Mouse Behavior on the Trial-Unique Nonmatching-to-Location (TUNL) Touchscreen Task Reflects a Mixture of Distinct Working Memory Codes and Response Biases.

The trial-unique nonmatching to location (TUNL) touchscreen task shows promise as a translational assay of working memory (WM) deficits in rodent models of autism, ADHD, and schizophrenia. However, the low-level neurocognitive processes that drive behavior in the TUNL task have not been fully elucidated. In particular, it is commonly assumed that the TUNL task predominantly measures spatial WM dependent on hippocampal pattern separation, but this proposition has not previously been tested. In this project, we tested this question using computational modeling of behavior from male and female mice performing the TUNL task (N = 163 across three datasets; 158,843 trials). Using this approach, we empirically tested whether TUNL behavior solely measured retrospective WM, or whether it was possible to deconstruct behavior into additional neurocognitive subprocesses. Overall, contrary to common assumptions, modeling analyses revealed that behavior on the TUNL task did not primarily reflect retrospective spatial WM. Instead, behavior was best explained as a mixture of response strategies, including both retrospective WM (remembering the spatial location of a previous stimulus) and prospective WM (remembering an anticipated future behavioral response) as well as animal-specific response biases. These results suggest that retrospective spatial WM is just one of a number of cognitive subprocesses that contribute to choice behavior on the TUNL task. We suggest that findings can be understood within a resource-rational framework, and use computational model simulations to propose several task-design principles that we predict will maximize spatial WM and minimize alternative behavioral strategies in the TUNL task.SIGNIFICANCE STATEMENT Touchscreen tasks represent a paradigm shift for assessment of cognition in nonhuman animals by automating large-scale behavioral data collection. Their main relevance, however, depends on the assumption of functional equivalence to cognitive domains in humans. The trial-unique, delayed nonmatching to location (TUNL) touchscreen task has revolutionized the study of rodent spatial working memory. However, its assumption of functional equivalence to human spatial working memory is untested. We leveraged previously untapped single-trial TUNL data to uncover a novel set of hierarchically ordered cognitive processes that underlie mouse behavior on this task. The strategies used demonstrate multiple cognitive approaches to a single behavioral outcome and the requirement for more precise task design and sophisticated data analysis in interpreting rodent spatial working memory.

Understanding the role of the NMDA receptor subunit, GluN2D, in mediating NMDA receptor antagonist-induced behavioral disruptions in male and female mice.

Noncompetitive NMDA receptor (NMDAR) antagonists like phencyclidine (PCP) and ketamine cause psychosis-like symptoms in healthy humans, exacerbate schizophrenia symptoms in people with the disorder, and disrupt a range of schizophrenia-relevant behaviors in rodents, including hyperlocomotion. This is negated in mice lacking the GluN2D subunit of the NMDAR, suggesting the GluN2D subunit mediates the hyperlocomotor effects of these drugs. However, the role of GluN2D in mediating other schizophrenia-relevant NMDAR antagonist-induced behavioral disturbances, and in both sexes, is unclear. This study aimed to investigate the role of the GluN2D subunit in mediating schizophrenia-relevant behaviors induced by a range of NMDA receptor antagonists. Using both male and female GluN2D knockout (KO) mice, we examined the effects of the NMDAR antagonist's PCP, the S-ketamine enantiomer (S-ket), and the ketamine metabolite R-norketamine (R-norket) on locomotor activity, anxiety-related behavior, and recognition and short-term spatial memory. GluN2D-KO mice showed a blunted locomotor response to R-norket, S-ket, and PCP, a phenotype present in both sexes. GluN2D-KO mice of both sexes showed an anxious phenotype and S-ket, R-norket, and PCP showed anxiolytic effects that were dependent on sex and genotype. S-ket disrupted spatial recognition memory in females and novel object recognition memory in both sexes, independent of genotype. This datum identifies a role for the GluN2D subunit in sex-specific effects of NMDAR antagonists and on the differential effects of the R- and S-ket enantiomers.

ComBating inter-site differences in field strength: harmonizing preclinical traumatic brain injury MRI data.

