seizure induction by cobalt remains unknown. Homocysteine appears to be an NMDA agonist, and thus the mechanism whereby homocysteine induces seizures is most likely by activation of excitatory amino acid receptors.⁷⁹

Flurothyl Model

Flurothyl (bis-2,2,2-triflurothyl ether) is a convulsant gas²²⁵ that has been used by a number of investigators to induce experimental status epilepticus.

Methods

Nevander et al.¹⁵⁹ developed a model of SE to study the effect of SE on neuronal necrosis. They induced continuous seizure activity in anesthetized (60% nitrous oxide/40% oxygen), paralyzed, mechanically ventilated rats by injecting 80 μL of flurothyl directly into the rebreathing system. Additional boluses of flurothyl were used to maintain a burst-suppression pattern on the EEG during the seizure period. When flurothyl was discontinued after 15 minutes, seizure activity resolved spontaneously; a single dose of intravenous thiopental, 15 mg/kg, also arrested seizure activity. Acute seizures and SE have also been produced in initially awake, freely moving rats by dripping liquid flurothyl (1.2–3 mL/hr) onto filter paper suspended in a closed plastic box²⁵² or infusing 20 μL/min into an airtight chamber. Immature rats can survive 60 minutes of such repetitive seizures without mechanical ventilation; adult rats cannot.¹⁰³^,²⁰²^,²¹⁵

Neuropathology

Nevander et al.¹⁵⁹ observed that infarction of the pars reticulata of the substantia nigra occurred in five of the six animals with seizure duration of 30 minutes and in all animals with longer seizure durations when brains were examined for histologic damage 1 week after the episode of SE. The central part of the globus pallidus was also commonly affected. Neocortical, amygdaloid, thalamic nuclear, and hippocampal pyramidal cell damage was seen in SE of longer duration. However, Sperber and Moshe²⁰⁵ found no evidence of flurothyl SE-induced histologic damage when SE was induced in 14-day-old rats, suggesting that in this model, as well as others, SE-induced neuronal damage is age dependent.

Natural History and Clinical Phenomenology

The behavioral characteristics of flurothyl-induced seizures are age specific.²⁰⁶ Swimming movements and tonic posturing are seen during the first week, and clonic seizures developed after P10, initially preceded by a few myoclonic jerks and then evolving into a clonic–tonic seizure. By the third postnatal week, episodes of motionless staring associated with spike-wave discharges are seen. These seizures have been suggested as a model of absence seizures and the clonic and tonic–clonic seizures as models of primarily generalized convulsive seizures. In adult rats, early seizures during flurothyl exposure are similar to those seen in P14 rats: myoclonic, then clonic, then tonic–clonic seizures. However, because freely moving, nonventilated adult

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rats cannot be kept alive during flurothyl-induced status epilepticus, there are no descriptions of the evolution of behavioral changes during status epilepticus in adult rats.

Electroencephalogram

Sperber and Moshe²⁰⁵ reported discrete electrographic seizures initially, followed by a waxing and waning of seizure discharges throughout the 60 minutes of EEG recording during flurothyl-induced status epilepticus in rat pups. These descriptions correspond to what Treiman et al.²²¹ labeled SE EEG stages I and II. However, EEG recordings were not continued long enough in Sperber’s studies to determine whether the pups would have exhibited SE EEG stages III to V. EEG changes during flurothyl-induced SE have not been described in adult rats.

Pharmacology

No pharmacologic studies have been reported using flurothyl-induced experimental status epilepticus.

Pathophysiology

Woodbury suggested that flurothyl induces seizures by opening sodium channels in neuronal membranes.^250a More recently, antagonism of GABA-mediated inhibition⁷ and activation of cholinergic transmission⁶⁵ have been proposed as possible mechanisms.

Kainic Acid Model

Kainic acid is a potent agonist for the AMPA/kainate subtype of ionotropic glutamate receptor that has been used extensively to induce status epilepticus as a means of generating a model of chronic epilepsy. Thus this model is discussed in detail earlier in this chapter. The model has not been nearly as popular for the specific study of status epilepticus. Nonetheless, it has certain advantages for this purpose, which are discussed here.

Methods

Kainic acid (KA) usually is dissolved in phosphate-buffered saline and administered intraperitoneally. Doses in the rat are age and strain dependent.²⁰⁷ For rat pups, 1 to 8 mg/kg may be sufficient to induce SE; for adults, 8 to 15 mg/kg is usually necessary. The model has a relatively low efficiency. Status induction is frequently inconsistent, and acute mortality may be high. For this reason, some have advocated multiple repeated small doses tailored to the individual rat.¹⁰¹^,¹⁴⁸ Focal onset SE can also be induced by direct injection of 0.4 to 1.6 μg into the amygdala or hippocampus.¹⁸ It is important to use fresh kainic acid because of a loss of potency over time, which may result in high mortality when the bottle is first opened and low efficacy subsequently. Furthermore, kainic acid has been difficult to obtain at times, and is currently very expensive.

Neuropathology

See the section under chronic models for a discussion of status-induced chronic pathology. There have been no studies of the time course of neuropathologic changes during kainate-induced experimental status epilepticus.

Natural History and Clinical Phenomenology

See the section under chronic models for a discussion of the natural history and behavioral changes during kainate-induced status.

Electroencephalogram

Experimental status epilepticus induced by kainic acid injection in adult rats results in the same sequence of EEG changes originally described by Treiman et al.²²¹ However, in the kainic acid model, 5 to 6 hours are necessary to progress to stage V. This is similar to the time course reported for soman-induced SE, and contrasts with the 2 to 2½ hours required to reach stage V in the cobalt-homocysteine and lithium/pilocarpine models of experimental status epilepticus.

Pharmacology

Several compounds have been studied for their efficacy at suppressing kainate-induced status epilepticus. Nefiracetam (100 mg/kg intravenously) suppressed focal seizures induced by focal infusion of kainate into the amygdala.⁹⁷ Zonisamide (100 mg/kg intravenously) only suppressed seizure spread but not the epileptic focus.²¹⁴ MK801 [(+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]-cyclohepten-5,10-imine] is anticonvulsant against KA-induced seizures in adult rats: It reduces their severity and protects against neuronal damage, although it may worsen electrographic seizures.²²^,⁴¹^,⁷²^,¹¹⁷ A similar effect of MK801 has been reported in lithium/pilocarpine-induced late SE.²⁴⁰ In neonatal rats (P11–P12) pretreated with MK801, there is prevention of neither seizures nor KA-induced death.²⁰⁸ Pretreatment with neuroactive steroids¹¹³ or vigabatrin⁹² prevents the development of status epilepticus, but these drugs have not been tested against ongoing SE.

Pathophysiology

Acute seizures in this model of status epilepticus are presumably due to the direct excitatory effects of kainate on AMPA/KA-type glutamate receptors, which are of greatest abundance in the hippocampus, amygdala, perirhinal cortex, and entorhinal cortex.¹⁵⁵

Lithium/Pilocarpine and High-Dose Pilocarpine Models

Pilocarpine- and lithium/pilocarpine-induced SE has been used both to create models of chronic epilepsy and specifically to study various aspects of status epilepticus. Many features of the model are discussed earlier in this chapter; here we focus on elements specific to their role as SE models. Lithium/pilocarpine-induced SE is almost uniformly fatal within 24 hours and can be viewed as an experimental model of a severe form of generalized convulsive SE, perhaps approximating the clinical condition when GCSE develops as a complication of a severe systemic illness or generalized encephalopathy in patients without a prior history of epilepsy.

Methods

Status epilepticus can be induced by the intraperitoneal administration of 300 to 400 mg/kg pilocarpine (usually preceded by scopolamine 1 mg/kg given subcutaneously 30 minutes before pilocarpine to minimize peripheral cholinergic effects). Peripheral cholinergic effects can also be reduced by administering lithium chloride (3 mEq/kg) 24 hours before SE induction. With LiCl pretreatment, 20 to 30 mg/kg pilocarpine intraperitoneally is sufficient to induce SE. There is a high mortality rate, especially with lithium/pilocarpine administration, which can be largely eliminated by giving 10 mg/kg acepromazine 1 hour after the pilocarpine injection.

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Neuropathology

Neuropathologic changes following pilocarpine-induced status epilepticus in the rat have been reviewed in detail by Turski et al.²²⁶ and are summarized in the discussion of chronic models. Fujikawa⁸² studied the relationship between duration of lithium/pilocarpine-induced SE and the extent and severity of pathologic changes and observed progressively severe and increasingly widespread neuropathologic changes during the initial 3 hours of the SE episode.

