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| <?xml version="1.0" encoding="UTF-8"?><!DOCTYPE article PUBLIC "-//NLM//DTD JATS (Z39.96) Journal Archiving and Interchange DTD v1.1d1 20130915//EN" "JATS-archivearticle1.dtd"><article article-type="research-article" dtd-version="1.1d1" xmlns:xlink="http://www.w3.org/1999/xlink"><front><journal-meta><journal-id journal-id-type="nlm-ta">elife</journal-id><journal-id journal-id-type="hwp">eLife</journal-id><journal-id journal-id-type="publisher-id">eLife</journal-id><journal-title-group><journal-title>eLife</journal-title></journal-title-group><issn publication-format="electronic">2050-084X</issn><publisher><publisher-name>eLife Sciences Publications, Ltd</publisher-name></publisher></journal-meta><article-meta><article-id pub-id-type="publisher-id">01267</article-id><article-id pub-id-type="doi">10.7554/eLife.01267</article-id><article-categories><subj-group subj-group-type="display-channel"><subject>Research article</subject></subj-group><subj-group subj-group-type="heading"><subject>Cell biology</subject></subj-group><subj-group subj-group-type="heading"><subject>Neuroscience</subject></subj-group></article-categories><title-group><article-title>The transcription factor NRSF contributes to epileptogenesis by selective repression of a subset of target genes</article-title></title-group><contrib-group><contrib contrib-type="author" equal-contrib="yes" id="author-6795"><name><surname>McClelland</surname><given-names>Shawn</given-names></name><xref ref-type="aff" rid="aff1"/><xref ref-type="aff" rid="aff2"/><xref ref-type="aff" rid="aff4"/><xref ref-type="fn" rid="equal-contrib">†</xref><xref ref-type="fn" rid="con1"/><xref ref-type="fn" rid="conf1"/></contrib><contrib contrib-type="author" equal-contrib="yes" id="author-6796"><name><surname>Brennan</surname><given-names>Gary P</given-names></name><xref ref-type="aff" rid="aff1"/><xref ref-type="aff" rid="aff2"/><xref ref-type="aff" rid="aff4"/><xref ref-type="fn" rid="equal-contrib">†</xref><xref ref-type="fn" rid="con2"/><xref ref-type="fn" rid="conf1"/></contrib><contrib contrib-type="author" id="author-16571"><name><surname>Dubé</surname><given-names>Celine</given-names></name><xref ref-type="aff" rid="aff1"/><xref ref-type="aff" rid="aff2"/><xref ref-type="aff" rid="aff4"/><xref ref-type="fn" rid="con3"/><xref ref-type="fn" rid="conf1"/></contrib><contrib contrib-type="author" id="author-6798"><name><surname>Rajpara</surname><given-names>Seeta</given-names></name><xref ref-type="aff" rid="aff1"/><xref ref-type="aff" rid="aff2"/><xref ref-type="aff" rid="aff4"/><xref ref-type="fn" rid="con4"/><xref ref-type="fn" rid="conf1"/></contrib><contrib contrib-type="author" id="author-15965"><name><surname>Iyer</surname><given-names>Shruti</given-names></name><xref ref-type="aff" rid="aff1"/><xref ref-type="aff" rid="aff2"/><xref ref-type="aff" rid="aff4"/><xref ref-type="fn" rid="con5"/><xref ref-type="fn" rid="conf1"/></contrib><contrib contrib-type="author" id="author-6861"><name><surname>Richichi</surname><given-names>Cristina</given-names></name><xref ref-type="aff" rid="aff1"/><xref ref-type="aff" rid="aff2"/><xref ref-type="aff" rid="aff4"/><xref ref-type="fn" rid="con6"/><xref ref-type="fn" rid="conf1"/></contrib><contrib contrib-type="author" id="author-6800"><name><surname>Bernard</surname><given-names>Christophe</given-names></name><xref ref-type="aff" rid="aff3"/><xref ref-type="other" rid="par-2"/><xref ref-type="fn" rid="con7"/><xref ref-type="fn" rid="conf1"/></contrib><contrib contrib-type="author" corresp="yes" id="author-6644"><name><surname>Baram</surname><given-names>Tallie Z</given-names></name><xref ref-type="aff" rid="aff1"/><xref ref-type="aff" rid="aff2"/><xref ref-type="aff" rid="aff4"/><xref ref-type="corresp" rid="cor1">*</xref><xref ref-type="other" rid="par-1"/><xref ref-type="fn" rid="con8"/><xref ref-type="fn" rid="conf1"/></contrib><aff id="aff1"><institution content-type="dept">Department of Anatomy and Neurobiology</institution>, <institution>University of California, Irvine</institution>, <addr-line><named-content content-type="city">Irvine</named-content></addr-line>, <country>United States</country></aff><aff id="aff2"><institution content-type="dept">Department of Pediatrics</institution>, <institution>University of California, Irvine</institution>, <addr-line><named-content content-type="city">Irvine</named-content></addr-line>, <country>United States</country></aff><aff id="aff3"><institution content-type="dept">Laboratoire Epilepsie et Cognition</institution>, <institution>Institut National de la Santé et de la Recherche Médicale</institution>, <addr-line><named-content content-type="city">Marseille</named-content></addr-line>, <country>France</country></aff><aff id="aff4"><institution content-type="dept">Department of Neurology</institution>, <institution>University of California, Irvine</institution>, <addr-line><named-content content-type="city">Irvine</named-content></addr-line>, <country>United States</country></aff></contrib-group><contrib-group content-type="section"><contrib contrib-type="editor"><name><surname>Levitt</surname><given-names>Pat</given-names></name><role>Reviewing editor</role><aff><institution>Children's Hospital Los Angeles</institution>, <country>United States</country></aff></contrib></contrib-group><author-notes><corresp id="cor1"><label>*</label>For correspondence: <email>tallie@uci.edu</email></corresp><fn fn-type="con" id="equal-contrib"><label>†</label><p>These authors contributed equally to this work</p></fn></author-notes><pub-date date-type="pub" publication-format="electronic"><day>12</day><month>08</month><year>2014</year></pub-date><pub-date pub-type="collection"><year>2014</year></pub-date><volume>3</volume><elocation-id>e01267</elocation-id><history><date date-type="received"><day>20</day><month>07</month><year>2013</year></date><date date-type="accepted"><day>17</day><month>07</month><year>2014</year></date></history><permissions><copyright-statement>© 2014, McClelland et al</copyright-statement><copyright-year>2014</copyright-year><copyright-holder>McClelland et al</copyright-holder><license xlink:href="http://creativecommons.org/licenses/by/4.0/"><license-p>This article is distributed under the terms of the <ext-link ext-link-type="uri" xlink:href="http://creativecommons.org/licenses/by/4.0/">Creative Commons Attribution License</ext-link>, which permits unrestricted use and redistribution provided that the original author and source are credited.</license-p></license></permissions><self-uri content-type="pdf" xlink:href="elife01267.pdf"/><abstract><object-id pub-id-type="doi">10.7554/eLife.01267.001</object-id><p>The mechanisms generating epileptic neuronal networks following insults such as severe seizures are unknown. We have previously shown that interfering with the function of the neuron-restrictive silencer factor (NRSF/REST), an important transcription factor that influences neuronal phenotype, attenuated development of this disorder. In this study, we found that epilepsy-provoking seizures increased the low NRSF levels in mature hippocampus several fold yet surprisingly, provoked repression of only a subset (∼10%) of potential NRSF target genes. Accordingly, the repressed gene-set was rescued when NRSF binding to chromatin was blocked. Unexpectedly, genes selectively repressed by NRSF had mid-range binding frequencies to the repressor, a property that rendered them sensitive to moderate fluctuations of NRSF levels. Genes selectively regulated by NRSF during epileptogenesis coded for ion channels, receptors, and other crucial contributors to neuronal function. Thus, dynamic, selective regulation of NRSF target genes may play a role in influencing neuronal properties in pathological and physiological contexts.</p><p><bold>DOI:</bold> <ext-link ext-link-type="doi" xlink:href="10.7554/eLife.01267.001">http://dx.doi.org/10.7554/eLife.01267.001</ext-link></p></abstract><abstract abstract-type="executive-summary"><object-id pub-id-type="doi">10.7554/eLife.01267.002</object-id><title>eLife digest</title><p>Epilepsy is a common brain disease that can cause disabling seizures. During a seizure, brain cells send out abnormal signals, which can mean that people having seizures may be unaware of their surroundings and may fall or otherwise injure themselves.</p><p>Individuals with epilepsy develop changes in their brain cells and in the circuits that connect these cells together. Some people develop epilepsy because they have mutations in genes. Others develop the condition after an injury or a long seizure, which leads to changes in gene expression and therefore changes to the brain's cells and circuits.</p><p>In 2011, researchers found that a protein that normally switches off the expression of certain genes during brain development, but which is almost absent in the adult brain, may run amok after a seizure. The level of this protein—a transcription factor called NRSF—increased in the brains of rats that had been caused to have a seizure. A long provoked seizure caused many of the rats to develop epilepsy. But, if NRSF was blocked after the original seizure, the rats were less likely to have further seizures later on. Now McClelland et al., including several of the researchers involved in the 2011 work, have examined what normally happens to the expression of genes after a seizure and what happens when the NRSF transcription factor is blocked.</p><p>McClelland et al. found that only a small subset—about 10%—of the genes that can theoretically be silenced by NRSF are switched off in the brain when this protein's levels increase after a seizure. The increased NRSF levels, unexpectedly, did not affect the genes that bind tightly to this transcription factor. Nor did NRSF affect genes that bind loosely. Instead, the genes that the transcription factor binds to with an intermediate strength were the ones that were switched off. McClelland et al. suggest that this ‘mid-range binding’ to NRSF allows the expression of these genes to be increased or decreased in response to there being more or less NRSF in the cell. Genes that bind tightly to NRSF are likely to already have a lot of NRSF bound and are therefore already switched off; and loosely-binding genes would likely need even more NRSF before they are switched off.</p><p>The subset of genes that were switched off by the increased levels of NRSF after a seizure code for a number of proteins that brain cells need to be able to effectively send and receive messages. Blocking the ability of NRSF to bind to these genes and switch them off may help to prevent the brain changes that cause epilepsy.</p><p><bold>DOI:</bold> <ext-link ext-link-type="doi" xlink:href="10.7554/eLife.01267.002">http://dx.doi.org/10.7554/eLife.01267.002</ext-link></p></abstract><kwd-group kwd-group-type="author-keywords"><title>Author keywords</title><kwd>neuron-restrictive silencing factor</kwd><kwd>epilepsy</kwd><kwd>gene set enrichment analysis</kwd></kwd-group><kwd-group kwd-group-type="research-organism"><title>Research organism</title><kwd>rat</kwd></kwd-group><funding-group><award-group id="par-1"><funding-source><institution-wrap><institution-id institution-id-type="FundRef">http://dx.doi.org/10.13039/100000002</institution-id><institution>National Institutes of Health</institution></institution-wrap></funding-source><award-id>R37 NS 35439, T32 NS45540</award-id><principal-award-recipient><name><surname>Baram</surname><given-names>Tallie Z</given-names></name></principal-award-recipient></award-group><award-group id="par-2"><funding-source><institution-wrap><institution-id institution-id-type="FundRef">http://dx.doi.org/10.13039/501100001677</institution-id><institution>Institut national de la santé et de la recherche médicale</institution></institution-wrap></funding-source><award-id>CURE</award-id><principal-award-recipient><name><surname>Bernard</surname><given-names>Christophe</given-names></name></principal-award-recipient></award-group><funding-statement>The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.</funding-statement></funding-group><custom-meta-group><custom-meta><meta-name>elife-xml-version</meta-name><meta-value>2</meta-value></custom-meta><custom-meta specific-use="meta-only"><meta-name>Author impact statement</meta-name><meta-value>Uncovering the mechanisms by which the transcription factor NRSF contributes to the development of epilepsy reveals that the regulation of gene expression by transcription factors in the brain is more finely-tuned than previously thought.</meta-value></custom-meta></custom-meta-group></article-meta></front><body><sec id="s1" sec-type="intro"><title>Introduction</title><p>Epileptic networks arise via numerous mechanisms (<xref ref-type="bibr" rid="bib70">Pitkänen et al., 2011</xref>; <xref ref-type="bibr" rid="bib98">Vezzani et al., 2011</xref>; <xref ref-type="bibr" rid="bib35">Kim et al., 2012</xref>; <xref ref-type="bibr" rid="bib26">Hildebrand et al., 2013</xref>; <xref ref-type="bibr" rid="bib80">Rowley and Patel, 2013</xref>). Changes in neuronal properties and resulting alterations of network behavior constitute one of the important mechanisms of epileptogenesis (<xref ref-type="bibr" rid="bib99">Yaari and Beck, 2002</xref>; <xref ref-type="bibr" rid="bib59">Noam et al., 2011</xref>; <xref ref-type="bibr" rid="bib23">Goldberg and Coulter, 2013</xref>). Neuronal properties may change because of genetic mutations in crucial neuronal genes (<xref ref-type="bibr" rid="bib53">Meisler and O'Brien, 2012</xref>; <xref ref-type="bibr" rid="bib62">Oliva et al., 2012</xref>; <xref ref-type="bibr" rid="bib36">Kingwell, 2013</xref>; <xref ref-type="bibr" rid="bib65">Papale et al., 2013</xref>) or from insult-related alterations of gene expression that affect the levels and function of their products. Whereas altered expression of individual ion channels (<xref ref-type="bibr" rid="bib10">Brewster et al., 2002</xref>; <xref ref-type="bibr" rid="bib34">Khirug et al., 2010</xref>; <xref ref-type="bibr" rid="bib71">Poolos, 2012</xref>; <xref ref-type="bibr" rid="bib97">van Loo et al., 2012</xref>; <xref ref-type="bibr" rid="bib49">Maslarova et al., 2013</xref>; <xref ref-type="bibr" rid="bib85">Shah et al., 2013</xref>), neurotransmitter receptors (<xref ref-type="bibr" rid="bib24">Gonzalez et al., 2013</xref>; <xref ref-type="bibr" rid="bib77">Rojas and Dingledine, 2013</xref>), and other cellular components (<xref ref-type="bibr" rid="bib104">Zeng et al., 2009</xref>; <xref ref-type="bibr" rid="bib50">Maroso et al., 2011</xref>; <xref ref-type="bibr" rid="bib30">Jimenez-Mateos et al., 2012</xref>; <xref ref-type="bibr" rid="bib45">Liu et al., 2013</xref>) have been extensively reported, the nature and mechanisms of the orchestration of these gene expression changes remain unknown: do epilepsy-provoking insults regulate individual genes via distinct cellular pathways? Alternatively, are there clusters of genes that are co-regulated based on specific common properties, and whose altered expression contributes to the generation of ‘epileptic neurons’? (<xref ref-type="bibr" rid="bib12">Brooks-Kayal et al., 2009</xref>; <xref ref-type="bibr" rid="bib52">McClelland et al., 2011b</xref>; <xref ref-type="bibr" rid="bib37">Kobow and Blümcke, 2012</xref>; <xref ref-type="bibr" rid="bib78">Roopra et al., 2012</xref>; <xref ref-type="bibr" rid="bib97">van Loo et al., 2012</xref>). These questions are critical for the understanding and future prevention of certain epilepsies.