Integrating datasets from multiple sites and scanners can increase statistical power for neuroimaging studies but can also introduce significant inter-site confounds. We evaluated the effectiveness of ComBat, an empirical Bayes approach, to combine longitudinal preclinical MRI data acquired at 4.7 or 9.4 T at two different sites in Australia. Male Sprague Dawley rats underwent MRI on Days 2, 9, 28, and 150 following moderate/severe traumatic brain injury (TBI) or sham injury as part of Project 1 of the NIH/NINDS-funded Centre Without Walls EpiBioS4Rx project. Diffusion-weighted and multiple-gradient-echo images were acquired, and outcomes included QSM, FA, and ADC. Acute injury measures including apnea and self-righting reflex were consistent between sites. Mixed-effect analysis of ipsilateral and contralateral corpus callosum (CC) summary values revealed a significant effect of site on FA and ADC values, which was removed following ComBat harmonization. Bland-Altman plots for each metric showed reduced variability across sites following ComBat harmonization, including for QSM, despite appearing to be largely unaffected by inter-site differences and no effect of site observed. Following harmonization, the combined inter-site data revealed significant differences in the imaging metrics consistent with previously reported outcomes. TBI resulted in significantly reduced FA and increased susceptibility in the ipsilateral CC, and significantly reduced FA in the contralateral CC compared with sham-injured rats. Additionally, TBI rats also exhibited a reversal in ipsilateral CC ADC values over time with significantly reduced ADC at Day 9, followed by increased ADC 150 days after injury. Our findings demonstrate the need for harmonizing multi-site preclinical MRI data and show that this can be successfully achieved using ComBat while preserving phenotypical changes due to TBI.

Epilepsy phenotype and its reproducibility after lateral fluid percussion-induced traumatic brain injury in rats: Multicenter EpiBioS4Rx study project 1.

OBJECTIVE: This study was undertaken to assess reproducibility of the epilepsy outcome and phenotype in a lateral fluid percussion model of posttraumatic epilepsy (PTE) across three study sites. METHODS: A total of 525 adult male Sprague Dawley rats were randomized to lateral fluid percussion-induced brain injury (FPI) or sham operation. Of these, 264 were assigned to magnetic resonance imaging (MRI cohort, 43 sham, 221 traumatic brain injury [TBI]) and 261 to electrophysiological follow-up (EEG cohort, 41 sham, 220 TBI). A major effort was made to harmonize the rats, materials, equipment, procedures, and monitoring systems. On the 7th post-TBI month, rats were video-EEG monitored for epilepsy diagnosis. RESULTS: A total of 245 rats were video-EEG phenotyped for epilepsy on the 7th postinjury month (121 in MRI cohort, 124 in EEG cohort). In the whole cohort (n = 245), the prevalence of PTE in rats with TBI was 22%, being 27% in the MRI and 18% in the EEG cohort (p > .05). Prevalence of PTE did not differ between the three study sites (p > .05). The average seizure frequency was .317 ± .725 seizures/day at University of Eastern Finland (UEF; Finland), .085 ± .067 at Monash University (Monash; Australia), and .299 ± .266 at University of California, Los Angeles (UCLA; USA; p < .01 as compared to Monash). The average seizure duration did not differ between UEF (104 ± 48 s), Monash (90 ± 33 s), and UCLA (105 ± 473 s; p > .05). Of the 219 seizures, 53% occurred as part of a seizure cluster (≥3 seizures/24 h; p >.05 between the study sites). Of the 209 seizures, 56% occurred during lights-on period and 44% during lights-off period (p > .05 between the study sites). SIGNIFICANCE: The PTE phenotype induced by lateral FPI is reproducible in a multicenter design. Our study supports the feasibility of performing preclinical multicenter trials in PTE to increase statistical power and experimental rigor to produce clinically translatable data to combat epileptogenesis after TBI.

Levetiracetam Pharmacokinetics and Brain Uptake in a Lateral Fluid Percussion Injury Rat Model.

Post-traumatic epilepsy (PTE) occurs in some patients after moderate/severe traumatic brain injury (TBI). Although there are no approved therapies to prevent epileptogenesis, levetiracetam (LEV) is commonly given for seizure prophylaxis due to its good safety profile. This led us to study LEV as part of the Epilepsy Bioinformatics Study for Antiepileptogenic Therapy (EpiBioS4Rx) Project. The objective of this work is to characterize the pharmacokinetics (PK) and brain uptake of LEV in naïve control rats and in the lateral fluid percussion injury (LFPI) rat model of TBI after either single intraperitoneal doses or a loading dose followed by a 7-day subcutaneous infusion. Sprague-Dawley rats were used as controls and for the LFPI model induced at the left parietal region using injury parameters optimized for moderate/severe TBI. Naïve and LFPI rats received either a bolus injection (intraperitoneal) or a bolus injection followed by subcutaneous infusion over 7 days. Blood and parietal cortical samples were collected at specified time points throughout the study. LEV concentrations in plasma and brain were measured using validated high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) methods. Noncompartmental analysis and a naive-pooled compartmental PK modeling approach were used. Brain-to-plasma ratios ranged from 0.54 to 1.4 to 1. LEV concentrations were well fit by one-compartment, first-order absorption PK models with a clearance of 112 ml/h per kg and volume of distribution of 293 ml/kg. The single-dose pharmacokinetic data were used to guide dose selection for the longer-term studies, and target drug exposures were confirmed. Obtaining LEV PK information early in the screening phase allowed us to guide optimal treatment protocols in EpiBioS4Rx. SIGNIFICANCE STATEMENT: The characterization of levetiracetam pharmacokinetics and brain uptake in an animal model of post-traumatic epilepsy is essential to identify target concentrations and guide optimal treatment for future studies.

Metyrapone abolishes spike-wave discharge seizures in genetic absence epilepsy rats from Strasbourg by reducing stress hormones.

OBJECTIVE: Stress is one of the most commonly reported triggers for seizures in patients with epilepsy, although the mechanisms that mediate this effect are not established. The clinical evidence supporting this is derived from patients' subjective experience of stress, and how this influences their own seizures. Animal models can be used to explore this phenomenon in controlled environments, free from subjective bias. Here, we used genetic absence epilepsy rats from Strasbourg (GAERS), a genetic rat model of absence epilepsy, to explore the influence of stress and stress hormones on spontaneous seizures. METHODS: Adult male GAERS (n = 38) and nonepileptic control (NEC) rats (n = 4) were used. First, rats were subjected to 30-min restraint stress to assess hypothalamic-pituitary-adrenal axis function. Next, we assessed the effects of 30-min noise stress, and cage tilt stress, on spike-wave discharge seizures in GAERS. We then performed pharmacological experiments to assess the direct effects of stress hormones on seizures, including corticosterone, metyrapone, and deoxycorticosterone. RESULTS: GAERS exhibited elevated baseline corticosterone levels, compared to NEC rats. Noise stress and cage tilt stress significantly enhanced seizure incidence (p < .05), but only during stress periods. Exogenous corticosterone administration also significantly increased seizure occurrence (p < .05). Metyrapone, an inhibitor of corticosterone synthesis, completely abolished seizures in GAERS, and seizures remained suppressed for >2 h. However, deoxycorticosterone, the precursor of corticosterone, increased seizures. SIGNIFICANCE: These results suggest that GAERS exhibit elevations in stress hormones, and this may contribute to seizures. Inhibiting corticosterone synthesis with metyrapone prevents seizures in GAERS, and shows potential for repurposing this drug as a future antiseizure medication.

E2730, an uncompetitive γ-aminobutyric acid transporter-1 inhibitor, suppresses epileptic seizures in a rat model of chronic mesial temporal lobe epilepsy.

OBJECTIVE: More than one third of mesial temporal lobe epilepsy (MTLE) patients are resistant to current antiseizure medications (ASMs), and half experience mild-to-moderate adverse effects of ASMs. There is therefore a strong need to develop and test novel ASMs. The objective of this work is to evaluate the pharmacokinetics and neurological toxicity of E2730, a novel uncompetitive inhibitor of γ-aminobutyric acid transporter-1, and to test its seizure suppression effects in a rat model of chronic MTLE. METHODS: We first examined plasma levels and adverse neurological effects of E2730 in healthy Wistar rats. Adult male rats were implanted with osmotic pumps delivering either 10, 20, or 100 mg/kg/day of E2730 subcutaneously for 1 week. Blood sampling and behavioral assessments were performed at several timepoints. We next examined whether E2730 suppressed seizures in rats with chronic MTLE. These rats were exposed to kainic acid-induced status epilepticus, and 9 weeks later, when chronic epilepsy was established, were assigned to receive one of the three doses of E2730 or vehicle for 1 week in a randomized crossover design. Continuous video-electroencephalographic monitoring was acquired during the treatment period to evaluate epileptic seizures. RESULTS: Plasma levels following continuous infusion of E2730 showed a clear dose-related increase in concentration. The drug was well tolerated at all doses, and any sedation or neuromotor impairment was mild and transient, resolving within 48 h of treatment initiation. Remarkably, E2730 treatment in chronically epileptic rats led to seizure suppression in a dose-dependent manner, with 65% of rats becoming seizure-free at the highest dose tested. Mean seizure class did not differ between the treatment groups. SIGNIFICANCE: This study shows that continuous subcutaneous infusion of E2730 over 7 days results in a marked, dose-dependent suppression of spontaneous recurrent seizures, with minimal adverse neurological effects, in a rat model of chronic MTLE. E2730 shows strong promise as an effective new ASM to be translated into clinical trials.

