The role of dendritic spines in epileptogenesis.

Epilepsy is a chronic central nervous system (CNS) disease associated with high morbidity. To date, there is no known disease-modifying therapy for epilepsy. A leading hypothesis for a mechanism of epileptogenesis is the generation of aberrant neuronal networks. Although the underlying biological mechanism is not clear, scientific evidence indicates that it is associated with a hyperexcitable synchronous neuronal network and active dendritic spine plasticity. Changes in dendritic spine morphology are related to altered expression of synaptic cytoskeletal proteins, inflammatory molecules, neurotrophic factors, and extracellular matrix signaling. However, it remains to be determined if these aberrant dendritic spine formations lead to neuronal hyperexcitability and abnormal synaptic connections or whether they constitute an underlying mechanism of seizure susceptibility. Focusing on dendritic spine machinery as a potential target for medications could limit or reverse the development of epilepsy.

Scoping review of disease-modifying effect of drugs in experimental epilepsy.

Objective: Epilepsy affects ~50 million people worldwide causing significant medical, financial, and sociologic concerns for affected patients and their families. To date, treatment of epilepsy is primarily symptomatic management because few effective preventative or disease-modifying interventions exist. However, recent research has identified neurobiological mechanisms of epileptogenesis, providing new pharmacologic targets to investigate. The current scientific evidence remains scattered across multiple studies using different model and experimental designs. The review compiles different models of anti-epileptogenic investigation and highlights specific compounds with potential epileptogenesis-modifying experimental drugs. It provides a platform for standardization of future epilepsy research to allow a more robust compound analysis of compounds with potential for epilepsy prevention. Methods: PubMed, Ovid MEDLINE, and Web of Science were searched from 2007 to 2021. Studies with murine models of epileptogenesis and explicitly detailed experimental procedures were included in the scoping review. In total, 51 articles were selected from 14,983 and then grouped by five core variables: (1) seizure frequency, (2) seizure severity, (3) spontaneous recurrent seizures (SRS), (4) seizure duration, and (5) mossy fiber sprouting (MFS). The variables were differentiated based on experimental models including methods of seizure induction, treatment schedule and timeline of data collection. Data was categorized by the five core variables and analyzed by converting original treatment values to units of percent of its respective control. Results: Discrepancies in current epileptogenesis models significantly complicate inter-study comparison of potential anti-epileptogenic interventions. With our analysis, many compounds showed a potential to reduce epileptogenic characteristics defined by the five core variables. WIN55,212-2, aspirin, rapamycin, 1400W, and LEV + BQ788 were identified compounds with the potential of effective anti-epileptic properties. Significance: Our review highlights the need for consistent methodology in epilepsy research and provides a novel approach for future research. Inconsistent experimental designs hinder study comparison, slowing the progression of treatments for epilepsy. If the research community can optimize and standardize parameters such as methods of seizure induction, administration schedule, sampling time, and aniMal models, more robust meta-analysis and collaborative research would follow. Additionally, some compounds such as rapamycin, WIN 55,212-2, aspirin, 1400W, and LEV + BQ788 showed anti-epileptogenic modulation across multiple variables. We believe they warrant further study both individually and synergistically.

CD40-CD40L in Neurological Disease.

Immune-inflammatory conditions in the central nervous system (CNS) rely on molecular and cellular interactions which are homeostatically maintained to protect neural tissue from harm. The CD40-CD40L interaction upregulates key proinflammatory molecules, a function best understood in the context of infection, during which B-cells are activated via CD40 signaling to produce antibodies. However, the role of CD40 in neurological disease of non-infectious etiology is unclear. We review the role of CD40-CD40L in traumatic brain injury, Alzheimer's Disease, Parkinson's Disease, stroke, epilepsy, nerve injury, multiple sclerosis, ALS, myasthenia gravis and brain tumors. We also highlight therapeutic advancements targeting the CD40 system to either attenuate the neuroinflammatory response or leverage the downstream effects of CD40 signaling for direct tumor cell lysis.

Brain injuries can set up an epileptogenic neuronal network.