Natural History and Clinical Manifestations

The temporal evolution of pilocarpine-induced behavioral changes and of seizure activity is discussed earlier in this chapter. Age-related deficits in visual-spatial learning after prolonged lithium/pilocarpine-induced SE have been reported by Holmes and colleagues,³⁹ which, it is interesting to note, can be improved by enriched environments.¹⁸² Marsh et al.¹³¹ recently showed that the degree of impairment of visual-spatial learning and memory in adult rats is predicted by the EEG stage at which SE is stopped. SE that progresses to EEG stage IV or V results in profound deficits in both memory and learning when tested in the Morris water maze.

Electroencephalogram

The progressive sequence of five EEG stages originally reported by Treiman et al.,²²¹ which was described in detail earlier, has been observed in lithium/pilocarpine-induced SE in adult rats²²¹^,²³⁶ and in pilocarpine-induced SE in adult¹¹¹ and juvenile²²² rats. The initial seizure activity on the EEG is seen about 20 minutes after pilocarpine injection, and stage V is reached in about 2 hours.

Pharmacology

Diazepam (5–10 mg/kg) is effective at stopping early lithium/pilocarpine-induced SE.²³⁶ A cocktail of diazepam, 10 mg/kg, and phenobarbital, 25 mg/kg, can stop later SE at some stages, but is not always effective unless combined with isoflurane. Pilocarpine models of SE have not been used systematically to study pharmacotherapy of status epilepticus. The seizures are too severe and pharmacoresistant to be useful for predicting efficacy of antistatus drugs for the initial treatment of generalized convulsive status epilepticus, but they may be useful as a model for the development of drugs for the treatment of refractory status epilepticus in humans.

Pathophysiology

Pilocarpine is a muscarinic acetylcholine receptor agonist and, like acetylcholine esterase inhibitors, induces status epilepticus by its initial excitatory role. However, the inability of anticholinergic agents, such as atropine, to stop ongoing pilocarpine-induced SE suggests the recruitment of other excitatory mechanisms, so that the episode of SE becomes self-sustaining. The mechanism whereby pretreatment with lithium markedly reduces the dose of pilocarpine necessary to induce SE is not known.

Soman and Other Cholinesterase Inhibitor (Nerve Agent) Models

Soman (pinacolyl methylphosphonofluoridate) is an organo-phosphorus cholinesterase inhibitor nerve agent that causes peripheral signs of cholinergic poisoning, convulsions, central neuronal damage, respiratory arrest, and death.⁴⁴^,⁸⁸^,¹⁴⁷ Soman and other organophosphorus agents have been studied extensively by military research facilities because of the potential of these agents to be used in warfare or terrorist attacks.

Methods

Almost all studies of these agents have been done with rats, guinea pigs, or rhesus monkeys, and most have been conducted by military investigators. Agents studied include tabun, cyclosarin, sarin, soman, VR, and VX. Median lethal doses (LD₅₀) range from 8 to 300 μg/kg, depending on the agent and the species in which it is studied.¹⁹²^,¹⁹³^,¹⁹⁴ Administration is usually subcutaneous.¹⁹³ In pharmacologic studies, guinea pigs are usually pretreated with pyridostigmine Br (0.026 mg/kg, intramuscularly) and 30 minutes later challenged with two times the LD₅₀ of the test agent, followed 1 minute later by treatment with atropine SO4 (2 mg/kg, intramuscularly) and pralidoxime chloride (2-PAM Cl; 25 mg/kg, intramuscularly).¹⁴³

Neuropathology

After early seizures in organophosphorus-induced status epilepticus, when anticholinergics readily terminate seizures, no neuropathology is evident. However, if the seizures are not stopped early, anticholinergics become less effective, and mild neuropathology is occasionally observed. With prolonged epileptiform activity, neuropathologic changes are observed in multiple brain regions, probably due to excessive influx of calcium due to repeated seizure-induced depolarization and prolonged stimulation of NMDA receptors, as seen in other models of prolonged status epilepticus.¹⁴¹

Natural History and Clinical Phenomenology

The initial behavioral change after soman administration is purposeless chewing, followed by head tremor and then lordosis-like posturing. Seizure activity begins with rhythmic movement of the ears and facial musculature and sometimes forepaw clonus, and sometimes progresses to class IV limbic seizures.¹⁴⁰

Electroencephalogram

Koplovitz and Skvorak¹¹⁵ reported the same sequence of five progressive EEG changes during soman-induced experimental status epilepticus in the rat that Treiman et al.²²¹ initially described in humans and three experimental models of SE in the rat; they pointed out that McDonough and Shih¹⁴¹ had previously described EEG changes corresponding to stages I, III, IV, and V in soman-intoxicated rats. McDonough has also observed a similar sequence of EEG changes following soman administration to rhesus monkeys (J. H. McDonough Jr., personal communication). In the Koplovitz and Skvorak study,¹¹⁵ the mean time of onset of stage I after soman administration (180 μg/kg subcutaneously) was 3.1 minutes, and it was 231.4 minutes for stage V.

Pharmacology

Diazepam can block soman-induced convulsions, electrographic seizure activity, and neuronal damage.⁹⁹^,¹²⁰^,¹³²^,¹⁴² Soman has been used to induce experimental SE in guinea pigs to test the ability of the anticholinergic agent scopolamine³¹ and the noncompetitive NMDA-receptor blocker dizocilpine (MK-801)²⁰⁴ to prevent or arrest seizure activity and thus to prevent neuronal necrosis. The dose of atropine can significantly affect the toxicity of the nerve agent and the efficacy of anticonvulsants.¹⁹⁵

Pathophysiology

Organophosphorus agents are potent irreversible inhibitors of acetylcholine esterase, the enzyme responsible for degradation

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of acetylcholine. Inhibition of acetylcholine esterase increases the availability of acetylcholine at all subtypes of acetylcholine receptors. Elevated acetylcholine levels in brain can be detected as early as 3 minutes after soman administration.¹⁹¹ McDonough and Shih¹⁴¹ proposed a three-phase “model” of the neuropharmacologic processes responsible for the seizures and neuropathology produced by nerve agent intoxication. Initiation and early expression of the seizures are cholinergic phenomena. If not checked, a transition phase occurs during which the neuronal excitation of the seizure per se perturbs other neurotransmitter systems. With prolonged epileptiform activity, the seizure enters a predominantly noncholinergic phase and becomes progressively refractory to all pharmacologic treatment, as is seen in other models of status epilepticus.

Electrogenic Models

Electrogenic models of SE have the advantage that the inducing stimulus immediately ceases when the electrical stimulation is stopped. Thus, response to drug therapy and evaluation of effects of status are not compromised by an ongoing exogenous stimulus.

Although several investigators had used repeated electroconvulsive shocks to produce prolonged seizure activity to study metabolic and biochemical consequences of prolonged seizures and SE,⁶¹^,²⁴⁸^,²⁴⁹ the first electrogenic models of self-sustaining SE were an outgrowth of kindling studies. Following Goddard’s⁹⁰ initial description of the kindling phenomenon, Pinel and Van Oot¹⁶⁷ demonstrated that SE could be produced in rats when kindling stimuli were administered over several months. Subsequently, Taber et al.²¹³ found that by using an interstimulus interval of 1 minute applied to hippocampal electrodes they could produce long-term, self-sustained, limbic or generalized SE in mice and subsequent deficits in an inhibitory avoidance task. McIntyre et al.¹⁴⁶ used a kindling-based electrical stimulation model of SE in the rat to study the pathologic consequences of partial-onset SE induced by 60 minutes of continuous electrical stimulation of the amygdala. The extent of pathologic change was determined by the duration of the episode of SE. Untreated rats showed massive gliosis and neuronal degeneration of the ipsilateral hemisphere; barbiturate-treated rats demonstrated less pathology. Sloviter and colleagues¹⁶¹^,¹⁹⁸^,²⁰¹ used electrical stimulation of the perforant pathway (the main excitatory pathway to the hippocampus) to replicate limbic SE and study seizure-induced neuronal damage independent of the metabolic effects associated with generalized convulsions. Sloviter¹⁹⁹ demonstrated a persistent loss of recurrent inhibition and irreversibly damaged adjacent interneurons following granule cell seizure activity. GABA-containing neurons survived, but there was a profound loss of adjacent somatostatin-containing interneurons and mossy cells, thus suggesting that seizure-induced loss of a basket cell–activating system may cause disinhibition. Vicedomini and Nadler²²⁹ also used stimulation of hippocampal afferent pathways to study the effects of prolonged seizure activity. They used a stimulus current administered through electrodes implanted in the angular bundle or fimbria to induce self-sustaining seizure activity that persisted after cessation of the electrical stimulation. In this model, the development of self-sustained seizure activity was essential for the production of neuronal damage: As little as 17 minutes of self-sustained seizure activity was sufficient to cause at least some neuronal loss, whereas as many as 759 stimulus trains (2.1 hours of evoked synaptic activity) produced no evidence of neuronal degeneration. EEG changes suggestive of a waxing and waning pattern (stage II) and of periodic epileptiform discharges (stage V) are illustrated in the description of this model, but whether continuous ictal discharges with or without flat periods (stages III and IV), as described in other models,⁹⁵^,²²¹ occurred with this model cannot be determined from the data provided. Lothman and colleagues,¹²⁵ on the other hand, described a model of self-sustained limbic SE that did exhibit the progressive EEG changes originally reported by Treiman et al.²²⁰^,²²¹ In this model, as little as 30 minutes of continuous focal electrical stimulation of the hippocampus elicited self-sustaining SE that persisted for many hours after discontinuing the electrical stimulation. Subsequently, Bertram and colleagues,²³^,¹⁷¹ using this model, noted that rats that progressed to stage V (periodic epileptiform discharges) were likely to develop chronic epilepsy, whereas animals in which SE stopped by early in stage III were not.

