Project description:Morphological changes in mesial Temporal Lobe Epilepsy with Hippocampal Sclerosis (mTLE-HS) are well characterized. Yet it remains elusive whether these are a consequence of seizures or originate from a hitherto unknown underlying pathology. We recently published data on changes in gene expression (GE) and DNA methylation (DNAm) in the ipsilateral hippocampus (ILH), using the intracortical kainate mouse model of mTLE-HS. In order to explore the effects of epileptic activity alone and also to further disentangle what triggers morphological alterations, we investigated glial and neuronal changes in GE and DNAm in the contralateral hippocampus (CLH). The intracortical kainic acid mouse model of mTLE-HS was used to elicit status epilepticus. Hippocampi contralateral to injection site from 8 kainate injected (KA) and 8 sham (SH) mice were extracted, and shock frozen at 24 hours post injection. Glial- and neuronal nuclei were sorted by flow cytometry. Alterations in GE and DNAm were assessed using reduced representation bisulfite sequencing (RRBS) and RNA sequencing (RNAseq). The R package edgeR was used for statistical analysis. The CLH featured substantial, mostly cell-specific changes in both GE and DNAm in glia and neurons. While changes in GE overlapped to a great degree between CLH and ILH, alterations in DNAm did not. In the CLH we found a significantly lower number of glial genes up- and downregulated compared to previous results from the ILH. Furthermore, several genes and pathways potentially involved in anti-epileptogenic effects were upregulated in the CLH. By comparing GE data from the CLH to previous results from the ILH (featuring hippocampal sclerosis), we derive potential upstream targets for epileptogenesis, including glial Cox2 and Cxcl10. Despite the absence of morphological changes, the CLH displays substantial changes in GE and DNAm. We find that GE changes related to potential anti-epileptogenic effects seem to dominate compared to pro-epileptogenic effects in the CLH and speculate whether this imbalance contributes to prevent morphological alterations like neuronal death and reactive gliosis.

Project description:Neither the molecular basis of the pathologic tendency of neuronal circuits to generate spontaneous seizures (epileptogenicity) nor anti-epileptogenic mechanisms that maintain a seizure-free state are well understood. Here, we performed transcriptomic analysis in the intrahippocampal kainate model of temporal lobe epilepsy in rats using both Agilent and Codelink microarray platforms to characterize the epileptic processes. The experimental design allowed subtraction of the confounding effects of the lesion, identification of expression changes associated with epileptogenicity, and genes upregulated by seizures with potential homeostatic anti-epileptogenic effects. Using differential expression analysis, we identified several hundred expression changes in chronic epilepsy, including candidate genes associated with epileptogenicity such as Bdnf and Kcnj13. To analyze these data from a systems perspective, we applied weighted gene co-expression network analysis (WGCNA) to identify groups of co-expressed genes (modules) and their central (hub) genes. One such module contained genes upregulated in the epileptogenic region, including multiple epileptogenicity candidate genes, and was found to be involved the protection of glial cells against oxidative stress, implicating glial oxidative stress in epileptogenicity. Another distinct module corresponded to the effects of chronic seizures and represented changes in neuronal synaptic vesicle trafficking. We found that the network structure and connectivity of one hub gene, Sv2a, showed significant changes between normal and epileptogenic tissue, becoming more highly connected in epileptic brain. Since Sv2a is a target of the antiepileptic levetiracetam, this module may be important in controlling seizure activity. Bioinformatic analysis of this module also revealed a potential mechanism for the observed transcriptional changes via generation of longer alternatively polyadenlyated transcripts through the upregulation of the RNA binding protein HuD. In summary, combining conventional statistical methods and network analysis allowed us to interpret the differentially regulated genes from a systems perspective, yielding new insight into several biological pathways underlying homeostatic anti-epileptogenic effects and epileptogenicity. Four condition experiment with five samples per condition. The samples include right dentate gyrus from animals with seizures, left dentate gyrus from animals with seizures, right dentate gyrus from animals without seizures, and left dentate gyrus from animals without seizures. A dye swap was included.

Project description:Neither the molecular basis of the pathologic tendency of neuronal circuits to generate spontaneous seizures (epileptogenicity) nor anti-epileptogenic mechanisms that maintain a seizure-free state are well understood. Here, we performed transcriptomic analysis in the intrahippocampal kainate model of temporal lobe epilepsy in rats using both Agilent and Codelink microarray platforms to characterize the epileptic processes. The experimental design allowed subtraction of the confounding effects of the lesion, identification of expression changes associated with epileptogenicity, and genes upregulated by seizures with potential homeostatic anti-epileptogenic effects. Using differential expression analysis, we identified several hundred expression changes in chronic epilepsy, including candidate genes associated with epileptogenicity such as Bdnf and Kcnj13. To analyze these data from a systems perspective, we applied weighted gene co-expression network analysis (WGCNA) to identify groups of co-expressed genes (modules) and their central (hub) genes. One such module contained genes upregulated in the epileptogenic region, including multiple epileptogenicity candidate genes, and was found to be involved the protection of glial cells against oxidative stress, implicating glial oxidative stress in epileptogenicity. Another distinct module corresponded to the effects of chronic seizures and represented changes in neuronal synaptic vesicle trafficking. We found that the network structure and connectivity of one hub gene, Sv2a, showed significant changes between normal and epileptogenic tissue, becoming more highly connected in epileptic brain. Since Sv2a is a target of the antiepileptic levetiracetam, this module may be important in controlling seizure activity. Bioinformatic analysis of this module also revealed a potential mechanism for the observed transcriptional changes via generation of longer alternatively polyadenlyated transcripts through the upregulation of the RNA binding protein HuD. In summary, combining conventional statistical methods and network analysis allowed us to interpret the differentially regulated genes from a systems perspective, yielding new insight into several biological pathways underlying homeostatic anti-epileptogenic effects and epileptogenicity. Four condition experiment with five samples per condition. The samples include right dentate gyrus from animals with seizures, left dentate gyrus from animals with seizures, right dentate gyrus from animals without seizures, and left dentate gyrus from animals without seizures.