</p><p>Gene expression is dynamically regulated throughout life and influences neuronal phenotype and behavior (<xref ref-type="bibr" rid="bib5">Bale et al., 2010</xref>; <xref ref-type="bibr" rid="bib75">Robison and Nestler, 2011</xref>; <xref ref-type="bibr" rid="bib93">Sweatt, 2013</xref>). This dynamic regulation is mediated by transcriptional enhancers and repressors that bind to specific sequences of target genes and modify their expression (<xref ref-type="bibr" rid="bib6">Ballas et al., 2005</xref>; <xref ref-type="bibr" rid="bib39">Kohyama et al., 2008</xref>; <xref ref-type="bibr" rid="bib29">Hwang et al., 2010</xref>). The repressor neuron-restrictive silencer factor (NRSF/ REST) functions by binding a DNA sequence called neuron-restrictive silencer element (NRSE) to mediate long-term, cell-specific gene repression (<xref ref-type="bibr" rid="bib83">Schoenherr and Anderson, 1995</xref>) via complex interactions with co-repressors such as mSin3 and Co-Rest (<xref ref-type="bibr" rid="bib14">Chen et al., 1998</xref>; <xref ref-type="bibr" rid="bib3">Andres et al., 1999</xref>; <xref ref-type="bibr" rid="bib56">Naruse et al., 1999</xref>; <xref ref-type="bibr" rid="bib79">Roopra et al., 2001</xref>; <xref ref-type="bibr" rid="bib102">Wood et al., 2003</xref>; <xref ref-type="bibr" rid="bib8">Belyaev et al., 2004</xref>; <xref ref-type="bibr" rid="bib41">Kuwabara et al., 2004</xref>; <xref ref-type="bibr" rid="bib55">Mortazavi et al., 2006</xref>; <xref ref-type="bibr" rid="bib31">Johnson et al., 2007</xref>; <xref ref-type="bibr" rid="bib1">Abrajano et al., 2009</xref>). NRSF is highly expressed in non-neuronal tissues where it represses neuronal genes (<xref ref-type="bibr" rid="bib7">Ballas and Mandel, 2005</xref>) and has important roles in neurogenesis and neuronal differentation within the brain (<xref ref-type="bibr" rid="bib27">Hsieh et al., 2004</xref>; <xref ref-type="bibr" rid="bib87">Singh et al., 2008</xref>; <xref ref-type="bibr" rid="bib100">Westbrook et al., 2008</xref>; <xref ref-type="bibr" rid="bib48">Mandel et al., 2011</xref>; <xref ref-type="bibr" rid="bib4">Aoki et al., 2012</xref>; <xref ref-type="bibr" rid="bib17">Ernsberger, 2012</xref>; <xref ref-type="bibr" rid="bib38">Kok et al., 2012</xref>; <xref ref-type="bibr" rid="bib76">Rodenas-Ruano et al., 2012</xref>).</p><p>NRSF levels are low in mature hippocampus where ∼600 expressed genes contain NRSEs and are thus potentially regulated by the repressor. Hippocampal NRSF expression increases several-fold following seizures (<xref ref-type="bibr" rid="bib63">Palm et al., 1998</xref>; <xref ref-type="bibr" rid="bib19">Garriga-Canut et al., 2006</xref>; <xref ref-type="bibr" rid="bib51">McClelland et al., 2011a</xref>) and following other insults that promote epilepsy (<xref ref-type="bibr" rid="bib13">Calderone et al., 2003</xref>; <xref ref-type="bibr" rid="bib60">Noh et al., 2012</xref>; <xref ref-type="bibr" rid="bib33">Kaneko et al., 2014</xref>). We found that blocking NRSF function attenuated the epileptic phenotype induced by severe seizures (<xref ref-type="bibr" rid="bib51">McClelland et al., 2011a</xref>). However, it is unclear ‘how’ augmented NRSF levels promote selective neuronal phenotypic changes that provoke epilepsy. In this study, we used large-scale transcriptome arrays, qPCR validation, chromatin immunoprecipitation (ChIP), and gene set enrichment analysis (GSEA) to examine the basis of NRSF-dependent gene expression changes during epileptogenesis. Surprisingly, a twofold to threefold increase of NRSF levels, induced by epilepsy-provoking seizures, led to repression of only a subset of potentially regulated genes, and these were rescued by interfering with NRSF function. Unexpectedly, genes found to be regulated by NRSF were characterized by mid-range NRSF-binding probabilities during the naive state. This moderate baseline binding enabled a significant increase of NRSF binding to these genes upon modest increases in NRSF tissue levels. NRSF-regulated hippocampal genes included ion channels, receptors, calcium-related molecules, and other transcription factors that govern neuronal function and plasticity, suggesting that their repression might contribute to the pathogenesis of epilepsy.</p></sec><sec id="s2" sec-type="results"><title>Results</title><sec id="s2-1"><title>NRSE-containing genes are preferentially repressed by seizure-provoked increase of hippocampal NRSF levels</title><p>NRSF mRNA and protein expression in mature hippocampus were increased following long seizures induced by systemic administration of the glutamate receptor agonist, kainic acid (KA) (<xref ref-type="fig" rid="fig1">Figure 1A,B</xref>). The direct dependence of increased NRSF expression on network activity (seizures) was tested by examining NRSF levels in vitro, using organotypic hippocampal slice cultures, where most potential confounders were excluded. KA provoked seizure-like electrophysiological events in the hippocampal slice (<xref ref-type="bibr" rid="bib74">Richichi et al., 2008</xref>) and led a to time-dependent increase of NRSF mRNA and protein levels (<xref ref-type="fig" rid="fig1">Figure 1C,D</xref>).<fig id="fig1" position="float"><object-id pub-id-type="doi">10.7554/eLife.01267.003</object-id><label>Figure 1.</label><caption><title>Kainic acid (KA)-seizure-induced increase of NRSF expression.</title><p>(<bold>A</bold>) Time-course of NRSF mRNA expression levels following KA-induced seizures, n = 4/group. (<bold>B</bold>) Representative western blot image of NRSF protein levels in control (ctrl) animals and animals at 72 hr and 1 week post KA-induced seizures. Quantification of NRSF protein levels using optical density measurements (ctrl 1.96 ± 0.18, n = 6; KA+72hr 6.75 ± 0.54, n = 3; KA+1 week: 5.42 ± 0.36, n = 3). (<bold>C</bold> and <bold>D</bold>) In situ hybridization and quantification of NRSF mRNA in organotypic hippocampal slice cultures which had undergone KA-induced seizure-like events. Quantification of mRNA in pyramidal cell layer was performed in control cultures as well as cultures 4 hr, 12 hr, 24 hr, and 1 week following seizure-like events in CA1 and CA3 region of the hippocampus, n = 4–8/group, *p<0.05.</p><p><bold>DOI:</bold> <ext-link ext-link-type="doi" xlink:href="10.7554/eLife.01267.003">http://dx.doi.org/10.7554/eLife.01267.003</ext-link></p></caption><graphic xlink:href="elife01267f001"/></fig></p><p>Because cellular plasticity often derives from large-scale changes in gene expression (<xref ref-type="bibr" rid="bib75">Robison and Nestler, 2011</xref>), we examined for such large-scale expression changes here using a microarray analysis. We initially screened the microarray data for candidate genes whose expression was repressed following KA-induced seizures and examined whether these genes contained functional NRSEs. Expression of 12,996 genes was detected in the CA1 region of the rat hippocampus of which 371 (2.8%) contained NRSE sites (<xref ref-type="bibr" rid="bib31">Johnson et al., 2007</xref>). 470 genes whose expression was significantly repressed by KA-induced seizures were identified by this initial candidate gene analysis that employed routine statistical significance (<xref ref-type="bibr" rid="bib92">Subramanian et al., 2005</xref>; <xref ref-type="bibr" rid="bib103">Yukhananov and Kissin, 2008</xref>; <xref ref-type="bibr" rid="bib9">Borg et al., 2010</xref>; <xref ref-type="bibr" rid="bib61">Oberbauer et al., 2010</xref>; <xref ref-type="bibr" rid="bib90">Stark et al., 2010</xref>). Of these, 39 (8.3%) contained NRSF binding sites (<xref ref-type="fig" rid="fig2">Figure 2A</xref>). The significant enrichment in the fraction of NRSE-containing genes in the repressed vs total gene population (8.3% vs 2.8%) indicated that NRSF-regulated genes were preferentially repressed by epilepsy-provoking network hyperactivity (<xref ref-type="fig" rid="fig2">Figure 2B</xref>).<fig id="fig2" position="float"><object-id pub-id-type="doi">10.7554/eLife.01267.004</object-id><label>Figure 2.</label><caption><title>NRSE-containing genes are enriched among hippocampal genes repressed after network activity.</title><p>(<bold>A</bold>) Heat map representing genes with repressed mRNA expression levels in two representative samples from hippocampi derived from control (Ctrl) and from hippocampi from KA-seizures-experiencing rats. Expression levels are represented by color using a scale from 50% to 150% of expression, where yellow is the highest and blue is the lowest. (<bold>B</bold>) Relative abundance of NRSF target genes among total detected genes vs those repressed by KA-seizures.</p><p><bold>DOI:</bold> <ext-link ext-link-type="doi" xlink:href="10.7554/eLife.01267.004">http://dx.doi.org/10.7554/eLife.01267.004</ext-link></p></caption><graphic xlink:href="elife01267f002"/></fig></p></sec><sec id="s2-2"><title>Only a subset of NRSE-containing genes are regulated by NRSF in mature hippocampus</title><p>Whereas 371 of the detected hippocampal genes contained an NRSE and were therefore potential NRSF targets, only 39 of these were repressed by KA-induced seizures when NRSF levels increased twofold to threefold. To examine if the correlation between NRSF levels and the repression of this subset (∼10%) of NRSE-containing genes was causal, we employed a decoy oligonucleotide strategy to block NRSF binding to the NRSE sequences of genomic DNA (<xref ref-type="bibr" rid="bib2">Akhtar et al., 1991</xref>; <xref ref-type="bibr" rid="bib94">Szklarczyk and Kaczmarek, 1995</xref>; <xref ref-type="bibr" rid="bib21">Gilar et al., 1998</xref>; <xref ref-type="bibr" rid="bib89">Soldati et al., 2011</xref>; <xref ref-type="bibr" rid="bib84">Sedaghat et al., 2013</xref>). We generated oligodeoxynucleotides (ODNs) comprised of the NRSF binding sequence (NRSE), modified their backbone for stability and infused them into the brain. These ODNs acted as ‘decoys’, binding to cellular NRSF and inhibiting its ability to bind to target genes (<xref ref-type="fig" rid="fig3">Figure 3A</xref>) (<xref ref-type="bibr" rid="bib51">McClelland et al., 2011a</xref>).<fig id="fig3" position="float"><object-id pub-id-type="doi">10.7554/eLife.01267.005</object-id><label>Figure 3.</label><caption><title>Abrogation of NRSF binding to target genes rescues the majority of NRSE-containing genes repressed by KA-seizures.</title><p>(<bold>A</bold>) A schematic illustrating the mechanism of action of NRSF following a seizure-induced increase and the decoy oligodeoxynucleotide (ODN) intervention strategy with the expected outcome. (<bold>B</bold>) A heat map representation of the changes in mRNA expression levels of genes that contain a putative NRSE site that were down-regulated 48 hr after KA-induced seizures. Heat map compares representative samples from two hippocampi, from each of four experimental conditions: 'controls' receiving random ODNs (n = 4); ‘controls’ receiving NRSE ODNs (n = 4); ‘KA-seizures’, rats sustaining KA induced seizure activity and receiving random ODNs (n = 3); ‘KA-seizures + NRSE-ODN’, rats sustaining KA-induced seizure activity and receiving NRSE-ODNs (n = 4). Samples and genes are plotted using hierarchical clustering using Euclidean distance and average linkage. Expression level is depicted by color using a scale from 50% to 150% of expression, where yellow is the highest and blue is the lowest. (<bold>C</bold>) Independent analysis of gene expression using qPCR. Several genes that were both repressed by seizure activity and rescued by interference with NRSF function were tested (Glra2, <italic>Myo5B, Stmn2</italic>), and results analyzed using two way ANOVA. <italic>Myo5B</italic> F<sub>(1,18)</sub> = 9.35, p = 0.007; <italic>Glra2</italic> F<sub>(1,17)</sub> = 46.89, p = 0.0001; <italic>Stmn2</italic> F<sub>(1,18)</sub> = 1.97, p = 0.047, n = 4/group.</p><p><bold>DOI:</bold> <ext-link ext-link-type="doi" xlink:href="10.7554/eLife.01267.005">http://dx.doi.org/10.7554/eLife.01267.005</ext-link></p></caption><graphic xlink:href="elife01267f003"/></fig></p><p>Consistent with the low baseline levels of NRSF in naive hippocampus, preventing the binding of the repressor to target genes had little effects on the expression of most of the 39 genes (compare “random ODN” and NRSE–ODN columns in the control brain, left side of <xref ref-type="fig" rid="fig3">Figure 3B</xref>). Of the 39 NRSE-containing genes that were repressed by seizures in the presence of random ODNs, (depicted in blue in <xref ref-type="fig" rid="fig3">Figure 3B</xref>) the majority (28) were no longer significantly repressed when NRSF binding to target genes was inhibited (compare the [KA-seizures + NRSE ODN] group to either of the control groups; <xref ref-type="fig" rid="fig3">Figure 3B</xref>). These microarray data supported the idea that in mature hippocampus a twofold to threefold change in NRSF levels influenced the expression of only a subset of putative NRSF target genes and this repression required binding of NRSF to the chromatin. To examine the validity of the microarray data supporting this observation, we performed qPCR on a subset of genes to confirm both their seizure-induced repression as well as their ‘rescue’ by NRSE-ODN administration (<xref ref-type="fig" rid="fig3">Figure 3C</xref>).</p></sec><sec id="s2-3"><title>Physical binding of NRSF co-varies with tissue levels specifically at genes that are regulated by the repressor</title><p>We next examined the microarray data by performing qPCR analysis of a subset of NRSE-containing genes that were found to be repressed on the array and a second subset of genes whose expression was not altered on the array following KA-seizures (<xref ref-type="fig" rid="fig4">Figure 4A,B</xref>). These qPCR analyses revealed excellent correlation with the microarray and both methods distinguished between the NRSE-containing genes that were or were not repressed by seizures.<fig id="fig4" position="float"><object-id pub-id-type="doi">10.7554/eLife.01267.006</object-id><label>Figure 4.</label><caption><title>qPCR validation of microarray results.</title><p>(<bold>A</bold>) A selection of NRSE containing genes (<italic>Atp2b, Crhr2, Ep300, Hcn2, Kcnh2, P2xr5, Pcsk1, Xpo6)</italic> whose expression was unchanged according to the microarray following KA-seizures were measured using qPCR to validate the microarray n = 4/group. (<bold>B</bold>) qPCR measurement of a selection of the NRSE containing genes (<italic>Calb1, Glra2, Grin2a, Hcn1, Kcnc2, Klf9, Lrp11, Myo5b, Stmn2)</italic> whose expression was down-regulated following KA-seizures and rescued by NRSE-ODNs n = 4/group, p*<0.05.</p><p><bold>DOI:</bold> <ext-link ext-link-type="doi" xlink:href="10.7554/eLife.01267.006">http://dx.doi.org/10.7554/eLife.01267.006</ext-link></p></caption><graphic xlink:href="elife01267f004"/></fig></p><p>To probe the basis of the differential regulation of these two gene groups, we compared the binding of NRSF to the repressed and non-repressed genes in both the naive hippocampus and following KA-seizures. <xref ref-type="fig" rid="fig5">Figure 5A</xref> depicts the relative binding of NRSF to 13 genes that are presented in the order of increasing NRSF binding in the naive hippocampus. Genes repressed by the seizures are shown in green. <xref ref-type="fig" rid="fig5">Figure 5B</xref> shows the relative binding of NRSF to the same genes in hippocampi taken from rats undergoing seizures. For each of these genes, we calculated the increment in NRSF binding after the seizures (when NRSF levels are increased) relative to the naive state (<xref ref-type="fig" rid="fig5">Figure 5C</xref>). As apparent in this figure, (a) genes repressed by seizure-induced increases of NRSF levels were those that had a large increment in NRSF binding. In addition, (b) this increment arose on the background of moderate binding levels in the naive state.<fig id="fig5" position="float"><object-id pub-id-type="doi">10.7554/eLife.01267.007</object-id><label>Figure 5.</label><caption><title>Physical binding of NRSF co-varies with tissue levels specifically at genes that are regulated by the repressor.