Development of epilepsy or epileptogenesis promotes recurrent seizures. As of today, there are no effective prophylactic therapies to prevent the onset of epilepsy. Contributing to this deficiency of preventive therapy is the lack of clarity in fundamental neurobiological mechanisms underlying epileptogenesis and lack of reliable biomarkers to identify patients at risk for developing epilepsy. This limits the development of prophylactic therapies in epilepsy. Here, neural network dysfunctions reflected by oscillopathies and microepileptiform activities, including neuronal hyperexcitability and hypersynchrony, drawn from both clinical and experimental epilepsy models, have been reviewed. This review suggests that epileptogenesis reflects a progressive and dynamic dysfunction of specific neuronal networks which recruit further interconnected groups of neurons, with this resultant pathological network mediating seizure occurrence, recurrence, and progression. In the future, combining spatial and temporal resolution of neuronal non-invasive recordings from patients at risk of developing epilepsy, together with analytics and computational tools, may contribute to determining whether the brain is undergoing epileptogenesis in asymptomatic patients following brain injury.

Downregulation of CD40L-CD40 attenuates seizure susceptibility and severity of seizures.

Unregulated neuro-inflammation mediates seizures in temporal lobe epilepsy (TLE). Our aim was to determine the effect of CD40-CD40L activation in experimental seizures. CD40 deficient mice (CD40KO) and control mice (wild type, WT) received pentenyltetrazole (PTZ) or pilocarpine to evaluate seizures and status epilepticus (SE) respectively. In mice, anti-CD40L antibody was administered intranasally before PTZ. Brain samples from human TLE and post-seizure mice were processed to determine CD40-CD40L expression using histological and molecular techniques. CD40 expression was higher in hippocampus from human TLE and in cortical neurons and hippocampal neural terminals after experimental seizures. CD40-CD40L levels increased after seizures in the hippocampus and in the cortex. After SE, CD40L/CD40 levels increased in cortex and showed an upward trend in the hippocampus. CD40KO mice demonstrated reduction in seizure severity and in latency compared to WT mice. Anti-CD40L antibody limited seizure susceptibility and seizure severity. CD40L-CD40 interaction can serve as a target for an immuno-therapy for TLE.

Cutis verticis gyrata: Two cases associated with drug-resistant epilepsy.

Cutis verticis gyrata (CVG) is a neurocutaneous syndrome characterized by the formation of folds in the scalp that resembles the cerebral cortex. We present two cases of CVG and intellectual disability with drug-resistant epilepsy. Recognizing CVG is necessary to provide interdisciplinary support for the treatment of comorbidities associated with this entity.

The role of inflammation in the development of epilepsy.

Epilepsy, a neurological disease characterized by recurrent seizures, is often associated with a history of previous lesions in the nervous system. Impaired regulation of the activation and resolution of inflammatory cells and molecules in the injured neuronal tissue is a critical factor to the development of epilepsy. However, it is still unclear as to how that unbalanced regulation of inflammation contributes to epilepsy. Therefore, one of the goals in epilepsy research is to identify and elucidate the interconnected inflammatory pathways in systemic and neurological disorders that may further develop epilepsy progression. In this paper, inflammatory molecules, in neurological and systemic disorders (rheumatoid arthritis, Crohn's, Type I Diabetes, etc.) that could contribute to epilepsy development, are reviewed.Understanding the neurobiology of inflammation in epileptogenesis will contribute to the development of new biomarkers for better screening of patients at risk for epilepsy and new therapeutic targets for both prophylaxis and treatment of epilepsy.

Impaired Attentional Set-Shifting Performance after Exposure to 5 cGy of 600 MeV/n 28Si Particles.

Astronauts on deep space missions will be required to work more autonomously than on previous missions, and thus their ability to perform executive functions could be critical to mission success. One of the most common measures of executive function in humans is the ability to perform attentional set shifting, which requires contributions from working memory, discrimination, reversal learning, attentional set shifting and attention. Rodent attentional set shifting assays require rats to form an association between the presence of the food reward and an associative cue, which is either the digging media or the scent that is placed in the bowl; by altering the combination of scent and digging media, progressively more complex cognitive processes can be tested. In this study, we have determined the effect that exposure to 5-20 cGy of 600 MeV/n 28Si particles has on the ability of male retired breeder Wistar rats to perform attentional set shifting at three months postirradiation. All doses of Si resulted in a significant impairment in the ability of the rats to perform the first and most simple step of the ATSET assay, the simple discrimination (SD) task. If astronauts were to experience HZE-induced SD impairments, they would be unable to identify key factors to successfully resolve a situation. Performance in at least one other component of the ATSET test was impaired at all doses studied, however, these varied according to the dose. Compared with our previous studies using 1 GeV/n 56Fe and 48Ti particles, 600 MeV/n 28Si ions impaired attentional set-shifting performance at lower doses than the heavier ions. However, when the effect of isofluences of the three HZE ions were compared, there were no significant differences in the severity of the impaired performance; there were, however, ion-specific decrements in the ability of rats to perform within the various stages of the test. This study further supports the notion that "mission-relevant" doses of HZE particles (<20 cGy) can impair certain aspects of attentional set-shifting performance in retired breeder rats, but there may be some ion-specific changes in the specific cognitive domains impaired.

Intranasal Therapy to Stop Status Epilepticus in Prehospital Settings.

Status epilepticus (SE) is a medical emergency characterized by uncontrolled, prolonged seizures with rapid and widespread neuronal damage. Patients that suffer from longer episodes of SE are more likely to have poorer clinical outcomes and a higher cost of healthcare. Understanding novel molecular mechanisms that regulate inhibitory and excitatory neurotransmission that initiate SE and the necessary medical infrastructure to stop SE could help identify targets for early intervention. Intranasal administration of benzodiazepines may shorten the time between initiation and cessation of seizures when compared to other routes of administration. Current pharmaceutical administration guidelines are appropriate for sporadic incidences of SE, but exploring other approaches is necessary to prepare for situations involving multiple patients outside of a hospital, such as a massive chemical weapons attack. Intranasal drug delivery helps to circumvent the blood-brain barrier and offers a noninvasive way to quickly administer drugs in settings that require an immediate response, such as nerve agent exposure. In addition, examining the intranasal delivery of new drugs, such as nanotherapeutics, may lead to more effective, noninvasive, scalable, and portable methods of treating SE.

Dysfunctional epileptic neuronal circuits and dysmorphic dendritic spines are mitigated by platelet-activating factor receptor antagonism.

Temporal lobe epilepsy or limbic epilepsy lacks effective therapies due to a void in understanding the cellular and molecular mechanisms that set in motion aberrant neuronal network formations during the course of limbic epileptogenesis (LE). Here we show in in vivo rodent models of LE that the phospholipid mediator platelet-activating factor (PAF) increases in LE and that PAF receptor (PAF-r) ablation mitigates its progression. Synthetic PAF-r antagonists, when administered intraperitoneally in LE, re-establish hippocampal dendritic spine density and prevent formation of dysmorphic dendritic spines. Concomitantly, hippocampal interictal spikes, aberrant oscillations, and neuronal hyper-excitability, evaluated 15-16 weeks after LE using multi-array silicon probe electrodes implanted in the dorsal hippocampus, are reduced in PAF-r antagonist-treated mice. We suggest that over-activation of PAF-r signaling induces aberrant neuronal plasticity in LE and leads to chronic dysfunctional neuronal circuitry that mediates epilepsy.

Hippocampal neuro-networks and dendritic spine perturbations in epileptogenesis are attenuated by neuroprotectin d1.

PURPOSE: Limbic epileptogenesis triggers molecular and cellular events that foster the establishment of aberrant neuronal networks that, in turn, contribute to temporal lobe epilepsy (TLE). Here we have examined hippocampal neuronal network activities in the pilocarpine post-status epilepticus model of limbic epileptogenesis and asked whether or not the docosahexaenoic acid (DHA)-derived lipid mediator, neuroprotectin D1 (NPD1), modulates epileptogenesis. METHODS: Status epilepticus (SE) was induced by intraperitoneal administration of pilocarpine in adult male C57BL/6 mice. To evaluate simultaneous hippocampal neuronal networks, local field potentials were recorded from multi-microelectrode arrays (silicon probe) chronically implanted in the dorsal hippocampus. NPD1 (570 µg/kg) or vehicle was administered intraperitoneally daily for five consecutive days 24 hours after termination of SE. Seizures and epileptiform activity were analyzed in freely-moving control and treated mice during epileptogenesis and epileptic periods. Then hippocampal dendritic spines were evaluated using Golgi-staining. RESULTS: We found brief spontaneous microepileptiform activity with high amplitudes in the CA1 pyramidal and stratum radiatum in epileptogenesis. These aberrant activities were attenuated following systemic NPD1 administration, with concomitant hippocampal dendritic spine protection. Moreover, NPD1 treatment led to a reduction in spontaneous recurrent seizures. CONCLUSIONS: Our results indicate that NPD1 displays neuroprotective bioactivity on the hippocampal neuronal network ensemble that mediates aberrant circuit activity during epileptogenesis. Insight into the molecular signaling mediated by neuroprotective bioactivity of NPD1 on neuronal network dysfunction may contribute to the development of anti-epileptogenic therapeutic strategies.

