mTOR and neuroinflammation in epilepsy: implications for disease progression and treatment.

Epilepsy remains a major health concern as anti-seizure medications frequently fail, and there is currently no treatment to stop or prevent epileptogenesis, the process underlying the onset and progression of epilepsy. The identification of the pathological processes underlying epileptogenesis is instrumental to the development of drugs that may prevent the generation of seizures or control pharmaco-resistant seizures, which affect about 30% of patients. mTOR signalling and neuroinflammation have been recognized as critical pathways that are activated in brain cells in epilepsy. They represent a potential node of biological convergence in structural epilepsies with either a genetic or an acquired aetiology. Interventional studies in animal models and clinical studies give strong support to the involvement of each pathway in epilepsy. In this Review, we focus on available knowledge about the pathophysiological features of mTOR signalling and the neuroinflammatory brain response, and their interactions, in epilepsy. We discuss mitigation strategies for each pathway that display therapeutic effects in experimental and clinical epilepsy. A deeper understanding of these interconnected molecular cascades could enhance our strategies for managing epilepsy. This could pave the way for new treatments to fill the gaps in the development of preventative or disease-modifying drugs, thus overcoming the limitations of current symptomatic medications.

Identification of an epilepsy-linked gut microbiota signature in a pediatric rat model of acquired epilepsy.

A dysfunctional gut microbiota-brain axis is emerging as a potential pathogenic mechanism in epilepsy, particularly in pediatric forms of epilepsy. To add new insights into gut-related changes in acquired epilepsy that develops early in life, we used a multi-omics approach in a rat model with a 56% incidence of epilepsy. The presence of spontaneous seizures was assessed in adult rats (n = 46) 5 months after status epilepticus induced by intra-amygdala kainate at postnatal day 13, by 2 weeks (24/7) ECoG monitoring. Twenty-six rats developed epilepsy (Epi) while the remaining 20 rats (No-Epi) did not show spontaneous seizures. At the end of ECoG monitoring, all rats and their sham controls (n = 20) were sacrificed for quantitative histopathological and immunohistochemical analyses of the gut structure, glia and macrophages, as well as RTqPCR analysis of inflammation/oxidative stress markers. By comparing Epi, No-Epi rats, and sham controls, we found structural, cellular, and molecular alterations reflecting a dysfunctional gut, which were specifically associated with epilepsy. In particular, the villus height-to-crypt depth ratio and number of Goblet cells were reduced in the duodenum of Epi rats vs both No-Epi rats and sham controls (p < 0.01). Villus height and crypt depth in the duodenum and jejunum (p < 0.01) were increased in No-Epi vs both Epi and sham controls. We also detected enhanced Iba1-positive macrophages, together with increased IL1b and NFE2L2 transcripts and TNF protein, in the small intestine of Epi vs both No-Epi and sham control rats (p < 0.01), denoting the presence of inflammation and oxidative stress. Astroglial GFAP-immunostaining was similar in all experimental groups. Metagenomic analysis in the feces collected 5 months after status epilepticus showed that the ratio of two dominant phyla (Bacteroidota-to-Firmicutes) was similarly increased in Epi and No-Epi rats vs sham control rats. Notably, the relative abundance of families, genera, and species associated with SCFA production differed in Epi vs No-Epi rats, describing a bacterial imprint associated with epilepsy. Furthermore, Epi rats showed a blood metabolic signature characterized by changes in lipid metabolism compared to both No-Epi and sham control rats. Our study provides new evidence of long-term gut alterations, along with microbiota-related metabolic changes, occurring specifically in rats that develop epilepsy after brain injury early in life.

Early treatment with rifaximin during epileptogenesis reverses gut alterations and reduces seizure duration in a mouse model of acquired epilepsy.