Integrative genomics reveals pathogenic mediator of valproate-induced neurodevelopmental disability.

Prenatal exposure to the anti-seizure medication sodium valproate (VPA) is associated with an increased risk of adverse postnatal neurodevelopmental outcomes, including lowered intellectual ability, autism spectrum disorder and attention-deficit hyperactivity disorder. In this study, we aimed to clarify the molecular mechanisms underpinning the neurodevelopmental consequences of gestational VPA exposure using integrative genomics. We assessed the effect of gestational VPA on foetal brain gene expression using a validated rat model of valproate teratogenicity that mimics the human scenario of chronic oral valproate treatment during pregnancy at doses that are therapeutically relevant to the treatment of epilepsy. Two different rat strains were studied-inbred Genetic Absence Epilepsy Rats from Strasbourg, a model of genetic generalized epilepsy, and inbred non-epileptic control rats. Female rats were fed standard chow or VPA mixed in standard chow for 2 weeks prior to conception and then mated with same-strain males. In the VPA-exposed rats maternal oral treatment was continued throughout pregnancy. Foetuses were extracted via C-section on gestational Day 21 (1 day prior to birth) and foetal brains were snap-frozen and genome-wide gene expression data generated. We found that gestational VPA exposure via chronic maternal oral dosing was associated with substantial drug-induced differential gene expression in the pup brains, including dysregulated splicing, and observed that this occurred in the absence of evidence for significant neuronal gain or loss. The functional consequences of VPA-induced gene expression were explored using pathway analysis and integration with genetic risk data for psychiatric disease and behavioural traits. The set of genes downregulated by VPA in the pup brains were significantly enriched for pathways related to neurodevelopment and synaptic function and significantly enriched for heritability to human intelligence, schizophrenia and bipolar disorder. Our results provide a mechanistic link between chronic foetal VPA exposure and neurodevelopmental disability mediated by VPA-induced transcriptional dysregulation.

Does air pollution confound associations between environmental noise and cardiovascular outcomes? - A systematic review.

BACKGROUND: Exposure to environmental noise is associated with adverse health effects, but there is potential for confounding and interaction with air pollution, particularly where both exposures arise from the same source, such as transport. OBJECTIVES: To review evidence on confounding and interaction of air pollution in relation to associations between environmental noise and cardiovascular outcomes. METHODS: Papers were identified from similar reviews published in 2013 and 2015, from the systematic reviews supporting the WHO 2018 noise guidelines, and from a literature search covering the period 2016-2022 using Medline and PubMed databases. Additional papers were identified from colleagues. Study selection was according to PECO inclusion criteria. Studies were evaluated against the WHO checklist for risk of bias. RESULTS: 52 publications, 36 published after 2015, were identified that assessed associations between transportation noise and cardiovascular outcomes, that also considered potential confounding (49 studies) or interaction (23 studies) by air pollution. Most, but not all studies, suggested that the associations between traffic noise and cardiovascular outcomes are independent of air pollution. NO2 or PM2.5 were the most commonly included air pollutants and we observed no clear differences across air pollutants in terms of the potential confounding role. Most papers did not appear to suggest an interaction between noise and air pollution. Eight studies found the largest noise effect estimates occurring within the higher noise and air pollution exposure categories, but were not often statistically significant. CONCLUSION: Whilst air pollution does not appear to confound associations of noise and cardiovascular health, more studies on potential interactions are needed. Current methods to assess quality of evidence are not optimal when evaluating evidence on confounding or interaction.

Potential Roles for the GluN2D NMDA Receptor Subunit in Schizophrenia.

Glutamate N-methyl-D-aspartate receptor (NMDAR) hypofunction has been proposed to underlie schizophrenia symptoms. This theory arose from the observation that administration of NMDAR antagonists, which are compounds that inhibit NMDAR activity, reproduces behavioural and molecular schizophrenia-like phenotypes, including hallucinations, delusions and cognitive impairments in healthy humans and animal models. However, the role of specific NMDAR subunits in these schizophrenia-relevant phenotypes is largely unknown. Mounting evidence implicates the GluN2D subunit of NMDAR in some of these symptoms and pathology. Firstly, genetic and post-mortem studies show changes in the GluN2D subunit in people with schizophrenia. Secondly, the psychosis-inducing effects of NMDAR antagonists are blunted in GluN2D-knockout mice, suggesting that the GluN2D subunit mediates NMDAR-antagonist-induced psychotomimetic effects. Thirdly, in the mature brain, the GluN2D subunit is relatively enriched in parvalbumin (PV)-containing interneurons, a cell type hypothesized to underlie the cognitive symptoms of schizophrenia. Lastly, the GluN2D subunit is widely and abundantly expressed early in development, which could be of importance considering schizophrenia is a disorder that has its origins in early neurodevelopment. The limitations of currently available therapies warrant further research into novel therapeutic targets such as the GluN2D subunit, which may help us better understand underlying disease mechanisms and develop novel and more effective treatment options.

Characterising seizure development, behavioural comorbidities and neuroinflammation in a self-sustained electrical status epilepticus model of mesial temporal lobe epilepsy in C57BL/6J mice.

OBJECTIVE: Status epilepticus (SE) models in rodents are commonly used to research mesial temporal lobe epilepsy (mTLE) in translational epilepsy research. However, due to differences in susceptibility of mice strains to chemoconvulsants, developing this model in mice is challenging. Mice offer experimental advantages; in particular, the ability to use transgenic strains could provide novel insights about neurobiological mechanisms or ease of genetic modification to test potential therapeutic targets. This study aimed to characterise the neuroinflammation, epileptic seizures and behavioural comorbidities after self-sustained Electrical Status Epilepticus (SSSE) in C57BL/6J mice. METHODS: SSSE was induced in C57BL/6J mice via prolonged electrical stimulation through a bipolar electrode implanted in the ventral hippocampus. Video electroencephalography (vEEG) monitoring was then performed between 1st month (acute timepoint) and 4th month (chronic timepoint). Brain tissues were collected at two timepoints for gene expression and immunohistochemical analysis: 7-days and 16-weeks post-SE. Additionally, at the chronic timepoint, animals underwent a series of neurobehavioural tests. RESULTS: Sixty percent of animals that underwent SSSE developed spontaneous seizures within the first month, and an additional 25% developed seizures at the chronic timepoint. The number of seizures per week during the chronic period ranged from 0.2 to 15.7. Mortality rate was ~9% during or after SSSE. SSSE animals displayed significant spatial memory impairment and depression-like behaviour compared to sham animals. mRNA expression of inflammatory cytokines was upregulated at 7-days following SE, but equal to sham levels at 16-weeks. SIGNIFICANCE: This study provides evidence that SSSE in C57BL/6J mice induces epileptic seizures consistent with those seen in patients with mTLE, along with cognitive and behavioural comorbidities. This model therefore has the potential to be used experimentally to uncover mechanisms to target against epileptogenesis, or to test novel treatment approaches.

11ß-HSD1 participates in epileptogenesis and the associated cognitive impairment by inhibiting apoptosis in mice.

BACKGROUND: Glucocorticoid signalling is closely related to both epilepsy and associated cognitive impairment, possibly through mechanisms involving neuronal apoptosis. As a critical enzyme for glucocorticoid action, the role of 11ß-hydroxysteroid dehydrogenase 1 (11ß-HSD1) in epileptogenesis and associated cognitive impairment has not previously been studied. METHODS: We first investigated the expression of 11ß-HSD1 in the pentylenetetrazole (PTZ) kindling mouse model of epilepsy. We then observed the effect of overexpressing 11ß-HSD1 on the excitability of primary cultured neurons in vitro using whole-cell patch clamp recordings. Further, we assessed the effects of adeno-associated virus (AAV)-induced hippocampal 11ß-HSD1 knockdown in the PTZ model, conducting behavioural observations of seizures, assessment of spatial learning and memory using the Morris water maze, and biochemical and histopathological analyses. RESULTS: We found that 11ß-HSD1 was primarily expressed in neurons but not astrocytes, and its expression was significantly (p < 0.05) increased in the hippocampus of PTZ epilepsy mice compared to sham controls. Whole-cell patch clamp recordings showed that overexpression of 11ß-HSD1 significantly decreased the threshold voltage while increasing the frequency of action potential firing in cultured hippocampal neurons. Hippocampal knockdown of 11ß-HSD1 significantly reduced the severity score of PTZ seizures and increased the latent period required to reach the fully kindled state compared to control knockdown. Knockdown of 11ß-HSD1 also significantly mitigated the impairment of spatial learning and memory, attenuated hippocampal neuronal damage and increased the ratio of Bcl-2/Bax, while decreasing the expression of cleaved caspase-3. CONCLUSIONS: 11ß-HSD1 participates in the pathogenesis of both epilepsy and the associated cognitive impairment by elevating neuronal excitability and contributing to apoptosis and subsequent hippocampal neuronal damage. Inhibition of 11ß-HSD1, therefore, represents a promising strategy to treat epilepsy and cognitive comorbidity.

Evolution, structure and emerging roles of C1ORF112 in DNA replication, DNA damage responses, and cancer.

The C1ORF112 gene initially drew attention when it was found to be strongly co-expressed with several genes previously associated with cancer and implicated in DNA repair and cell cycle regulation, such as RAD51 and the BRCA genes. The molecular functions of C1ORF112 remain poorly understood, yet several studies have uncovered clues as to its potential functions. Here, we review the current knowledge on C1ORF112 biology, its evolutionary history, possible functions, and its potential relevance to cancer. C1ORF112 is conserved throughout eukaryotes, from plants to humans, and is very highly conserved in primates. Protein models suggest that C1ORF112 is an alpha-helical protein. Interestingly, homozygous knockout mice are not viable, suggesting an essential role for C1ORF112 in mammalian development. Gene expression data show that, among human tissues, C1ORF112 is highly expressed in the testes and overexpressed in various cancers when compared to healthy tissues. C1ORF112 has also been shown to have altered levels of expression in some tumours with mutant TP53. Recent screens associate C1ORF112 with DNA replication and reveal possible links to DNA damage repair pathways, including the Fanconi anaemia pathway and homologous recombination. These insights provide important avenues for future research in our efforts to understand the functions and potential disease relevance of C1ORF112.

An Integrated Multi-Omic Network Analysis Identifies Seizure-Associated Dysregulated Pathways in the GAERS Model of Absence Epilepsy.

Absence epilepsy syndromes are part of the genetic generalized epilepsies, the pathogenesis of which remains poorly understood, although a polygenic architecture is presumed. Current focus on single molecule or gene identification to elucidate epileptogenic drivers is unable to fully capture the complex dysfunctional interactions occurring at a genetic/proteomic/metabolomic level. Here, we employ a multi-omic, network-based approach to characterize the molecular signature associated with absence epilepsy-like phenotype seen in a well validated rat model of genetic generalized epilepsy with absence seizures. Electroencephalographic and behavioral data was collected from Genetic Absence Epilepsy Rats from Strasbourg (GAERS, n = 6) and non-epileptic controls (NEC, n = 6), followed by proteomic and metabolomic profiling of the cortical and thalamic tissue of rats from both groups. The general framework of weighted correlation network analysis (WGCNA) was used to identify groups of highly correlated proteins and metabolites, which were then functionally annotated through joint pathway enrichment analysis. In both brain regions a large protein-metabolite module was found to be highly associated with the GAERS strain, absence seizures and associated anxiety and depressive-like phenotype. Quantitative pathway analysis indicated enrichment in oxidative pathways and a downregulation of the lysine degradation pathway in both brain regions. GSTM1 and ALDH2 were identified as central regulatory hubs of the seizure-associated module in the somatosensory cortex and thalamus, respectively. These enzymes are involved in lysine degradation and play important roles in maintaining oxidative balance. We conclude that the dysregulated pathways identified in the seizure-associated module may be involved in the aetiology and maintenance of absence seizure activity. This dysregulated activity could potentially be modulated by targeting one or both central regulatory hubs.

Altered cardiac structure and function is related to seizure frequency in a rat model of chronic acquired temporal lobe epilepsy.

OBJECTIVE: This study aimed to prospectively examine cardiac structure and function in the kainic acid-induced post-status epilepticus (post-KA SE) model of chronic acquired temporal lobe epilepsy (TLE), specifically to examine for changes between the pre-epileptic, early epileptogenesis and the chronic epilepsy stages. We also aimed to examine whether any changes related to the seizure frequency in individual animals. METHODS: Four hours of SE was induced in 9 male Wistar rats at 10 weeks of age, with 8 saline treated matched control rats. Echocardiography was performed prior to the induction of SE, two- and 10-weeks post-SE. Two weeks of continuous video-EEG and simultaneous ECG recordings were acquired for two weeks from 11 weeks post-KA SE. The video-EEG recordings were analyzed blindly to quantify the number and severity of spontaneous seizures, and the ECG recordings analyzed for measures of heart rate variability (HRV). PicroSirius red histology was performed to assess cardiac fibrosis, and intracellular Ca2+ levels and cell contractility were measured by microfluorimetry. RESULTS: All 9 post-KA SE rats were demonstrated to have spontaneous recurrent seizures on the two-week video-EEG recording acquired from 11 weeks SE (seizure frequency ranging from 0.3 to 10.6 seizures/day with the seizure durations from 11 to 62 s), and none of the 8 control rats. Left ventricular wall thickness was thinner, left ventricular internal dimension was shorter, and ejection fraction was significantly decreased in chronically epileptic rats, and was negatively correlated to seizure frequency in individual rats. Diastolic dysfunction was evident in chronically epileptic rats by a decrease in mitral valve deceleration time and an increase in E/E` ratio. Measures of HRV were reduced in the chronically epileptic rats, indicating abnormalities of cardiac autonomic function. Cardiac fibrosis was significantly increased in epileptic rats, positively correlated to seizure frequency, and negatively correlated to ejection fraction. The cardiac fibrosis was not a consequence of direct effect of KA toxicity, as it was not seen in the 6/10 rats from separate cohort that received similar doses of KA but did not go into SE. Cardiomyocyte length, width, volume, and rate of cell lengthening and shortening were significantly reduced in epileptic rats. SIGNIFICANCE: The results from this study demonstrate that chronic epilepsy in the post-KA SE rat model of TLE is associated with a progressive deterioration in cardiac structure and function, with a restrictive cardiomyopathy associated with myocardial fibrosis. Positive correlations between seizure frequency and the severity of the cardiac changes were identified. These results provide new insights into the pathophysiology of cardiac disease in chronic epilepsy, and may have relevance for the heterogeneous mechanisms that place these people at risk of sudden unexplained death.