Cain et al.³⁰ also used a continuous stimulation paradigm as a simple and rapid procedure to induce limbic SE. These investigators administered 3-Hz biphasic square wave pulses via an electrode placed in the amygdala to reliably induce SE in almost all rats studied.

Almost all electrogenic models of self-sustained SE have resulted in limbic seizures, although occasional generalized convulsions are sometimes observed. However, Handforth and Treiman⁹⁵ described a nonpharmacologic model of limb clonic convulsions in the rat. Status epilepticus was induced by pulsed trains of suprathreshold electric current administered bilaterally to one of four forebrain sites: (a) orbital cortex, (b) medial prefrontal cortex, (c) deep prepiriform cortex, or (d) rostral caudate-putamen. Phenobarbital at very high serum concentrations stopped behavioral and electrical seizure activity; phenytoin, even at extremely high concentrations, did not. Handforth and Treiman⁹⁵ thus suggested this model for evaluation of the mechanisms and treatment of refractory SE. Mazarati, Wasterlain, and colleagues have used a perforant path stimulation model of self-sustaining status epilepticus (SSSE) in adult rats to study the response to a number of drugs. Diazepam and phenytoin are effective early, but much less so later in the episode.¹³⁷ NMDA-receptor blockers but not an AMPA-receptor blocker (NBQX) stop SSSE,¹³⁶ as does felbamate.¹³⁸ Levetiracetam in combination with diazepam appears to have some efficacy at high doses.¹³⁹ Pitkanen and colleagues developed an amygdala stimulation model of SSSE¹⁵⁷ and have used it to study consequences of pharmacologic treatment during status epilepticus. Treatment with diazepam within 2 hours reduces subsequent chronic epilepsy,¹⁶⁸ but lamotrigine does not.¹⁵⁸

Summary and Conclusions

Models can be used to understand basic mechanisms of a disease process, its consequences, and its treatment. Focal or systemic administration of chemical convulsants and direct cerebral electrical stimulation have been used to create a large variety of experimental models of epilepsy and of SE. Recently, sustained seizure activity in hippocampal slices has also been used as a model of SE.¹⁷⁸

The mechanisms of SE are now understood to be the result of failure of seizure-terminating mechanisms or of the mechanisms that make the brain refractory to subsequent seizures after a single discrete seizure, although many of the details of these mechanisms remain to be elucidated. Experimental models provide a method with which to study such phenomena. Furthermore, experimental models can be used effectively to test new drugs or new combinations of drugs for their utility in the treatment of chronic epilepsy and of SE before incurring the risk and expense of clinical trials.

Investigations on status epilepticus–induced neuronal damage and chronic epilepsy that occur after prolonged seizure activity are contributing to a better understanding the epileptogenic mechanisms underlying focal human epilepsies. In particular, the experimental results obtained in animal models of

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chronic epilepsies induced by pilocarpine, kainic acid, tetanus toxin, and focal electrical stimulation have cast some light on the mechanisms by which acute epileptic conditions can induce lesion- and seizure-dependent epileptogenic changes.

The animal models reviewed in this chapter have proven to be particularly suitable for testing the efficacy of antiepileptic drugs and may open new perspectives to the development of novel antiepileptogenic strategies.

References

1. Acampora D, Barone P, Simeone A. Otx genes in corticogenesis and brain development. Cereb Cortex. 1999;9:533–542.

2. Acampora D, Mazan S, Avantaggato V, et al. Epilepsy and brain abnormalities in mice lacking the Otx1 gene. Nat Genet. 1996;14:218–222.

3. Adrian ED. The spread of activity in the cerebral cortex. J Physiol. 1936;88:127–161.

4. Ajmone Marsan C. Focal electrical stimulation. In: Purpura DP, Penry JK, Tower D, et al., eds. Experimental Models of Epilepsy. A Manual for the Laboratory Worker. New York: Raven Press; 1972:147–172.

5. Alonso-De Florida F, Delgado JMR. Lasting behavioral and EEG changes in cats induced by prolonged stimulation of the amygdala. Am J Physiol. 1958;193:223–229.

6. Amano S, Ihara N, Uemura S, et al. Microdysgenesis in the hippocampal formation in a newly developed Ihara’s genetically epileptic rat (IGER). Epilepsia. 1996;37(Suppl 3):100–101.

7. Araki H, Kobayashi Y, Hashimoto Y, et al. Characteristics of flurothyl-induced seizures and the effect of antiepileptic drugs on flurothyl-induced seizures in Mongolian gerbils. Pharmacol Biochem Behav. 2002;74:141–147.

8. Avanzini G. Animal models relevant to human epilepsies. Ital J Neurol Sci. 1995;16:5–8.

9. Avanzini G, Spreafico R, Cipelletti B, et al. Synaptic properties of neocortical neurons in epileptic mice lacking the Otx1 gene. Epilepsia. 2000;41(Suppl 6):S200–S205.

10. Baglioni S, Magnini M. Azione di alcune sostanze chimiche sulle zone eccitabili della corteccia del cane. Arch Fisiol. 1909;6:240–249.

11. Baraban SC, Schwartzkroin PA. Electrophysiology of CA1 pyramidal neurons in an animal model of neuronal migration disorders: prenatal methylazoxymethanol treatment. Epilepsy Res. 1995;22:145–156.

12. Baraban SC, Schwartzkroin PA. Fluorothyl seizure susceptibility in rats following prenatal methylazoxymethanol treatment. Epilepsy Res. 1996;23:189–194.

13. Battaglia G, Colacitti C, Di Luca M, et al. The methylazoxymethanol-treated rat: an animal model for human developmental dysgeneses. Epilepsia. 1996;37(Suppl 5):72.

14. Battaglia G, Granata G, Farina L, et al. Periventricular nodular heterotopia: epileptogenic findings. Epilepsia. 1997;38:1173–1182.

15. Benardo LS, Prince DA. Acetylcholine induced modulation of hippocampal pyramidal neurons. Brain Res. 1981;211:227–234.

16. Benardo LS, Prince DA. Cholinergic excitation of mammalian hippocampal pyramidal cells. Brain Res. 1982;249:315–331.

17. Ben-Ari Y, Represa A. Brief seizure episodes induce long-term potentiation and mossy fibre sprouting in the hippocampus. Trends Neurosci. 1990;13:312–318.

18. Ben-Ari Y, Lagowska J, Tremblay E, et al. A new model of focal status epilepticus: intra-amygdaloid application of kainic acid elicits repetitive secondarily generalized convulsive seizures. Brain Res. 1979;163:176–179.

19. Ben-Ari Y, Tremblay E, Ottersen OP. Injections of kainic acid into the amygdaloid complex of the rat: an electrographic, clinical and histological study in relation to the pathology of epilepsy. Neuroscience. 1980;5:515–528.

20. Ben-Ari Y, Tremblay E, Ottersen OP, et al. The role of epileptic activity in hippocampal and “remote” cerebral lesions induced by kainic acid. Brain Res. 1980;191:79–97.

21. Ben-Ari Y, Tremblay E, Riche D. Electrographic, clinical and pathological alterations following systemic administration of kainic acid, bicuculline or pentetrazole: metabolic mapping using the deoxyglucose method with special reference to the pathology of epilepsy. Neuroscience. 1981;6:1361–1391.

22. Berg M, Bruhn T, Johansen FF, et al. Kainic acid–induced seizures and brain damage in the rat: different effects of NMDA- and AMPA receptor antagonists. Pharmacol Toxicol. 1993;73:262–68.

23. Bertram EH, Cornett J. The ontogeny of seizures in a rat model of limbic epilepsy: evidence for a kindling process in the development of chronic spontaneous seizures. Brain Res. 1993;625:295–300.