Project description:Early childhood convulsions have been correlated with hippocampal neuron loss in patients with intractable temporal lobe epilepsy. Using a "two-hit" rat seizure model, we have shown that animals subjected to kainate (KA)- or hypoxia-induced seizures during early postnatal period showed no cell death, yet sustained more extensive neuronal death after second seizures in adulthood. An early life seizure, without causing overt cellular injury, predisposes the brain to the damaging effect of seizures in later life. Cellular and molecular changes that accompany early seizures and that lead to subsequent epileptogenesis and increased susceptibility to seizure-induced neuronal injury, however, remain poorly understood. We propose to investigate age-specific, time-dependent changes in gene expression that may underlie this priming effect of early-life seizures. We will determine the sequence of gene expression pattern in the hippocampus at various times following KA induced seizures at postnatal day (P) 15. Previous studies have shown that AMPA receptor subtype of glutamate receptors play a crucial role in the age-specific vulnerability and in the long-term epileptogenic effects of perinatal hypoxia seizures. We found that AMPA receptor antagonists block the increased susceptibility caused by early life seizures to later seizures and seizure-induced brain damage. We hypothesize that an alteration of AMPA receptor composition is one of many changes caused by early-life seizures that leads to an increase in Ca2+ permeability, which then results in cascade of downstream events and modifies array of gene expression that promote epileptogenesis and susceptibility to neuronal death in later life. We will examine three time points: 1hr, 72 hr, and 15 days following systemic KA-induced seizures at P15 as we have previously observed structural changes within the hippocampus at these time points. Within an hour of KA seizures, a marked swelling of dendrites, disassembly of dendritic microtubules and glycogen depletion are observed by electron microscopy. Within 5 days, basal dendrites of CA3 hippocampal pyramidal neurons show abnormal spine morphology and decreased branching pattern. 15 days after the seizures, aberrant growth of mossy fibers in the CA3 stratum oriens is observed in animals exposed to KA. Ten hippocampi will be pooled from five animals treated with KA (3mg/kg i.p.) and from five littermate controls injected with PBS. Animals will be decapitated and hippocampi will be rapidly dissected from the brain, flash frozen in liquid nitrogen, and stored at -80C until extraction of total RNA, which will be sent to the center. We will provide 4 tissue samples-2 controls and 2 KA, each a pool of five animals - for each time points. Mixing tissues from multiple rats will normalize single nucleotide polymorphisms and tissue heterogeneity.

Project description:Neither the molecular basis of the pathologic tendency of neuronal circuits to generate spontaneous seizures (epileptogenicity) nor anti-epileptogenic mechanisms that maintain a seizure-free state are well understood. Here, we performed transcriptomic analysis in the intrahippocampal kainate model of temporal lobe epilepsy in rats using both Agilent and Codelink microarray platforms to characterize the epileptic processes. The experimental design allowed subtraction of the confounding effects of the lesion, identification of expression changes associated with epileptogenicity, and genes upregulated by seizures with potential homeostatic anti-epileptogenic effects. Using differential expression analysis, we identified several hundred expression changes in chronic epilepsy, including candidate genes associated with epileptogenicity such as Bdnf and Kcnj13. To analyze these data from a systems perspective, we applied weighted gene co-expression network analysis (WGCNA) to identify groups of co-expressed genes (modules) and their central (hub) genes. One such module contained genes upregulated in the epileptogenic region, including multiple epileptogenicity candidate genes, and was found to be involved the protection of glial cells against oxidative stress, implicating glial oxidative stress in epileptogenicity. Another distinct module corresponded to the effects of chronic seizures and represented changes in neuronal synaptic vesicle trafficking. We found that the network structure and connectivity of one hub gene, Sv2a, showed significant changes between normal and epileptogenic tissue, becoming more highly connected in epileptic brain. Since Sv2a is a target of the antiepileptic levetiracetam, this module may be important in controlling seizure activity. Bioinformatic analysis of this module also revealed a potential mechanism for the observed transcriptional changes via generation of longer alternatively polyadenlyated transcripts through the upregulation of the RNA binding protein HuD. In summary, combining conventional statistical methods and network analysis allowed us to interpret the differentially regulated genes from a systems perspective, yielding new insight into several biological pathways underlying homeostatic anti-epileptogenic effects and epileptogenicity.

Dataset Information

Transcriptomic profile of the hippocampus after status epilepticus in wildtype and EZH2 neuronal knockout mice.

Dataset's files

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets