Pablo M. Casillas Espinosa

Objective: Traumatic brain injury (TBI) induces widespread and persistent neuroinflammation, which is increasingly recognized as a key contributor to chronic sequelae including posttraumatic epilepsy (PTE), cognitive deficits, and neuropsychiatric disturbances. However, reliable biomarkers for identifying individuals at risk for these long‐term outcomes remain limited. In this study, we evaluated in vivo positron emission tomography (PET) imaging of the 18‐kDa translocator protein (TSPO) as a predictive biomarker of epileptogenesis and neurobehavioral dysfunction in a rat model of moderate–severe TBI. Methods: Adult rats received lateral fluid percussion injury ( n = 61) or sham surgery ( n = 14). TSPO PET was performed using [ ¹⁸ F]‐PBR06 at 1 and 4 weeks postinjury. Long‐term behavioral outcomes were assessed at 5–6 months post‐TBI, followed by continuous 4‐week video‐electroencephalographic monitoring during 7 ^th month to identify epileptic rats. Results: TBI rats demonstrated robust and sustained increases in TSPO binding across multiple brain regions at both time points compared with sham animals ( p < .001). At 4 weeks postinjury, however, TSPO binding in the cortex, hippocampus, and thalamus was significantly lower in rats that later developed epilepsy (PTE ⁺ ) than in those that did not (PTE ⁻ ; p < .05), with receiver operating characteristic (ROC) analysis showing good predictive performance (area under the curve [AUC] = .76, p < .01). Behavioral testing identified deficits in spatial learning, memory, and open‐field activity in TBI rats ( p < .05). Data‐driven stratification identified behaviorally impaired (TBI‐BI ⁺ ) and unimpaired (TBI‐BI ⁻ ) subgroups, with cortical TSPO moderately distinguishing impaired animals (AUC ≈ .7). Among impaired rats, those with multidomain behavioral deficits ( n = 33) showed elevated thalamic TSPO compared to those with a single impairment ( p = .032), with ROC analysis confirming discrimination (AUC = .72, p = .018). Significance: Our findings support TSPO PET imaging as a promising early biomarker for predicting PTE and neurobehavioral outcomes following TBI.

Epilepsy is a heterogeneous neurological disorder affecting more than 70 million people worldwide, posing significant challenges for clinicians due to its complex etiology, diverse manifestations, variable treatment responses, and the inability to predict seizures or disease onset reliably. Despite advances in antiseizure medications, approximately 30% of patients remain treatment‐resistant, highlighting the urgent need for therapies with antiepileptogenic or disease‐modifying effects. To optimize and individualize strategies to predict and treat seizures and epilepsy, efforts to identify biomarkers of epilepsy risk, epileptogenesis, seizures, and therapy response are ongoing. This article reports key presentations and discussions from the 2023 Workshop on Neurobiology of Epilepsy (WONOEP XVII) in Kilkea, Ireland on novel epilepsy biomarkers and treatment strategies beyond the synapse and does not constitute a comprehensive review of biomarkers or treatment strategies. Much of the focus in epilepsy research has centered on identifying primarily neuronal processes or components. The 2023 WONOEP presentations discussed advances in plasma biomarkers for posttraumatic seizures and outcomes, perivascular spaces in posttraumatic epilepsy and poststroke epilepsy, imaging biomarkers of astrogliosis, and plasma microRNA biomarkers of intellectual disability and autism in tuberous sclerosis complex. Furthermore, research on immuno‐ and anti‐inflammatory therapies and blood–brain barrier in drug‐resistant focal epilepsies and infantile epileptic spasms syndrome was presented, as well as on the effects of antiseizure and cardioprotective medications on cardiac injury in temporal lobe epilepsy. The review also emphasizes the need for further interdisciplinary collaboration to accelerate the translation of these findings into clinical practice, ultimately improving outcomes and quality of life for people with epilepsy.

There is a wealth of data indicating that the immune system plays an important role in seizure disorders. This includes autoimmune encephalitis, in which an immune response directed against neuronal antigens results in brain inflammation and subsequent seizure activity, as well as autoimmune‐associated epilepsy and neuroinflammatory changes detected in the tissues of patients with epilepsy, in which context it is often difficult to distinguish whether the inflammation is causal or a consequence of the seizures. Here, we summarize the discussion on this topic held during the XVII Workshop on Neurobiology of Epilepsy (WONOEP XVII), organized in 2023 by the Neurobiology Commission of the International League Against Epilepsy on the topic of the extracellular space in epilepsy. This critical appraisal explores the progress, emerging concepts, and discussion on the immunopathogenesis of epilepsy, with a particular focus on the interplay between the immune system and the central nervous system, highlighting the role of autoimmunity, neuroinflammation, and neuroimmunological processes in the etiology of various epileptic disorders and the prospects for new therapies.