24. Boyle R. New experiments physico-mechanical, touching the spring of air, and its effects. Oxford: H. Hall; 1660. Quoted by Wood JD. Systemic oxygen derangements. In: Purpura DP, Penry JK, Tower D, et al., eds. Experimental Models of Epilepsy. A Manual for the Laboratory Worker. New York: Raven Press; 1972:459–476.

25. Bragin A, Engel J Jr, Wilson CL, et al. Electrophysiologic analysis of a chronic seizure model after unilateral hippocampal KA injection. Epilepsia. 1999;40:1210–1221.

26. Bragin A, Wilson CL, Almajano J, et al. High frequency oscillations after status epilepticus: epileptogenesis and seizure genesis. Epilepsia. 2004;45:1017–1023.

27. Brener K, Amitai Y, Jefferys JGR, et al. Chronic epileptic foci in neocortex: in vivo and in vitro effects of tetanus toxin. Eur J Neurosci. 1991;3:47–54.

28. Brooks VB, Asanuma H. Action of tetanus toxin in the cerebral cortex. Science. 1962;137:674–676.

29. Bugiani O, Ghetti B. Progressing encephalomyelopathy with muscular atrophy, induced by aluminum powder. Neurobiol Aging. 1982;3:209–222.

30. Cain DP, McKitrick DJ, Boon F. Rapid and reliable induction of partial status epilepticus in naive rats by low-frequency (3-Hz) stimulation of the amygdala. Epilepsy Res. 1992;12:51–55.

31. Capacio BR, Shih TM. Anticonvulsant actions of anticholinergic drugs in soman poisoning. Epilepsia. 1991;32:604–615.

32. Carrea R, Lanari A. Chronic effect of tetanus toxin applied locally to the cerebral cortex of the dog. Science. 1962;137:342–343.

33. Cavalheiro, EA. The pilocarpine model of epilepsy. Ital J Neurol Sci. 1995;16:33–37.

34. Cavalheiro EA, Riche DA, La Salle GLG. Long-term effects of intrahippocampal kainic acid injection in rats: a method for inducing spontaneous recurrent seizures. Electroencephalogr Clin Neurophysiol. 1982;53:581–589.

35. Cavalheiro EA, Leite JP, Bortolotto ZA, et al. Long-term effects of pilocarpine in rats: structural damage of the brain triggers kindling and spontaneous recurrent seizures. Epilepsia. 1991;32:778–782.

36. Caviness VS Jr. Patterns of cells and fiber distribution in the neocortex of reeler mutant mouse. J Comp Neurol. 1976;170:435–448.

37. Chen ZF, Schottler F, Bertram E, et al. Distribution and initiation of seizure activity in a rat brain with subcortical band heterotopia. Epilepsia. 2000;41:493–501.

38. Chevassous-au-Luis N, Rafiki A, Jorquera I, et al. Neocortex in the hippocampus: an anatomical and functional study of CA1 heterotopias after prenatal treatment with methylazoxymethanol in rats. J Compl Neurol. 1998;394:520–536.

39. Cilio MR, Sogawa Y, Cha BH, et al. Long-term effects of status epilepticus in the immature brain are specific for age and model. Epilepsia. 2003;44:518–528.

40. Clifford DB, Olney JW, Maniotis A, et al. The functional anatomy and pathology of lithium-pilocarpine and high-dose pilocarpine seizures. Neuroscience. 1987;23:953–968.

41. Clifford DB, Olney JW, Benz AM, et al. Ketamine, phencyclidine, and MK-801 protect against kainic acid–induced seizure-related brain damage. Epilepsia. 1990;31:382–390.

42. Colacitti C, Sancini G, De Biasi S, et al. Prenatal methylazoxymethanol treatment in rats produces brain abnormalities with morphological similarities to human developmental brain dysgeneses. J Neuropathol Exp Neurol. 1998;58:92–106.

43. Colasanti BK, Hartman ER, Craig CR. Electrocorticogram and behavioral correlates during the development of chronic cobalt experimental epilepsy in the rat. Epilepsia. 1974;15:361–373.

44. Coleman IW, Little PE, Bannard RAB. Cholinolytics in the treatment of anti-cholinesterase poisoning. I. The effectiveness of certain cholinolytics in combination with an oxime for treatment of sarin poisoning. Can J Biochem Physiol. 1962;40:814–826.

45. Collier PA, Ashwell KW. Distribution of neuronal heterotopiae following prenatal exposure to methylazoxymethanol. Neurotoxicol Teratol. 1993;15:439–444.

46. Collingridge GL, Thompson PA, Davies J, et al. In vitro effect of tetanus toxin on GABA release from rat hippocampal slices. J Neurochem. 1981;37:1039–1041.

47. Collins RC, McLean M, Olney J. Cerebral metabolic response to systemic kainic acid: 14-C-deoxyglucose studies. Life Sci. 1980;27:855–862.

48. Commission on Classification and Terminology of the International League Against Epilepsy. Proposal for revised classification of epilepsies and epileptic syndromes. Epilepsia. 1989;30:389–399.

49. Corcoran M, Moshé S, eds. Kindling 5. New York: Plenum Press; 1998.

50. Corcoran ME, Moshé SL, eds. Kindling 6. New York: Springer Science + Business Media; 2005.

51. Curtis DR, Duggan AW, Felix D, et al. GABA, bicuculline and central inhibition. Nature. 1970;226:1222–1224.

52. Curtis DR, Duggan AW, Felix D, et al. Bicuculline and central GABA receptors. Nature. 1970;228:676–677.

53. Daly DD, Pedley TA. Current Practice of Clinical Electroencephalography. New York: Raven Press; 1990.

54. Danscher G. Histochemical demonstration of heavy metals. A revised version of the sulfide silver method suitable for light and electron microscopy. Histochemistry. 1981;71:1–16.

55. De Deyn PP, D’Hooge R, Marescau B, et al. Chemical models of epilepsy with some reference to their applicability in the development of anticonvulsants. Epilepsy Res. 1992;12:87–110.

P.441

56. de Feo MR, Mecarelli O, Ricci GF. Bicuculline- and allylglycine-induced epilepsy in developing rats. Exp Neurol. 1985;90:411–421.

57. de Feo MR, Mecarelli O, Ricci GF. Seizure susceptibility in immature rats with microencephaly induced by prenatal exposure to methylazoxymethanol acetate. Pharmacol Res. 1995;31:109–114.

58. DeLorenzo RJ, Pellock JM, Towne AR, et al. Epidemiology of status epilepticus. J Clin Neurophysiol. 1995;12:316–325.

59. Des Portes V, Pinard JM, Billuart P, et al. Identification of a novel CNS gene required for neuronal migration and involved in X-linked subcortical heterotopias and lissencephaly syndrome. Cell. 1998;92:51–61.

60. Dubeau F, Tampieri D, Lee N, et al. Periventricular and subcortical nodular heterotopia. A study of 33 patients. Brain. 1995;118:1273–1287.

61. Duffy TE, Howse DC, Plum F. Cerebral energy metabolism during experimental status epilepticus. J Neurochem. 1975;24:925–934.

62. Dvorak K, Feit J. Migration of neuroblasts through partial necrosis of the cerebral cortex in newborn rats. Contribution to the problems of morphological development and developmental period of cerebral microgyria. Acta Neuropathol. 1977;38:203–212.

63. Dvorak K, Feit J, Jurankova Z. Experimentally induced focal microgyria and status verrucosus deformis in rats. Pathogenesis and interrelation histological and autoradiographical study. Acta Neuropathol. 1978;44:121–129.

64. Edmonds HL, Bellin SI, Chen FCM, et al. Anticonvulsant properties of ropizine in epileptic and non-epileptic beagle dogs. Epilepsia. 1978;19:139–146.

65. Eger EI, Gong D, Xing Y, et al. Acetylcholine receptors and thresholds for convulsions from flurothyl and 1,2-dichlorohexafluorocyclobutane. Anesth Analg. 2002;95:1611–1615.

66. Eksioglu YZ, Scheffer IE, Cardenas P, et al. Periventricular heterotopia: an X-linked dominant epilepsy locus causing aberrant cerebral cortical development. Neuron. 1996;16:77–87.

67. Empson RM, Jefferys JGR. Inhibitory synaptic function in primary and secondary chronic epileptic foci induced by intrahippocampal tetanus toxin in the rat. J Physiol (London). 1993;465:595–614.

68. Engel J Jr. Seizure and Epilepsy. Philadelphia: FA Davis;, 1989.

69. Engel J Jr. Report of the ILAE Classification Core Group. Epilepsia. 2006;47:1558–1568.

70. Engel J Jr, Ackermann RF. Interictal EEG spikes correlate with decreased rather than increased epileptogenicity in amygdaloid kindled rats. Brain Res. 1980;190:543–548.