Therapeutic efficacy of aldoxorubicin in an intracranial xenograft mouse model of human glioblastoma.

Glioblastoma multiforme (GBM) is the most aggressive primary brain tumor with a median survival of 12 to 15 months after diagnosis. Acquired chemoresistance, high systemic toxicity, and low penetration of the blood brain barrier by many anticancer drugs contribute to the failure of anti-GBM therapies. To circumvent some of these obstacles, we tested a novel prodrug approach to evaluate anti-GBM efficacy by utilizing serum albumin-binding doxorubicin (Doxo), aldoxorubicin (Aldoxo), which is less toxic, is released from albumin in an acidic environment and accumulates in tumor tissues. A human GBM cell line that expresses a luciferase reporter (U87-luc) was stereotactically injected into the left striatum of the brain of immunodeficient mice. Following initial tumor growth for 12 days, mice were injected once a week in the tail-vein with Aldoxo [24 mg/kg or 18 mg/kg of doxorubicin equivalents-3/4 maximum tolerated dose (MTD)], Doxo [6 mg/kg (3/4 MTD)], or vehicle. Aldoxo-treated mice demonstrated significantly slower growth of the tumor when compared to vehicle-treated or Doxo-treated mice. Five out of eight Aldoxo-treated mice remained alive more than 60 days with a median survival of 62 days, while the median survival of vehicle- and Doxo-treated mice was only 26 days. Importantly, Aldoxo-treated mice exhibited high levels of Doxo within the tumor tissue, accompanied by low tumor cell proliferation (Ki67) and abundant intratumoral programmed cell death (cleaved caspase-3). Effective accumulation of Aldoxo in brain tumor tissues but not normal brain, its anti-tumor efficacy, and low toxicity, provide a strong rationale for evaluating this novel drug conjugate as a treatment for patients afflicted with GBM.

All-trans-retinoid acid induces the differentiation of encapsulated mouse embryonic stem cells into GABAergic neurons.

Embryonic stem (ES) cells are pluripotent cells that can differentiate into all three main germ layers: endoderm, mesoderm, and ectoderm. Although a number of methods have been developed to differentiate ES cells into neuronal phenotypes such as sensory and motor neurons, the efficient generation of GABAergic interneurons from ES cells still presents an ongoing challenge. Because the main output of inhibitory GABAergic interneurons is the gamma-aminobutyric-acid (GABA), a neurotransmitter whose controlled homeostasis is required for normal brain function, the efficient generation in culture of functional interneurons may have future implications on the treatment of neurological disorders such as epilepsy, autism, and schizophrenia. The goal of this work was to examine the generation of GABAergic neurons from mouse ES cells by comparing an embryoid body-based methodology versus a hydrogel-based encapsulation protocol that involves the use of all-trans-retinoid acid (RA). We observed that (1) there was a 2-fold increase in neuronal differentiation in encapsulated versus non-encapsulated cells and (2) there was an increase in the specificity for interneuronal differentiation in encapsulated cells, as assessed by mRNA expression and electrophysiology approaches. Furthermore, our results indicate that most of the neurons obtained from encapsulated mouse ES cells are GABA-positive (∼87%). Thus, these results suggest that combining encapsulation of ES cells and RA treatment provide a more efficient and scalable differentiation strategy for the generation in culture of functional GABAergic interneurons. This technology may have implications for future cell replacement therapies and the treatment of CNS disorders.

Combined process automation for large-scale EEG analysis.

Epileptogenesis is a dynamic process producing increased seizure susceptibility. Electroencephalography (EEG) data provides information critical in understanding the evolution of epileptiform changes throughout epileptic foci. We designed an algorithm to facilitate efficient large-scale EEG analysis via linked automation of multiple data processing steps. Using EEG recordings obtained from electrical stimulation studies, the following steps of EEG analysis were automated: (1) alignment and isolation of pre- and post-stimulation intervals, (2) generation of user-defined band frequency waveforms, (3) spike-sorting, (4) quantification of spike and burst data and (5) power spectral density analysis. This algorithm allows for quicker, more efficient EEG analysis.

Endogenous signaling by omega-3 docosahexaenoic acid-derived mediators sustains homeostatic synaptic and circuitry integrity.

The harmony and function of the complex brain circuits and synapses are sustained mainly by excitatory and inhibitory neurotransmission, neurotrophins, gene regulation, and factors, many of which are incompletely understood. A common feature of brain circuit components, such as dendrites, synaptic membranes, and other membranes of the nervous system, is that they are richly endowed in docosahexaenoic acid (DHA), the main member of the omega-3 essential fatty acid family. DHA is avidly retained and concentrated in the nervous system and known to play a role in neuroprotection, memory, and vision. Only recently has it become apparent why the surprisingly rapid increases in free (unesterified) DHA pool size take place at the onset of seizures or brain injury. This phenomenon began to be clarified by the discovery of neuroprotectin D1 (NPD1), the first-uncovered bioactive docosanoid formed from free DHA through 15-lipoxygenase-1 (15-LOX-1). NPD1 synthesis includes, as agonists, oxidative stress and neurotrophins. The evolving concept is that DHA-derived docosanoids set in motion endogenous signaling to sustain homeostatic synaptic and circuit integrity. NPD1 is anti-inflammatory, displays inflammatory resolving activities, and induces cell survival, which is in contrast to the pro-inflammatory actions of the many of omega-6 fatty acid family members. We highlight here studies relevant to the ability of DHA to sustain neuronal function and protect synapses and circuits in the context of DHA signalolipidomics. DHA signalolipidomics comprises the integration of the cellular/tissue mechanism of DHA uptake, its distribution among cellular compartments, the organization and function of membrane domains containing DHA phospholipids, and the precise cellular and molecular events revealed by the uncovering of signaling pathways regulated by docosanoids endowed with prohomeostatic and cell survival bioactivity. Therefore, this approach offers emerging targets for prevention, pharmaceutical intervention, and clinical translation involving DHA-mediated signaling.

The omega-3 fatty acid-derived neuroprotectin D1 limits hippocampal hyperexcitability and seizure susceptibility in kindling epileptogenesis.

PURPOSE: Temporal lobe epilepsy, one of the most common epilepsy syndromes, is characterized by hippocampal hyperexcitability and progressive seizure susceptibility. Omega-3 fatty acids are involved in neuronal excitability and have anticonvulsant properties. We studied the effect of docosahexaenoic acid (DHA) or its derived lipid mediator, neuroprotectin D1 (NPD1, 10R,17S-dihydroxy-docosa-4Z,7Z,11E,13E,15Z,19Z-hexaenoic acid), in evoked seizures using a rapid kindling model of temporal lobe epilepsy. METHODS: DHA or NPD1 was administered in rodents with or without kindling acquisition. Locomotor seizures and evoked epileptiform hippocampal activity immediately after hippocampal stimulations were analyzed. KEY FINDINGS: DHA or NPD1 limits hippocampal electrically induced hyperexcitability. Seizures induced by kindling triggered NPD1 synthesis in the hippocampus. Supplying its precursor, DHA, or direct injection of NPD1 into the third ventricle resulted in attenuation of kindling progression and hippocampal hyperexcitability. SIGNIFICANCE: The significance of NPD1 in temporal lobe epilepsy could open new pathways for understanding the initiation and propagation of seizures and the role this lipid mediator plays in the neuronal network.

Platelet-activating factor receptor antagonism targets neuroinflammation in experimental epilepsy.

PURPOSE: Temporal lobe epilepsy is associated with the inflammatory process related to the basic mechanisms that lead to seizure susceptibility and brain damage. Platelet-activating factor (PAF), a potent, short-lived phospholipid mediator of inflammation, participates in physiologic signaling in the brain. However, after seizures, PAF accumulates in the brain and activates intracellular signaling related with inflammation-mediated excitotoxicity and hippocampal hyperexcitability. The objective of this study is to evaluate the effect of PAF antagonism on hippocampal hyperexcitability, seizure susceptibility, and neuroprotection using the kindling paradigm and pilocarpine-induced seizure damage models. METHODS: The PAF antagonist, LAU-0901 (60 mg/kg, i.p.), or vehicle, was administrated each day of kindling or daily during the 4 weeks after status epilepticus (SE). We analyzed seizure severity, electrical activity, cellular damage, and inflammation in the hippocampi of both treated groups. KEY FINDINGS: LAU-0901 limits the progression of kindling and attenuates seizure susceptibility 1 week after the kindling procedure. In addition, under the seizure-damage conditions studied here, we observed that LAU-0901 induces hippocampal neuroprotection and limits somatostatin interneuronal cell loss and inflammation. SIGNIFICANCE: Our results indicate that modulation of PAF overactivity attenuates seizure susceptibility, hippocampal hyperexcitability, and neuroinflammation.