</title><p>(<bold>A</bold>) NRSF binding (expressed as percent of input) to selected NRSE-containing genes in naive hippocampus with genes whose expression is repressed by seizure-induced NRSF increase represented in green, genes where NRSF occupancy was low are depicted in white, while genes where NRSF binding was abundant are depicted in black. (<bold>B</bold>) NRSF occupancy (percent input) at the same gene set in the hippocampus 48 hr following KA-induced seizures. (<bold>C</bold>) Graphical depiction of the changes (Delta) in NRSF occupancy at NRSE-containing gene sets comparing occupancy following KA-induced seizures to occupancy in the naive state. Genes whose expression was repressed are represented in green, n = 4–6/group, p*<0.05.</p><p><bold>DOI:</bold> <ext-link ext-link-type="doi" xlink:href="10.7554/eLife.01267.007">http://dx.doi.org/10.7554/eLife.01267.007</ext-link></p></caption><graphic xlink:href="elife01267f005"/></fig></p><p>Specifically, the effects of seizure-induced increase of NRSF levels on NRSF binding to the chromatin correlated with the degree of repressor binding in the naive state: binding of NRSF to certain genes (<italic>Ep300, Hcn2, P2xr5, Xpo6</italic>) was modest and did not increase significantly upon seizure-induced increase of NRSF levels (white columns of <xref ref-type="fig" rid="fig5">Figure 5A–C</xref>). NRSF binding was very robust at NRSE sites of other genes (<italic>Atp2b, Crhr2, Htr1a, Pcsk1</italic>), and the degree of binding was relatively independent of NRSF tissue levels (black columns of <xref ref-type="fig" rid="fig5">Figure 5A–C</xref>). In contrast, seizure-induced increase of NRSF levels markedly augmented NRSF binding to genes such as <italic>Calb1, Glra2, Grin2A, Hcn1, and Kcnc2</italic>, where binding was moderate in the naive hippocampus. The latter set of genes was those preferentially repressed by NRSF in mature hippocampus (<xref ref-type="fig" rid="fig4">Figure 4B</xref> and <xref ref-type="fig" rid="fig5">Figure 5C</xref>).</p></sec><sec id="s2-4"><title>A potential ‘dynamic range’ of repressor binding might enable gene regulation by moderate fluctuations of NRSF levels</title><p>As shown above, only a subset of NRSE-containing genes in mature hippocampus was repressed by seizure-dependent increase of hippocampal NRSF levels and ‘rescued’ when NRSF binding was prevented. Interestingly, this sub-population of putative NRSF target genes had moderate repressor binding in the naive state which increased markedly after seizures. This led us to propose a simple model to explain these findings. According to the model (<xref ref-type="fig" rid="fig6">Figure 6</xref>), in genes with very low binding to NRSF, a moderate increase of NRSF levels would not alter repressor binding appreciably. Similarly, in genes with high binding frequency to NRSF, NRSEs would likely be occupied even by the low NRSF levels in the naive hippocampus. Increased NRSF levels would be unlikely to augment occupancy or promote suppression of these genes. In contrast, when NRSF binding is ‘mid-range’, then relatively small fluctuation of NRSF levels would result in major changes of repressor binding and gene repression.<fig id="fig6" position="float"><object-id pub-id-type="doi">10.7554/eLife.01267.008</object-id><label>Figure 6.</label><caption><title>A potential ‘dynamic range’ of repressor binding might enable gene regulation by moderate fluctuations of NRSF levels.</title><p>A graphical representation of our proposed hypothesis based on our observations that only a subset of NRSE-containing genes are functionally repressed by seizure-induced increases in NRSF levels and that these genes appear to have moderate NRSF binding in the naive brain.</p><p><bold>DOI:</bold> <ext-link ext-link-type="doi" xlink:href="10.7554/eLife.01267.008">http://dx.doi.org/10.7554/eLife.01267.008</ext-link></p></caption><graphic xlink:href="elife01267f006"/></fig></p><p>We sought to refute or support this broad supposition by examining available information about binding frequencies of NRSF to a large gene set. A comprehensive analysis of NRSF binding probabilities has been reported, based on ChIP-sequencing data (ChIP-Seq) (<xref ref-type="bibr" rid="bib31">Johnson et al., 2007</xref>). The published binding frequencies were derived from a cell line, and it is likely that several cell- and tissue-specific parameters influence NRSF binding to the chromatin (<xref ref-type="bibr" rid="bib43">Lawinger et al., 2000</xref>; <xref ref-type="bibr" rid="bib105">Zuccato et al., 2007</xref>; <xref ref-type="bibr" rid="bib22">Gillies et al., 2011</xref>; <xref ref-type="bibr" rid="bib46">Lovén et al., 2013</xref>; <xref ref-type="bibr" rid="bib101">Whyte et al., 2013</xref>). Cognizant of these potential caveats, we simply employed the data set to estimate broadly the binding frequencies of NRSE-containing genes to NRSF. We partitioned these putative NRSF target genes into 8 sets (bins) based on their NRSF binding-frequency rank (<xref ref-type="fig" rid="fig7">Figure 7A</xref>). We then partitioned (binned) all hippocampal NRSE-containing genes that were detectable in the transcriptome arrays into 8 sets based on their predicted NRSF binding-frequency rank using the Johnson data set.<fig id="fig7" position="float"><object-id pub-id-type="doi">10.7554/eLife.01267.009</object-id><label>Figure 7.</label><caption><title>Genes regulated by seizure-dependent changes in NRSF function possess a distinct range of NRSF binding frequencies.</title><p>(<bold>A</bold>) Diagram illustrating sets of genes that were binned according to binding frequency based on <xref ref-type="bibr" rid="bib31">Johnson et al., 2007</xref>. Gene sets were numbered in increasing order of their NRSF binding frequencies. Mid-range NRSF-binding frequencies are in green. (<bold>B</bold> and <bold>C</bold>) A binding frequency metric was established for each NRSF-binding gene based on the number of ChIP-Seq reads from published data (<xref ref-type="bibr" rid="bib31">Johnson et al., 2007</xref>). The distribution of (<bold>B</bold>) all microarray-detectable NRSF-binding genes was compared to the distribution of (<bold>C</bold>) genes regulated by seizure-dependent NRSF changes, that is, those genes significantly repressed by network hyperactivity and rescued by NRSE–ODN treatment. Presented are scatter dot plots, with median with interquartile ranges. Below them are regression fit histogram plots. (<bold>D</bold>) Comparing microarray data from control rats to that of rats experiencing KA-seizures (both with scrambled ODN), using GSEA, illustrates that three gene-sets were significantly enriched in the control rats (repressed in the seizure + scrambled-ODN rats) and these fell in the mid-frequency category (see <xref ref-type="table" rid="tbl1">Table 1</xref> for numeric values).</p><p><bold>DOI:</bold> <ext-link ext-link-type="doi" xlink:href="10.7554/eLife.01267.009">http://dx.doi.org/10.7554/eLife.01267.009</ext-link></p></caption><graphic xlink:href="elife01267f007"/></fig></p><p>We compared the binding frequency distribution of all of the hippocampal NRSE-containing genes to that of the subset of genes that were functionally regulated by NRSF (<xref ref-type="fig" rid="fig7">Figure 7B</xref>). In the overall population of NRSE-containing hippocampal genes, binding frequencies ranged from 19 to 5544 (mean 279; median 153; standard deviation 429). In contrast, NRSF-binding scores of the genes that were functionally regulated by NRSF ranged from 27 to 663 (mean 214; median 172; standard deviation 166). Whereas the frequency medians of all NRSE-containing hippocampal genes and NRSF-regulated NRSE-containing genes were not significantly different (Mann–Whitney p = 0.86), the scatter plots and regression-fit histograms demonstrated that the gene population regulated by NRSF in the context of epileptogenesis was more tightly clustered and had distinct distribution properties. Skewness was 6.5 in all detected NRSE-containing genes vs 1.2 in the regulated genes; Kurtosis was 66 in all of the genes and 0.98 in the regulated genes. Notably, whereas NRSF-binding distribution of the total NRSE-containing hippocampal gene population was exceedingly different from normal (all: Kolmogorov–Smirnov test p < 0.0001, D'Agostino and Pearson omnibus normality test p < 0.0001), the distribution pattern of the regulated gene population was much closer to normal (Kolmogorov–Smirnov test p > 0.10, D'Agostino and Pearson omnibus normality test p = 0.012). Together, these clustering patterns suggest that the population of NRSF-regulated hippocampal genes is distinct and characterized by a relatively narrow, normally-distributed range of NRSF-binding frequencies.</p><p>To further test this idea, we analyzed the array data using gene set enrichment analysis (GSEA, MIT Broad Institute) (<xref ref-type="bibr" rid="bib54">Mootha et al., 2003</xref>; <xref ref-type="bibr" rid="bib92">Subramanian et al., 2005</xref>). GSEA is a computational method that determines whether an a priori defined set of genes shows statistically significant, concordant differences between two experimental groups. Comparison of the control group to the seizures group for each binned gene set as described above resulted in a graphic representation depicting the GSEA-normalized Enrichment Scores (NES), obtained from the Broad Institute's molecular signature database (MSigDB), for each set of partitioned NRSE-containing genes (<xref ref-type="fig" rid="fig7">Figure 7D</xref>). The GSEA graphs are shown in <xref ref-type="fig" rid="fig8">Figure 8</xref>, and the analysis parameters, including false discovery rate [FDR] and family-wise error rates [FWER] are found in <xref ref-type="table" rid="tbl1">Table 1</xref>. The expression of three of these ‘bins’ (gene sets partitioned by binding frequency) was significantly enriched in the control group (i.e., repressed in the seizure group) when compared to the levels of enrichment of gene sets composed of random permutations of genes (FWER *p < 0.05). Notably, the three significantly enriched gene-sets were not composed of genes with the highest NRSF binding frequency; rather, they consisted of genes between the 37.5<sup>th</sup> and 75<sup>th</sup> binding frequency rank percentile (<xref ref-type="fig" rid="fig7">Figure 7D</xref>; <xref ref-type="table" rid="tbl1">Table 1</xref>; <xref ref-type="fig" rid="fig8">Figure 8</xref>). In addition, when binding-frequency binned GSEA was used to compare the control group to the KA-seizure group receiving the NRSE–ODN treatment (i.e., with blocked NRSF function), the three binned gene sets were no longer significantly different from those in the control group (<xref ref-type="table" rid="tbl1">Table 1</xref>).<fig id="fig8" position="float"><object-id pub-id-type="doi">10.7554/eLife.01267.010</object-id><label>Figure 8.</label><caption><title>Gene set enrichment analysis (GSEA, Broad Institute, MIT) curves of hippocampal NRSE-containing genes.</title><p>These genes have been classified (‘binned’) into eight groups by increasing order of their NRSF binding frequency rank percentile. Hence, these genes might be categorized as low-binding frequency (<bold>A</bold>–<bold>C</bold>), mid-binding frequency (<bold>D</bold>–<bold>F</bold>), and high-binding frequency (<bold>G</bold>–<bold>H</bold>). Each graph shows enrichment plots comparing gene expression in KA-seizures vs controls in the presence of random ODNs. Please see <xref ref-type="table" rid="tbl1">Table 1</xref>, top, for the numeric values and parameters of the analyses.</p><p><bold>DOI:</bold> <ext-link ext-link-type="doi" xlink:href="10.7554/eLife.01267.010">http://dx.doi.org/10.7554/eLife.01267.010</ext-link></p></caption><graphic xlink:href="elife01267f008"/></fig><table-wrap id="tbl1" position="float"><object-id pub-id-type="doi">10.7554/eLife.01267.011</object-id><label>Table 1.</label><caption><p>Gene-set enrichment analyses focusing on NRSF</p><p><bold>DOI:</bold> <ext-link ext-link-type="doi" xlink:href="10.7554/eLife.01267.011">http://dx.doi.org/10.7554/eLife.01267.011</ext-link></p></caption><table frame="hsides" rules="groups"><thead><tr><th>Gene set name</th><th>NES</th><th>NOM p-val</th><th>FDR q-val</th><th>FWER p-val</th></tr></thead><tbody><tr><td>Ctrl vs Activity (both with scrambled ODN)</td><td/><td/><td/><td/></tr><tr><td> NRSF(0-12.5)(19-44)</td><td>1.1449</td><td>0.2891</td><td>0.3824</td><td>0.8710</td></tr><tr><td> NRSF(12.5-25)(45-66)</td><td>1.2296</td><td>0.5170</td><td>0.5150</td><td>0.9870</td></tr><tr><td> NRSF(25-37.5)(67-99)</td><td>1.1865</td><td>0.2140</td><td>0.2020</td><td>0.2940</td></tr><tr><td> <bold>NRSF(37.5-50)(100-151)</bold></td><td><bold>1.6817</bold></td><td><bold>0.0102</bold></td><td><bold>0.0212</bold></td><td><bold>0.0160</bold></td></tr><tr><td> <bold>NRSF(50-62.5)(153-221)</bold></td><td><bold>1.6487</bold></td><td><bold>0.0258</bold></td><td><bold>0.0175</bold></td><td><bold>0.0210</bold></td></tr><tr><td> <bold>NRSF(62.5-75)(223-341)</bold></td><td><bold>2.0243</bold></td><td><bold>0.0000</bold></td><td><bold>0.0000</bold></td><td><bold>0.0000</bold></td></tr><tr><td> NRSF(75-87.5)(348-533)</td><td>1.1396</td><td>0.2777</td><td>0.5563</td><td>0.8600</td></tr><tr><td> NRSF(87.5-100)(535-5544)</td><td>0.9720</td><td>0.1868</td><td>0.7621</td><td>0.7200</td></tr><tr><td>Ctrl vs Activity (both with NRSE-ODN)</td><td/><td/><td/><td/></tr><tr><td> NRSF(0-12.5)(19-44)</td><td>1.1485</td><td>0.2634</td><td>0.3881</td><td>0.6590</td></tr><tr><td> NRSF(12.5-25)(45-66)</td><td>1.1842</td><td>0.2000</td><td>0.2071</td><td>0.6050</td></tr><tr><td> NRSF(25-37.5)(67-99)</td><td>1.3069</td><td>0.1165</td><td>0.1323</td><td>0.3540</td></tr><tr><td> NRSF(37.5-50)(100-151)</td><td>1.4666</td><td>0.0458</td><td>0.0646</td><td>0.1450</td></tr><tr><td> NRSF(50-62.5)(153-221)</td><td>1.4762</td><td>0.0270</td><td>0.0880</td><td>0.1300</td></tr><tr><td> NRSF(62.5-75)(223-341)</td><td>1.5241</td><td>0.0294</td><td>0.1276</td><td>0.0970</td></tr><tr><td> NRSF(75-87.5)(348-533)</td><td>1.2320</td><td>0.4121</td><td>0.4184</td><td>0.8400</td></tr><tr><td> NRSF(87.5-100)(535-5544)</td><td>1.0232</td><td>0.1824</td><td>0.5207</td><td>0.5190</td></tr></tbody></table><table-wrap-foot><fn><p>NRSF is significantly enriched in gene sets with moderate binding probability even after false discovery corrections (FDR q-val), when the control and activity groups are compared (top; in bold). When the function of NRSF is blocked, that is, when comparison groups are the control and seizure-activity groups both treated with the NRSE-ODN, NRSF target gene-sets the bins (gene-sets) with moderate binding-frequencies are no longer enriched among differentially expressed genes.</p></fn></table-wrap-foot></table-wrap></p></sec><sec id="s2-5"><title>Genes regulated by NRSF are key molecules that contribute to neuronal properties</title><p>To better identify genes regulated by moderate fluctuations of NRSF in mature hippocampus, we followed the screen of microarray data for candidate genes with further analyses to control for false discovery (type I error). This approach uncovered genes that were significantly different in the (seizure + random-ODN) hippocampi vs the other groups (<xref ref-type="table" rid="tbl2">Table 2</xref>; p < 0.05, one-way ANOVA, Benjamini–Hochberg FDR p < 0.25). Genes that were both repressed by the seizures and ‘rescued’ by blocking NRSF–NRSE interactions encoded primarily molecules that are well-known to contribute to neuronal function and plasticity. These included ion channels and their accessory subunits, neurotransmitter receptors, genes involved in calcium-mediated cellular cascades, and phospho-enzymes (<xref ref-type="table" rid="tbl2">Table 2</xref>). Several of these genes encode for transcription factors (e.g., Kruppel-like factor 9 [<italic>Klf9</italic>]), or proteins involved in signaling cascades that might influence downstream gene expression (<italic>Grin2A</italic>, <italic>PKC</italic>, <italic>Nell1</italic>), suggesting that they might mediate secondary (indirect) effects of NRSF. Notably, this NRSF-regulated gene set is crucial in influencing the function of both individual neurons (<xref ref-type="bibr" rid="bib19">Garriga-Canut et al., 2006</xref>) and neuronal network behavior. Their importance is supported by the fact that preventing their repression attenuated seizure-provoked neuroplasticity that promoted epilepsy (<xref ref-type="bibr" rid="bib51">McClelland et al., 2011a</xref>).<table-wrap id="tbl2" position="float"><object-id pub-id-type="doi">10.7554/eLife.01267.012</object-id><label>Table 2.</label><caption><p>Candidate list of seizure-dependent NRSF regulated genes</p><p><bold>DOI:</bold> <ext-link ext-link-type="doi" xlink:href="10.7554/eLife.01267.012">http://dx.doi.org/10.7554/eLife.01267.012</ext-link></p></caption><table frame="hsides" rules="groups"><tbody><tr><td colspan="5">Ion channels, accessory subunits or receptors</td></tr><tr><td>HCN1</td><td>hyperpolarization-activated cyclic nucleotide-gated channel 1 (Hcn1)</td><td><xref ref-type="table-fn" rid="tblfn1">*</xref></td><td><xref ref-type="table-fn" rid="tblfn2">†</xref></td><td><xref ref-type="table-fn" rid="tblfn3">‡</xref></td></tr><tr><td>LRP11</td><td>low density lipoprotein receptor-related protein 11</td><td><xref ref-type="table-fn" rid="tblfn1">*</xref></td><td><xref ref-type="table-fn" rid="tblfn2">†</xref></td><td><xref ref-type="table-fn" rid="tblfn3">‡</xref></td></tr><tr><td>KCNC2</td><td>potassium voltage gated channel, Shaw-related subfamily, member 2</td><td><xref ref-type="table-fn" rid="tblfn1">*</xref></td><td><xref ref-type="table-fn" rid="tblfn2">†</xref></td><td><xref ref-type="table-fn" rid="tblfn3">‡</xref></td></tr><tr><td>KCNIP2</td><td>Kv channel-interacting protein 2 (Kcnip2), transcript variant a</td><td><xref ref-type="table-fn" rid="tblfn1">*</xref></td><td><xref ref-type="table-fn" rid="tblfn2">†</xref></td><td/></tr><tr><td>BAI2</td><td>brain-specific angiogenesis inhibitor 2 (predicted)</td><td><xref ref-type="table-fn" rid="tblfn1">*</xref></td><td><xref ref-type="table-fn" rid="tblfn2">†</xref></td><td/></tr><tr><td>SCN3B</td><td>sodium channel, voltage-gated, type III, beta (Scn3b)</td><td><xref ref-type="table-fn" rid="tblfn1">*</xref></td><td/><td/></tr><tr><td>SLC12A5</td><td>solute carrier family 12 member 5 (Slc12a5), (KCC2)</td><td><xref ref-type="table-fn" rid="tblfn1">*</xref></td><td/><td/></tr><tr><td>GRIN2A</td><td>Glutamate receptor, ionotropic, N-methyl D-aspartate 2A (Grin2a)</td><td><xref ref-type="table-fn" rid="tblfn1">*</xref></td><td/><td/></tr><tr><td>KCNC1</td><td>K voltage gated channel, Shaw-related subfamily, member 1, transcript 2 (Kcnc1)</td><td><xref ref-type="table-fn" rid="tblfn1">*</xref></td><td/><td/></tr><tr><td>GLRA2</td><td>glycine receptor, alpha 2 (Glra2)</td><td><xref ref-type="table-fn" rid="tblfn1">*</xref></td><td/><td/></tr><tr><td>NTRK3</td><td>neurotrophic tyrosine kinase, receptor, type 3 (Ntrk3)</td><td><xref ref-type="table-fn" rid="tblfn1">*</xref></td><td/><td/></tr><tr><td colspan="5">Calcium-mediated cellular cascades</td></tr><tr><td>CADPS</td><td>Ca++-dependent secretion activator (Cadps)</td><td><xref ref-type="table-fn" rid="tblfn1">*</xref></td><td><xref ref-type="table-fn" rid="tblfn2">†</xref></td><td><xref ref-type="table-fn" rid="tblfn3">‡</xref></td></tr><tr><td>CALB1</td><td>calbindin 1 (Calb1)</td><td><xref ref-type="table-fn" rid="tblfn1">*</xref></td><td><xref ref-type="table-fn" rid="tblfn2">†</xref></td><td/></tr><tr><td>HPCA</td><td>hippocalcin (Hpca)</td><td><xref ref-type="table-fn" rid="tblfn1">*</xref></td><td><xref ref-type="table-fn" rid="tblfn2">†</xref></td><td/></tr><tr><td>CABP7</td><td>calcium binding protein 7 (Cabp7)</td><td><xref ref-type="table-fn" rid="tblfn1">*</xref></td><td><xref ref-type="table-fn" rid="tblfn2">†</xref></td><td/></tr><tr><td>MYO5B</td><td>myosin Vb (Myo5b)</td><td><xref ref-type="table-fn" rid="tblfn1">*</xref></td><td/><td/></tr><tr><td>CAMKV</td><td>CaM kinase-like vesicle-associated (Camkv)</td><td><xref ref-type="table-fn" rid="tblfn1">*</xref></td><td/><td/></tr><tr><td colspan="5">Phosphoenzymes</td></tr><tr><td>GNAO1</td><td>guanine nucleotide binding protein, alpha O (Gnao1)</td><td><xref ref-type="table-fn" rid="tblfn1">*</xref></td><td><xref ref-type="table-fn" rid="tblfn2">†</xref></td><td><xref ref-type="table-fn" rid="tblfn3">‡</xref></td></tr><tr><td>PRKCG</td><td>protein kinase C, gamma (Prkcg)</td><td><xref ref-type="table-fn" rid="tblfn1">*</xref></td><td/><td/></tr><tr><td>PRKCB1</td><td>protein kinase C, beta 1 (Prkcb1)</td><td><xref ref-type="table-fn" rid="tblfn1">*</xref></td><td/><td/></tr><tr><td>NELL1</td><td>NEL-like 1 (chicken) (Nell1)</td><td><xref ref-type="table-fn" rid="tblfn1">*</xref></td><td/><td/></tr><tr><td colspan="5">Others</td></tr><tr><td>KLF9</td><td>Kruppel-like factor 9 (Klf9)</td><td><xref ref-type="table-fn" rid="tblfn1">*</xref></td><td><xref ref-type="table-fn" rid="tblfn2">†</xref></td><td><xref ref-type="table-fn" rid="tblfn3">‡</xref></td></tr><tr><td>STMN2</td><td>stathmin-like 2 (Stmn2)</td><td><xref ref-type="table-fn" rid="tblfn1">*</xref></td><td><xref ref-type="table-fn" rid="tblfn2">†</xref></td><td/></tr><tr><td>OLFM3</td><td>olfactomedin 3 (Olfm3)</td><td><xref ref-type="table-fn" rid="tblfn1">*</xref></td><td><xref ref-type="table-fn" rid="tblfn2">†</xref></td><td/></tr><tr><td>ICA1</td><td>islet cell autoantigen 1 (Ica1)</td><td><xref ref-type="table-fn" rid="tblfn1">*</xref></td><td><xref ref-type="table-fn" rid="tblfn2">†</xref></td><td/></tr><tr><td>STMN3</td><td>stathmin-like 3 (Stmn3)</td><td><xref ref-type="table-fn" rid="tblfn1">*</xref></td><td/><td/></tr><tr><td>OLFM1</td><td>olfactomedin 1 (Olfm1)</td><td><xref ref-type="table-fn" rid="tblfn1">*</xref></td><td/><td/></tr><tr><td>NXPH1</td><td>neurexophilin 1 (Nxph1)</td><td><xref ref-type="table-fn" rid="tblfn1">*</xref></td><td/><td/></tr></tbody></table><table-wrap-foot><fn><p>Genes were designated as ‘NRSE-containing’ based on NRSF ChIP-sequencing data from previous studies (<xref ref-type="bibr" rid="bib31">Johnson et al., 2007</xref>). For inclusion in the initial list of candidate genes for NRSF regulation, genes were considered significantly repressed if their expression was reduced by more than 20% after KA-induced seizures (p < 0.05). Genes considered ‘rescued’ were significantly repressed after seizures but not significantly repressed after ODN infusions [seizures + NRSE-ODN treatment]. This candidate gene approach was followed by measures to correct for FDR (see ‘Materials and methods’).</p></fn><fn id="tblfn1"><label>*</label><p>genes (28) with expression reduced by > 20% after KA-induced activity, and rescued by NRSF ODN; p < 0.05.</p></fn><fn id="tblfn2"><label>†</label><p>genes (14) found with ANOVA, Benjamini–Hochberg FDR <0.25.</p></fn><fn id="tblfn3"><label>‡</label><p>genes (6) found with one-way ANOVA, unadjusted p value <0.05.</p></fn></table-wrap-foot></table-wrap></p></sec></sec><sec id="s3" sec-type="discussion"><title>Discussion</title><p>The key findings of these studies are: (1) NRSF contributes to epileptogenesis by regulating a group of genes that critically influence neuronal function; (2) whereas hundreds of hippocampal genes are potential NRSF targets because they contain NRSEs, only a few dozen are actually repressed by the transcription factor in the context of seizure-induced epilepsy as evident from their rescue when the function of NRSF is blocked; (3) the basis of the selective regulation of a subset of NRSE-containing genes includes the mid-range binding frequencies of these genes to the repressor, so that moderate fluctuations in NRSF levels influence the degree of their binding to the repressor. Whereas these findings were observed in the context of epileptogenesis, a neuroplasticity that leads to disease, they may apply to a broad spectrum of physiological and pathological activity-dependent plasticities in the mature hippocampus.</p><p>NRSF is a transcription factor with protean effects. Whereas much is known about the role of this molecule in non-neuronal tissues and in neuronal differentiation, much less information is available about the function of NRSF in mature brain (<xref ref-type="bibr" rid="bib40">Korosi et al., 2010</xref>; <xref ref-type="bibr" rid="bib52">McClelland et al., 2011b</xref>). More recently, a protective role of NRSF has been proposed in aging and dementia (<xref ref-type="bibr" rid="bib47">Lu et al., 2014</xref>). In addition, NRSF may be involved in neurological disorders including Huntington disease (<xref ref-type="bibr" rid="bib105">Zuccato et al., 2007</xref>; <xref ref-type="bibr" rid="bib89">Soldati et al., 2011</xref>) and stroke (<xref ref-type="bibr" rid="bib13">Calderone et al., 2003</xref>; <xref ref-type="bibr" rid="bib33">Kaneko et al., 2014</xref>).</p><p>The contribution of NRSF to epilepsy and its development (epileptogenesis) is currently the focus of intense study (<xref ref-type="bibr" rid="bib78">Roopra et al., 2012</xref>; <xref ref-type="bibr" rid="bib23">Goldberg and Coulter 2013</xref>; <xref ref-type="bibr" rid="bib72">Pozzi et al., 2013</xref>). We previously found that interfering with NRSF binding to the chromatin-attenuated epileptogenesis. Interestingly, Hu et al. found that the conditional deletion of NRSF in the forebrain resulted in increased susceptibility to kindling and enhanced mossy fiber sprouting (<xref ref-type="bibr" rid="bib28">Hu et al., 2011</xref>). The apparent discrepancy between these studies is likely a result of the significantly different approaches. Complete deletion of NRSF in ‘naive’ neurons may induce large cascades of transcriptional changes rendering neurons more susceptible to seizures. In contrast, acute disruption of NRSF function ‘after’ an epilepsy-provoking insult, as performed in our current and previous studies, provides a more direct approach to elucidate the epileptogenic functions of NRSF because it does not interfere with NRSF function prior to the epilepsy-inducing insult. This point is important also because the role of NRSF in mature neurons is not fully understood, as discussed above.</p><p>In previous studies, we focused on the rescue of a single gene coding for the ion channel HCN1 (<xref ref-type="bibr" rid="bib51">McClelland et al., 2011a</xref>). However, whereas it was clear that NRSF likely regulated numerous additional genes that are involved in the epileptogenic process, the identity of these genes was unclear. In this study, we used large-scale transcriptome arrays and found that NRSE-containing genes were enriched among seizure-suppressed genes and that the majority of these genes were restored to control levels after treatment with decoy NRSE–ODNs. The fact that preventing the repression of (‘rescuing’) these genes led to attenuation of epilepsy strongly supports a role for these genes in the transformation of normal neurons (and neuronal networks) into epileptic ones.</p><p>Investigating the identity of these genes, we were pleased to find that the majority were already known to contribute to crucial neuronal properties, and many had been previously implicated in pathological states including epilepsy (<xref ref-type="bibr" rid="bib88">Sloviter, 1989</xref>; <xref ref-type="bibr" rid="bib64">Palm et al., 1999</xref>; <xref ref-type="bibr" rid="bib66">Pathak et al., 2007</xref>; <xref ref-type="bibr" rid="bib105">Zuccato et al., 2007</xref>; <xref ref-type="bibr" rid="bib96">van Gassen et al., 2009</xref>; <xref ref-type="bibr" rid="bib34">Khirug et al., 2010</xref>; <xref ref-type="bibr" rid="bib89">Soldati et al., 2011</xref>; <xref ref-type="bibr" rid="bib51">McClelland et al., 2011a</xref>; <xref ref-type="bibr" rid="bib58">Nikitidou et al., 2012</xref>; <xref ref-type="bibr" rid="bib36">Kingwell, 2013</xref>). This was not surprising, because these genes included ion channels, neurotransmitter receptors, calcium-dependent effectors, and phosphoenzymes (<xref ref-type="bibr" rid="bib68">Pernhorst et al., 2011</xref>; <xref ref-type="bibr" rid="bib16">Dingledine, 2012</xref>; <xref ref-type="bibr" rid="bib76">Rodenas-Ruano et al., 2012</xref>; <xref ref-type="bibr" rid="bib49">Maslarova et al., 2013</xref>).</p><p>Whereas NRSF-mediated repression might theoretically involve hundreds of hippocampal NRSE-containing genes, it was intriguing to discover that in fact only a small subset of detected NRSE-containing genes were repressed by seizures. Most were rescued by blocking NRSF function—as evident using a differential array that included control and insult groups as well as groups where NRSF function was blocked—thus ascertaining that genes that are repressed during epileptogenesis are indeed repressed by the augmented levels of endogenous NRSF. To our knowledge this is the first study demonstrating the specificity of NRSF-mediated gene regulation in mature hippocampus in vivo.</p><p>What might be the basis for the apparent selectivity of NRSF repression of only a minority of putative target genes? The binding of NRSF to a gene is required (though not always sufficient; [<xref ref-type="bibr" rid="bib8">Belyaev et al., 2004</xref>]) for its repression. A number of mechanisms may influence both the degree of NRSF binding as well as the degree of its repressive effect on genes. These include the specific NRSE sequence (<xref ref-type="bibr" rid="bib55">Mortazavi et al., 2006</xref>; <xref ref-type="bibr" rid="bib31">Johnson et al., 2007</xref>) that is unlikely to be the cause in the current studies (<xref ref-type="supplementary-material" rid="SD1-data">Supplementary file 1A</xref>). Other important mechanisms include the interaction of NRSF with co-factors (<xref ref-type="bibr" rid="bib3">Andres et al., 1999</xref>; <xref ref-type="bibr" rid="bib56">Naruse et al., 1999</xref>; <xref ref-type="bibr" rid="bib102">Wood et al., 2003</xref>; <xref ref-type="bibr" rid="bib18">Gao et al., 2011</xref>), as well as additional, still enigmatic effects of the chromatin environment of individual genes (<xref ref-type="bibr" rid="bib8">Belyaev et al., 2004</xref>), which might be influenced by ‘super-enhancers’ (<xref ref-type="bibr" rid="bib46">Lovén et al., 2013</xref>; <xref ref-type="bibr" rid="bib101">Whyte et al., 2013</xref>).</p><p>Here, we examined the binding of NRSF to hippocampal genes before and after an epilepsy-provoking insult and found significant increment in genes that were repressed by the factor. This suggested a correlation between repressor occupancy and gene repression (<xref ref-type="bibr" rid="bib25">Hammar et al., 2014</xref>). To our surprise, basal binding of NRSF in regulated hippocampal genes was moderate. By contrast, most non-regulated genes had either high or low NRSF occupancy in naive hippocampus, with little change after an insult that augmented NRSF levels twofold to threefold. We examined our finding in mature hippocampus by comparing the binding-frequency distribution found in hippocampus to a larger set of NRSE target genes as determined by ChIP-Seq analysis (<xref ref-type="bibr" rid="bib31">Johnson et al., 2007</xref>). Whereas many factors, as discussed above, may lead to different NRSF binding in cell lines and hippocampus, it was encouraging to find that our estimates for the relative mid-range binding of regulated genes among those expressed in hippocampus held when superimposed on a large NRSE-containing gene set (<xref ref-type="bibr" rid="bib31">Johnson et al., 2007</xref>). Traditionally, the focus of studies on gene–repressor interaction has been on genes with the strongest binding probability (<xref ref-type="bibr" rid="bib57">Nichols et al., 1992</xref>; <xref ref-type="bibr" rid="bib14">Chen et al., 1998</xref>; <xref ref-type="bibr" rid="bib86">Singh et al., 2002</xref>). However, the genes with the strongest binding for NRSF would likely be constitutively bound to—and suppressed by—NRSF. These genes are unlikely to respond to moderate changes in NRSF levels. Rather, relatively modest seizure-induced changes in the tissue levels of NRSF regulate a subset of NRSE-containing genes that fall within a moderate ‘dynamic range’ of NRSF binding probabilities. This concept may be important for other forms of insult-induced plasticity in the mature brain including ischemia (<xref ref-type="bibr" rid="bib60">Noh et al., 2012</xref>; <xref ref-type="bibr" rid="bib33">Kaneko et al., 2014</xref>), peripheral nerve injury (<xref ref-type="bibr" rid="bib95">Uchida et al., 2010</xref>), and seizure-induced epilepsy (<xref ref-type="bibr" rid="bib51">McClelland et al., 2011a</xref>; <xref ref-type="bibr" rid="bib23">Goldberg and Coulter, 2013</xref>) where changes in NRSF levels are moderate.</p><p>These findings support a role of NRSF in mature brain that is distinct from its role in development, when NRSF contributes to neuronal identity by repressing neuronal genes in non-neuronal tissues (<xref ref-type="bibr" rid="bib83">Schoenherr and Anderson, 1995</xref>; <xref ref-type="bibr" rid="bib15">Coulson, 2005</xref>), and specifies neuronal fate (<xref ref-type="bibr" rid="bib42">Kuwabara et al., 2005</xref>). In that context, large changes in the expression of NRSF take place: NRSF levels plunge during neuronal differentiation (<xref ref-type="bibr" rid="bib20">Gates et al., 2010</xref>; <xref ref-type="bibr" rid="bib17">Ernsberger, 2012</xref>), de-repressing genes that contribute to neuronal identity (<xref ref-type="bibr" rid="bib102">Wood et al., 2003</xref>; <xref ref-type="bibr" rid="bib8">Belyaev et al., 2004</xref>; <xref ref-type="bibr" rid="bib1">Abrajano et al., 2009</xref>). Remarkably, the gene set regulated by moderate changes in NRSF expression in the current study consisted of molecules which are extremely important for neuronal function and plasticity not only during development (<xref ref-type="bibr" rid="bib20">Gates et al., 2010</xref>; <xref ref-type="bibr" rid="bib32">Juliandi et al., 2010</xref>; <xref ref-type="bibr" rid="bib76">Rodenas-Ruano et al., 2012</xref>; <xref ref-type="bibr" rid="bib69">Pernhorst et al., 2013</xref>), but also in the adult brain (<xref ref-type="bibr" rid="bib44">Lepagnol-Bestel et al., 2009</xref>; <xref ref-type="bibr" rid="bib18">Gao et al., 2011</xref>; <xref ref-type="bibr" rid="bib68">Pernhorst et al., 2011</xref>).</p><p>In summary, insult-induced increases of NRSF levels and activity contribute to epileptogenesis via NRSF-mediated repression of a group of genes that critically influence neuronal function. Of the hundreds of NRSE-containing hippocampal genes only a subset are repressed by the transcription factor in the context of seizure-induced epilepsy as evident from their rescue when the function of NRSF is blocked. The basis of the selective regulation of a subset of NRSE-containing genes includes their mid-range binding frequencies to NRSF, so that moderate fluctuations in NRSF levels influence the degree of their binding. These findings inform us about the mechanisms of neuroplasticity in the mature hippocampus.</p></sec><sec id="s4" sec-type="materials|methods"><title>Materials and methods</title><p>All experiments were performed according to NIH guidelines and approved by the UCI Institutional Animal Care and Use Committee.</p><sec id="s4-1"><title>Organotypic slice cultures</title><p>Organotypic hippocampal slice cultures were prepared and maintained using the interface technique (<xref ref-type="bibr" rid="bib91">Stoppini et al., 1991</xref>). Briefly, hippocampi from P8 rat pups of both genders were resected and cut into 400-µm slices using a McIlwain tissue chopper (The Mickle Laboratory Engineering Co. Ltd, Surrey, UK). The slices were collected in ice-cold preparation buffer (100% Minimal Essential Medium, containing 30 mM glucose, and 3 mM glutamine, pH 7.3; Life Technologies, Rockville, MD), then placed onto moistened membrane inserts (Millicell-CM, 30 mm, 0.4 µm pore diameter; Millipore, Bedford, MA), transferred to sterile six-well plates filled with 1 ml culture medium (50% Minimal Essential Medium, 25% Hank's balanced salt solution, 20% heat-inactivated horse serum, 30 mM HEPES, 30 mM glucose, 3 mM glutamine, 0.5 mM ascorbic acid, 1 mg/ml insulin, 5 mM NaHCO<sub>3</sub>, pH 7.3) and incubated in a humidified CO<sub>2</sub>-enriched atmosphere at 36°C. Pairs of adjacent slices (sister cultures) were always compared (i.e., adjacent slices assigned to control and experimental conditions, n = 4–6/group for each time-point). Seizure-like activity was induced after 3 days in vitro by incubating cultures for 3 hr in a medium containing KA (6 µM; Sigma, St. Louis, MO). This seizure-like activity was halted by changing the medium after 3 hr (<xref ref-type="bibr" rid="bib74">Richichi et al., 2008</xref>).</p></sec><sec id="s4-2"><title>Surgery, induction of KA-seizures, and oligonucleotide (ODN) infusion in vivo</title><p>Male Sprague–Dawley rats (n = 36) were anaesthetized with inhalation of 4% isoflurane. The rats were shaved and placed in a stereotaxic frame, and their eyes were protected and hydrated with Ocry-gel. A bolus of 2% Lidocaine was injected subcutaneously prior to a midsagittal incision. The skull was exposed and cleaned of blood and periost. Bilateral infusion cannulae were positioned on the cortical surface (−1.0 mm posterior, ±1.5 mm lateral from Bregma) directly above the lateral ventricles, using the coordinates of Paxinos and Watson (<xref ref-type="bibr" rid="bib67">Paxinos et al., 1985</xref>). The incision was then sutured closed as necessary. All rats received ∼5 ml of 0.9% saline I.P. to rehydrate and aid in recovery from surgery. Rats were allowed 7 days post-surgical recovery before any further experimental procedures were conducted. Status epilepticus (SE) was induced as previously described (<xref ref-type="bibr" rid="bib51">McClelland et al., 2011a</xref>). Controls received saline. Seizures were observed and scored using the Racine scale (<xref ref-type="bibr" rid="bib73">Racine, 1972</xref>). One day later, rats were anaesthetized with inhalation of 4% isoflurane. An infusion needle was lowered through the guide cannulae until it reached the cortical surface. The needle was lowered an additional 3.0 mm so the tip was within the lateral ventricle. On the first day, rats received an infusion of 2.0 nmol/µl of ordered NRSE or random ODNs at a volume of 5 µl and a rate of 0.1 µl/min (NRSE: GGA GCT GTC CAC AGT TCT GAA; Random: AGG TCG TAC GTT AAT CGT CGC). The needle remained in place for an additional 10 min to allow for thorough diffusion of the solutions. On the following day, the procedure was identical except that each rat received a volume of 2.5 µl. This infusion protocol was chosen to ensure that an adequate level of ODNs remained in the brain over the course of the experiment (<xref ref-type="bibr" rid="bib11">Bristol and Rothstein, 1998</xref>).</p></sec><sec id="s4-3"><title>In situ hybridization (ISH)</title><p>Organotypic hippocampal slice cultures were used at different time points after treatment with KA, and rats receiving KA–SE were decapitated at different time points. Brains were dissected and placed on powdered dry ice. Semi-quantitative analysis of NRSF mRNA levels were accomplished using in situ hybridization (ISH) (<xref ref-type="bibr" rid="bib10">Brewster et al., 2002</xref>; <xref ref-type="bibr" rid="bib51">McClelland et al., 2011a</xref>) using S<sup>35</sup>-cRNA probes and slide-mounted frozen sections (20 µm) with the most stringent wash at 0.03 SSC, at 62°C for 60 min. Sections were then dehydrated and apposed to Kodak BioMax film. Exposure time was monitored using 14C standards to maintain the signal in the linear range. Analyses compared control and KA-treated sister cultures derived from the same hippocampus at every time point (n = 4–8 per group per time point) and were performed without the knowledge of treatment group.</p></sec><sec id="s4-4"><title>Western blot protein analysis</title><p>Animals were decapitated and the hippocampi were rapidly dissected. Tissues were placed in pre-chilled microcentrifuge tubes and processed immediately. The tissue was homogenized in glass/Teflon homogenizers in a cold solution containing 0.32 M sucrose, 0.01 M Tris–HCl (pH 7.4), and Protease Inhibitor Cocktail (PIC Complete; di; Roche, Alameda, CA). Samples were then centrifuged at 800×<italic>g</italic> for 10 min at 4°C, and the pellet was retained for processing to obtain a nuclear-enriched fraction. The supernatant was centrifuged at 16,000×<italic>g</italic> for 40 min at 4°C, and the pellet containing membrane fractions resuspended in RIPA buffer (50 mM Tris–HCl, pH 7.4, 1% NP-40, 1% Triton X-100, 1 mM EDTA, 150 mM NaCl, 1X PIC).</p><p>Pellets retained for the nuclear-enriched fraction described above were resuspended in 1 ml hypotonic buffer (10 mM KCl, 10 mM Tris pH 8, 1.5 mM MgCl<sub>2</sub>). Samples were then incubated on ice for 15 min, followed by the addition of 100 μl of 10% NP-40, samples were then spun for 1 min at 16,000×g. The supernatant was then discarded, and 360 μl of RIPA buffer was added to the remaining nuclear pellet. Samples were then sonicated for 10 min on the highest setting (30 s on/30 s off) using a Bioruptor sonicator (Diagenode, Sparta, NJ, USA), to disrupt nuclei. Protein concentration was determined using Bio-Rad Protein assay (Bio-Rad, Hercules, CA).</p><p>Protein samples (30 μg) were suspended in Laemmli buffer, separated by 4–12% SDS-PAGE (Lonza, Rockland, ME, USA), transferred to PVDF membrane and blocked in 5% milk for 2 hr at room temperature (RT). Membranes were probed with anti-Actin (1:10,000; Sigma), anti-NRSF (1:2000, sc-25398X; Santa Cruz Biotechnology, Santa Cruz, CA, USA), in PBS with 5% non-fat milk. Membranes were then incubated with secondary IgG-horseradish peroxidase conjugates (1:10,000; AmerCtrl Pharmacia Biotech, Piscataway, NJ, USA) for 1 hr at room temperature. After final washes in PBS-T (6 × 5 min), membranes were incubated with ECL-Plus (Pierce Scientific, Rockford, IL, USA) for 5 min, and immunoreactive bands were visualized by apposing membrane to hyperfilmTM ECL.</p><p>A series of ECL exposures were carried out to ensure that non-saturated bands were used for quantification. Western blot data acquisition and analysis were accomplished by measuring the pixel density and area of immunoreactive bands from the ECL films using the ImageJ software (NIH). Values were normalized to actin. Significance level for unpaired t-tests was set at 0.05, and data are presented as mean with standard errors.</p></sec><sec id="s4-5"><title>Transcriptome gene array</title><p>Hippocampal CA1 regions were dissected using instruments bathed in RNALater (Ambion, Life Technologies, Rockford, IL, USA). The CA1 region was obtained 48 hr after SE from rats of the following groups: ctrl + random ODN (n = 4), ctrl + NRSE–ODN (n = 5), KA-seizures + NRSE–ODN (n = 4), and KA-seizures + random ODN (n = 3). Samples were processed for RNA extraction by SABiosciences using the Array Grade total RNA isolation kit. RNA from an individual rat was applied to separate Illumina RatRef-12. Expression BeadChips for transcriptome analysis of over 22,000 genes. GenomeStudio software was used by SABiosciences (an Illumina CSPro certified service provider) to perform quality control, quantile normalization and background correction, and to execute two-tailed Student's t-tests on the microarray expression results for all genes.</p><p>The microarray data complied with ‘minimum information about a microarray experiment’ (MIAME), and the raw data have been deposited in the Gene Expression Omnibus (GEO) (<ext-link ext-link-type="uri" xlink:href="http://www.ncbi.nlm.nih.gov/geo">http://www.ncbi.nlm.nih.gov/geo</ext-link>) as GSE 22899.</p></sec><sec id="s4-6"><title>qRT-PCR</title><p>The CA1 region was micro-dissected from dorsal rat hippocampus or whole hippocampus was dissected using pre-chilled RNase free instruments under a light microscope. Dissected tissue was placed immediately into pre-chilled sterile centrifuge tubes on powdered dry ice and stored at −80°C until use. Total RNA was extracted from CA1 tissue using the RNeasy kit (Qiagen, Sussex, UK) per protocol and quantified using a nanodrop (Thermo Scientific). RNA purity was determined. Double-stranded cDNA was synthesized from total RNA using Roche 1<sup>st</sup> strand cDNA synthesis kit (Cat # 04379012001; Roche) utilizing random hexamer primers. PCR analysis was performed using cDNA samples in triplicate on a Roche Lightcycler 96 system (Roche). Samples were normalized to Gapdh and relative quantitative amounts were analyzed using the cycle threshold method (2^-ΔΔCt). Minus-reverse transcription and non-template controls were routinely used to eliminate the possibility of genomic contamination or false positive analyses. Primers are provided in <xref ref-type="supplementary-material" rid="SD1-data">Supplementary file 1B</xref>.</p></sec><sec id="s4-7"><title>Chromatin immunoprecipitation</title><p>Whole hippocampal tissue was homogenized in 1% formaldehyde (Sigma) and incubated at room temperature (RT) for 20 min. Samples were centrifuged and the supernatant was discarded. Samples were then subjected to a number of incubations in glycine (Sigma) and hypotonic buffers, all containing a protease inhibitor cocktail (Roche). Samples were then transferred to a Dounce homogenizer and further homogenized before the addition of 10% NP-40 (MP Biochemicals, Solon, OH, USA), then centrifuged and the supernatant was once more discarded. The pellet was dissolved in RIPA buffer with PIC, vortexed, and sonicated (Diagenode) to shear DNA. Samples were then centrifuged for 45 min at 15,000 rpm, and the supernatant was collected and added to tubes containing 1 ml of magnetic DynaBeads (Life Technologies) pre-incubated with 5 µg NRSF antibody/sample or IgG control for 24 hr. Beads and DNA were left to incubate for 1 hr at 4°C. The beads were then accumulated using a magnet and the liquid was aspirated. The beads were washed 5 times with RIPA to remove loosely or non-specifically bound DNA. DNA and beads were separated by adding a Chelex solution (Bio-Rad) and heating samples to 100°C for 10 min. Samples were then centrifuged at 15,000 rpm, and the DNA containing supernatant was transferred to a new PCR tube and stored at 4°C, then quantified using qPCR. NRSF occupancy at NRSE sites was normalized to IgG binding to the DNA and calculated as a percentage of total input. Primers are provided in <xref ref-type="supplementary-material" rid="SD1-data">Supplementary file 1C</xref>.</p></sec><sec id="s4-8"><title>Transcriptome array analyses</title><p>Gene set enrichment analysis (GSEA version 2.5; Broad Institute, Cambridge, MA) (<xref ref-type="bibr" rid="bib54">Mootha et al., 2003</xref>; <xref ref-type="bibr" rid="bib92">Subramanian et al., 2005</xref>) was employed to compare the KA-seizures + NRSE–ODN and KA-seizures + random ODN groups to the ctrl + random ODN and ctrl + NRSE–ODN groups using the entire array data set (see <xref ref-type="table" rid="tbl1">Table 1</xref> and <xref ref-type="fig" rid="fig8">Figure 8</xref>). The differential expression for each gene was calculated using GSEA software's Signal2Noise metric, which is based on the difference in the mean of the two groups divided by the sum of the two standard deviations. All genes were then ranked and an enrichment score (ES) for each gene set was generated for different gene set collections in the Broad Institute's Molecular Signatures Database (MSigDB—a collection of annotated gene sets for use with GSEA software). The ES was adjusted based on the size of each gene set to obtain the normalized enrichment score (NES) which was used to rank the gene sets according to their over-representation in one group when compared to another. For each gene set the false discovery rate was calculated to determine the probability that its NES represented a false positive finding. The more conservative family-wise error rate (FWER) statistic was also generated. Subsets of NRSF-binding genes from ChIP-Sequencing data previously obtained were populated according to binding affinity rank (<xref ref-type="bibr" rid="bib31">Johnson et al., 2007</xref>). GSEA was run to determine if any specific subset was significantly contributing to the enrichment and the NES of each subset was plotted.