Altered metabolic pathways in a transgenic mouse model suggest mechanistic role of amyloid precursor protein overexpression in Alzheimer's disease.

INTRODUCTION: The mechanistic role of amyloid precursor protein (APP) in Alzheimer's disease (AD) remains unclear. OBJECTIVES: Here, we aimed to identify alterations in cerebral metabolites and metabolic pathways in cortex, hippocampus and serum samples from Tg2576 mice, a widely used mouse model of AD. METHODS: Metabolomic profilings using liquid chromatography-mass spectrometry were performed and analysed with MetaboAnalyst and weighted correlation network analysis (WGCNA). RESULTS: Expressions of 11 metabolites in cortex, including hydroxyphenyllactate-linked to oxidative stress-and phosphatidylserine-lipid metabolism-were significantly different between Tg2576 and WT mice (false discovery rate < 0.05). Four metabolic pathways from cortex, including glycerophospholipid metabolism and pyrimidine metabolism, and one pathway (sulphur metabolism) from hippocampus, were significantly enriched in Tg2576 mice. Network analysis identified five pathways, including alanine, aspartate and glutamate metabolism, and mitochondria electron transport chain, that were significantly correlated with AD genotype. CONCLUSIONS: Changes in metabolite concentrations and metabolic pathways are present in the early stage of APP pathology, and may be important for AD development and progression.

Daprodustat Compared with Epoetin Beta Pegol for Anemia in Japanese Patients Not on Dialysis: A 52-Week Randomized Open-Label Phase 3 Trial.

BACKGROUND: Daprodustat is an oral agent that stimulates erythropoiesis by inhibiting the prolyl hydroxylases which mark hypoxia-inducible factor for degradation through hydroxylation. Its safety and efficacy (noninferiority) were assessed in this 52-week, open-label study. METHODS: Japanese patients not on dialysis (ND) (N = 299) with anemia of CKD (stages G3, G4, and G5) with iron parameters of ferritin >100 ng/mL or transferrin saturation >20% at screening were randomized to daprodustat or epoetin beta pegol (continuous erythropoietin receptor activator [CERA], also known as methoxy polyethylene glycol-epoetin beta). After initiation of the study, the daprodustat starting dose for erythropoiesis-stimulating agent (ESA)-naïve participants was revised, and daprodustat was started at 2 or 4 mg once daily depending on baseline hemoglobin. ESA users switched to daprodustat 4 mg once daily. CERA was started at 25 µg every 2 weeks for ESA-naïve patients and 25-250 µg every 4 weeks for ESA users based on previous ESA dose. In both treatment groups, dose was adjusted every 4 weeks based on hemoglobin level and changed according to a prespecified algorithm. The primary endpoint was mean hemoglobin level during weeks 40-52 in the intention-to-treat (ITT) population. ESA-naïve patients who entered before the protocol amendment revising the daprodustat starting dose were excluded from the ITT population. RESULTS: Mean hemoglobin levels during weeks 40-52 were 12.0 g/dL in the daprodustat group (n = 108; 95% confidence interval [CI], 11.8-12.1) and 11.9 g/dL for CERA (n = 109; 95% CI 11.7-12.0); the difference between the groups was 0.1 g/dL (95% CI -0.1 to 0.3 g/dL). The lower limit of the 95% CI of the difference was greater than the prespecified margin of -1.0 g/dL. The mean hemoglobin level was within the target range (11.0-13.0 g/dL) during weeks 40-52 for 92% of participants in both groups. There was no meaningful difference in the frequencies of adverse events. CONCLUSIONS: Oral daprodustat was noninferior to CERA in achieving and maintaining target hemoglobin levels in Japanese ND patients. Daprodustat was well tolerated, with no new safety concerns identified.