The gut microbiota is altered in epilepsy and is emerging as a potential target for new therapies. We studied the effects of rifaximin, a gastrointestinal tract-specific antibiotic, on seizures and neuropathology and on alterations in the gut and its microbiota in a mouse model of temporal lobe epilepsy (TLE). Epilepsy was induced by intra-amygdala kainate injection causing status epilepticus (SE) in C57Bl6 adult male mice. Sham mice were injected with vehicle. Two cohorts of SE mice were fed a rifaximin-supplemented diet for 21 days, starting either at 24 h post-SE (early disease stage) or at day 51 post-SE (chronic disease stage). Corresponding groups of SE mice (one each disease stage) were fed a standard (control) diet. Cortical ECoG recording was done at each disease stage (24/7) for 21 days in all SE mice to measure the number and duration of spontaneous seizures during either rifaximin treatment or control diet. Then, epileptic mice ± rifaximin and respective sham mice were sacrificed and brain, gut and feces collected. Biospecimens were used for: (i) quantitative histological analysis of the gut structural and cellular components; (ii) markers of gut inflammation and intestinal barrier integrity by RTqPCR; (iii) 16S rRNA metagenomics analysis in feces. Hippocampal neuronal cell loss was assessed in epileptic mice killed in the early disease phase. Rifaximin administered for 21 days post-SE (early disease stage) reduced seizure duration (p < 0.01) and prevented hilar mossy cells loss in the hippocampus compared to epileptic mice fed a control diet. Epileptic mice fed a control diet showed a reduction of both villus height and villus height/crypt depth ratio (p < 0.01) and a decreased number of goblet cells (p < 0.01) in the duodenum, as well as increased macrophage (Iba1)-immunostaining in the jejunum (p < 0.05), compared to respective sham mice. Rifaximin's effect on seizures was associated with a reversal of gut structural and cellular changes, except for goblet cells which remained reduced. Seizure duration in epileptic mice was negatively correlated with the number of mossy cells (p < 0.01) and with villus height/crypt depth ratio (p < 0.05). Rifaximin-treated epileptic mice also showed increased tight junctions (occludin and ZO-1, p < 0.01) and decreased TNF mRNA expression (p < 0.01) in the duodenum compared to epileptic mice fed a control diet. Rifaximin administered for 21 days in chronic epileptic mice (chronic disease stage) did not change the number or duration of seizures compared to epileptic mice fed a control diet. Chronic epileptic mice fed a control diet showed an increased crypt depth (p < 0.05) and reduced villus height/crypt depth ratio (p < 0.01) compared to respective sham mice. Rifaximin treatment did not affect these intestinal changes. At both disease stages, rifaximin modified α- and ß-diversity in epileptic and sham mice compared to respective mice fed a control diet. The microbiota composition in epileptic mice, as well as the effects of rifaximin at the phylum, family and genus levels, depended on the stage of the disease. During the early disease phase, the abundance of specific taxa was positively correlated with seizure duration in epileptic mice. In conclusion, gut-related alterations reflecting a dysfunctional state, occur during epilepsy development in a TLE mouse model. A short-term treatment with rifaximin during the early phase of the disease, reduced seizure duration and neuropathology, and reversed some intestinal changes, strengthening the therapeutic effects of gut-based therapies in epilepsy.

Neuroinflammation microenvironment sharpens seizure circuit.

A large set of inflammatory molecules and their receptors are induced in epileptogenic foci of patients with pharmacoresistant epilepsies of structural etiologies or with refractory status epilepticus. Studies in animal models mimicking these clinical conditions have shown that the activation of specific inflammatory signallings in forebrain neurons or glial cells may modify seizure thresholds, thus contributing to both ictogenesis and epileptogenesis. The search for mechanisms underlying these effects has highlighted that inflammatory mediators have CNS-specific neuromodulatory functions, in addition to their canonical activation of immune responses for pathogen recognition and clearance. This review reports the neuromodulatory effects of inflammatory mediators and how they contribute to alter the inhibitory/excitatory balance in neural networks that underlie seizures. In particular, we describe key findings related to the ictogenic role of prototypical inflammatory cytokines (IL-1ß and TNF) and danger signals (HMGB1), their modulatory effects of neuronal excitability, and the mechanisms underlying these effects. It will be discussed how harnessing these neuromodulatory properties of immune mediators may lead to novel therapies to control drug-resistant seizures.

ECoG spiking activity and signal dimension are early predictive measures of epileptogenesis in a translational mouse model of traumatic brain injury.

The latency between traumatic brain injury (TBI) and the onset of epilepsy (PTE) represents an opportunity for counteracting epileptogenesis. Antiepileptogenesis trials are hampered by the lack of sensitive biomarkers that allow to enrich patient's population at-risk for PTE. We aimed to assess whether specific ECoG signals predict PTE in a clinically relevant mouse model with ∼60% epilepsy incidence. TBI was provoked in adult CD1 male mice by controlled cortical impact on the left parieto-temporal cortex, then mice were implanted with two perilesional cortical screw electrodes and two similar electrodes in the hemisphere contralateral to the lesion site. Acute seizures and spikes/sharp waves were ECoG-recorded during 1 week post-TBI. These early ECoG events were analyzed according to PTE incidence as assessed by measuring spontaneous recurrent seizures (SRS) at 5 months post-TBI. We found that incidence, number and duration of acute seizures during 3 days post-TBI were similar in PTE mice and mice not developing epilepsy (No SRS mice). Control mice with cortical electrodes (naïve, n = 5) or with electrodes and craniotomy (sham, n = 5) exhibited acute seizures but did not develop epilepsy. The daily number of spikes/sharp waves at the perilesional electrodes was increased similarly in PTE (n = 15) and No SRS (n = 8) mice vs controls (p < 0.05, n = 10) from day 2 post-injury. Differently, the daily number of spikes/sharp waves at both contralateral electrodes showed a progressive increase in PTE mice vs No SRS and control mice. In particular, spikes number was higher in PTE vs No SRS mice (p < 0.05) at 6 and 7 days post-TBI, and this measure predicted epilepsy development with high accuracy (AUC = 0.77, p = 0.03; CI 0.5830-0.9670). The cut-off value was validated in an independent cohort of TBI mice (n = 12). The daily spike number at the contralateral electrodes showed a circadian distribution in PTE mice which was not observed in No SRS mice. Analysis of non-linear dynamics at each electrode site showed changes in dimensionality during 4 days post-TBI. This measure yielded the best discrimination between PTE and No SRS mice (p < 0.01) at the cortical electrodes contralateral to injury. Data show that epileptiform activity contralateral to the lesion site has the the highest predictive value for PTE in this model reinforcing the hypothesis that the hemisphere contralateral to the lesion core may drive epileptogenic networks after TBI.

Lipid mediator n-3 docosapentaenoic acid-derived protectin D1 enhances synaptic inhibition of hippocampal principal neurons by interaction with a G-protein-coupled receptor.

Epilepsy is a severe neurological disease manifested by spontaneous recurrent seizures due to abnormal hyper-synchronization of neuronal activity. Epilepsy affects about 1% of the population and up to 40% of patients experience seizures that are resistant to currently available drugs, thus highlighting an urgent need for novel treatments. In this regard, anti-inflammatory drugs emerged as potential therapeutic candidates. In particular, specific molecules apt to resolve the neuroinflammatory response occurring in acquired epilepsies have been proven to counteract seizures in experimental models, and humans. One candidate investigational molecule has been recently identified as the lipid mediator n-3 docosapentaenoic acid-derived protectin D1 (PD1n-3DPA ) which significantly reduced seizures, cell loss, and cognitive deficit in a mouse model of acquired epilepsy. However, the mechanisms that mediate the PD1n-3DPA effect remain elusive. We here addressed whether PD1n-3DPA has direct effects on neuronal activity independent of its anti-inflammatory action. We incubated, therefore, hippocampal slices with PD1n-3DPA and investigated its effect on excitatory and inhibitory synaptic inputs to the CA1 pyramidal neurons. We demonstrate that inhibitory drive onto the perisomatic region of the pyramidal neurons is increased by PD1n-3DPA , and this effect is mediated by pertussis toxin-sensitive G-protein coupled receptors. Our data indicate that PD1n-3DPA acts directly on inhibitory transmission, most likely at the presynaptic site of inhibitory synapses as also supported by Xenopus oocytes and immunohistochemical experiments. Thus, in addition to its anti-inflammatory effects, PD1n-3DPA anti-seizure and neuroprotective effects may be mediated by its direct action on neuronal excitability by modulating their synaptic inputs.

Review and standard operating procedures for collection of biospecimens and analysis of biomarkers in new onset refractory status epilepticus.

New onset refractory status epilepticus (NORSE), including its subtype with a preceding febrile illness known as febrile infection-related epilepsy syndrome (FIRES), is one of the most severe forms of status epilepticus. The exact causes of NORSE are currently unknown, and there is so far no disease-specific therapy. Identifying the underlying pathophysiology and discovering specific biomarkers, whether immunologic, infectious, genetic, or other, may help physicians in the management of patients with NORSE. A broad spectrum of biomarkers has been proposed for status epilepticus patients, some of which were evaluated for patients with NORSE. Nonetheless, none has been validated, due to significant variabilities in study cohorts, collected biospecimens, applied analytical methods, and defined outcome endpoints, and to small sample sizes. The NORSE Institute established an open NORSE/FIRES biorepository for health-related data and biological samples allowing the collection of biospecimens worldwide, promoting multicenter research and sharing of data and specimens. Here, we suggest standard operating procedures for biospecimen collection and biobanking in this rare condition. We also propose criteria for the appropriate use of previously collected biospecimens. We predict that the widespread use of standardized procedures will reduce heterogeneity, facilitate the future identification of validated biomarkers for NORSE, and provide a better understanding of the pathophysiology and best clinical management for these patients.

Neuroimmunology of status epilepticus.

Status epilepticus (SE) is a very heterogeneous clinical condition often refractory to available treatment options. Evidence in animal models shows that neuroinflammation arises in the brain during SE due to the activation of innate immune mechanisms in brain parenchyma cells. Intervention studies in animal models support the involvement of neuroinflammation in SE onset, duration, and severity, refractoriness to treatments, and long-term neurological consequences. Clinical evidence shows that neuroinflammation occurs in patients with SE of diverse etiologies likely representing a common phenomenon, thus broadening the involvement of the immune system beyond the infective and autoimmune etiologies. There is urgent need for novel therapies for refractory SE that rely upon a better understanding of the basic mechanisms underlying this clinical condition. Preclinical and clinical evidence encourage consideration of specific anti-inflammatory treatments for controlling SE and its consequences in patients.

Drug Resistance in Epilepsy: Clinical Impact, Potential Mechanisms, and New Innovative Treatment Options.

Epilepsy is a chronic neurologic disorder that affects over 70 million people worldwide. Despite the availability of over 20 antiseizure drugs (ASDs) for symptomatic treatment of epileptic seizures, about one-third of patients with epilepsy have seizures refractory to pharmacotherapy. Patients with such drug-resistant epilepsy (DRE) have increased risks of premature death, injuries, psychosocial dysfunction, and a reduced quality of life, so development of more effective therapies is an urgent clinical need. However, the various types of epilepsy and seizures and the complex temporal patterns of refractoriness complicate the issue. Furthermore, the underlying mechanisms of DRE are not fully understood, though recent work has begun to shape our understanding more clearly. Experimental models of DRE offer opportunities to discover, characterize, and challenge putative mechanisms of drug resistance. Furthermore, such preclinical models are important in developing therapies that may overcome drug resistance. Here, we will review the current understanding of the molecular, genetic, and structural mechanisms of ASD resistance and discuss how to overcome this problem. Encouragingly, better elucidation of the pathophysiological mechanisms underpinning epilepsies and drug resistance by concerted preclinical and clinical efforts have recently enabled a revised approach to the development of more promising therapies, including numerous potential etiology-specific drugs ("precision medicine") for severe pediatric (monogenetic) epilepsies and novel multitargeted ASDs for acquired partial epilepsies, suggesting that the long hoped-for breakthrough in therapy for as-yet ASD-resistant patients is a feasible goal. SIGNIFICANCE STATEMENT: Drug resistance provides a major challenge in epilepsy management. Here, we will review the current understanding of the molecular, genetic, and structural mechanisms of drug resistance in epilepsy and discuss how the problem might be overcome.

Cholesterol 24-hydroxylase is a novel pharmacological target for anti-ictogenic and disease modification effects in epilepsy.

Therapies for epilepsy mainly provide symptomatic control of seizures since most of the available drugs do not target disease mechanisms. Moreover, about one-third of patients fail to achieve seizure control. To address the clinical need for disease-modifying therapies, research should focus on targets which permit interventions finely balanced between optimal efficacy and safety. One potential candidate is the brain-specific enzyme cholesterol 24-hydroxylase. This enzyme converts cholesterol to 24S-hydroxycholesterol, a metabolite which among its biological roles modulates neuronal functions relevant for hyperexcitability underlying seizures. To study the role of cholesterol 24-hydroxylase in epileptogenesis, we administered soticlestat (TAK-935/OV935), a potent and selective brain-penetrant inhibitor of the enzyme, during the early disease phase in a mouse model of acquired epilepsy using a clinically relevant dose. During soticlestat treatment, the onset of epilepsy was delayed and the number of ensuing seizures was decreased by about 3-fold compared to vehicle-treated mice, as assessed by EEG monitoring. Notably, the therapeutic effect was maintained 6.5 weeks after drug wash-out when seizure number was reduced by about 4-fold and their duration by 2-fold. Soticlestat-treated mice showed neuroprotection of hippocampal CA1 neurons and hilar mossy cells as assessed by post-mortem brain histology. High throughput RNA-sequencing of hippocampal neurons and glia in mice treated with soticlestat during epileptogenesis showed that inhibition of cholesterol 24-hydroxylase did not directly affect the epileptogenic transcriptional network, but rather modulated a non-overlapping set of genes that might oppose the pathogenic mechanisms of the disease. In human temporal lobe epileptic foci, we determined that cholesterol 24-hydroxylase expression trends higher in neurons, similarly to epileptic mice, while the enzyme is ectopically induced in astrocytes compared to control specimens. Soticlestat reduced significantly the number of spontaneous seizures in chronic epileptic mice when was administered during established epilepsy. Data show that cholesterol 24-hydroxylase contributes to spontaneous seizures and is involved in disease progression, thus it represents a novel target for chronic seizures inhibition and disease-modification therapy in epilepsy.

High-mobility group box 1 as a predictive biomarker for drug-resistant epilepsy: A proof-of-concept study.

Currently no sensitive and specific biomarkers exist to predict drug-resistant epilepsy. We determined whether blood levels of high-mobility group box 1 (HMGB1), a mediator of neuroinflammation implicated in drug-resistant epilepsies, identifies patients with drug-resistant seizures. Patients with drug-resistant epilepsy express significantly higher levels of blood HMGB1 than those with drug-responsive, well-controlled seizures and healthy controls. No correlation existed between blood HMGB1 levels and total pretreatment seizure count or days since last seizure at new epilepsy diagnosis, indicating that blood HMGB1 does not solely reflect ongoing seizures. HMGB1 distinguishes with high specificity and selectivity drug-resistant versus drug-responsive patients. This protein therefore has potential clinical utility to act as a biomarker for predicting response to therapy, which should be addressed in prospective clinical studies.

The association between systemic autoimmune disorders and epilepsy and its clinical implications.

Systemic autoimmune disorders occur more frequently in patients with epilepsy than in the general population, suggesting shared disease mechanisms. The risk of epilepsy is elevated across the spectrum of systemic autoimmune disorders but is highest in systemic lupus erythematosus and type 1 diabetes mellitus. Vascular and metabolic factors are the most important mediators between systemic autoimmune disorders and epilepsy. Systemic immune dysfunction can also affect neuronal excitability, not only through innate immune activation and blood-brain barrier dysfunction in most epilepsies but also adaptive immunity in autoimmune encephalitis. The presence of systemic autoimmune disorders in subjects with acute seizures warrants evaluation for infectious, vascular, toxic and metabolic causes of acute symptomatic seizures, but clinical signs of autoimmune encephalitis should not be missed. Immunosuppressive medications may have antiseizure properties and trigger certain drug interactions with antiseizure treatments. A better understanding of mechanisms underlying the co-existence of epilepsy and systemic autoimmune disorders is needed to guide new antiseizure and anti-epileptogenic treatments. This review aims to summarize the epidemiological evidence for systemic autoimmune disorders as comorbidities of epilepsy, explore potential immune and non-immune mechanisms, and provide practical implications on diagnostic and therapeutic approach to epilepsy in those with comorbid systemic autoimmune disorders.

CXCL1-CXCR1/2 signaling is induced in human temporal lobe epilepsy and contributes to seizures in a murine model of acquired epilepsy.

CXCL1, a functional murine orthologue of the human chemokine CXCL8 (IL-8), and its CXCR1 and CXCR2 receptors were investigated in a murine model of acquired epilepsy developing following status epilepticus (SE) induced by intra-amygdala kainate. CXCL8 and its receptors were also studied in human temporal lobe epilepsy (TLE). The functional involvement of the chemokine in seizure generation and neuronal cell loss was assessed in mice using reparixin (formerly referred to as repertaxin), a non-competitive allosteric inhibitor of CXCR1/2 receptors. We found a significant increase in hippocampal CXCL1 level within 24 h of SE onset that lasted for at least 1 week. No changes were measured in blood. In analogy with human TLE, immunohistochemistry in epileptic mice showed that CXCL1 and its two receptors were increased in hippocampal neuronal cells. Additional expression of these molecules was found in glia in human TLE. Mice were treated with reparixin or vehicle during SE and for additional 6 days thereafter, using subcutaneous osmotic minipumps. Drug-treated mice showed a faster SE decay, a reduced incidence of acute symptomatic seizures during 48 h post-SE, and a delayed time to spontaneous seizures onset compared to vehicle controls. Upon reparixin discontinuation, mice developed spontaneous seizures similar to vehicle mice, as shown by EEG monitoring at 14 days and 2.5 months post-SE. In the same epileptic mice, reparixin reduced neuronal cell loss in the hippocampus vs vehicle-injected mice, as assessed by Nissl staining at completion of EEG monitoring. Reparixin administration for 2 weeks in mice with established chronic seizures, reduced by 2-fold on average seizure number vs pre-treatment baseline, and this effect was reversible upon drug discontinuation. No significant changes in seizure number were measured in vehicle-injected epileptic mice that were EEG monitored in parallel. Data show that CXCL1-IL-8 signaling is activated in experimental and human epilepsy and contributes to acute and chronic seizures in mice, therefore representing a potential new target to attain anti-ictogenic effects.

Chromosome 14 deletions, rings, and epilepsy genes: A riddle wrapped in a mystery inside an enigma.

The ring 14 syndrome is a rare condition caused by the rearrangement of one chromosome 14 into a ring-like structure. The formation of the ring requires two breakpoints and loss of material from the short and long arms of the chromosome. Like many other chromosome syndromes, it is characterized by multiple congenital anomalies and developmental delays. Typical of the condition are retinal anomalies and drug-resistant epilepsy. These latter manifestations are not found in individuals who are carriers of comparable 14q deletions without formation of a ring (linear deletions). To find an explanation for this apparent discrepancy and gain insight into the mechanisms leading to seizures, we reviewed and compared literature cases of both ring and linear deletion syndrome with respect to both their clinical manifestations and the role and function of potentially epileptogenic genes. Knowledge of the epilepsy-related genes in chromosome 14 is an important premise for the search of new and effective drugs to combat seizures. Current clinical and molecular evidence is not sufficient to explain the known discrepancies between ring and linear deletions.

Microglia proliferation plays distinct roles in acquired epilepsy depending on disease stages.

OBJECTIVE: Microgliosis occurs in animal models of acquired epilepsy and in patients. It includes cell proliferation that is associated with seizure frequency and decreased neuronal cells in human epilepsy. The role of microglia proliferation in the development of acquired epilepsy is unknown; thus, we examined its contribution to spontaneous seizure, neurodegeneration, and cognitive deficits in different disease phases. METHODS: We used a model of acquired epilepsy triggered by intra-amygdala kainic acid in C57BL6N adult male mice. Mice were electroencephalographically (EEG) monitored (24/7) during status epilepticus and in early and chronic disease. Microglia proliferation was blocked by GW2580, a selective CSF1 receptor inhibitor, supplemented in the diet for 21 days from status epilepticus onset. Then, mice were returned to placebo diet until experiment completion. Control mice were exposed to status epilepticus and fed with placebo diet. Experimental mice were tested in the novel object recognition test (NORT) and in Barnes maze, and compared to control and sham mice. At the end of the behavioral test, mice were killed for brain histopathological analysis. Additionally, seizure baseline was monitored in chronic epileptic mice, then mice were fed for 14 days with GW2580 or placebo diet under 24/7 EEG recording. RESULTS: GW2580 prevented microglia proliferation in mice undergoing epilepsy, whereas it did not affect microglia or basal excitatory neurotransmission in the hippocampus of naive mice. Mice with occluded microglia proliferation during early disease development underwent status epilepticus and subsequent epilepsy similar to placebo diet mice, and were similarly impaired in NORT, with improvement in Barnes maze. GW2580-treated mice displayed neuroprotection in the hippocampus. In contrast, blockade of microglia proliferation in chronic epileptic mice resulted in spontaneous seizure reduction versus placebo mice. SIGNIFICANCE: Microglia proliferation during early disease contributes to neurodegeneration, whereas in late chronic disease it contributes to seizures. Timely pharmacological interference with microglia proliferation may offer a potential target for improving disease outcomes.

Climate change and epilepsy: Insights from clinical and basic science studies.

Climate change is with us. As professionals who place value on evidence-based practice, climate change is something we cannot ignore. The current pandemic of the novel coronavirus, SARS-CoV-2, has demonstrated how global crises can arise suddenly and have a significant impact on public health. Global warming, a chronic process punctuated by acute episodes of extreme weather events, is an insidious global health crisis needing at least as much attention. Many neurological diseases are complex chronic conditions influenced at many levels by changes in the environment. This review aimed to collate and evaluate reports from clinical and basic science about the relationship between climate change and epilepsy. The keywords climate change, seasonal variation, temperature, humidity, thermoregulation, biorhythm, gene, circadian rhythm, heat, and weather were used to search the published evidence. A number of climatic variables are associated with increased seizure frequency in people with epilepsy. Climate change-induced increase in seizure precipitants such as fevers, stress, and sleep deprivation (e.g. as a result of more frequent extreme weather events) or vector-borne infections may trigger or exacerbate seizures, lead to deterioration of seizure control, and affect neurological, cerebrovascular, or cardiovascular comorbidities and risk of sudden unexpected death in epilepsy. Risks are likely to be modified by many factors, ranging from individual genetic variation and temperature-dependent channel function, to housing quality and global supply chains. According to the results of the limited number of experimental studies with animal models of seizures or epilepsy, different seizure types appear to have distinct susceptibility to seasonal influences. Increased body temperature, whether in the context of fever or not, has a critical role in seizure threshold and seizure-related brain damage. Links between climate change and epilepsy are likely to be multifactorial, complex, and often indirect, which makes predictions difficult. We need more data on possible climate-driven altered risks for seizures, epilepsy, and epileptogenesis, to identify underlying mechanisms at systems, cellular, and molecular levels for better understanding of the impact of climate change on epilepsy. Further focussed data would help us to develop evidence for mitigation methods to do more to protect people with epilepsy from the effects of climate change.

Targeting Oxidative Stress with Antioxidant Duotherapy after Experimental Traumatic Brain Injury.

We assessed the effect of antioxidant therapy using the Food and Drug Administration-approved respiratory drug N-acetylcysteine (NAC) or sulforaphane (SFN) as monotherapies or duotherapy in vitro in neuron-BV2 microglial co-cultures and validated the results in a lateral fluid-percussion model of TBI in rats. As in vitro measures, we assessed neuronal viability by microtubule-associated-protein 2 immunostaining, neuroinflammation by monitoring tumor necrosis factor (TNF) levels, and neurotoxicity by measuring nitrite levels. In vitro, duotherapy with NAC and SFN reduced nitrite levels to 40% (p < 0.001) and neuroinflammation to -29% (p < 0.001) compared with untreated culture. The treatment also improved neuronal viability up to 72% of that in a positive control (p < 0.001). The effect of NAC was negligible, however, compared with SFN. In vivo, antioxidant duotherapy slightly improved performance in the beam walking test. Interestingly, duotherapy treatment decreased the plasma interleukin-6 and TNF levels in sham-operated controls (p < 0.05). After TBI, no treatment effect on HMGB1 or plasma cytokine levels was detected. Also, no treatment effects on the composite neuroscore or cortical lesion area were detected. The robust favorable effect of duotherapy on neuroprotection, neuroinflammation, and oxidative stress in neuron-BV2 microglial co-cultures translated to modest favorable in vivo effects in a severe TBI model.

The ENIGMA-Epilepsy working group: Mapping disease from large data sets.

Epilepsy is a common and serious neurological disorder, with many different constituent conditions characterized by their electro clinical, imaging, and genetic features. MRI has been fundamental in advancing our understanding of brain processes in the epilepsies. Smaller-scale studies have identified many interesting imaging phenomena, with implications both for understanding pathophysiology and improving clinical care. Through the infrastructure and concepts now well-established by the ENIGMA Consortium, ENIGMA-Epilepsy was established to strengthen epilepsy neuroscience by greatly increasing sample sizes, leveraging ideas and methods established in other ENIGMA projects, and generating a body of collaborating scientists and clinicians to drive forward robust research. Here we review published, current, and future projects, that include structural MRI, diffusion tensor imaging (DTI), and resting state functional MRI (rsfMRI), and that employ advanced methods including structural covariance, and event-based modeling analysis. We explore age of onset- and duration-related features, as well as phenomena-specific work focusing on particular epilepsy syndromes or phenotypes, multimodal analyses focused on understanding the biology of disease progression, and deep learning approaches. We encourage groups who may be interested in participating to make contact to further grow and develop ENIGMA-Epilepsy.

Repurposed molecules for antiepileptogenesis: Missing an opportunity to prevent epilepsy?

Prevention of epilepsy is a great unmet need. Acute central nervous system (CNS) insults such as traumatic brain injury (TBI), cerebrovascular accidents (CVA), and CNS infections account for 15%-20% of all epilepsy. Following TBI and CVA, there is a latency of days to years before epilepsy develops. This allows treatment to prevent or modify postinjury epilepsy. No such treatment exists. In animal models of acquired epilepsy, a number of medications in clinical use for diverse indications have been shown to have antiepileptogenic or disease-modifying effects, including medications with excellent side effect profiles. These include atorvastatin, ceftriaxone, losartan, isoflurane, N-acetylcysteine, and the antiseizure medications levetiracetam, brivaracetam, topiramate, gabapentin, pregabalin, vigabatrin, and eslicarbazepine acetate. In addition, there are preclinical antiepileptogenic data for anakinra, rapamycin, fingolimod, and erythropoietin, although these medications have potential for more serious side effects. However, except for vigabatrin, there have been almost no translation studies to prevent or modify epilepsy using these potentially "repurposable" medications. We may be missing an opportunity to develop preventive treatment for epilepsy by not evaluating these medications clinically. One reason for the lack of translation studies is that the preclinical data for most of these medications are disparate in terms of types of injury, models within different injury type, dosing, injury-treatment initiation latencies, treatment duration, and epilepsy outcome evaluation mode and duration. This makes it difficult to compare the relative strength of antiepileptogenic evidence across the molecules, and difficult to determine which drug(s) would be the best to evaluate clinically. Furthermore, most preclinical antiepileptogenic studies lack information needed for translation, such as dose-blood level relationship, brain target engagement, and dose-response, and many use treatment parameters that cannot be applied clinically, for example, treatment initiation before or at the time of injury and dosing higher than tolerated human equivalent dosing. Here, we review animal and human antiepileptogenic evidence for these medications. We highlight the gaps in our knowledge for each molecule that need to be filled in order to consider clinical translation, and we suggest a platform of preclinical antiepileptogenesis evaluation of potentially repurposable molecules or their combinations going forward.

Targeting oxidative stress improves disease outcomes in a rat model of acquired epilepsy.

Epilepsy therapy is based on antiseizure drugs that treat the symptom, seizures, rather than the disease and are ineffective in up to 30% of patients. There are no treatments for modifying the disease-preventing seizure onset, reducing severity or improving prognosis. Among the potential molecular targets for attaining these unmet therapeutic needs, we focused on oxidative stress since it is a pathophysiological process commonly occurring in experimental epileptogenesis and observed in human epilepsy. Using a rat model of acquired epilepsy induced by electrical status epilepticus, we show that oxidative stress occurs in both neurons and astrocytes during epileptogenesis, as assessed by measuring biochemical and histological markers. This evidence was validated in the hippocampus of humans who died following status epilepticus. Oxidative stress was reduced in animals undergoing epileptogenesis by a transient treatment with N-acetylcysteine and sulforaphane, which act to increase glutathione levels through complementary mechanisms. These antioxidant drugs are already used in humans for other therapeutic indications. This drug combination transiently administered for 2 weeks during epileptogenesis inhibited oxidative stress more efficiently than either drug alone. The drug combination significantly delayed the onset of epilepsy, blocked disease progression between 2 and 5 months post-status epilepticus and drastically reduced the frequency of spontaneous seizures measured at 5 months without modifying the average seizure duration or the incidence of epilepsy in animals. Treatment also decreased hippocampal neuron loss and rescued cognitive deficits. Oxidative stress during epileptogenesis was associated with de novo brain and blood generation of high mobility group box 1 (HMGB1), a neuroinflammatory molecule implicated in seizure mechanisms. Drug-induced reduction of oxidative stress prevented HMGB1 generation, thus highlighting a potential novel mechanism contributing to therapeutic effects. Our data show that targeting oxidative stress with clinically used drugs for a limited time window starting early after injury significantly improves long-term disease outcomes. This intervention may be considered for patients exposed to potential epileptogenic insults.