71. Falconer DS. Two mutants, “trembler” and “reeler” with neurological actions in the house mouse (Mus musculus L.). J Genet. 1951;50:192–201.

72. Fariello RG, Golden GT, Smith GG, et al. Potentiation of kainic acid epileptogenicity and sparing from neuronal damage by an NMDA receptor antagonist. Epilepsy Res. 1989;3:206–213.

73. Ferrier D. Experimental researches in cerebral physiology and pathology. West Riding Lunatic Asylum Medical Report (London). 1873;3:1–50.

74. Fisher RS. Animal models of the epilepsies. Brain Res Rev. 1989;14:245–278.

75. Fisher RS, van Emde Boas W, Blume W, et al. Epileptic seizures and epilepsy. Definitions proposed by the International League Against Epilepsy (ILAE) and the International Bureau for Epilepsy. Epilepsia. 2005;46:470–472.

76. Forrester TM, Yokel RA. Comparative toxicity of intracerebroventricular and subcutaneous aluminum in the rabbit. Neurotoxicology. 1985;6:75–80.

77. Franceschetti S, Bugiani O, Panzica F, et al. Changes in excitability of CA1 pyramidal neurons in slices prepared from AlCl₃-treated rabbits. Epilepsy Res. 1990;6:39–48.

78. Franceschetti S, Sancini G, Colacitti C, et al. The methylazoxymethanol (MAM) treated rat as animal model for the neuronal migration disorders: electrophysiological findings in identified pyramidal neurones. In: Spreafico R, Avanzini G, Andermann F, eds. Abnormal Cortical Development and Epilepsy. London: John Libbey; 1999:121–132.

79. Freed WJ. Selective inhibition of homocysteine-induced seizures by glutamic acid diethyl ester and other glutamate esters. Epilepsia. 1985;26:30–36.

80. French JD, Gernandt BE, Livingston RB. Regional differences in seizure susceptibility in monkey cortex. AMA Arch Neurol Psychiatry. 1956;75:260–274.

81. Fritsch G, Hitzig E. Uber die elektrische Erregbarkeit des Grosshirns. Arch Anat Physiol. 1870;37:300–332.

82. Fujikawa DG. The temporal evolution of neuronal damage from pilocarpine-induced status epilepticus. Brain Res. 1996;725:11–22.

83. Gastaut HA, Meyer A, Naquet R, et al. Experimental psychomotor epilepsy in the cat: electro-clinical and anatomo-pathological correlations. J Neuropathol Exp Neurol. 1959;18:270–292.

84. Germano I, Sperber EF. Increased seizure susceptibility in adult rats with neuronal migration disorders. Brain Res. 1997;777:219–222.

85. Germano I, Sperber EF. Transplacentally induced neuronal migration disorders: an animal model for the study of the epilepsies. J Neurosci Res. 1998;51:473–488.

86. Germano IM, Zhang YF, Sperber EF, et al. Neuronal migration disorders increase susceptibility to hyperthermia-induced seizures in developing rats. Epilepsia. 1996;37:902–910.

87. Gilbert ME, Goodman JH. Chemical kindling. In: Pitkänen A, Schwartzkroin PA, Moshé S, eds. Models of Seizures and Epilepsy. San Diego, CA: Academic Press; 2006:379–393.

88. Glenn JF, Hinman DJ, McMaster SB. Electroencephalographic correlates of nerve agent poisoning. In: Dun NJ, Perlman RL, Karczman AG, eds. Neurobiology of Acetylcholine. New York: Plenum Press; 1987:503–534.

89. Gloor P. Mesial temporal sclerosis: historical background and an overview from a modern perspective. In: Lüders H, ed. Epilepsy Surgery. New York: Raven Press; 1991:689–703.

90. Goddard GV. Development of epileptic seizures through brain stimulation at low intensity. Nature. 1967;214:1020–1021.

91. Goddard GV, McIntyre DC, Leech CK. A permanent change in brain function resulting from daily electrical stimulation. Exp Neurol. 1969;25:295–330.

92. Halonen T, Miettinen R, Toppinen A, et al. Vigabatrin protects against kainic acid–induced neuronal damage in the rat hippocampus. Neurosci Lett. 1995;195:13–16.

93. Handforth A, Ackermann RF. Hierarchy of seizure states in the electrogenic limbic status epilepticus model: behavioral and electrographic observations of initial states and temporal progression. Epilepsia. 1992;33:589–600.

94. Handforth A, Ackermann RF. Electrogenic status epilepticus induced from numerous limbic sites. Epilepsy Res. 1993;15:21–26.

95. Handforth A, Treiman DM. A new, non-pharmacologic model of convulsive status epilepticus induced by electrical stimulation: behavioral/electroencephalographic observations and response to phenytoin and phenobarbital. Epilepsy Res. 1994;19:15–25.

96. Harris AB, Lockard JS. Macaca fascicularis: alternative epileptic model. Life Sci. 1979;25:281–286.

97. Hashizume K, Kunimoto M, Maeda T, et al. Antiepileptic effect of nefiracetam on kainic acid–induced limbic seizure in rats. Epilepsy Res. 2000;39:221–228.

98. Hauser WA. Status epilepticus: epidemiologic considerations. Neurology 1990;40(Suppl 2):9–13.

99. Hayward IJ, Wall HG, Jaax NK, et al. Decreased brain pathology in organophosphate-exposed rhesus monkeys following benzodiazepine therapy. J Neurol Sci. 1990;98:99–106.

100. Heinemann U, Dietzel I. Extracellular potassium concentration in chronic alumina cream foci of cats. J Neurophysiol. 1984;52:421–434.

101. Hellier JL, Dudek FE. Chemoconvulsant model of chronic spontaneous seizures. In: Crawley E, Gerfen CR, Rogawski MA, eds. Current Protocols in Neuroscience: Models of Chronic Spontaneous Seizures. Hoboken, NJ: Wiley.

102. Hoeppner TJ, Morrell F. Control of scar formation in experimentally induced epilepsy. Exp Neurol. 1986;94:519–536.

103. Huang L, Cilio MR, Silveira DC, et al. Long-term effects of neonatal seizures: a behavioral, electrophysiological, and histological study. Brain Res Dev Brain Res. 1999;118:99–107.

104. Jacobs KM, Gutnick MJ, Prince DA. Hyperexcitability in a model of cortical maldevelopment. Cereb Cortex. 1996;6:514–523.

105. Jefferys JGR. Chronic epileptic foci in vitro in hippocampal slices from rats with the tetanus toxin epileptic syndrome. J Neurophysiol. 1989;62:458–468.

106. Jefferys JGR, Borck C, Mellanby J. Chronic focal epilepsy induced by intracerebral tetanus toxin. Ital J Neurol Sci. 1995;16:27–32.

107. Johnston MV, Coyle JT. Histological and neurochemical effects of fetal treatment with methylazoxymethanol on rat neocortex in adulthood. Brain Res. 1979;170:135–55.

108. Kairiss EW, Racine RJ, Smith GK. The development of the interictal spike during kindling in the rat. Brain Res. 1984;322:101–110.

109. Kaufmann PY. Electrical phenomena in the cerebral cortex (Part 1) [in Russian]. Obozr Psikh Nevrol Eksper Psikhol. 1912;7–8:403–424.

110. Kaufmann PY. Electrical phenomena in the cerebral cortex (Part 2) [in Russian]. Obozr Psikh Nevrol Eksper Psikhol. 1912;9:513–535.

111. Kim JM, Walton NY, Treiman DM. EEG patterns of high-dose pilocarpine-induced status epilepticus in rats. Epilepsia. 1997;38(Suppl 8):225–226.

112. Klatzo I, Wisniewski H, Streicher E. Experimental production of neurofibrillary degeneration. Light microscopic observations. J Neuropathol Exp Neurol. 1965;24:187–199.

113. Kokate TG, Cohen AL, Karp E, et al. Neuroactive steroids protect against pilocarpine- and kainic acid–induced limbic seizures and status epilepticus in mice. Neuropharmacology. 1996;35:1049–1056.

114. Kopeloff LM, Barrera SE, Kopeloff N. Recurrent convulsive seizures in animals produced by immunologic and chemical means. Am J Psychiatry. 1942;98:881–902.

115. Koplovitz I, Skvorak JP. Electrocorticographic changes during generalized convulsive status epilepticus in soman intoxicated rats. Epilepsy Res. 1998;30:159–164.

116. Lee KS, Schottler F, Collins JL, et al. A genetic animal model of human neocortical heterotopia associated with seizures. J Neurosci. 1997;17:6236–6242.

117. Lees GJ. Effects of anaesthetics, anticonvulsants and glutamate antagonists on kainic acid–induced local and distal neuronal loss. J Neurol Sci. 1992;108:221–228.

P.442

118. Leite JP, Cavalheiro EA. Antiepileptic drugs and their antagonism of spontaneous seizures in rats. Epilepsia. 1991;32(Suppl 1):35.

119. Lemos T, Cavalheiro EA. Reorganizaçao sináptica e ocorrência de crises espontaneas em ratos. J Liga Bras Epil. 1991;5:135–138.

120. Lipp JA. Effect of diazepam upon soman-induced seizure activity and convulsions. Electroencephalogr Clin Neurophysiol. 1972;32:557–560.

121. Liu Z, Nagao T, Desjardins GC, et al. Quantitative evaluation of neuronal loss in the dorsal hippocampus in rats with long-term pilocarpine seizures. Epilepsy Res. 1994;17:237–247.

122. Lockard JS. A primate model of clinical epilepsy: mechanism of action through quantification of therapeutic effects. In: Lockard JS, Ward AA. Epilepsy: A Window to Brain Mechanisms. New York: Raven Press, 1980;11–49.

123. Lombardo AJ, Kuzniecky R, Powers RE, et al. Altered brain sodium channel transcript levels in human epilepsy. Brain Res Mol Brain Res. 1996;35:84–90.

124. Löscher W, Honack D, Rundfeldt C. Antiepileptogenic effects of the novel anticonvulsant levitaracetam (ucb L059) in the kindling model of temporal lobe epilepsy. J Pharmacol Exp Ther. 1998;284:474–479.

125. Lothman EW, Collins RC, Ferrendelli JA. Kainic acid–induced limbic seizures: electrophysiologic studies. Neurology. 1981;31:806–812.

126. Lothman EW, Bertram EH, Bekenstein JW, et al. Self-sustaining limbic status epilepticus induced by “continuous” hippocampal stimulation: electrographic and behavioral characteristics. Epilepsy Res. 1989;3:107–119.

127. Louis ED, Williamson PD, Darcey TM. Chronic focal epilepsy induced by microinjection of tetanus toxin into the cat motor cortex. Electroencephalogr Clin Neurophysiol. 1990;75:548–557.

128. Luciani, L. Sulla patogenesi dell’epilessia. Riv Sperimentale Freniatria Medicina Legale. 1878;4:617–646.

129. Luhmann HJ, Raabe K. Characterization of neuronal migration disorders in neocortical structures. I. Expression of epileptiform activity in an animal model. Epilepsy Res. 1996;26:67–74.

130. Luhmann HJ, Karpuk N, Qu M, et al. Characterization of neuronal migration disorders in neocortical structures. II. Intracellular in vitro recordings. J Neurophysiol. 1998;80:92–102.

131. Marsh ST, Good LB, Garvey KJ, et al. EEG stage predicts impairment of spatial memory and learning after experimental status epilepticus in the rat. Epilepsia. 2006;47(Suppl 4):314–315.

132. Martin LJ, Doebler JA, Shih TM, et al. Protective effect of diazepam pretreatment on soman-induced brain lesion formation. Brain Res. 1985;325:287–289.

133. Mathern GW, Cifuentes F, Leite JP, et al. Hippocampal EEG excitability and chronic spontaneous seizures are associated with aberrant synaptic reorganization in the rat intrahippocampal kainate model. Electroencephalogr Clin Neurophysiol. 1993;87:326–339.

134. Matsumoto H, Ajmone Marsan C. Cortical cellular phenomena in experimental epilepsy. Interictal manifestations. Exp Neurol. 1964;9:286–304.

135. Matsumoto H, Ajmone Marsan C. Cortical cellular phenomena in experimental epilepsy. Ictal manifestations. Exp Neurol. 1964;9:305–326.

136. Mazarati AM, Wasterlain CG. N-methyl-d-aspartate receptor antagonists abolish the maintenance phase of self-sustaining status epilepticus in rat. Neurosci Lett. 1999;265:187–190.

137. Mazarati AM, Baldwin RA, Sankar R, et al. Time-dependent decrease in the effectiveness of antiepileptic drugs during the course of self-sustaining status epilepticus. Brain Res. 1998;814:179–185.

138. Mazarati AM, Baldwin RA, Sofia RD, et al. Felbamate in experimental model of status epilepticus. Epilepsia. 2000;41:123–127.

139. Mazarati AM, Baldwin R, Klitgaard H, et al. Anticonvulsant effects of levetiracetam and levetiracetam–diazepam combinations in experimental status epilepticus. Epilepsy Res. 2004;58:167–174.

140. McDonough JH Jr, Shih TM. Pharmacological modulation of soman-induced seizures. Neurosci Biobehav Rev. 1993;17:203–215.

141. McDonough JH Jr, Shih TM. Neuropharmacological mechanisms of nerve agent–induced seizure and neuropathology. Neurosci Biobehav Rev. 1997;21:559–579.

142. McDonough JH Jr, Jaax NK, Crowley RA, et al. Atropine and/or diazepam therapy protects against soman-induced neural and cardiac pathology. Fund Appl Toxicol. 1989;13:256–276.

143. McDonough JH Jr, Zoeffel LD, McMonagle J, et al. Anticonvulsant treatment of nerve agent seizures: anticholinergics versus diazepam in soman–intoxicated guinea pigs. Epilepsy Res. 2000;38:1–14.

144. McIntyre DC. Split brain rat: transfer and interference of kindled amygdala convulsions. Can J Neurol Sci. 1975;2:429–437.

145. McIntyre DC. The kindling phenomenon. In: Pitkänen A, Schwartzkroin PA, Moshé S, eds. Models of Seizures and Epilepsy. San Diego, CA: Academic Press; 2006:351–363.

146. McIntyre DC, Nathanson D, Edson N. A new model of partial status epilepticus based on kindling. Brain Res. 1982;250:53–63.

147. McLeod CG, Jr, Singer AW, Harrington DG. Acute neuropathology in soman poisoned rats. Neurotoxicology. 1984;5:53–57.

148. Meier CL, Dudek FE. Spontaneous and stimulation-induced synchronized burst afterdischarges in the isolated CA1 of kainate-treated rats. J Neurophysiol. 1996;76:2231–2239.

149. Meldrum BS, Brierley JB. Prolonged epileptic seizures in primates: ischemic cell change and its relation to ictal physiological events. Arch Neurol. 1973;28:10–17.

150. Meldrum BS, Horton RW. Convulsive effects of 4-deoxypyridoxine and of bicuculline in photosensitive baboons (Papio papio) and in rhesus monkeys (Macaca mulatta). Brain Res. 1971;35:419–436.

151. Meldrum BS, Vigouroux RA, Brierley JB. Systemic factors and epileptic brain damage: prolonged seizures in paralyzed artificially ventilated baboons. Arch Neurol. 1973;29:82–87.

152. Mellanby J, George G, Robinson A, et al. Epileptiform syndrome in rats produced by injecting tetanus toxin into the hippocampus. J Neurol Neurosurg Psychiatry. 1977;40:404–414.

153. Mello LEAM, Cavalheiro EA, Tan AM, et al. Circuit mechanisms of seizures in the pilocarpine model of chronic epilepsy: cell loss and mossy fiber sprouting. Epilepsia. 1993;39:985–995.

154. Mikati MA, Werner S, Shehab L, et al. Stages of status epilepticus in the developing brain. Epilepsy Res. 2003;55:9–19.

155. Miller LP, Johnson AE, Gelhard RE, et al. The ontogeny of excitatory amino acid receptors in the rat forebrain––II. Kainic acid receptors. Neuroscience. 1990;35:45–51.

156. Mucha RF, Pinel JPJ. Postseizure inhibition of kindled seizures. Exp Neurol. 1977;54:266–282.

157. Nissinen J, Halonen T, Koivisto E, et al. A new model of chronic temporal lobe epilepsy induced by electrical stimulation of the amygdala in rat. Epilepsy Res. 2000;38:177–205.

158. Nissinen J, Large CH, Stratton SC, et al. Effect of lamotrigine treatment on epileptogenesis: an experimental study in rat. Epilepsy Res. 2004;58:119–132.

159. Nevander G, Ingvar M, Auer R, et al. Status epilepticus in well-oxygenated rats causes neuronal necrosis. Ann Neurol. 1985;18:281–290.

160. Olney JW, Rhee V, Ho OL. Kainic acid: a powerful neurotoxic, analog of glutamate. Brain Res. 1974;77:501–512.

161. Olney JW, de Gubareff T, Sloviter RS. “Epileptic” brain damage in rats induced by sustained electrical stimulation of the perforant path. II. Ultrastructural analysis of acute hippocampal pathology. Brain Res Bull. 1983;10:699–712.

162. Olney JW, Collins RC, Sloviter RS. Excitotoxic mechanisms of epileptic brain damage. In: Delgado-Escueta AV, Ward AA, Woodbury DM, et al., eds. Basic Mechanisms of the Epilepsies. Molecular and Cellular Approaches. New York: Raven Press; 1986:857–877.

163. Openchowski P. Sur l’action localisée du froid, appliqué à la surface de la région corticale du cerveau. C R Seances Mem Soc Biol. 1883;35:38–43.

164. Palmini A, Andermann F, Olivier A, et al. Focal neuronal migration disorders and intractable partial epilepsy: a study of 30 patients. Ann Neurol. 1991;30:741–749.

165. Peterson CJ, Vinayak S, Pazos A, et al. A rodent model of focally evoked self-sustaining status epilepticus. Eur J Pharmacol. 1992;221:151–155.

166. Pinel JPJ, Rovner LI. Experimental epileptogenesis: Kindling-induced epilepsy in rats. Exp Neurol. 1978;58:190–202.

167. Pinel JP, Van Oot PH. Generality of the kindling phenomenon: some clinical implications. Can J Neurol Sci. 1975;2:467–475.

168. Pitkänen A, Kharatishvili I, Narkilahti S, et al. Administration of diazepam during status epilepticus reduces development and severity of epilepsy in rat. Epilepsy Res. 2005;63:27–42.

169. Pitkänen A, Schwartzkroin PA, Moshé SL, eds. Models of Seizures and Epilepsy. San Diego, CA: Elsevier; 2006.

170. Post RM, Zhang ZJ, Weiss SRB, et al. Contingent tolerance and cross tolerance to anticonvulsant effects in amygdala-kindled seizures: mechanistic and clinical implications. In: Corcoran ME, Moshé SL, eds. Kindling 6. New York: Springer Science + Business Media; 2005:305–314.

171. Prasad A, Williamson JM, Bertram EH. Phenobarbital and MK-801, but not phenytoin, improve the long-term outcome of status epilepticus. Ann Neurol. 2002;51:175–181.

172. Prince DA. Topical convulsant drugs and metabolic antagonists. In: Purpura DP, Penry JK, Tower D, Woodbury DM, et al., eds. Experimental Models of Epilepsy. A Manual for the Laboratory Worker. New York: Raven Press; 1972:51–83.

173. Prince, DA. Basic mechanisms of focal epileptogenesis. In: Avanzini G, Fariello R, Heinemann U, et al., eds. Epileptogenic and Excitotoxic Mechanisms. London: John Libbey; 1993:17–27.

174. Prince DA, Futamachi KJ. Intracellular recordings from chronic epileptogenic foci in the monkey. Electroencephalogr Clin Neurophysiol. 1970;29:496–510.

175. Prince DA, Jacobs KM. Epileptogenesis in the freeze model of cortical microgyria. In: Spreafico R, Avanzini G, Andermann F, eds. Abnormal Cortical Development and Epilepsy. London: John Libbey; 1999:133–143.

176. Racine RJ. Modification of seizure activity by electrical stimulation. I. Afterdischarge threshold. Electroencephalogr Clin Neurophysiol. 1972;32:269–279.

177. Racine RJ. Modification of seizure activity by electrical stimulation. II. Motor seizure. Electroencephalogr Clin Neurophysiol. 1972;32:281–294.

178. Rafiq A, Zhang YF, DeLorenzo RJ, et al. Long-duration self-sustained epileptiform activity in the hippocampal parahippocampal slice: A model of status epilepticus. J Neurophysiol. 1995;74:2028–2042.

P.443

179. Ribak CE, Harris AB, Vaughn JE, et al. Inhibitory, GABAergic nerve terminals decrease at sites of focal epilepsy. Science. 1979;205:211–214.

180. Rosen GD, Jacobs KM, Prince DA. Effects of neonatal freeze lesions on expression of parvalbumin in rat neocortex. Cereb Cortex. 1998;8/8:753–761.

181. Roux E, Borrel A. Tétanos cérébral et immunité contre le tétanos. Ann Inst Pasteur. 1898;4:225–239.

182. Rutten A, van Albada M, Silveira DC, et al. Memory impairment following status epilepticus in immature rats: time-course and environmental effects. Eur J Neurosci. 2002;16:501–513.

183. Sancini G, Franceschetti S, Battaglia G, et al. Dysplastic neocortex and subcortical heterotopias in methylazoxymethanol-treated rats: an intracellular study of identified pyramidal neurons. Neurosci Lett. 1998;246:181–185.

184. Sancini G, Franceschetti S, Lavazza T, et al. Potentially epileptogenic dysfunction of cortical NMDA- and GABA-mediated neurotransmission in Otx1^-/- mice. Eur J Neurosci. 2001;14:1065–1074.

185. Scantlebury MH, Ouellet PL, Psarropoulou C, et al. Freeze lesion–induced focal cortical dysplasia predisposes to atypical hyperthermic seizures in the immature rat. Epilepsia. 2004;45:592–600.

186. Schield LK, Griffen JW, Drachman DB, et al. Retrograde axonal transport: a direct method for measurement of rate. Neurology. 1977;27:393.

187. Schwartzkroin PA, Turner DA, Knowles WD, et al. Studies of human and monkey “epileptic” neocortex in the in vitro slice preparation. Ann Neurol. 1983;13:249–257.

188. Schwarz P, Stichel CC, Luhmann HJ. Characterization of neuronal migration disorders in neocortical structures: loss or preservation of inhibitory interneurons? Epilepsia. 2000;41:781–787.

189. Sekiguchi M, Abe H, Shimai K, et al. Disruption of neuronal migration in the neocortex of the dreher mutant mouse. Dev Brain Res. 1994;77:37–43.

190. Seutin V, Johnson SW. Recent advances in the pharmacology of quaternary salts of bicuculline. Trends Pharmacol Sci. 1999;20:268–270.

191. Shih TM, McDonough JH Jr. Neurochemical mechanisms in soman-induced seizures. J Appl Toxicol. 1997;17:255–264.

192. Shih TM, McDonough JH Jr. Organophosphorus nerve agents–induced seizures and efficacy of atropine sulfate as anticonvulsant treatment. Pharmacol Biochem Behav. 1999;64:147–153.

193. Shih TM, McDonough JH Jr, Koplovitz I. Anticonvulsants for soman-induced seizure activity. J Biomed Sci. 1999;6:86–96.

194. Shih TM, Duniho SM, McDonough JH. Control of nerve agent–induced seizures is critical for neuroprotection and survival. Toxicol Appl Pharmacol. 2003;188:69–80.

195. Shih TM, Rowland TC, McDonough JH. Anticonvulsants for nerve agent–induced seizures: the influence of the therapeutic dose of atropine. J Pharmacol Exp Ther. 2007;320:154–161.

196. Shinozaki H, Konishi S. Actions of several anthelmintics and insecticides on rat cortical neurones. Brain Res. 1970;24:368–371.

197. Singh SC. Ectopic neurones in hippocampus of the post-natal rat exposed to methylazoxymethanol during foetal development. Acta Neuropathol. 1977;40:111–116.

198. Sloviter RS. “Epileptic” brain damage in rats induced by sustained electrical stimulation of the perforant path. I. Acute electrophysiological and light microscopic studies. Brain Res Bull. 1983;10:675–697.

199. Sloviter RS. Decreased hippocampal inhibition and a selective loss of interneurons in experimental epilepsy. Science. 1987;235:73–76.

200. Sloviter, RS. The functional organization of the hippocampal dentate gyrus and its relevance to the pathogenesis of temporal lobe epilepsy. Ann Neurol. 1994;35:640–654.

201. Sloviter RS, Damiano BP. Sustained electrical stimulation of the perforant path duplicates kainate-induced electrophysiological effects and hippocampal damage in rats. Neurosci Lett. 1981;24:279–284.

202. Sogawa Y, Monokoshi M, Silveira DC, et al. Timing of cognitive deficits following neonatal seizures: relationship to histological changes in the hippocampus. Brain Res Dev Brain Res. 2001;131:73–83.

203. Soper HV, Strain GM, Babb TL, et al. Chronic alumina temporal lobe seizures in monkeys. Exp Neurol. 1978;62:99–121.

204. Sparenborg S, Brennecke LH, Jaax NK, et al. Dizocilpine (MK-801) arrests status epilepticus and prevents brain damage induced by soman. Neuropharmacology. 1992;31:357–368.

205. Sperber EF, Moshe SL. Age-related differences in seizure susceptibility to flurothyl. Brain Res. 1988;467:295–297.

206. Sperber EF, Haas KZ, Romero MT, et al. Flurothyl status epilepticus in developing rats: behavioral, electrographic histological and electrophysiological studies. Brain Res Dev Brain Res. 1999;116:59–68.

207. Stafstrom CE, Thompson JL, Holmes GL. Kainic acid seizures in the developing brain: status epilepticus and spontaneous recurrent seizures. Brain Res Dev Brain Res. 1992;65:227–236.

208. Stafstrom CE, Tandon P, Hori A, et al. Acute effects of MK801 on kainic acid–induced seizures in neonatal rats. Epilepsy Res. 1997;26:335–344.

209. Stevens JR. An anatomy of schizophrenia. Arch Gen Psychiatry. 1973;29:177–189.

210. Stringer JL, Lothman EW. Maximal dentate activation: a tool to screen compounds for activity against limbic seizures. Epilepsy Res. 1990;5:169–176.

211. Sutula T, He XX, Cavazos J, et al. Synaptic reorganization in the hippocampus induced by abnormal functional activity. Science. 1988;239:1147–1150.

212. Sutula TP, Ockuly J. Kindling, spontaneous seizures, and the consequences of epilepsy: more than a model. In: Pitkänen A, Schwartzkroin PA, Moshé S, eds. Models of Seizures and Epilepsy. San Diego, CA: Academic Press; 2006:395–406.

213. Taber KH, McNamara JJ, Zornetzer SF. Status epilepticus: a new rodent model. Electroencephalogr Clin Neurophysiol. 1977;43:707–724.

214. Takano K, Tanaka T, Fujita T, et al. Zonisamide: electrophysiological and metabolic changes in kainic acid–induced limbic seizures in rats. Epilepsia. 1995;36:644–648.

215. Tanaka T, Kaijima M, Yonemasu Y, et al. Spontaneous secondarily generalized seizures induced by a single microinjection of kainic acid into unilateral amygdala in cats. Electroencephalogr Clin Neurophysiol. 1985;61:422–429.

216. Tauck D, Nadler JV. Evidence of functional mossy fiber sprouting in hippocampal formation of kainic acid–treated rats. J Neurosci. 1985;5:1016–1022.

217. Timm, F. Zur Histochemie der Schwermetalle. Das Sulfid Silber-Verfahren. Dtsch Z Ges Gerichtl Med. 1958;46:706–711.

218. Treiman DM. Generalized convulsive status epilepticus in the adult. Epilepsia. 1993;34(Suppl 1):S2–S11.

219. Treiman DM. Status epilepticus. In: Wyllie E, ed. Treatment of Epilepsy: Principles and Practice, 3rd ed. Philadelphia: Lippincott Williams & Wilkins; 2001:681–697.

220. Treiman DM, Walton NY, Wickboldt C, et al. Predictable sequence of EEG changes during generalized convulsive status epilepticus in man and three experimental models of status epilepticus in the rat. Neurology. 1987;7(Suppl 1):244.

221. Treiman DM, Walton NY, Kendrick C. A progressive sequence of electroencephalographic changes during generalized convulsive status epilepticus. Epilepsy Res. 1990;5:49–60.

222. Treiman DM, Good LB, Garvey KJ. A predictable sequence of EEG changes during pilocarpine-induced status epilepticus in juvenile rats. Epilepsia. 2004;45(Suppl 7):50.

223. Treiman MA, Wu J, Marsh ST, et al. EEG stage predicts electrophysiological changes in hippocampal slice recordings during experimental status epilepticus. Epilepsia. 2006;47(Suppl 4):326–327.

224. Trotter SA, Kapur J, Anzivino MJ, et al. GABAergic synaptic inhibition is reduced before seizure onset in a genetic model of cortical malformation. J Neurosci. 2006;26:10756–10767.

225. Truitt EB, Ebesberg EM, Ling ASG. Measurement of brain excitability by use of hexaflurodiethyl ether (Indoclon). J Pharmacol Exp Ther. 1960;129:445–453.

226. Turski L, Ikonomidou C, Turski WA, et al. Review: cholinergic mechanisms and epileptogenesis. The seizures induced by pilocarpine: a novel experimental model of intractable epilepsy. Synapse. 1989;3:154–171.

227. Turski L, Niemann W, Stephens DN. Differential effects of antiepileptic drugs and β-carbolines on seizures induced by excitatory amino acids. Neuroscience. 1990;39:799–807.

228. Veliskova J, Velisek L, Mares P, et al. Ketamine suppresses both bicuculline- and picrotoxin-induced generalized tonic–clonic seizures during ontogenesis. Pharmacol Biochem Behav. 1990;37:667–674.

229. Vicedomini JP, Nadler JV. A model of status epilepticus based on electrical stimulation of hippocampal afferent pathways. Exp Neurol. 1987;96:681–691.

230. Wada JA, ed. Kindling. New York: Raven Press; 1976.

231. Wada JA, ed. Kindling 2. New York: Raven Press; 1981.

232. Wada JA, ed. Kindling 3. New York: Raven Press; 1986.

233. Wada JA, ed. Kindling 4. New York: Plenum Press; 1989.

234. Wada JA, Osawa T. Spontaneous recurrent seizure state induced by daily electrical amygdaloid stimulation in Senegalese baboons, Papio papio. Neurology. 1976;26:273–286.

235. Walton NY, Treiman DM. Experimental secondarily generalized convulsive status epilepticus induced by d,l-homocysteine thiolactone. Epilepsy Res. 1988;2:79–86.

236. Walton NY, Treiman DM. Response of status epilepticus induced by lithium and pilocarpine to treatment with diazepam. Exp Neurol. 1988;101:267–275.

237. Walton NY, Treiman DM. Phenobarbital treatment of status epilepticus in a rodent model. Epilepsy Res. 1989;4:216–221.

238. Walton NY, Treiman DM. Efficacy of ACC-9653 (a phenytoin prodrug) in experimental status epilepticus in the rat. Epilepsy Res. 1990;5:165–168.

239. Walton NY, Treiman DM. Lorazepam treatment of experimental status epilepticus in the rat: relevance to clinical practice. Neurology. 1990;40:990–994.

240. Walton NY, Treiman DM. Motor and electroencephalographic response of refractory experimental status epilepticus in rats to treatment with MK-801, diazepam, or MK-801 plus diazepam. Brain Res. 1991;553:97–104.

241. Walton NY, Treiman DM. Valproic acid treatment of experimental status epilepticus. Epilepsy Res. 1992;12:199–205.

P.444

242. Walton NY, Treiman DM. Efficacy of a new NMDA receptor antagonist in treatment of experimental status epilepticus. Epilepsia. 1993;34:52.

243. Walton NY, Treiman DM. Remacemide versus phenytoin for treatment of experimental status epilepticus. Epilepsia. 1996;37(Suppl 5):212.

244. Walton NY, Treiman DM. Rational polytherapy in the treatment of status epilepticus. Epilepsy Res Suppl. 1996;11:123–139.

245. Walton NY, Gunawan S, Treiman DM. Treatment of experimental status epilepticus with the GABA uptake inhibitor, tiagabine. Epilepsy Res. 1994;19:237–244.

246. Walton NY, Jaing QB, Hyun B, et al. Lamotrigine vs phenytoin for treatment of status epilepticus: comparison in an experimental model. Epilepsy Res. 1996;24:19–28.

247. Ward AA. Topical convulsant metals. In: Purpura DP, Penry JK, Tower D, et al., eds. Experimental Models of Epilepsy. A Manual for the Laboratory Worker. New York: Raven Press; 1972:13–35.

248. Wasterlain CG. Breakdown of brain polysomes in status epilepticus. Brain Res. 1972;39:278–284.

249. Wasterlain CG. Mortality and morbidity from serial seizures: an experimental study. Epilepsia. 1974;15:155–176.

250. Westrum LE, White LE, Ward AA. Morphology of the experimental epileptic focus. J Neurosurg. 1964;21:1033–1046.

250a. Woodbury DM. Convulsant drugs: mechanism of action. In: Glaser GH, Penry JK, Woodburg DM, eds. Antiepileptic Drugs. New York: Raven Press; 1980:249–303.

251. Youmans JR. Experimental production of seizures in the macaque by temporal lobe lesions. Neurology. 1956;6:179–186.

252. Zhao Q, Holmes GL. Repetitive seizures in the immature brain. In: Pitkänen A, Schwartzkroin PA, Moshé S, eds. Models of Seizures and Epilepsy. San Diego, CA: Academic Press; 2006:341–350.

253. Zouhar A, Mares P, Liskova-Bernaskova K, et al. Motor and electrocorticographic epileptic activity induced by bicuculline in developing rats. Epilepsia. 1989;30:501–510.

	Epilepsy: A Comprehensive Textbook 2nd Edition
	© 2008 Lippincott Williams & Wilkins

Epilepsy: A Comprehensive Textbook2nd Edition

Epilepsy: A Comprehensive Textbook
2nd Edition