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The interaction between basic science epilepsy researchers and clinical epileptologists is a longstanding issue. Efforts to provide opportunities for a dialogue between preclinical and clinical epilepsy professionals are crucial to reduce the knowledge gap between them and improve the translational success of neurobiology-based research. The International League Against Epilepsy (ILAE) Research and Innovation Task Force circulated a survey to investigate the need for an update on new clinical epilepsy concepts within the basic science community. The 336 respondents included basic scientists (BS), preclinical scientists (PCSs), and/or clinical scientists (CSs). The majority of the 237 BSs/PCSs were engaged in preclinical studies in translational epilepsy research and declared translational research as a priority research interest. Fewer respondents from low-middle-income countries than from upper-middle or high-income countries (40.7% vs 65%) considered translational research a critical aspect of their research. A broad understanding of both clinical and neurobiological aspects of epilepsy was declared by 48% of BSs/PCSs; 96% of CSs declared a superficial knowledge of neurobiology of epilepsy. Most BSs/PCSs were aware that epilepsy is a complex condition that should be investigated with the help of clinical epileptologists, even though concerns were expressed on the relationship with clinicians. A focused training program on emerging clinical epileptological aspects tailored for BSs/PCSs was recommended by 81% of the participants; the majority of respondents preferred either 1- or 2-week in-presence tutoring or continuous online training coordinated by ILAE at the regional/national level. The survey also underscored the value of educational programs on neurobiology of epilepsy targeting CSs and low-middle-income countries (LMIC) investigators., (© 2024 The Author(s). Epilepsia published by Wiley Periodicals LLC on behalf of International League Against Epilepsy.)

Objective: Epilepsy surgery is a treatment option for patients with seizures that do not respond to pharmacotherapy. The histopathological characterization of the resected tissue has an important prognostic value to define postoperative seizure outcome in these patients. However, the diagnostic classification process based on microscopic assessment remains challenging, particularly in the case of focal cortical dysplasia (FCD). Imaging mass spectrometry is a spatial omics technique that could improve tissue phenotyping and patient stratification by investigating hundreds of biomolecules within a single tissue sample, without the need for target-specific reagents., Methods: An in situ proteomic technique called matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) is here investigated as a potential new tool to expand conventional diagnosis on standard paraffin brain tissue sections. Unsupervised and region of interest-based MALDI-MSI analyses of sections from 10 FCD type IIb (FCDIIb) cases were performed, and the results were validated by immunohistochemistry., Results: MALDI-MSI identified distinct histopathological features and the boundaries of the dysplastic lesion. The capability to visualize the spatial distribution of well-known diagnostic markers enabling multiplex measurements on single tissue sections was demonstrated. Finally, a fingerprint list of potential discriminant peptides that distinguish FCD core from peri-FCD tissue was generated., Significance: This is the first study that explores the potential application of MALDI-MSI in epilepsy postsurgery fixed tissue, by utilizing the well-characterized FCDIIb features as a model. Extending these preliminary analyses to a larger cohort of patients will generate spectral libraries of molecular signatures that discriminate tissue features and will contribute to patient phenotyping., (© 2024 The Author(s). Epilepsia published by Wiley Periodicals LLC on behalf of International League Against Epilepsy.)

The development of seizures in epilepsy syndromes associated with malformations of cortical development (MCDs) has traditionally been attributed to intrinsic cortical alterations resulting from abnormal network excitability. However, recent analyses at single-cell resolution of human brain samples from MCD patients have indicated the possible involvement of adaptive immunity in the pathogenesis of these disorders. By exploiting the MethylAzoxyMethanol (MAM)/pilocarpine (MP) rat model of drug-resistant epilepsy associated with MCD, we show here that the occurrence of status epilepticus and subsequent spontaneous recurrent seizures in the malformed, but not in the normal brain, are associated with the outbreak of a destructive autoimmune response with encephalitis-like features, involving components of both cell-mediated and humoral immune responses. The MP brain is characterized by blood-brain barrier dysfunction, marked and persisting CD8+ T cell invasion of the brain parenchyma, meningeal B cell accumulation, and complement-dependent cytotoxicity mediated by antineuronal antibodies. Furthermore, the therapeutic treatment of MP rats with the immunomodulatory drug fingolimod promotes both antiepileptogenic and neuroprotective effects. Collectively, these data show that the MP rat could serve as a translational model of epileptogenic cortical malformations associated with a central nervous system autoimmune response. This work indicates that a preexisting brain maldevelopment predisposes to a secondary autoimmune response, which acts as a precipitating factor for epilepsy and suggests immune intervention as a therapeutic option to be further explored in epileptic syndromes associated with MCDs., Competing Interests: Competing interests statement:The authors declare no competing interest.

Objective: Encephaloceles (ENCs) may cause clinical complications, including drug-resistant epilepsy that can be cured with epilepsy surgery., Methods: We describe clinical, diagnostic, and neuropathological findings of 12 patients with temporal ENC and epilepsy evaluated for surgery and compare them with a control group of 26 temporal lobe epilepsy (TLE) patients., Results: Six patients had unilateral and 6 bilateral temporal ENCs. Compared to TLEs, ENCs showed i) later epilepsy onset, ii) higher prevalence of psychiatric comorbidities, iii) no history of febrile convulsions, and iv) ictal semiology differences. Seven patients had MRI signs of gliosis, and 9 of intracranial hypertension. Interictal EEG analysis in ENCs demonstrated significant differences with controls: prominent activity in the beta/gamma frequency bands in frontal regions, interictal short sequences of low-voltage fast activity, and less frequent and more localized interictal epileptiform discharges. Ictal EEG patterns analyzed in 9 ENCs showed delayed and slower contralateral spread compared to TLEs. All ENCs that underwent surgery (7 lobectomies and 1 lesionectomy) are in Engel class I. Neuropathological examination revealed 4 patterns: herniated brain fragments, focal layer I distortion, white matter septa extending into the cortex, and altered gyral profile., Conclusions and Significance: The described peculiarities might help clinicians to suspect the presence of largely underdiagnosed ENCs., (© 2023. Fondazione Società Italiana di Neurologia.)

In vitro preparations (defined here as cultured cells, brain slices, and isolated whole brains) offer a variety of approaches to modeling various aspects of seizures and epilepsy. Such models are particularly amenable to the application of anti-seizure compounds, and consequently are a valuable tool to screen the mechanisms of epileptiform activity, mode of action of known anti-seizure medications (ASMs), and the potential efficacy of putative new anti-seizure compounds. Despite these applications, all disease models are a simplification of reality and are therefore subject to limitations. In this review, we summarize the main types of in vitro models that can be used in epilepsy research, describing key methodologies as well as notable advantages and disadvantages of each. We argue that a well-designed battery of in vitro models can form an effective and potentially high-throughput screening platform to predict the clinical usefulness of ASMs, and that in vitro models are particularly useful for interrogating mechanisms of ASMs. To conclude, we offer several key recommendations that maximize the potential value of in vitro models in ASM screening. This includes the use of multiple in vitro tests that can complement each other, carefully combined with in vivo studies, the use of tissues from chronically epileptic (rather than naïve wild-type) animals, and the integration of human cell/tissue-derived preparations., (© 2023 The Authors. Epilepsia published by Wiley Periodicals LLC on behalf of International League Against Epilepsy.)

Dendritic spines are the postsynaptic sites for most excitatory glutamatergic synapses. We previously demonstrated a severe spine loss and synaptic reorganization in human neocortices presenting Type II focal cortical dysplasia (FCD), a developmental malformation and frequent cause of drug-resistant focal epilepsy. We extend the findings, investigating the potential role of complement components C1q and C3 in synaptic pruning imbalance. Data from Type II FCD were compared with those obtained in focal epilepsies with different etiologies. Neocortical tissues were collected from 20 subjects, mainly adults with a mean age at surgery of 31 years, admitted to epilepsy surgery with a neuropathological diagnosis of: cryptogenic, temporal lobe epilepsy with hippocampal sclerosis, and Type IIa/b FCD. Dendritic spine density quantitation, evaluated in a previous paper using Golgi impregnation, was available in a subgroup. Immunohistochemistry, in situ hybridization, electron microscopy, and organotypic cultures were utilized to study complement/microglial activation patterns. FCD Type II samples presenting dendritic spine loss were characterized by an activation of the classical complement pathway and microglial reactivity. In the same samples, a close relationship between microglial cells and dendritic segments/synapses was found. These features were consistently observed in Type IIb FCD and in 1 of 3 Type IIa cases. In other patient groups and in perilesional areas outside the dysplasia, not presenting spine loss, these features were not observed. In vitro treatment with complement proteins of organotypic slices of cortical tissue with no sign of FCD induced a reduction in dendritic spine density. These data suggest that dysregulation of the complement system plays a role in microglia-mediated spine loss. This mechanism, known to be involved in the removal of redundant synapses during development, is likely reactivated in Type II FCD, particularly in Type IIb; local treatment with anticomplement drugs could in principle modify the course of disease in these patients., (© 2022 The Authors. Brain Pathology published by John Wiley & Sons Ltd on behalf of International Society of Neuropathology.)

Epilepsies affecting the limbic regions are common and generate seizures often resistant to pharmacological treatment. Clinical evidence demonstrates that diverse regions of the mesial portion of the temporal lobe participate in limbic seizures; these include the hippocampus, the entorhinal, perirhinal and parahippocampal regions and the piriform cortex. The network mechanisms involved in the generation of olfactory-limbic epileptiform patterns will be here examined, with particular emphasis on acute interictal and ictal epileptiform discharges obtained by treatment with pro-convulsive drugs and by high-frequency stimulations on in vitro preparations, such as brain slices and the isolated guinea pig brain. The interactions within olfactory-limbic circuits can be summarized as follows: independent, region-specific seizure-like events (SLE) are generated in the olfactory and in the limbic cortex; SLEs generated in the hippocampal-parahippocampal regions tend to remain within these areas; the perirhinal region controls the neocortical propagation and the generalization of limbic seizures; interictal spiking in the olfactory regions prevents the invasion by SLEs generated in limbic regions. The potential relevance of these observations for human focal epilepsy is discussed., (Copyright © 2023. Published by Elsevier Inc.)

Under physiological conditions, neuronal network synchronization leads to different oscillatory EEG patterns that are associated with specific behavioral and cognitive functions. Excessive synchronization can, however, lead to focal or generalized epileptiform activities. It is indeed well established that in both epileptic patients and animal models, focal epileptiform EEG patterns are characterized by interictal and ictal (seizure) discharges. Over the last three decades, employing in vitro and in vivo recording techniques, several experimental studies have firmly identified a paradoxical role of GABA A signaling in generating interictal discharges, and in initiating-and perhaps sustaining-focal seizures. Here, we will review these experiments and we will extend our appraisal to evidence suggesting that GABA A signaling may also contribute to epileptogenesis, i.e., the development of plastic changes in brain excitability that leads to the chronic epileptic condition. Overall, we anticipate that this information should provide the rationale for developing new specific pharmacological treatments for patients presenting with focal epileptic disorders such as mesial temporal lobe epilepsy (MTLE)., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Avoli, de Curtis, Lévesque, Librizzi, Uva and Wang.)

Epilepsy surgery is recommended in selected patients with Tuberous Sclerosis Complex (TSC). However, reports on predictive factors of seizure outcome are variable. Here we report on seizure and cognitive outcome of 35 TSC patients who received surgery for refractory epilepsy in 7 Italian centers over a period of 22 years (1997-2019). The rate of seizure-free individuals at last follow-up (mean 7.5 years, range 1-21 years) was 51%. Patients with longer follow-up (≥10 years) had a lower rate of Engel I outcome (11.1%) than those who received surgery in the last 10 years (65.4%, p = 0.003). Factors associated with Engel II, III, IV outcome in our cohort included: high number of cortical tubers (≥5); presence of subependymal nodules (SENs); seizure onset before age 1 year; and multifocal interictal epileptic discharges (IEDs) on electroencephalogram (EEG). A subset of patients evaluated with Vineland Adaptive Behaviour Scales (VABS) showed developmental gains, in line with their developmental trajectories, but no improvement in standard scores after surgery was noted. Our study demonstrates that the rates of successful seizure outcome of epilepsy surgery in TSC have improved in the last 10 years. More than half of the patients achieved seizure freedom, and a high proportion of affected individuals experienced a reduction in seizure burden and in antiseizure medications. A comprehensive assessment after surgery should be performed in TSC patients to evaluate the overall neurodevelopmental outcome, as measures that are based only on seizure control do not adequately identify the benefits of surgery on global functioning in these patients., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Epilepsy is a pathologic condition with high prevalence and devastating consequences for the patient and its entourage. Means for accurate diagnosis of type, patient monitoring for predicting seizures and follow up, and efficacious treatment are desperately needed. To improve this adverse outcome, miRNAs and the chaperone system (CS) are promising targets to understand pathogenic mechanisms and for developing theranostics applications. miRNAs implicated in conditions known or suspected to favor seizures such as neuroinflammation, to promote epileptic tolerance and neuronal survival, to regulate seizures, and others showing variations in expression levels related to seizures are promising candidates as useful biomarkers for diagnosis and patient monitoring, and as targets for developing novel therapies. Components of the CS are also promising as biomarkers and as therapeutic targets, since they participate in epileptogenic pathways and in cytoprotective mechanisms in various epileptogenic brain areas, even if what they do and how is not yet clear. The data in this review should help in the identification of molecular targets among the discussed miRNAs and CS components for research aiming at understanding epileptogenic mechanisms and, subsequently, develop means for predicting/preventing seizures and treating the disease.