Astrocyte and glutamate involvement in the pathogenesis of epilepsy in Alzheimer's disease.

Alzheimer's disease (AD) can increase the risk of epilepsy by up to 10-fold compared to healthy age-matched controls. However, the pathological mechanisms that underlie this increased risk are poorly understood. Because disruption in brain glutamate homeostasis has been implicated in both AD and epilepsy, this might play a mechanistic role in the pathogenesis of epilepsy in AD. Prior to the formation of amyloid beta (Aß) plaques, the brain can undergo pathological changes as a result of increased production of amyloid precursor protein (APP) and Aß oligomers. Impairments in the glutamate uptake ability of astrocytes due to astrogliosis are hypothesized to be an early event occurring before Aß plaque formation. Astrogliosis may increase the susceptibility to epileptogenesis of the brain via accumulation of extracellular glutamate and resulting excitotoxicity. Here we hypothesize that Aß oligomers and proinflammatory cytokines can cause astrogliosis and accumulation of extracellular glutamate, which then contribute to the pathogenesis of epilepsy in AD. In this review article, we consider the evidence supporting a potential role of dysfunction of the glutamate-glutamine cycle and the astrocyte in the pathogenesis of epilepsy in AD.

Low prevalence of amyloid and tau pathology in drug-resistant temporal lobe epilepsy.

OBJECTIVE: Cognitive impairment is common in patients with chronic drug-resistant temporal lobe epilepsy (TLE). Hyperphosphorylated tau (pTau) and amyloid-ß (Aß) plaques, pathological hallmarks of Alzheimer disease, have been hypothesized to play a mechanistic role. We investigated Aß plaques and pTau prevalence in TLE patients who underwent resective surgery and correlated their presence with preoperative psychometric test scores and clinical factors. METHODS: Patients were retrospectively selected from the epilepsy surgery register of the Royal Melbourne Hospital, Australia. Sections from the resected temporal lobe were immunostained for pTau and Aß plaques (antibodies: AT8, 1E8). The presence and severity of pathology were correlated with clinical characteristics, and verbal and visual learning functions as measured by the Verbal Pair Associates (VPA) test and Rey Complex Figure Test. RESULTS: Fifty-six patients (55% female) aged 20-68 years (median = 34 years) at surgery were included. Aß plaques were detected in four patients (7%), all at the moderate level. There was no difference in duration, age at onset of epilepsy, or side of resection between patients with and without Aß plaques. Sparse pTau was found in two patients (3.5%). Both had moderate Aß plaques and were >50 years of age. Patients with Aß plaques had a lower median score for the VPA hard assessment compared to those without (0 vs. 4; p = .02). There was otherwise no correlation between pathology and psychometric test scores. SIGNIFICANCE: Aß plaques and pTau were uncommon in the resected brain tissue of patients who have undergone temporal lobectomy, and did not correlate with clinical characteristics or preoperative psychometric test scores, except for a lower VPA median score in patients with Aß plaques. Therefore, considering the low prevalence of Aß plaques and pTau herein observed, it is unlikely that cognitive impairment in TLE is driven by the same mechanisms as in Alzheimer disease.

Antagonism of the ATP-gated P2X7 receptor: a potential therapeutic strategy for cancer.

The P2X receptor 7 (P2X7R) is a plasma membrane receptor sensing extracellular ATP associated with a wide variety of cellular functions. It is most commonly expressed on immune cells and is highly upregulated in a number of human cancers where it can play a trophic role in tumorigenesis. Activation of this receptor leads to the formation of a non-selective cation channel, which has been associated with several cellular functions mediated by the PI3K/Akt pathway and protein kinases. Due to its broad range of functions, the receptor represents a potential therapeutic target for a number of cancers. This review describes the range of mechanisms associated with P2X7R activation in cancer settings and highlights the potential of targeted inhibition of P2X7R as a therapy. It also describes in detail a number of key P2X7R antagonists currently in pre-clinical and clinical development, including oxidised ATP, Brilliant Blue G (BBG), KN-62, KN-04, A740003, A438079, GSK1482160, CE-224535, JNJ-54175446, JNJ-55308942, and AZ10606120. Lastly, it summarises the in vivo studies and clinical trials associated with the use and development of these P2X7R antagonists in different disease contexts.