</p><p>Statistical analyses of the microarray were stepwise (see <xref ref-type="table" rid="tbl2">Table 2</xref>). An initial discovery phase employed direct p values, and this was followed by False discovery rate (FDR) adjustments. This step-wise approach enabled initial candidate gene discovery by decreasing Type II error (and clearly increasing the initial possibility of Type I error). This was important in the context of the moderate seizure-dependent changes of NRSF expression in mature hippocampus where amplified Type II error may result in early rejection of biologically important candidate genes.</p><p>Benjamini–Hochberg FDR was employed in subsequent ANOVAs. Genes were considered significantly repressed if they had a greater than 20% reduction in expression at a p-value less than 0.05. Genes were designated as ‘NRSE-containing’ based on NRSF ChIP-sequencing data from previous studies (<xref ref-type="bibr" rid="bib31">Johnson et al., 2007</xref>). One-way ANOVA was performed using MeV 4.7 (MultiExperiment Viewer from the TM4 Microarray Software Suite) (<xref ref-type="bibr" rid="bib82">Saeed et al., 2003</xref>, <xref ref-type="bibr" rid="bib81">2006</xref>), and post hoc Benjamini–Hochberg FDRadjustments were made to the results using Excel (Microsoft). To avoid Type I error in the context of moderate changes of gene repression in mature hippocampus (see above), we set the FDR at 0.25. To avoid spurious discoveries (Type II error), we validated the repression of a sample of these genes using an independent method (qPCR). Heat maps were generated using MeV 4.7.</p></sec><sec id="s4-9"><title>Statistical analysis of qRT-PCR and ChIP</title><p>qPCR data were analyzed using a two-way ANOVA to answer the following questions; (a) did ODN treatment have the same effect at all values of status, that is was there any interaction? (b) did the NRSE ODN affect the result? (c) did the long seizures affect the result? ChIP data were analyzed by normalizing NRSF immunoprecipitation to IgG and calculating as a percentage of total input. Control and KA-treated groups were then compared using standard t-test.</p></sec></sec></body><back><ack id="ack"><title>Acknowledgements</title><p>The authors thank Annie Koh for technical support. This work was supported by NIH Grants R37 NS35439; T32 NS45540, RO1 NS78279 and by awards from INSERM and CURE. GPB is a recipient of a postdoctoral scholarship from the George E Hewitt Foundation for Medical Research. The authors thank Barbara Cartwright for excellent editorial assistance.</p></ack><sec sec-type="additional-information"><title>Additional information</title><fn-group content-type="competing-interest"><title>Competing interests</title><fn fn-type="conflict" id="conf1"><p>The authors declare that no competing interests exist.</p></fn></fn-group><fn-group content-type="author-contribution"><title>Author contributions</title><fn fn-type="con" id="con1"><p>SMC, Conception and design, Acquisition of data, Analysis and interpretation of data, Drafting or revising the article</p></fn><fn fn-type="con" id="con2"><p>GPB, Conception and design, Acquisition of data, Analysis and interpretation of data, Drafting or revising the article</p></fn><fn fn-type="con" id="con3"><p>CD, Conception and design, Acquisition of data</p></fn><fn fn-type="con" id="con4"><p>SR, Acquisition of data</p></fn><fn fn-type="con" id="con5"><p>SI, Acquisition of data</p></fn><fn fn-type="con" id="con6"><p>CR, Acquisition of data</p></fn><fn fn-type="con" id="con7"><p>CB, Drafting or revising the article</p></fn><fn fn-type="con" id="con8"><p>TZB, Conception and design, Analysis and interpretation of data, Drafting or revising the article</p></fn></fn-group><fn-group content-type="ethics-information"><title>Ethics</title><fn fn-type="other"><p>Animal experimentation: All experiments were performed according to NIH guidelines and approved by the UCI Institutional Animal Care and Use Committee. All animals were handled according to approved institutional animal car and use committee (IACUC) protocols (#1998-1665) All surgeries were performed under anesthesia and every effort was made to minimize suffering.</p></fn></fn-group></sec><sec sec-type="supplementary-material"><title>Additional files</title><supplementary-material id="SD1-data"><object-id pub-id-type="doi">10.7554/eLife.01267.013</object-id><label>Supplementary file 1.</label><caption><p>(<bold>A</bold>) NRSE sequences for NRSF regulated and non-regulated genes. (<bold>B</bold>) Primers used in qPCR. (<bold>C</bold>) Primers used in ChIP.</p><p><bold>DOI:</bold> <ext-link ext-link-type="doi" xlink:href="10.7554/eLife.01267.013">http://dx.doi.org/10.7554/eLife.01267.013</ext-link></p></caption><media mime-subtype="docx" mimetype="application" xlink:href="elife01267s001.docx"/></supplementary-material></sec><ref-list><title>References</title><ref id="bib1"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Abrajano</surname><given-names>JJ</given-names></name><name><surname>Qureshi</surname><given-names>IA</given-names></name><name><surname>Gokhan</surname><given-names>S</given-names></name><name><surname>Zheng</surname><given-names>D</given-names></name><name><surname>Bergman</surname><given-names>A</given-names></name><name><surname>Mehler</surname><given-names>MF</given-names></name></person-group><year>2009</year><article-title>Differential deployment of REST and CoREST promotes glial subtype specification and oligodendrocyte lineage maturation</article-title><source>PLOS ONE</source><volume>4</volume><fpage>e7665</fpage><pub-id pub-id-type="doi">10.1371/journal.pone.0007665</pub-id></element-citation></ref><ref id="bib2"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Akhtar</surname><given-names>S</given-names></name><name><surname>Kole</surname><given-names>R</given-names></name><name><surname>Juliano</surname><given-names>RL</given-names></name></person-group><year>1991</year><article-title>Stability of antisense DNA oligodeoxynucleotide analogs in cellular extracts and sera</article-title><source>Life Sciences</source><volume>49</volume><fpage>1793</fpage><lpage>1801</lpage><pub-id pub-id-type="doi">10.1016/0024-3205(91)90480-Y</pub-id></element-citation></ref><ref id="bib3"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Andres</surname><given-names>ME</given-names></name><name><surname>Burger</surname><given-names>C</given-names></name><name><surname>Peral-Rubio</surname><given-names>MJ</given-names></name><name><surname>Battaglioli</surname><given-names>E</given-names></name><name><surname>Anderson</surname><given-names>ME</given-names></name><name><surname>Grimes</surname><given-names>J</given-names></name><name><surname>Dallman</surname><given-names>J</given-names></name><name><surname>Ballas</surname><given-names>N</given-names></name><name><surname>Mandel</surname><given-names>G</given-names></name></person-group><year>1999</year><article-title>CoREST: a functional corepressor required for regulation of neural specific gene expression</article-title><source>Proceedings of the National Academy of Sciences of USA</source><volume>96</volume><fpage>9873</fpage><lpage>9878</lpage><pub-id pub-id-type="doi">10.1073/pnas.96.17.9873</pub-id></element-citation></ref><ref id="bib4"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Aoki</surname><given-names>H</given-names></name><name><surname>Hara</surname><given-names>A</given-names></name><name><surname>Era</surname><given-names>T</given-names></name><name><surname>Kunisada</surname><given-names>T</given-names></name><name><surname>Yamada</surname><given-names>Y</given-names></name></person-group><year>2012</year><article-title>Genetic ablation of Rest leads to in vitro-specific derepression of neuronal genes during neurogenesis</article-title><source>Development</source><volume>139</volume><fpage>667</fpage><lpage>677</lpage><pub-id pub-id-type="doi">10.1242/dev.072272</pub-id></element-citation></ref><ref id="bib5"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Bale</surname><given-names>TL</given-names></name><name><surname>Baram</surname><given-names>TZ</given-names></name><name><surname>Brown</surname><given-names>AS</given-names></name><name><surname>Goldstein</surname><given-names>JM</given-names></name><name><surname>Insel</surname><given-names>TR</given-names></name><name><surname>McCarthy</surname><given-names>MM</given-names></name><name><surname>Nemeroff</surname><given-names>CB</given-names></name><name><surname>Reyes</surname><given-names>TM</given-names></name><name><surname>Simerly</surname><given-names>RB</given-names></name><name><surname>Susser</surname><given-names>ES</given-names></name><name><surname>Nestler</surname><given-names>EJ</given-names></name></person-group><year>2010</year><article-title>Early life programming and neurodevelopmental disorders</article-title><source>Biological Psychiatry</source><volume>68</volume><fpage>314</fpage><lpage>319</lpage><pub-id pub-id-type="doi">10.1016/j.biopsych.2010.05.028</pub-id></element-citation></ref><ref id="bib6"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Ballas</surname><given-names>N</given-names></name><name><surname>Grunseich</surname><given-names>C</given-names></name><name><surname>Lu</surname><given-names>DD</given-names></name><name><surname>Speh</surname><given-names>JC</given-names></name><name><surname>Mandel</surname><given-names>G</given-names></name></person-group><year>2005</year><article-title>REST and its corepressors mediate plasticity of neuronal gene chromatin throughout neurogenesis</article-title><source>Cell</source><volume>121</volume><fpage>645</fpage><lpage>657</lpage><pub-id pub-id-type="doi">10.1016/j.cell.2005.03.013</pub-id></element-citation></ref><ref id="bib7"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Ballas</surname><given-names>N</given-names></name><name><surname>Mandel</surname><given-names>G</given-names></name></person-group><year>2005</year><article-title>The many faces of REST oversee epigenetic programming of neuronal genes</article-title><source>Current Opinion in Neurobiology</source><volume>15</volume><fpage>500</fpage><lpage>506</lpage><pub-id pub-id-type="doi">10.1016/j.conb.2005.08.015</pub-id></element-citation></ref><ref id="bib8"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Belyaev</surname><given-names>ND</given-names></name><name><surname>Wood</surname><given-names>IC</given-names></name><name><surname>Bruce</surname><given-names>AW</given-names></name><name><surname>Street</surname><given-names>M</given-names></name><name><surname>Trinh</surname><given-names>JB</given-names></name><name><surname>Buckley</surname><given-names>NJ</given-names></name></person-group><year>2004</year><article-title>Distinct RE-1 silencing transcription factor-containing complexes interact with different target genes</article-title><source>The Journal of Biological Chemistry</source><volume>279</volume><fpage>556</fpage><lpage>561</lpage><pub-id pub-id-type="doi">10.1074/jbc.M310353200</pub-id></element-citation></ref><ref id="bib9"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Borg</surname><given-names>J</given-names></name><name><surname>Papadopoulos</surname><given-names>P</given-names></name><name><surname>Georgitsi</surname><given-names>M</given-names></name><name><surname>Gutierrez</surname><given-names>L</given-names></name><name><surname>Grech</surname><given-names>G</given-names></name><name><surname>Fanis</surname><given-names>P</given-names></name><name><surname>Phylactides</surname><given-names>M</given-names></name><name><surname>Verkerk</surname><given-names>AJ</given-names></name><name><surname>van der Spek</surname><given-names>PJ</given-names></name><name><surname>Scerri</surname><given-names>CA</given-names></name><name><surname>Cassar</surname><given-names>W</given-names></name><name><surname>Galdies</surname><given-names>R</given-names></name><name><surname>van Ijcken</surname><given-names>W</given-names></name><name><surname>Ozgür</surname><given-names>Z</given-names></name><name><surname>Gillemans</surname><given-names>N</given-names></name><name><surname>Hou</surname><given-names>J</given-names></name><name><surname>Bugeja</surname><given-names>M</given-names></name><name><surname>Grosveld</surname><given-names>FG</given-names></name><name><surname>von Lindern</surname><given-names>M</given-names></name><name><surname>Felice</surname><given-names>AE</given-names></name><name><surname>Patrinos</surname><given-names>GP</given-names></name><name><surname>Philipsen</surname><given-names>S</given-names></name></person-group><year>2010</year><article-title>Haploinsufficiency for the erythroid transcription factor KLF1 causes hereditary persistence of fetal hemoglobin</article-title><source>Nature Genetics</source><volume>42</volume><fpage>801</fpage><lpage>805</lpage><pub-id pub-id-type="doi">10.1038/ng.630</pub-id></element-citation></ref><ref id="bib10"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Brewster</surname><given-names>A</given-names></name><name><surname>Bender</surname><given-names>RA</given-names></name><name><surname>Chen</surname><given-names>Y</given-names></name><name><surname>Dube</surname><given-names>C</given-names></name><name><surname>Eghbal-Ahmadi</surname><given-names>M</given-names></name><name><surname>Baram</surname><given-names>TZ</given-names></name></person-group><year>2002</year><article-title>Developmental febrile seizures modulate hippocampal gene expression of hyperpolarization-activated channels in an isoform- and cell-specific manner</article-title><source>The Journal of Neuroscience</source><volume>22</volume><fpage>4591</fpage><lpage>4599</lpage></element-citation></ref><ref id="bib11"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Bristol</surname><given-names>LA</given-names></name><name><surname>Rothstein</surname><given-names>JD</given-names></name></person-group><year>1998</year><article-title>Use of antisense oligodeoxynucleotides to inhibit expression of glutamate transporter subtypes</article-title><source>Methods in Enzymology</source><volume>296</volume><fpage>514</fpage><lpage>529</lpage><pub-id pub-id-type="doi">10.1016/S0076-6879(98)96037-2</pub-id></element-citation></ref><ref id="bib12"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Brooks-Kayal</surname><given-names>AR</given-names></name><name><surname>Raol</surname><given-names>YH</given-names></name><name><surname>Russek</surname><given-names>SJ</given-names></name></person-group><year>2009</year><article-title>Alteration of epileptogenesis genes</article-title><source>Neurotherapeutics</source><volume>6</volume><fpage>312</fpage><lpage>318</lpage><pub-id pub-id-type="doi">10.1016/j.nurt.2009.01.019</pub-id></element-citation></ref><ref id="bib13"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Calderone</surname><given-names>A</given-names></name><name><surname>Jover</surname><given-names>T</given-names></name><name><surname>Noh</surname><given-names>KM</given-names></name><name><surname>Tanaka</surname><given-names>H</given-names></name><name><surname>Yokota</surname><given-names>H</given-names></name><name><surname>Lin</surname><given-names>Y</given-names></name><name><surname>Grooms</surname><given-names>SY</given-names></name><name><surname>Regis</surname><given-names>R</given-names></name><name><surname>Bennett</surname><given-names>MV</given-names></name><name><surname>Zukin</surname><given-names>RS</given-names></name></person-group><year>2003</year><article-title>Ischemic insults derepress the gene silencer REST in neurons destined to die</article-title><source>The Journal of Neuroscience</source><volume>23</volume><fpage>2112</fpage><lpage>2121</lpage></element-citation></ref><ref id="bib14"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Chen</surname><given-names>ZF</given-names></name><name><surname>Paquette</surname><given-names>AJ</given-names></name><name><surname>Anderson</surname><given-names>DJ</given-names></name></person-group><year>1998</year><article-title>NRSF/REST is required in vivo for repression of multiple neuronal target genes during embryogenesis</article-title><source>Nature Genetics</source><volume>20</volume><fpage>136</fpage><lpage>142</lpage><pub-id pub-id-type="doi">10.1038/2431</pub-id></element-citation></ref><ref id="bib15"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Coulson</surname><given-names>JM</given-names></name></person-group><year>2005</year><article-title>Transcriptional regulation: cancer, neurons 4and the REST</article-title><source>Current Biology</source><volume>15</volume><fpage>R665</fpage><lpage>R668</lpage><pub-id pub-id-type="doi">10.1016/j.cub.2005.08.032</pub-id></element-citation></ref><ref id="bib16"><element-citation publication-type="book"><person-group person-group-type="author"><name><surname>Dingledine</surname><given-names>R</given-names></name></person-group><year>2012</year><article-title>Glutamatergic mechanisms related to epilepsy: ionotropic receptors</article-title><person-group person-group-type="editor"><name><surname>Noebel</surname><given-names>JL</given-names></name><name><surname>Avoli</surname><given-names>M</given-names></name><name><surname>Rogawski</surname><given-names>MA</given-names></name><name><surname>Olsen</surname><given-names>RW</given-names></name><name><surname>Delgado-Escueta</surname><given-names>AV</given-names></name></person-group><source>Jasper's basic mechanisms of the epilepsies</source><edition>4th edition</edition><publisher-loc>New York</publisher-loc><publisher-name>Oxford University Press</publisher-name><fpage>122</fpage><lpage>131</lpage></element-citation></ref><ref id="bib17"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Ernsberger</surname><given-names>U</given-names></name></person-group><year>2012</year><article-title>Regulation of gene expression during early neuronal differentiation: evidence for patterns conserved across neuron populations and vertebrate classes</article-title><source>Cell and Tissue Research</source><volume>348</volume><fpage>1</fpage><lpage>27</lpage><pub-id pub-id-type="doi">10.1007/s00441-012-1367-y</pub-id></element-citation></ref><ref id="bib18"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Gao</surname><given-names>Z</given-names></name><name><surname>Ure</surname><given-names>K</given-names></name><name><surname>Ding</surname><given-names>P</given-names></name><name><surname>Nashaat</surname><given-names>M</given-names></name><name><surname>Yuan</surname><given-names>L</given-names></name><name><surname>Ma</surname><given-names>J</given-names></name><name><surname>Hammer</surname><given-names>RE</given-names></name><name><surname>Hsieh</surname><given-names>J</given-names></name></person-group><year>2011</year><article-title>The master negative regulator REST/NRSF controls adult neurogenesis by restraining the neurogenic program in quiescent stem cells</article-title><source>The Journal of Neuroscience</source><volume>31</volume><fpage>9772</fpage><lpage>9786</lpage><pub-id pub-id-type="doi">10.1523/JNEUROSCI.1604-11.2011</pub-id></element-citation></ref><ref id="bib19"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Garriga-Canut</surname><given-names>M</given-names></name><name><surname>Schoenike</surname><given-names>B</given-names></name><name><surname>Qazi</surname><given-names>R</given-names></name><name><surname>Bergendahl</surname><given-names>K</given-names></name><name><surname>Daley</surname><given-names>TJ</given-names></name><name><surname>Pfender</surname><given-names>RM</given-names></name><name><surname>Morrison</surname><given-names>JF</given-names></name><name><surname>Ockuly</surname><given-names>J</given-names></name><name><surname>Stafstrom</surname><given-names>C</given-names></name><name><surname>Sutula</surname><given-names>T</given-names></name><name><surname>Roopra A</surname></name></person-group><year>2006</year><article-title>2-Deoxy-D-glucose reduces epilepsy progression by NRSF-CtBP-dependent metabolic regulation of chromatin structure</article-title><source>Nature Neuroscience</source><volume>9</volume><fpage>1382</fpage><lpage>1387</lpage><pub-id pub-id-type="doi">10.1038/nn1791</pub-id></element-citation></ref><ref id="bib20"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Gates</surname><given-names>KP</given-names></name><name><surname>Mentzer</surname><given-names>L</given-names></name><name><surname>Karlstrom</surname><given-names>RO</given-names></name><name><surname>Sirotkin</surname><given-names>HI</given-names></name></person-group><year>2010</year><article-title>The transcriptional repressor REST/NRSF modulates hedgehog signaling</article-title><source>Developmental Biology</source><volume>340</volume><fpage>293</fpage><lpage>295</lpage><pub-id pub-id-type="doi">10.1016/j.ydbio.2010.01.029</pub-id></element-citation></ref><ref id="bib21"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Gilar</surname><given-names>M</given-names></name><name><surname>Belenky</surname><given-names>A</given-names></name><name><surname>Budman</surname><given-names>Y</given-names></name><name><surname>Smisek</surname><given-names>DL</given-names></name><name><surname>Cohen</surname><given-names>S</given-names></name></person-group><year>1998</year><article-title>Study of phosphorothioate-modified oligonucleotide resistance to 3'-exonuclease using capillary electrophoresis</article-title><source>Journal of Chromatography B Biomedical Sciences and Appliances</source><volume>714</volume><fpage>13</fpage><lpage>20</lpage><pub-id pub-id-type="doi">10.1016/S0378-4347(98)00160-1</pub-id></element-citation></ref><ref id="bib22"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Gillies</surname><given-names>SG</given-names></name><name><surname>Haddley</surname><given-names>K</given-names></name><name><surname>Vasiliou</surname><given-names>SA</given-names></name><name><surname>Jacobson</surname><given-names>GM</given-names></name><name><surname>von Mentzer</surname><given-names>B</given-names></name><name><surname>Bubb</surname><given-names>VJ</given-names></name><name><surname>Quinn</surname><given-names>JP</given-names></name></person-group><year>2011</year><article-title>Distinct gene expression profiles directed by the isoforms of the transcription factor neuron-restrictive silencer factor in human SK-N-AS neuroblastoma cells</article-title><source>Journal of Molecular Neuroscience</source><volume>44</volume><fpage>77</fpage><lpage>90</lpage><pub-id pub-id-type="doi">10.1007/s12031-010-9420-3</pub-id></element-citation></ref><ref id="bib23"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Goldberg</surname><given-names>EM</given-names></name><name><surname>Coulter</surname><given-names>DA</given-names></name></person-group><year>2013</year><article-title>Mechanisms of epileptogenesis: a convergence on neural circuit dysfunction</article-title><source>Nature Reviews Neuroscience</source><volume>14</volume><fpage>337</fpage><lpage>349</lpage><pub-id pub-id-type="doi">10.1038/nrn3482</pub-id></element-citation></ref><ref id="bib24"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Gonzalez</surname><given-names>MI</given-names></name><name><surname>Cruz Del Angel</surname><given-names>Y</given-names></name><name><surname>Brooks-Kayal</surname><given-names>A</given-names></name></person-group><year>2013</year><article-title>Down-regulation of gephyrin and GABAA receptor subunits during epileptogenesis in the CA1 region of hippocampus</article-title><source>Epilepsia</source><volume>54</volume><fpage>616</fpage><lpage>624</lpage><pub-id pub-id-type="doi">10.1111/epi.12063</pub-id></element-citation></ref><ref id="bib25"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Hammar</surname><given-names>P</given-names></name><name><surname>Walldén</surname><given-names>M</given-names></name><name><surname>Fange</surname><given-names>D</given-names></name><name><surname>Persson</surname><given-names>F</given-names></name><name><surname>Baltekin</surname><given-names>O</given-names></name><name><surname>Ullman</surname><given-names>G</given-names></name><name><surname>Leroy</surname><given-names>P</given-names></name><name><surname>Elf</surname><given-names>J</given-names></name></person-group><year>2014</year><article-title>Direct measurement of transcription factor dissociation excludes a simple operator occupancy model for gene regulation</article-title><source>Nature Genetics</source><volume>46</volume><fpage>405</fpage><lpage>408</lpage><pub-id pub-id-type="doi">10.1038/ng.2905</pub-id></element-citation></ref><ref id="bib26"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Hildebrand</surname><given-names>MS</given-names></name><name><surname>Dahl</surname><given-names>HH</given-names></name><name><surname>Damiano</surname><given-names>JA</given-names></name><name><surname>Smith</surname><given-names>RJ</given-names></name><name><surname>Scheffer</surname><given-names>IE</given-names></name><name><surname>Berkovic</surname><given-names>SF</given-names></name></person-group><year>2013</year><article-title>Recent advances in the molecular genetics of epilepsy</article-title><source>Journal of Medical Genetics</source><volume>50</volume><fpage>271</fpage><lpage>279</lpage><pub-id pub-id-type="doi">10.1136/jmedgenet-2012-101448</pub-id></element-citation></ref><ref id="bib27"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Hsieh</surname><given-names>J</given-names></name><name><surname>Nakashima</surname><given-names>K</given-names></name><name><surname>Kuwabara</surname><given-names>T</given-names></name><name><surname>Mejia</surname><given-names>E</given-names></name><name><surname>Gage</surname><given-names>FH</given-names></name></person-group><year>2004</year><article-title>Histone deacetylase inhibition-mediated neuronal differentiation of multipotent adult neural progenitor cells</article-title><source>Proceedings of the National Academy of Sciences of USA</source><volume>101</volume><fpage>16659</fpage><lpage>16664</lpage><pub-id pub-id-type="doi">10.1073/pnas.0407643101</pub-id></element-citation></ref><ref id="bib28"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Hu</surname><given-names>XL</given-names></name><name><surname>Cheng</surname><given-names>X</given-names></name><name><surname>Cai</surname><given-names>L</given-names></name><name><surname>Tan</surname><given-names>GH</given-names></name><name><surname>Xu</surname><given-names>L</given-names></name><name><surname>Feng</surname><given-names>XY</given-names></name><name><surname>Lu</surname><given-names>TJ</given-names></name><name><surname>Xiong</surname><given-names>H</given-names></name><name><surname>Fei</surname><given-names>J</given-names></name><name><surname>Xiong</surname><given-names>ZQ</given-names></name></person-group><year>2011</year><article-title>Conditional deletion of NRSF in forebrain neurons accelerates epileptogenesis in the kindling model</article-title><source>Cerebral Cortex</source><volume>21</volume><fpage>2158</fpage><lpage>2165</lpage><pub-id pub-id-type="doi">10.1093/cercor/bhq284</pub-id></element-citation></ref><ref id="bib29"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Hwang</surname><given-names>CK</given-names></name><name><surname>Kim</surname><given-names>CS</given-names></name><name><surname>Kim do</surname><given-names>K</given-names></name><name><surname>Law</surname><given-names>PY</given-names></name><name><surname>Wei</surname><given-names>LN</given-names></name><name><surname>Loh</surname><given-names>HH</given-names></name></person-group><year>2010</year><article-title>Up-regulation of the mu-opioid receptor gene is mediated through chromatin remodeling and transcriptional factors in differentiated neuronal cells</article-title><source>Molecular Pharmacology</source><volume>78</volume><fpage>58</fpage><lpage>68</lpage><pub-id pub-id-type="doi">10.1124/mol.110.064311</pub-id></element-citation></ref><ref id="bib30"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Jimenez-Mateos</surname><given-names>EM</given-names></name><name><surname>Engel</surname><given-names>T</given-names></name><name><surname>Merino-Serrais</surname><given-names>P</given-names></name><name><surname>McKiernan</surname><given-names>RC</given-names></name><name><surname>Tanaka</surname><given-names>K</given-names></name><name><surname>Mouri</surname><given-names>G</given-names></name><name><surname>Sano</surname><given-names>T</given-names></name><name><surname>O'Tuathaigh</surname><given-names>C</given-names></name><name><surname>Waddington</surname><given-names>JL</given-names></name><name><surname>Prenter</surname><given-names>S</given-names></name><name><surname>Delanty</surname><given-names>N</given-names></name><name><surname>Farrell</surname><given-names>MA</given-names></name><name><surname>O'Brien</surname><given-names>DF</given-names></name><name><surname>Conroy</surname><given-names>RM</given-names></name><name><surname>Stallings</surname><given-names>RL</given-names></name><name><surname>DeFelipe</surname><given-names>J</given-names></name><name><surname>Henshall</surname><given-names>DC</given-names></name></person-group><year>2012</year><article-title>Silencing microRNA-134 produces neuroprotective and prolonged seizure-suppressive effects</article-title><source>Nature Medicine</source><volume>18</volume><fpage>1087</fpage><lpage>1094</lpage><pub-id pub-id-type="doi">10.1038/nm.2834</pub-id></element-citation></ref><ref id="bib31"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Johnson</surname><given-names>DS</given-names></name><name><surname>Mortazavi</surname><given-names>A</given-names></name><name><surname>Myers</surname><given-names>RM</given-names></name><name><surname>Wold</surname><given-names>B</given-names></name></person-group><year>2007</year><article-title>Genome-wide mapping of in vivo protein-DNA interactions</article-title><source>Science</source><volume>316</volume><fpage>1497</fpage><lpage>1502</lpage><pub-id pub-id-type="doi">10.1126/science.1141319</pub-id></element-citation></ref><ref id="bib32"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Juliandi</surname><given-names>B</given-names></name><name><surname>Abematsu</surname><given-names>M</given-names></name><name><surname>Nakashima</surname><given-names>K</given-names></name></person-group><year>2010</year><article-title>Chromatin remodeling in neural stem cell differentiation</article-title><source>Current Opinion in Neurobiology</source><volume>20</volume><fpage>408</fpage><lpage>415</lpage><pub-id pub-id-type="doi">10.1016/j.conb.2010.04.001</pub-id></element-citation></ref><ref id="bib33"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Kaneko</surname><given-names>N</given-names></name><name><surname>Hwang</surname><given-names>JY</given-names></name><name><surname>Gertner</surname><given-names>M</given-names></name><name><surname>Pontarelli</surname><given-names>F</given-names></name><name><surname>Zukin</surname><given-names>RS</given-names></name></person-group><year>2014</year><article-title>Casein kinase 1 suppresses activation of REST in insulted hippocampal neurons