Objective: Traumatic brain injury (TBI) is a leading cause of long‐term disability, and infections such as pneumonia represent a common and serious complication for patients with TBI in the acute and subacute post‐injury period. Although the acute effects of infections have been documented, their long‐term consequences on neurological and behavioral recovery as well as the potential precipitation of seizures after TBI remain unclear. This study aimed to investigate the chronic effects of Klebsiella pneumoniae infection following TBI, focusing on post‐traumatic seizure development and neurobehavioral changes. Methods: Using a mouse model, we assessed the long‐term effects of TBI and K. pneumoniae infection both in isolation and in combination. Results: We found that, although infection with K. pneumoniae resulted in loss of body weight and increased mortality compared to vehicle‐inoculated mice, there was no additional mortality in TBI animals. Furthermore, although TBI alone induced chronic hyperactivity and reduced anxiety‐like behaviors, K. pneumoniae lung infection had no lasting effect on these long‐term outcomes. Third, although TBI resulted in both spontaneous and evoked seizures long‐term post‐injury, early post‐injury K. pneumoniae infection did not affect late‐onset seizure susceptibility. Significance: Together with recent findings on acute outcomes in this combined insult model of TBI and K. pneumoniae infection, this study suggests that K. pneumoniae does not significantly alter long‐term neurobehavioral outcomes or the development of post‐traumatic epilepsy. This research highlights the need to further explore the interplay between additional immune insults such as infection that may influence long‐term recovery.

Analogous to DNA methylation and protein phosphorylation, it is now well understood that RNA is also subject to extensive processing and modification. N ⁶ -methyladenosine (m ⁶ A) is the most abundant internal RNA modification and regulates RNA fate in several ways, including stability and translational efficiency. The role of m ⁶ A in both experimental and human epilepsy remains unknown. Here, we used transcriptome-wide m ⁶ A arrays to obtain a detailed analysis of the hippocampal m ⁶ A-ome from both mouse and human epilepsy samples. We combined this with human proteomic analyses and show that epileptic tissue displays disrupted metabolic and autophagic pathways that may be directly linked to m ⁶ A processing. Specifically, our results suggest that m ⁶ A levels inversely correlate with protein pathway activation. Finally, we show that elevated levels of m ⁶ A decrease seizure susceptibility and severity in mice. Together, our findings indicate that m ⁶ A represents an additional layer of gene regulation complexity in epilepsy and may contribute to the pathomechanisms that drive the development and maintenance of hyperexcitable brain networks.

Stroke remains a predominant cause of death and long-term disability among adults worldwide. Emerging evidence suggests that proteinopathies, characterized by the aggregation and accumulation of misfolded proteins, may play a significant role in the aftermath of stroke and the progression of neurodegenerative disorders. In this review, we explore preclinical and clinical research on key proteinopathies associated with stroke, including tau, Aβ (amyloid-β), TDP-43 (TAR DNA-binding protein 43), α-synuclein, and UCH-L1 (ubiquitin C-terminal hydrolase-L1). We focus on their potential as biomarkers for recovery management and as novel treatment targets that may enhance neuronal repair and mitigate secondary neurodegeneration. The involvement of these proteinopathies in various aspects of stroke, including neuroinflammation, oxidative stress, neuronal damage, and vascular dysfunction, underscores their potential. However, further investigations are essential to validate the clinical utility of these biomarkers, elucidate the mechanisms connecting proteinopathies to poststroke neurodegeneration, and develop targeted interventions. Identifying specific protein signatures associated with stroke outcomes could facilitate the advancement of precision medicine tailored to individual patient needs, significantly enhancing the quality of life for stroke survivors.