Neuronal dendritic arborizations and dendritic spines are crucial for a normal synaptic transmission and may be critically involved in the pathophysiology of epilepsy. Alterations in dendritic morphology and spine loss mainly in hippocampal neurons have been reported both in epilepsy animal models and in human brain tissues from patients with epilepsy. However, it is still unclear whether these dendritic abnormalities relate to the cause of epilepsy or are generated by seizure recurrence. We investigated fine neuronal structures at the level of dendritic and spine organization using Golgi impregnation, and analysed synaptic networks with immunohistochemical markers of glutamatergic (vGLUT1) and GABAergic (vGAT) axon terminals in human cerebral cortices derived from epilepsy surgery. Specimens were obtained from 28 patients with different neuropathologically defined aetiologies: type Ia and type II focal cortical dysplasia, cryptogenic (no lesion) and temporal lobe epilepsy with hippocampal sclerosis. Autoptic tissues were used for comparison. Three-dimensional reconstructions of Golgi-impregnated neurons revealed severe dendritic reshaping and spine alteration in the core of the type II focal cortical dysplasia. Dysmorphic neurons showed increased dendritic complexity, reduction of dendritic spines and occasional filopodia-like protrusions emerging from the soma. Surprisingly, the intermingled normal-looking pyramidal neurons also showed severe spine loss and simplified dendritic arborization. No changes were observed outside the dysplasia (perilesional tissue) or in neocortical postsurgical tissue obtained in the other patient groups. Immunoreactivities of vGLUT1 and vGAT showed synaptic reorganization in the core of type II dysplasia characterized by the presence of abnormal perisomatic baskets around dysmorphic neurons, in particular those with filopodia-like protrusions, and changes in vGLUT1/vGAT expression. Ultrastructural data in type II dysplasia highlighted the presence of altered neuropil engulfed by glial processes. Our data indicate that the fine morphological aspect of neurons and dendritic spines are normal in epileptogenic neocortex, with the exception of type II dysplastic lesions. The findings suggest that the mechanisms leading to this severe form of cortical malformation interfere with the normal dendritic arborization and synaptic network organization. The data argue against the concept that long-lasting epilepsy and seizure recurrence per se unavoidably produce a dendritic pathology., (© The Author(s) (2020). Published by Oxford University Press on behalf of the Guarantors of Brain. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

During epidemic outbreaks, epilepsy course can be modified by different physical and psychological stressors and, most importantly, by irregular therapy intake. The effect of COVID-19 and quarantine isolation on the course of epilepsy and on incidence of new-onset seizures is still unclear. With the aim of managing epilepsy in quarantined patients, three Italian Epilepsy Centers set up telephone consultations using a semistructured interview, allowing a prospective collection of data on seizure course and other seizure-related problems during pandemic. The collected data on seizure course were compared with the analogous period of 2019. The level of patients' concern relating to the COVID-19 pandemic was also assessed using a numeric rating scale. To address the effect of COVID-19 pandemic on seizure incidence, data collection included the number of consultations for first seizures, relapse seizures, and status epilepticus (SE) in the emergency department of one of the participating centers. Clinical telephone interviews suggest the absence of quarantine effect on epilepsy course in our cohort. No differences in incidence of emergency consultations for seizures over a two-month period were also observed compared with a control period. As demonstrated in other infective outbreaks, good antiepileptic drug (AED) supplying, precise information, and reassurance are the most important factors in chronic conditions to minimize psychological and physical stress, and to avoid unplanned treatment interruptions., Competing Interests: Declaration of competing interest None. This research did not receive any specific grant from funding agencies in the public, commercial, or not-for-profit sectors., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Introduction: Focal Cortical Dysplasia (FCD) represents a broad spectrum of histopathological entities that cause drug-resistant epilepsy. Surgery has been shown to be the treatment of choice, but incomplete resection represents the leading cause of seizure persistence. Preliminary experiences with intraoperative ultrasound (ioUS) have proven its potential in defining and characterizing the lesion. In this study we analyzed the feasibility of advanced ultrasound techniques such as sono-elastography (SE) and contrast enhancement ultrasound (CEUS) in a small cohort of patients with FCD., Material and Methods: We retrospectively reviewed all clinical records and images of patients with drug resistant epilepsy who underwent at least one advanced sonographic technique (SE and/or CEUS) during ioUS guided surgery between November 2014 and October 2017. We excluded from our analysis all patients with lesions other than FCD or those who had FCD associated with other pathological entities., Results: Four patients with type IIb FCD in the right frontal lobe were evaluated. All of them underwent SE, which highlighted heterogeneous stiffness in the dysplastic foci, also multiple areas of higher consistency were detected in all patients. Three patients evaluated with CEUS had visible enhancement in the FCD. Neither SE nor CEUS were better than ioUS in the identification of lesion boundaries. In the three patients who underwent both SE and CEUS we found no correspondence between stiffer areas and enhancement in the dysplastic areas., Conclusion: Ourpreliminary report confirms the feasibility of SE and CEUS in FCD surgery and describes the imaging findings in this category of patients. Studies on larger cohorts of patients are warranted to better clarify the role of these advanced intraoperative ultrasound techniques in patients with FCD., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Objective: The current study aimed to describe quality of life (QoL) levels, psychiatric symptoms prevalence, and perceived stigma levels in persons with either drug-resistant epilepsy (DRE) or drug-sensitive epilepsy (DSE) and in persons with epilepsy (PwE) with DRE that underwent epilepsy surgery (DREES)., Methods: Persons with epilepsy diagnosed as having DRE according to International League Against Epilepsy (ILAE) criteria, DSE, and DREES were enrolled at the Epilepsy Unit of the Neurological Institute Carlo Besta of Milan. Sociodemographic and clinical data, Quality of Life in Epilepsy Inventory (QOLIE-31), Symptom Checklist-90 (SCL-90), and the Epilepsy Stigma Scale (ESS) were collected based on self-reported information and on medical records., Results: Sociodemographic, medical, and psychological data were obtained from 181 PwE: 80 with DRE, 31 with DSE, and 70 with DREES. We found that QoL is higher and psychiatric symptoms are lower in persons with DSE compared with DRE and that patients with DREES, who were drug-resistant before surgery, are in between DSE and DRE for both measures. Perceived stigma level is different in DSE and in DRE, that report the highest levels of stigma, and is between the other two groups in DREES., Significance: This study suggests that low QoL levels and high psychiatric symptoms prevalence in drug-resistant PwE may be significantly improved after epilepsy surgery and suggests the importance of a biopsychosocial approach when planning therapeutic intervention., Competing Interests: Declaration of competing interest The authors report no conflict of interest. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Objective: Patients with epilepsy often ask if recurrent seizures harm their brain and aggravate their epileptic condition. This crucial question has not been specifically addressed by dedicated experiments. We analyze here if intense bilateral seizure activity induced by local injection of kainic acid (KA) in the right hippocampus produces brain damage in the left hippocampus., Methods: Adult guinea pigs were bilaterally implanted with hippocampal electrodes for continuous video-electroencephalography (EEG) monitoring. Unilateral injection of 1 μg KA in the dorsal CA1 area induced nonconvulsive status epilepticus (ncSE) characterized by bilateral hippocampal seizure discharges. This treatment resulted in selective unilateral sclerosis of the KA-injected hippocampus. Three days after KA injection, the animals were killed, and the brains were submitted to ex vivo magnetic resonance imaging (MRI) and were processed for immunohistochemical analysis., Results: During ncSE, epileptiform activity was recorded for 27.6 ± 19.1 hours in both the KA-injected and contralateral hippocampi. Enhanced T1-weighted MR signal due to gadolinium deposition, mean diffusivity reduction, neuronal loss, gliosis, and blood-brain barrier permeability changes was observed exclusively in the KA-injected hippocampus. Despite the presence of a clear unilateral hippocampal sclerosis at the site of KA injection, no structural alterations were detected by MR and immunostaining analysis performed in the hippocampus contralateral to KA injection 3 days and 2 months after ncSE induction. Fluoro-Jade and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining at the same time points confirmed the absence of degenerating cells in the hippocampi contralateral to KA injection., Significance: We demonstrate that intense epileptiform activity during ncSE does not cause obvious brain damage in the hippocampus contralateral to unilateral hippocampal KA injection. These findings argue against the hypothesis that epileptiform activity per se contributes to focal brain injury in previously undamaged cortical regions., (Wiley Periodicals, Inc. © 2019 International League Against Epilepsy.)

The brain is a complex system composed of networks of interacting elements, from genes to circuits, whose function (and dysfunction) is not derivable from the superposition of individual components. Epilepsy is frequently described as a network disease, but to date, there is no standardized framework within which network concepts applicable to all levels from genes to whole brain can be used to generate deeper insights into the pathogenesis of seizures or the associated morbidities. To address this shortcoming, the Neurobiology Commission of the International League Against Epilepsy dedicated a Workshop on Neurobiology of Epilepsy (XIV WONOEP 2017) with the aim of formalizing network concepts as they apply to epilepsy and to critically discuss whether and how such concepts could augment current research endeavors. Here, we review concepts and strategies derived by considering epilepsy as a disease of different network hierarchies that range from genes to clinical phenotypes. We propose that the concept of networks is important for understanding epilepsy and is critical for developing new study designs. These approaches could ultimately facilitate the development of novel diagnostic and therapeutic strategies., (Wiley Periodicals, Inc. © 2018 International League Against Epilepsy.)

Potassium channels dysfunction and altered genes encoding for molecules involved in potassium homeostasis have been associated with human epilepsy. These observations are in agreement with a control role of extracellular potassium on neuronal excitability and seizure generation. Epileptiform activity, in turn, regulates potassium homeostasis through mechanisms that are still not well established. We review here how potassium-associated processes are regulated in the brain and examine the mechanisms that support the role of potassium in triggering epileptiform activities., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Epilepsy affects all age groups and is one of the most common and most disabling neurological disorders. The accurate diagnosis of seizures is essential as some patients will be misdiagnosed with epilepsy, whereas others will receive an incorrect diagnosis. Indeed, errors in diagnosis are common, and many patients fail to receive the correct treatment, which often has severe consequences. Although many patients have seizure control using a single medication, others require multiple medications, resective surgery, neuromodulation devices or dietary therapies. In addition, one-third of patients will continue to have uncontrolled seizures. Epilepsy can substantially impair quality of life owing to seizures, comorbid mood and psychiatric disorders, cognitive deficits and adverse effects of medications. In addition, seizures can be fatal owing to direct effects on autonomic and arousal functions or owing to indirect effects such as drowning and other accidents. Deciphering the pathophysiology of epilepsy has advanced the understanding of the cellular and molecular events initiated by pathogenetic insults that transform normal circuits into epileptic circuits (epileptogenesis) and the mechanisms that generate seizures (ictogenesis). The discovery of >500 genes associated with epilepsy has led to new animal models, more precise diagnoses and, in some cases, targeted therapies.

Electroencephalography (EEG)-the direct recording of the electrical activity of populations of neurons-is a tremendously important tool for diagnosing, treating, and researching epilepsy. Although standard procedures for recording and analyzing human EEG exist and are broadly accepted, there are no such standards for research in animal models of seizures and epilepsy-recording montages, acquisition systems, and processing algorithms may differ substantially among investigators and laboratories. The lack of standard procedures for acquiring and analyzing EEG from animal models of epilepsy hinders the interpretation of experimental results and reduces the ability of the scientific community to efficiently translate new experimental findings into clinical practice. Accordingly, the intention of this report is twofold: (1) to review current techniques for the collection and software-based analysis of neural field recordings in animal models of epilepsy, and (2) to offer pertinent standards and reporting guidelines for this research. Specifically, we review current techniques for signal acquisition, signal conditioning, signal processing, data storage, and data sharing, and include applicable recommendations to standardize collection and reporting. We close with a discussion of challenges and future opportunities, and include a supplemental report of currently available acquisition systems and analysis tools. This work represents a collaboration on behalf of the American Epilepsy Society/International League Against Epilepsy (AES/ILAE) Translational Task Force (TASK1-Workgroup 5), and is part of a larger effort to harmonize video-EEG interpretation and analysis methods across studies using in vivo and in vitro seizure and epilepsy models., (Wiley Periodicals, Inc. © 2017 International League Against Epilepsy.)

This paper is a result of work of the AES/ILAE Translational Task Force of the International League Against Epilepsy. The aim is to provide acceptable standards and interpretation of results of electrophysiological depth recordings in vivo in control rodents., (Wiley Periodicals, Inc. © 2017 International League Against Epilepsy.)

In vivo electrophysiological recordings are widely used in neuroscience research, and video-electroencephalography (vEEG) has become a mainstay of preclinical neuroscience research, including studies of epilepsy and cognition. Studies utilizing vEEG typically involve comparison of measurements obtained from different experimental groups, or from the same experimental group at different times, in which one set of measurements serves as "control" and the others as "test" of the variables of interest. Thus, controls provide mainly a reference measurement for the experimental test. Control rodents represent an undiagnosed population, and cannot be assumed to be "normal" in the sense of being "healthy." Certain physiological EEG patterns seen in humans are also seen in control rodents. However, interpretation of rodent vEEG studies relies on documented differences in frequency, morphology, type, location, behavioral state dependence, reactivity, and functional or structural correlates of specific EEG patterns and features between control and test groups. This paper will focus on the vEEG of standard laboratory rodent strains with the aim of developing a small set of practical guidelines that can assist researchers in the design, reporting, and interpretation of future vEEG studies. To this end, we will: (1) discuss advantages and pitfalls of common vEEG techniques in rodents and propose a set of recommended practices and (2) present EEG patterns and associated behaviors recorded from adult rats of a variety of strains. We will describe the defining features of selected vEEG patterns (brain-generated or artifactual) and note similarities to vEEG patterns seen in adult humans. We will note similarities to normal variants or pathological human EEG patterns and defer their interpretation to a future report focusing on rodent seizure patterns., (Wiley Periodicals, Inc. © 2017 International League Against Epilepsy.)

In vitro preparations are a powerful tool to explore the mechanisms and processes underlying epileptogenesis and ictogenesis. In this review, we critically review the numerous in vitro methodologies utilized in epilepsy research. We provide support for the inclusion of detailed descriptions of techniques, including often ignored parameters with unpredictable yet significant effects on study reproducibility and outcomes. In addition, we explore how recent developments in brain slice preparation relate to their use as models of epileptic activity., (Wiley Periodicals, Inc. © 2017 International League Against Epilepsy.)

Objective: The contribution of recurring seizures to the progression of epileptogenesis is debated. Seizure-induced brain damage is not conclusively demonstrated either in humans or in animal models of epilepsy. We evaluated the expression of brain injury biomarkers on postsurgical brain tissue obtained from 20 patients with frequent seizures and a long history of drug-resistant focal epilepsy., Methods: The expression patterns of specific glial, neuronal, and inflammatory molecules were evaluated by immunohistochemistry in the core of type II focal cortical dysplasias (FCD-II), at the FCD boundary (perilesion), and in the adjacent normal-appearing area included in the epileptogenic region. We also analyzed surgical specimens from cryptogenic patients not presenting structural alterations at imaging., Results: Astroglial and microglial activation, reduced neuronal density, perivascular CD3-positive T-lymphocyte clustering, and fibrinogen extravasation were demonstrated in the core of FCD-II lesions. No pathological immunoreactivity was observed outside the FCD-II or in cryptogenetic specimens, where the occurrence of interictal and ictal epileptiform activity was confirmed by either stereo-electroencephalography or intraoperative electrocorticography., Interpretation: Recurrent seizures do not induce the expression of brain damage markers in nonlesional epileptogenic cortex studied in postsurgical tissue from cryptogenic and FCD patients. This evidence argues against the hypothesis that epileptiform activity per se contributes to focal brain injury, at least in the neocortical epilepsies considered here. Ann Neurol 2017;82:331-341., (© 2017 American Neurological Association.)

Objective: Several studies have reported that inhibitory networks are altered in dysplastic tissue obtained from epilepsy surgery specimens. A consistent decrease in the number of inhibitory interneuronal subpopulation that expresses parvalbumin (PV) was reported in postsurgical tissue from patients with focal cortical dysplasia (FCD). We tested if the decrease in PV protein expression observed in epileptic tissue corresponds to a parallel impairment in the γ-aminobutyric acid (GABA)ergic compartment., Methods: We analyzed postsurgical tissue from 30 surgically treated patients who underwent surgery for intractable epilepsy including 26 patients with FCD (types I, II, and III) and 4 patients without any microscopic visible lesion (cryptogenic) as controls. Serial sections were processed using in situ hybridization with GAD-65 and GAD-67 probes and immunocytochemistry with antibody against PV. The density of inhibitory PV-immunoreactive interneurons in relation to GABAergic cells was estimated in controls and in all different pathologic groups by using a two- and three-dimensional (2D and 3D) cell-counting technique. Field fraction and line profile analyses were added to estimate immunostaining proportion and distribution of PV signal generated in gray matter., Results: A reduction of PV-positive cells and PV-immunoreactivity was observed exclusively in FCD type I/III specimens compared with cryptogenic tissue from control patients with a poor postsurgical outcome. In FCD type II, a profound rearrangement in the cortical distribution of PV immunoreactivity was observed, without a quantitative reduction of the number of neurons and terminals. In situ hybridization did not reveal significant variations of GAD expression in any FCD subtype., Significance: Our study suggests a preservation of inhibitory networks in FCD postsurgical tissue, demonstrated by a substantial normal count of GABAergic neurons. A selective PV expression impairment is demonstrated in FCD type I and III and an abnormal, but not reduced, distribution of PV cells and terminals is confirmed in type II FCD. Possible functional consequences are discussed., (Wiley Periodicals, Inc. © 2016 International League Against Epilepsy.)

Background: Research on ictogenesis is based on the study of activity between seizures and during seizures in animal models of epilepsy (chronic condition) or in in vitro slices obtained from naïve non-epileptic brains after treatment with pro-convulsive drugs, manipulations of the extracellular medium and specific stimulation protocols., New Method: The in vitro isolated guinea pig brain retains the functional connectivity between brain structures and maintains interactions between neuronal, glial and vascular compartments. It is a close-to-in vivo preparation that offers experimental advantages not achieved with the use of other experimental models. Neurophysiological and imaging techniques can be utilized in this preparation to study brain activity during and between seizures induced by pharmacological or functional manipulations., Results: Cellular and network determinants of interictal and ictal discharges that reproduce abnormal patterns observed in human focal epilepsies and the associated changes in extracellular ion and blood-brain permeability can be identified and analyzed in the isolated guinea pig brain., Comparison With Existing Methods: Ictal and interictal patterns recorded in in vitro slices may show substantial differences from seizure activity recorded in vivo due to slicing procedure itself. The isolated guinea pig brain maintained in vitro by arterial perfusion combines the typical facilitated access of in vitro preparations, that are difficult to approach during in vivo experiments, with the preservation of larger neuronal networks., Conclusions: The in vitro whole isolated guinea pig brain preparation offers an unique experimental model to study systemic and neurovascular changes during ictogenesis., (Published by Elsevier B.V.)

In patients diagnosed with pharmaco-resistant epilepsy, cerebral areas responsible for seizure generation can be defined by performing implantation of intracranial electrodes. The identification of the epileptogenic zone (EZ) is based on visual inspection of the intracranial electroencephalogram (IEEG) performed by highly qualified neurophysiologists. New computer-based quantitative EEG analyses have been developed in collaboration with the signal analysis community to expedite EZ detection. The aim of the present report is to compare different signal analysis approaches developed in four different European laboratories working in close collaboration with four European Epilepsy Centers. Computer-based signal analysis methods were retrospectively applied to IEEG recordings performed in four patients undergoing pre-surgical exploration of pharmaco-resistant epilepsy. The four methods elaborated by the different teams to identify the EZ are based either on frequency analysis, on nonlinear signal analysis, on connectivity measures or on statistical parametric mapping of epileptogenicity indices. All methods converge on the identification of EZ in patients that present with fast activity at seizure onset. When traditional visual inspection was not successful in detecting EZ on IEEG, the different signal analysis methods produced highly discordant results. Quantitative analysis of IEEG recordings complement clinical evaluation by contributing to the study of epileptogenic networks during seizures. We demonstrate that the degree of sensitivity of different computer-based methods to detect the EZ in respect to visual EEG inspection depends on the specific seizure pattern.

Objective: Cortical dysplasias (CDs) represent a wide range of cortical abnormalities that closely correlate with intractable epilepsy. Rats prenatally exposed to 1-3-bis-chloroethyl-nitrosurea (BCNU) represent an injury-based model that reproduces many histopathologic features of human CD. Previous studies reported in vivo hyperexcitability in this model, but in vivo epileptogenicity has not been confirmed., Methods: To determine whether cortical and hippocampal lesions lead to epileptiform discharges and/or seizures in the BCNU model, rats at three different ages (3, 5, and 9 months old) were implanted for long-term video electroencephalographic recording. At the end of the recording session, brain tissue was processed for histologic and immunohistochemical investigation including cAMP response element binding protein (CREB) phosphorylation, as a biomarker of epileptogenicity., Results: BCNU-treated rats showed spontaneous epileptiform activity (67%) in the absence of a second seizure-provoking hit. Such activity originated mainly from one hippocampus and propagated to the ipsilateral neocortex. No epileptiform activity was found in age-matched control rats. The histopathologic investigation revealed that all BCNU rats with epileptiform activity showed neocortical and hippocampal abnormalities; the presence and the severity of these lesions did not correlate consistently with the propensity to generate epileptiform discharges. Epileptiform activity was found only in cortical areas of BCNU-treated rats in which a correlation between brain abnormalities and increased pCREB expression was observed., Significance: This study demonstrates the in vivo occurrence of spontaneous epileptiform discharges in the BCNU model and shows that increased pCREB expression can be utilized as a reliable biomarker of epileptogenicity., (Wiley Periodicals, Inc. © 2015 International League Against Epilepsy.)

Epilepsy is a disease characterized by aberrant connections between brain areas. The altered activity patterns generated by epileptic networks can be analyzed with intracerebral electrodes during pre-surgical stereo-electroencephalographic (EEG) monitoring in patients candidate to epilepsy surgery. The responses to high frequency stimulation (HFS) at 50 Hz performed for diagnostic purposes during SEEG were analyzed with a new algorithm, to evaluate signal parameters that are masked to visual inspection and to define the boundaries of the epileptogenic network. The analysis was focused on 60-80 Hz activity that represented the largest frequency component evoked by HFS. The distribution of HFS-evoked fast activity across all (up to 162) recording contacts allowed to define different clusters of contacts that retrospectively correlated to the epileptogenic zone identified by the clinicians on the basis of traditional visual analysis. The study demonstrates that computer-assisted analysis of HFS-evoked activities may contribute to the definition of the epileptogenic network on intracranial recordings performed in a pre-surgical setting.

Relevant to the definition of epileptic encephalopathy (EE) is the concept that the epileptic activity itself may contribute to bad outcomes, both in terms of epilepsy and cognition, above and beyond what might be expected from the underlying pathology alone, and that these can worsen over time. The review of the clinical and experimental evidence that seizures or interictal electroencephalography (EEG) discharges themselves can induce a progression toward more severe epilepsy and a regression of brain function leads to the following conclusions: The possibility of seizure-dependent worsening is by no means a general one but is limited to some types of epilepsy, namely mesial temporal lobe epilepsy (MTLE) and EEs. Clinical and experimental data concur in indicating that prolonged seizures/status epilepticus (SE) are a risky initial event that can set in motion an epileptogenic process leading to persistent, possibly drug-refractory epilepsies. The mechanisms for SE-related epileptogenic process are incompletely known; they seem to involve inflammation and/or glutamatergic transmission. The evidence of the role of recurrent individual seizures in sustaining epilepsy progression is ambiguous. The correlation between high seizure frequency and bad outcome does not necessarily demonstrate a cause-effect relationship, rather high seizure frequency and bad outcome can both depend on a particularly aggressive epileptogenic process. The results of EE studies challenge the idea of a common seizure-dependent mechanism for epilepsy progression/intellectual deterioration., (Wiley Periodicals, Inc. © 2013 International League Against Epilepsy.)

Epilepsy has been historically seen as a functional brain disorder associated with excessive synchronization of large neuronal populations leading to a hypersynchronous state. Recent evidence showed that epileptiform phenomena, particularly seizures, result from complex interactions between neuronal networks characterized by heterogeneity of neuronal firing and dynamical evolution of synchronization. Desynchronization is often observed preceding seizures or during their early stages; in contrast, high levels of synchronization observed towards the end of seizures may facilitate termination. In this review we discuss cellular and network mechanisms responsible for such complex changes in synchronization. Recent work has identified cell-type-specific inhibitory and excitatory interactions, the dichotomy between neuronal firing and the non-local measurement of local field potentials distant to that firing, and the reflection of the neuronal dark matter problem in non-firing neurons active in seizures. These recent advances have challenged long-established views and are leading to a more rigorous and realistic understanding of the pathophysiology of epilepsy.

Objective: Epilepsy is a common neurological disorder characterized by recurrent seizures often unresponsive to pharmacological treatment. Brain inflammation is considered a crucial etiopathogenetic mechanism of epilepsy that could be targeted to control seizures. Specific inflammatory mediators overexpressed in human epileptogenic foci are known to promote seizures in animal models. We investigated whether seizures induce brain inflammation independently on extracerebral factors. We also investigated whether brain-borne inflammation is required and sufficient to maintain seizure activity and whether it causes blood-brain barrier (BBB) impairment. We addressed these questions by studying the relation between seizures, inflammation, and BBB permeability in a brain preparation isolated from extracerebral compartments., Methods: Epileptiform activity was induced by arterial perfusion of bicuculline in the in vitro isolated guinea pig brain. Seizure-induced brain inflammation was evaluated by quantitative immunohistochemical analysis of interleukin (IL)-1β in parenchymal cells. BBB damage was assessed by extravasation of intravascular fluorescein isothiocyanate-albumin. The effects of arterially perfused anakinra, a human recombinant IL-1β receptor antagonist, were investigated on epileptiform discharges, brain inflammation, and BBB damage., Results: Seizure induction in the absence of extracerebral factors promoted the release of IL-1β from brain resident cells and enhanced its biosynthesis in astrocytes. Anakinra rapidly terminated seizures, prevented their recurrence, and resolved seizure-associated BBB breakdown., Interpretation: Seizures initiate brain inflammation in glia and promote BBB damage that is independent of either leukocytes or blood-borne inflammatory molecules. Brain inflammation contributes to the duration and recurrence of seizures. This study supports the use of specific anti-inflammatory drugs in clinical conditions that present with intractable recurrent seizures., (Copyright © 2012 American Neurological Association.)

Purpose: Early seizures are a frequent consequence of stroke. The main goal of the present study is to verify whether anoxic ischemia per se is able to induce early changes in excitability that may be a prelude to the generation of seizures and, ultimately, to epileptogenesis. Excitability changes in the very acute postischemic phase are here analyzed in a new model of ischemia developed in the isolated guinea pig brain preparation., Methods: Permanent bilateral occlusion of the anterior cerebral arteries (ACAs) was performed in the isolated guinea pig brain maintained in vitro by arterial perfusion. Magnetic resonance imaging and immunohistochemistry were utilized to identify the penumbra and core regions induced by ACA occlusion (ACAo). Slow potentials and evoked responses recorded in olfactory cortices were utilized to evaluate excitability changes in the acute phase after ischemia., Key Findings: ACAo induces a core area located in the shell of the nucleus accumbens and a region of penumbra in the underlying olfactory cortices, where characteristic slow potential shifts, but no reduction of diffusion tensor magnetic resonance (MR) signal and microtubule associated protein 2 (MAP-2) immunostaining (typical of ischemic core) was observed. Recording of responses evoked by low- and high-frequency stimulations of the lateral olfactory tract showed no excitability changes in the early hours that follow ischemia in the olfactory cortical areas supplied by ACAs., Significance: The absence of early hyperexcitability changes in an isolated whole brain model of ischemia, strongly suggests that brain anoxia per se does not contribute to the generation of early seizures. These findings support the view that blood-borne events (such as hemorrhage and inflammation) may play a major role in early postischemic seizures., (Wiley Periodicals, Inc. © 2011 International League Against Epilepsy.)

Purpose: The piriform cortex (PC) is known to be epileptic-prone and it may be involved in the manifestation of limbic seizures. Herein, we have characterized some electrophysiologic and pharmacologic properties of the spontaneous epileptiform activity generated by PC networks maintained in vitro., Methods: We performed field potential recordings from the PC in coronal or sagittal rat brain slices along with pharmacologic manipulations of γ-aminobutyric acid (GABA)ergic and glutamatergic signaling during application of the convulsant drug 4-aminopyridine (4AP, 50 μm)., Key Findings: Coronal and sagittal preparations generated interictal-like and ictal-like epileptiform discharges with similar duration and frequency. Ictal-like discharges in sagittal slices were initiated mostly in the PC anterior subregion, whereas interictal activity did not have any preferential site of origin. In sagittal slices, high frequency oscillations (HFOs) at 80-200 Hz were detected mainly at the beginning of the ictal discharge in both posterior and anterior subregions. N-Methyl-d-aspartate (NMDA) receptor antagonism abolished ictal discharges, but failed to influence interictal activity. In the absence of ionotropic glutamatergic transmission, PC networks generated slow, GABA receptor-dependent events. Finally, GABA(A) receptor antagonism during application of 4AP only, abolished ictal discharges and disclosed recurrent interictal activity., Significance: Our findings demonstrate that PC networks can sustain in vitro epileptiform activity induced by 4AP. HFOs, which emerge at the onset of ictal activity, may be involved in PC ictogenesis. As reported in several cortical structures, ionotropic glutamatergic neurotransmission is necessary but not sufficient for ictal discharge generation, a process that also requires operative GABA(A) receptor-mediated signaling., (Wiley Periodicals, Inc. © 2012 International League Against Epilepsy.)

GABA is the main inhibitory neurotransmitter in the adult forebrain, where it activates ionotropic type A and metabotropic type B receptors. Early studies have shown that GABA(A) receptor-mediated inhibition controls neuronal excitability and thus the occurrence of seizures. However, more complex, and at times unexpected, mechanisms of GABAergic signaling have been identified during epileptiform discharges over the last few years. Here, we will review experimental data that point at the paradoxical role played by GABA(A) receptor-mediated mechanisms in synchronizing neuronal networks, and in particular those of limbic structures such as the hippocampus, the entorhinal and perirhinal cortices, or the amygdala. After having summarized the fundamental characteristics of GABA(A) receptor-mediated mechanisms, we will analyze their role in the generation of network oscillations and their contribution to epileptiform synchronization. Whether and how GABA(A) receptors influence the interaction between limbic networks leading to ictogenesis will be also reviewed. Finally, we will consider the role of altered inhibition in the human epileptic brain along with the ability of GABA(A) receptor-mediated conductances to generate synchronous depolarizing events that may lead to ictogenesis in human epileptic disorders as well., (Copyright © 2011. Published by Elsevier Ltd.)

In vitro studies performed on brain slices demonstrate that the potassium channel blocker 4-aminopyridine (4AP, 50 microM) discloses electrographic seizure activity and interictal discharges. These epileptiform patterns have been further analyzed here in a isolated whole guinea pig brain in vitro by using field potential recordings in olfactory and limbic structures. In 8 of 13 experiments runs of fast oscillatory activity (fast runs, FRs) in the piriform cortex (PC) propagated to the lateral entorhinal cortex (EC), hippocampus and occasionally to the medial EC. Early and late FRs were asynchronous in the hemispheres showed different duration [1.78 +/- 0.51 and 27.95 +/- 4.55 (SD) s, respectively], frequency of occurrence (1.82 +/- 0.49 and 34.16 +/- 6.03 s) and frequency content (20-40 vs. 40-60 Hz). Preictal spikes independent from the FRs appeared in the hippocampus/EC and developed into ictal-like discharges that did not propagate to the PC. Ictal-like activity consisted of fast activity with onset either in the hippocampus (n = 6) or in the mEC (n = 2), followed by irregular spiking and sequences of diffusely synchronous bursts. Perfusion of the N-methyl-d-aspartate receptor antagonist 2-amino-5-phosphonopentanoic acid (100 microM) did not prevent FRs, increased the duration of limbic ictal-like discharges and favored their propagation to olfactory structures. The AMPA receptor antagonist 6,7-dinitroquinoxaline-2,3-dione (50 microM) blocked ictal-like events and reduced FRs. In conclusion, 4AP-induced epileptiform activities are asynchronous and independent in olfactory and hippocampal-entorhinal regions. Epileptiform discharges in the isolated guinea pig brain show different pharmacological properties compared with rodent in vitro slices.

In this study we report that in the presence of normal buffer, epileptiform discharges occur spontaneously (duration = 2.60 +/- 0.49 s) or can be induced by electrical stimuli (duration = 2.50 +/- 0.62 s) in the entorhinal cortex (EC) of brain slices obtained from pilocarpine-treated rats but not in those from age-matched, nonepileptic control (NEC) animals. These network-driven epileptiform events consist of field oscillatory sequences at frequencies greater than 200 Hz that most often initiate in the lateral EC and propagate to the medial EC with 4-63 ms delays. The NMDA receptor antagonist CPP depresses the rate of occurrence (P < 0.01) of these spontaneous epileptiform discharges but fails in blocking them. Paradoxically, stimulus-induced epileptiform responses are enhanced in duration during CPP application. However, concomitant application of NMDA and non-NMDA glutamatergic antagonists abolishes spontaneous and stimulus-induced epileptiform events. Intracellular recordings from lateral EC layer V cells indicate a lower frequency of spontaneous hyperpolarizing postsynaptic potentials in pilocarpine-treated tissue than in NEC (P < 0.002) both under control conditions and with glutamatergic receptor blockade; the reversal potential of pharmacologically isolated GABA(A) receptor-mediated inhibitory postsynaptic potentials has similar values in the two types of tissue. Finally, immunohistochemical analysis shows that parvalbumin-positive interneurons are selectively reduced in number in EC deep layers. Collectively, these results indicate that reduced inhibition within the pilocarpine-treated EC layer V may promote network epileptic hyperexcitability.

Systemic application of the muscarinic agonist, pilocarpine, is commonly utilized to induce an acute status epilepticus that evolves into a chronic epileptic condition characterized by spontaneous seizures. Recent findings suggest that the status epilepticus induced by pilocarpine may be triggered by changes in the blood-brain barrier (BBB) permeability. We tested the role of the BBB in an acute pilocarpine model by using the in vitro model brain preparation and compared our finding with in vivo data. Arterial perfusion of the in vitro isolated guinea-pig brain with <1 mM pilocarpine did not cause epileptiform activity, but rather reduced synaptic transmission and induced steady fast (20-25 Hz) oscillatory activity in limbic cortices. These effects were reversibly blocked by co-perfusion of the muscarinic antagonist atropine sulfate (5 microM). Brain pilocarpine measurements in vivo and in vitro suggested modest BBB penetration. Pilocarpine induced epileptiform discharges only when perfused with compounds that enhance BBB permeability, such as bradykinin (n=2) or histamine (n=10). This pro-epileptic effect was abolished when the BBB-impermeable muscarinic antagonist atropine methyl bromide (5 microM) was co-perfused with histamine and pilocarpine. In the absence of BBB permeability enhancing drugs, pilocarpine induced epileptiform activity only after arterial perfusion at concentrations >10 mM. Ictal discharges correlated with a high intracerebral pilocarpine concentration measured by high pressure liquid chromatography. We propose that acute epileptiform discharges induced by pilocarpine treatment in the in vitro isolated brain preparation are mediated by a dose-dependent, atropine-sensitive muscarinic effect promoted by an increase in BBB permeability. Pilocarpine accumulation secondary to BBB permeability changes may contribute to in vivo ictogenesis in the pilocarpine epilepsy model.

Purpose: Brain inflammation has been recently considered in the pathogenesis of focal epilepsies. Synthesis of pro-inflammatory mediators in the brain was described both in experimental models of seizures and in human postsurgical tissue. Inflammatory mediators may up-regulate endothelial adhesion molecules, therefore promoting adhesion and homing of leucocytes into the brain. In the present study, expression of inducible adhesion factors in brain endothelium was verified after pharmacological induction of seizure-like activity in specific brain areas of the in vitro isolated guinea pig brain., Methods: Experiments were performed in isolated guinea-pig brains maintained in vitro by arterial perfusion. In this preparation, brief application of the GABAa receptor-antagonist, bicuculline, consistently induced focal ictal discharges in the limbic region that secondarily diffuse to the neocortex, as verified by simultaneous electrophysiological recording of extracellular activity. At the end of the electrophysiological experiment (after 5 h in vitro), brains were fixed and immunostaining for adhesion molecules P-selectin and ICAM-1 and for Fos protein was evaluated., Results: Immunohistochemical analysis of isolated brains in which seizure-like activity was induced revealed expression of inducible adhesion factors P-selectin and ICAM-1 in the endothelium of small-medium size brain vessels. In particular, the expression of these molecules was consistently observed in all areas involved in epileptic seizure-like ictal activity (limbic cortices and neocortex), and was infrequently found in regions that generated interictal spiking (piriform cortex), suggesting a trigger role played by seizures for endothelial activation. An increase in Fos protein expression was evident in all analyzed limbic areas and in the neocortex, indicating a correlation between the areas of neuronal and endothelial activation. In control brains maintained in vitro for comparable times without induction of epileptiform activity, no immunoreactivity for Fos and adhesion molecules was observed., Conclusions: Seizure-like activity in an in vitro isolated brain preparation induces the expression of adhesion molecules in the cerebral endothelium. These observations indicate that local endothelial activation may represent a crucial step for the development of an inflammatory response induced by seizures, and suggest a possible novel pathogenic mechanism during the process of epileptogenesis.

Mechanisms underlying epileptic activities recorded from entorhinal cortex (EC) were studied through a computational model based on review of cytoarchitectonic and neurobiological data about this structure. The purpose of this study is to describe and use this model to interpret epileptiform discharge patterns recorded in an experimental model of ictogenesis (guinea pig isolated brain perfused with bicuculline). A macroscopic modeling approach representing synaptic interactions between cells subpopulations in the EC was chosen for its adequacy to mimic field potentials reflecting overall dynamics rising from interconnected cells populations. Therefore intrinsic properties of neurons were not included in the modeling design. Model parameters were adjusted from an identification procedure based on quantitative comparison between real and simulated signals. For both EC deep and superficial layers, results show that the model generates very realistic signals regarding temporal dynamics, spectral features, and cross-correlation values. These simulations allowed us to infer information about the evolution of synaptic transmission between principal cell and interneuronal populations and about connectivity between deep and superficial layers during the transition from background to ictal activity. In the model, this transition was obtained for increased excitation in deep versus superficial layers. Transitions between epileptiform activities [interictal spikes, fast onset activity (25 Hz), ictal bursting activity] were explained by changes of parameters mainly related to GABAergic interactions. Notably, the model predicted an important role of GABAa,fast- and GABAb-receptor-mediated inhibition in the generation of ictal fast onset and burst activities, respectively. These findings are discussed with respect to experimental data.

Purpose: Aim of the study is to investigate the involvement of parahippocampal subregions in the generation and in the propagation of focal epileptiform discharges in an acute model of seizure generation in the temporal lobe induced by arterial application of bicuculline in the in vitro isolated guinea pig brain preparation., Methods: Electrophysiological recordings were simultaneously performed with single electrodes and multichannel silicon probes in the entorhinal, perirhinal, and piriform cortices and in the area CA1 of the hippocampus of the in vitro isolated guinea pig brain. Interictal and ictal epileptiform discharges restricted to the temporal region were induced by a brief (3-5 min) arterial perfusion of the GABA(A) receptor antagonist, bicuculline methiodide (50 microM). Current source density analysis of laminar field profiles performed with the silicon probes was carried out at different sites to establish network interactions responsible for the generation of epileptiform potentials. Nonlinear regression analysis was conducted on extracellular recordings during ictal onset in order to quantify the degree of interaction between fast activities generated at different sites, as well as time delays., Results: Experiments were performed in 31 isolated guinea pig brains. Bicuculline-induced interictal and ictal epileptiform activities that showed variability of spatial propagation and time course in the olfactory-temporal region. The most commonly observed pattern (n = 23) was characterized by the initial appearance of interictal spikes (ISs) in the piriform cortex (PC), which propagated to the lateral entorhinal region. Independent and asynchronous preictal spikes originated in the entorhinal cortex (EC)/hippocampus and progressed into ictal fast discharges (around 25 Hz) restricted to the entorhinal/hippocampal region. The local generation of fast activity was verified and confirmed both by CSD and phase shift analysis performed on laminar profiles. Fast activity was followed by synchronous afterdischarges that propagated to the perirhinal cortex (PRC) (but not to the PC). Within 1-9 min, the ictal discharge ceased and a postictal period of depression occurred, after which periodic ISs in the PC resumed. Unlike preictal ISs, postictal ISs propagated to the PRC., Conclusions: Several studies proposed that reciprocal connections between the entorhinal and the PRC are under a very efficient inhibitory control (1). We report that ISs determined by acute bicuculline treatment in the isolated guinea pig brain progress from the PC to the hippocampus/EC just before ictal onset. Ictal discharges are characterized by a peculiar pattern of fast activity that originates from the entorhinal/hippocampal region and only secondarily propagates to the PRC. Postictal propagation of ISs to the PRC occurred exclusively when an ictal discharge was generated in the hippocampal/entorhinal region. The results suggest that reiteration of ictal events may promote changes in propagation pattern of epileptiform discharges that could act as trigger elements in the development of temporal lobe epilepsy.

Interictal potentials are commonly observed between seizures in human epilepsies and in animal models of epilepsy. It is uncertain whether interictal spiking in partial epilepsies is causally related with the onset of an ictal discharge. To analyze the reciprocal correlation between interictal and ictal epileptiform events, we performed extracellular recordings in the limbic system of the in vitro isolated guinea pig brain preparation. Arterial perfusion of bicuculline (50 microM) in vitro consistently induced a focal ictal discharge in the hippocampal-entorhinal region that in one third of the experiments was associated with periodic interictal spikes in the piriform cortex. In the absence of active interictal spiking, the piriform cortex was secondarily invaded by the ictal discharge initiated in the hippocampal-entorhinal region, whereas no secondary ictal entrainment was observed in the presence of periodic piriform cortex spikes at circa 0.1 to 0.2 Hz. Similarly, ictal events never occurred when arterial perfusion of bicuculline was preceded by a local injection of the same drug in the piriform cortex, a procedure that induces a sustained interictal spiking. A reduced responsiveness to incoming paroxysmal discharges generated in the hippocampus was observed during the interval between two interictal spikes in the piriform cortex.

Interictal spikes (ISs) are typically observed between seizures in focal epilepsies. Whether ISs are causally involved or represent protective elements in the transition toward an ictal discharge is an open question. Previous studies suggested that inhibition or disfacilitation occurs during the period elapsing between two ISs induced by local application of either bicuculline or penicillin in the piriform cortex of the in vitro isolated guinea pig brain preparation. We further investigated this issue by studying responses to afferent stimulation during the interspike period (6.3 +/- 2.5 s; mean +/- SD). Properly set stimulation intensity of the lateral olfactory tract resets ISs exclusively (and not before) 4-10 s (5.6 +/- 2.0 s; mean +/- SD) after a preceding spontaneous spike. This finding demonstrates the existence of a period of enhanced threshold to stimulus-evoked activation that coincides with the interspike interval in the absence of stimulation. Current source density analysis of depth laminar profiles demonstrated that both stimulus-evoked and spontaneous ISs were generated by the activation of an identical cortical circuit. Our study suggests that interictal spiking could play a protective role or at least provide an effective restraint against the onset of a focal ictal discharge.