Objective: Cardiac structural and molecular changes are prevalent in people with chronic epilepsy, possibly contributing to an increased risk of premature mortality. However, understanding of the underlying pathophysiological mechanisms is limited. Here, we investigated the subacute and chronic changes in cardiac structure and ion channel/exchanger expression in different rodent models of temporal lobe epilepsy (TLE).Methods: Two models of TLE were used: the kainic acid‐induced post‐status epilepticus (KASE) model in Wistar rats and the electrical self‐sustained status epilepticus (SSSE) model in C57BL/6J mice. Heart tissue was collected at subacute (7 days post‐SE) and chronic (12–16 weeks post‐SE) timepoints from both models. Histological analysis for cardiac fibrosis and qPCR of ion channel/exchanger mRNA expression was performed.Results: Increased cardiac fibrosis was found in the KASE rats at the subacute (p = 0.016) and chronic (p = 0.003) timepoints compared with sham rats. In chronically epileptic KASE rats, mRNA expression analyses showed that Na_V1.5 and NCX1 were reduced in the septum (p = 0.026 and p = 0.020, respectively) compared with shams. In SSSE mice, Na_V1.5 was decreased in the right atrium (p = 0.039), and Ca_V3.2 and NCX1 were increased in the left ventricle subacutely (p = 0.033 and p = 0.003, respectively), and Na_V1.5 was increased in the septum at the chronic timepoint (p = 0.008), compared with the non‐epileptic sham group.Significance: Cardiac alterations at structural and molecular levels were found in both experimental rodent epilepsy models, subacutely post‐SE and during the chronically epileptic timepoint. The presence of similar cardiac changes across the models, despite being different species and having different modes of epilepsy indication, suggests that these changes are a direct or indirect result of the seizures.Plain Language Summary: Epilepsy may lead to heart problems, which could raise the risk of early death, but the exact causes are unclear. This study examined heart changes in two rodent models of epilepsy. In rats, heart scarring and stiffness (fibrosis) increased both shortly after seizures and during chronic epilepsy, and the ability to produce key heart proteins was altered. In mice, similar changes in heart proteins appeared in different heart areas. These findings suggest seizures can directly or indirectly cause harmful heart changes. Understanding these effects might help improve care for people with epilepsy and reduce related heart risks.

Objective: To test a hypothesis that acutely regulated plasma microRNAs (miRNAs) can serve as prognostic biomarkers for the development of post‐traumatic epilepsy (PTE). Methods: Adult male Sprague–Dawley rats ( n = 245) were randomized to lateral fluid‐percussion–induced traumatic brain injury (TBI) or sham operation at three study sites (Finland, Australia, United States). Video‐electroencephalography (vEEG) was performed on the seventh post‐injury month to detect spontaneous seizures. Tail vein plasma collected 48 h after TBI for miRNA analysis was available from 209 vEEG monitored animals (45 sham, 164 TBI [32 with epilepsy]). Based on small RNA sequencing and previous data, the seven most promising brain enriched miRNAs (miR‐183‐5p, miR‐323‐3p, miR‐434‐3p, miR‐9a‐3p, miR‐124‐3p, miR‐132‐3p, and miR‐212‐3p) were validated by droplet digital polymerase chain reaction (ddPCR). Results: All seven plasma miRNAs differentiated between TBI and sham‐operated rats. None of the seven miRNAs differentiated TBI rats that did and did not develop epilepsy ( p > .05), or rats with ≥3 vs .05). However, miR‐212‐3p differentiated rats that developed epilepsy with seizure clusters (i.e., ≥3 seizures within 24 h) from those without seizure clusters (.34 ± .14 vs .60 ± .34, adj. p < .05) with an area under the curve (AUC) of .81 (95% confidence interval [CI] .65–.97, p < .01, 64% sensitivity, 95% specificity). Lack of elevation in miR‐212‐3p also differentiated rats that developed epilepsy with seizure clusters from all other TBI rats ( n = 146, .34 ± .14 vs .55 ± .31, p < .01) with an AUC of .74 (95% CI .61–.87, p < .01, 82% sensitivity, 62% specificity). Glmnet analysis identified a combination of miR‐212‐3p and miR‐132‐3p as an optimal set to differentiate TBI rats with vs without seizure clusters (cross‐validated AUC .75, 95% CI .47–.92, p < .05). Significance: miR‐212‐3p alone or in combination with miR‐132‐3p shows promise as a translational prognostic biomarker for the development of severe PTE with seizure clusters.

The early onset epilepsies encompass a heterogeneous group of disorders, some of which result in drug‐resistant seizures, developmental delay, psychiatric comorbidities, and sudden death. Advancement in the widespread use of targeted gene panels as well as genome and exome sequencing has facilitated the identification of different causative genes in a subset of these patients. The ability to recognize the genetic basis of early onset epilepsies continues to improve, with de novo coding variants accounting for most of the genetic etiologies identified. Although current disease‐specific and disease‐modifying therapies remain limited, novel precision medicine approaches, such as small molecules, cell therapy, and other forms of genetic therapies for early onset epilepsies, have created excitement among researchers, clinicians, and caregivers. Here, we summarize the main findings of presentations and discussions on novel therapeutic strategies for targeted treatment of early onset epilepsies that occurred during the Workshop on Neurobiology of Epilepsy (WONOEP XVI, Talloires, France, July 2022). The presentations discussed the use of chloride transporter inhibitors for neonatal seizures, targeting orexinergic signaling for childhood absence epilepsy, targeting energy metabolism in Dravet syndrome, and the role of cannabinoid receptor type 2, reversible acetylcholinesterase inhibitors, cell therapies, and RNA‐based therapies in early life epilepsies.