and halts ischemia-induced neuronal death</article-title><source>The Journal of Neuroscience</source><volume>34</volume><fpage>6030</fpage><lpage>6039</lpage><pub-id pub-id-type="doi">10.1523/JNEUROSCI.4045-13.2014</pub-id></element-citation></ref><ref id="bib34"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Khirug</surname><given-names>S</given-names></name><name><surname>Ahmad</surname><given-names>F</given-names></name><name><surname>Puskarjov</surname><given-names>M</given-names></name><name><surname>Afzalov</surname><given-names>R</given-names></name><name><surname>Kaila</surname><given-names>K</given-names></name><name><surname>Blaesse</surname><given-names>P</given-names></name></person-group><year>2010</year><article-title>A single seizure episode leads to a rapid functional activation of KCC2 in the neonatal rat hippocampus</article-title><source>The Journal of Neuroscience</source><volume>30</volume><fpage>12028</fpage><lpage>12035</lpage><pub-id pub-id-type="doi">10.1523/JNEUROSCI.3154-10.2010</pub-id></element-citation></ref><ref id="bib35"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Kim</surname><given-names>SY</given-names></name><name><surname>Buckwalter</surname><given-names>M</given-names></name><name><surname>Soreq</surname><given-names>H</given-names></name><name><surname>Vezzani</surname><given-names>A</given-names></name><name><surname>Kaufer</surname><given-names>D</given-names></name></person-group><year>2012</year><article-title>Blood-brain barrier dysfunction-induced inflammatory signaling in brain pathology and epileptogenesis</article-title><source>Epilepsia</source><volume>53</volume><supplement>suppl 6</supplement><fpage>37</fpage><lpage>44</lpage><pub-id pub-id-type="doi">10.1111/j.1528-1167.2012.03701.x</pub-id></element-citation></ref><ref id="bib36"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Kingwell</surname><given-names>K</given-names></name></person-group><year>2013</year><article-title>GRIN2A mutations identified as key genetic drivers of epilepsy-aphasia spectrum disorders</article-title><source>Nature Reviews Neurology</source><volume>9</volume><fpage>541</fpage><pub-id pub-id-type="doi">10.1038/nrneurol.2013.181</pub-id></element-citation></ref><ref id="bib37"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Kobow</surname><given-names>K</given-names></name><name><surname>Blümcke</surname><given-names>I</given-names></name></person-group><year>2012</year><article-title>The emerging role of DNA methylation in epileptogenesis</article-title><source>Epilepsia</source><volume>53</volume><supplement>Suppl 9</supplement><fpage>11</fpage><lpage>20</lpage><pub-id pub-id-type="doi">10.1111/epi.12031</pub-id></element-citation></ref><ref id="bib38"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Kok</surname><given-names>FO</given-names></name><name><surname>Taibi</surname><given-names>A</given-names></name><name><surname>Wanner</surname><given-names>SJ</given-names></name><name><surname>Xie</surname><given-names>X</given-names></name><name><surname>Moravec</surname><given-names>CE</given-names></name><name><surname>Love</surname><given-names>CE</given-names></name><name><surname>Prince</surname><given-names>VE</given-names></name><name><surname>Mumm</surname><given-names>JS</given-names></name><name><surname>Sirotkin</surname><given-names>HI</given-names></name></person-group><year>2012</year><article-title>Zebrafish rest regulates developmental gene expression but not neurogenesis</article-title><source>Development</source><volume>139</volume><fpage>3838</fpage><lpage>3848</lpage><pub-id pub-id-type="doi">10.1242/dev.080994</pub-id></element-citation></ref><ref id="bib39"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Kohyama</surname><given-names>J</given-names></name><name><surname>Kojima</surname><given-names>T</given-names></name><name><surname>Takatsuka</surname><given-names>E</given-names></name><name><surname>Yamashita</surname><given-names>T</given-names></name><name><surname>Namiki</surname><given-names>J</given-names></name><name><surname>Hsieh</surname><given-names>J</given-names></name><name><surname>Gage</surname><given-names>FH</given-names></name><name><surname>Namihira</surname><given-names>M</given-names></name><name><surname>Okano</surname><given-names>H</given-names></name><name><surname>Sawamoto</surname><given-names>K</given-names></name><name><surname>Nakashima</surname><given-names>K</given-names></name></person-group><year>2008</year><article-title>Epigenetic regulation of neural cell differentiation plasticity in the adult mammalian brain</article-title><source>Proceedings of the National Academy of Sciences of USA</source><volume>105</volume><fpage>18012</fpage><lpage>18017</lpage><pub-id pub-id-type="doi">10.1073/pnas.0808417105</pub-id></element-citation></ref><ref id="bib40"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Korosi</surname><given-names>A</given-names></name><name><surname>Shanabrough</surname><given-names>M</given-names></name><name><surname>McClelland</surname><given-names>S</given-names></name><name><surname>Liu</surname><given-names>ZW</given-names></name><name><surname>Borok</surname><given-names>E</given-names></name><name><surname>Gao</surname><given-names>XB</given-names></name><name><surname>Horvath</surname><given-names>TL</given-names></name><name><surname>Baram</surname><given-names>TZ</given-names></name></person-group><year>2010</year><article-title>Early-life experience reduces excitation to stress-responsive hypothalamic neurons and reprograms the expression of corticotropin-releasing hormone</article-title><source>The Journal of Neuroscience</source><volume>30</volume><fpage>703</fpage><lpage>713</lpage><pub-id pub-id-type="doi">10.1523/JNEUROSCI.4214-09.2010</pub-id></element-citation></ref><ref id="bib41"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Kuwabara</surname><given-names>T</given-names></name><name><surname>Hsieh</surname><given-names>J</given-names></name><name><surname>Nakashima</surname><given-names>K</given-names></name><name><surname>Taira</surname><given-names>K</given-names></name><name><surname>Gage</surname><given-names>FH</given-names></name></person-group><year>2004</year><article-title>A small modulatory dsRNA specifies the fate of adult neural stem cells</article-title><source>Cell</source><volume>116</volume><fpage>779</fpage><lpage>793</lpage><pub-id pub-id-type="doi">10.1016/S0092-8674(04)00248-X</pub-id></element-citation></ref><ref id="bib42"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Kuwabara</surname><given-names>T</given-names></name><name><surname>Hsieh</surname><given-names>J</given-names></name><name><surname>Nakashima</surname><given-names>K</given-names></name><name><surname>Warashina</surname><given-names>M</given-names></name><name><surname>Taira</surname><given-names>K</given-names></name><name><surname>Gage</surname><given-names>FH</given-names></name></person-group><year>2005</year><article-title>The NRSE smRNA specifies the fate of adult hippocampal neural stem cells</article-title><source>Nucleic Acids Symposium Series</source><volume>49</volume><fpage>87</fpage><lpage>88</lpage><pub-id pub-id-type="doi">10.1093/nass/49.1.87</pub-id></element-citation></ref><ref id="bib43"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Lawinger</surname><given-names>P</given-names></name><name><surname>Venugopal</surname><given-names>R</given-names></name><name><surname>Guo</surname><given-names>ZS</given-names></name><name><surname>Immaneni</surname><given-names>A</given-names></name><name><surname>Sengupta</surname><given-names>D</given-names></name><name><surname>Lu</surname><given-names>W</given-names></name><name><surname>Rastelli</surname><given-names>L</given-names></name><name><surname>Marin Dias Carneiro</surname><given-names>A</given-names></name><name><surname>Levin</surname><given-names>V</given-names></name><name><surname>Fuller</surname><given-names>GN</given-names></name><name><surname>Echelard</surname><given-names>Y</given-names></name><name><surname>Majumder</surname><given-names>S</given-names></name></person-group><year>2000</year><article-title>The neuronal repressor REST/NRSF is an essential regulator in medulloblastoma cells</article-title><source>Nature Medicine</source><volume>6</volume><fpage>826</fpage><lpage>831</lpage><pub-id pub-id-type="doi">10.1038/77565</pub-id></element-citation></ref><ref id="bib44"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Lepagnol-Bestel</surname><given-names>AM</given-names></name><name><surname>Zvara</surname><given-names>A</given-names></name><name><surname>Maussion</surname><given-names>G</given-names></name><name><surname>Quignon</surname><given-names>F</given-names></name><name><surname>Ngimbous</surname><given-names>B</given-names></name><name><surname>Ramoz</surname><given-names>N</given-names></name><name><surname>Imbeaud</surname><given-names>S</given-names></name><name><surname>Loe-Mie</surname><given-names>Y</given-names></name><name><surname>Benihoud</surname><given-names>K</given-names></name><name><surname>Agier</surname><given-names>N</given-names></name><name><surname>Salin</surname><given-names>PA</given-names></name><name><surname>Cardona</surname><given-names>A</given-names></name><name><surname>Khung-Savatovsky</surname><given-names>S</given-names></name><name><surname>Kallunki</surname><given-names>P</given-names></name><name><surname>Delabar</surname><given-names>JM</given-names></name><name><surname>Puskas</surname><given-names>LG</given-names></name><name><surname>Delacroix</surname><given-names>H</given-names></name><name><surname>Aggerbeck</surname><given-names>L</given-names></name><name><surname>Delezoide</surname><given-names>AL</given-names></name><name><surname>Delattre</surname><given-names>O</given-names></name><name><surname>Gorwood</surname><given-names>P</given-names></name><name><surname>Moalic</surname><given-names>JM</given-names></name><name><surname>Simonneau</surname><given-names>M</given-names></name></person-group><year>2009</year><article-title>DYRK1A interacts with the REST/NRSF-SWI/SNF chromatin remodelling complex to deregulate gene clusters involved in the neuronal phenotypic traits of Down syndrome</article-title><source>Human Molecular Genetics</source><volume>18</volume><fpage>1405</fpage><lpage>1414</lpage><pub-id pub-id-type="doi">10.1093/hmg/ddp047</pub-id></element-citation></ref><ref id="bib45"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Liu</surname><given-names>G</given-names></name><name><surname>Gu</surname><given-names>B</given-names></name><name><surname>He</surname><given-names>XP</given-names></name><name><surname>Joshi</surname><given-names>R</given-names></name><name><surname>Wackerle</surname><given-names>HD</given-names></name><name><surname>Rodriguiz</surname><given-names>RM</given-names></name><name><surname>Wetsel</surname><given-names>WC</given-names></name><name><surname>McNamara</surname><given-names>JO</given-names></name></person-group><year>2013</year><article-title>Transient inhibition of TrkB Kinase after status epilepticus prevents development of temporal lobe epilepsy</article-title><source>Neuron</source><volume>79</volume><fpage>31</fpage><lpage>38</lpage><pub-id pub-id-type="doi">10.1016/j.neuron.2013.04.027</pub-id></element-citation></ref><ref id="bib46"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Lovén</surname><given-names>J</given-names></name><name><surname>Hoke</surname><given-names>HA</given-names></name><name><surname>Lin</surname><given-names>CY</given-names></name><name><surname>Lau</surname><given-names>A</given-names></name><name><surname>Orlando</surname><given-names>DA</given-names></name><name><surname>Vakoc</surname><given-names>CR</given-names></name><name><surname>Bradner</surname><given-names>JE</given-names></name><name><surname>Lee</surname><given-names>TI</given-names></name><name><surname>Young</surname><given-names>RA</given-names></name></person-group><year>2013</year><article-title>Selective inhibition of tumor oncogenes by disruption of super-enhancers</article-title><source>Cell</source><volume>153</volume><fpage>320</fpage><lpage>334</lpage><pub-id pub-id-type="doi">10.1016/j.cell.2013.03.036</pub-id></element-citation></ref><ref id="bib47"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Lu</surname><given-names>T</given-names></name><name><surname>Aron</surname><given-names>L</given-names></name><name><surname>Zullo</surname><given-names>J</given-names></name><name><surname>Pan</surname><given-names>Y</given-names></name><name><surname>Kim</surname><given-names>H</given-names></name><name><surname>Chen</surname><given-names>Y</given-names></name><name><surname>Yang</surname><given-names>TH</given-names></name><name><surname>Kim</surname><given-names>HM</given-names></name><name><surname>Drake</surname><given-names>D</given-names></name><name><surname>Liu</surname><given-names>XS</given-names></name><name><surname>Bennett</surname><given-names>DA</given-names></name><name><surname>Colaiácovo</surname><given-names>MP</given-names></name><name><surname>Yankner</surname><given-names>BA</given-names></name></person-group><year>2014</year><article-title>REST and stress resistance in ageing and Alzheimer's disease</article-title><source>Nature</source><volume>507</volume><fpage>448</fpage><lpage>454</lpage><pub-id pub-id-type="doi">10.1038/nature13163</pub-id></element-citation></ref><ref id="bib48"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Mandel</surname><given-names>G</given-names></name><name><surname>Fiondella</surname><given-names>CG</given-names></name><name><surname>Covey</surname><given-names>MV</given-names></name><name><surname>Lu</surname><given-names>DD</given-names></name><name><surname>Loturco</surname><given-names>JJ</given-names></name><name><surname>Ballas</surname><given-names>N</given-names></name></person-group><year>2011</year><article-title>Repressor element 1 silencing transcription factor (REST) controls radial migration and temporal neuronal specification during neocortical development</article-title><source>Proceedings of the National Academy of Sciences of USA</source><volume>108</volume><fpage>16789</fpage><lpage>16794</lpage><pub-id pub-id-type="doi">10.1073/pnas.1113486108</pub-id></element-citation></ref><ref id="bib49"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Maslarova</surname><given-names>A</given-names></name><name><surname>Salar</surname><given-names>S</given-names></name><name><surname>Lapilovar</surname><given-names>E</given-names></name><name><surname>Friedman</surname><given-names>A</given-names></name><name><surname>Veh</surname><given-names>RW</given-names></name><name><surname>Heinemann</surname><given-names>U</given-names></name></person-group><year>2013</year><article-title>Increased susceptibility to acetylcholine in the entorhinal cortex of pilocarpine treated rats involves alterations in KCNQ channels</article-title><source>Neurobiology of Disease</source><volume>56</volume><fpage>14</fpage><lpage>24</lpage><pub-id pub-id-type="doi">10.1016/j.nbd.2013.02.016</pub-id></element-citation></ref><ref id="bib50"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Maroso</surname><given-names>M</given-names></name><name><surname>Balosso</surname><given-names>S</given-names></name><name><surname>Ravizza</surname><given-names>T</given-names></name><name><surname>Liu</surname><given-names>J</given-names></name><name><surname>Bianchi</surname><given-names>ME</given-names></name><name><surname>Vezzani</surname><given-names>A</given-names></name></person-group><year>2011</year><article-title>Interleukin-1 type 1 receptor/Toll-like receptor signalling in epilepsy: the importance of IL-1beta and high-mobility group box 1</article-title><source>Journal of Internal Medicine</source><volume>270</volume><fpage>319</fpage><lpage>326</lpage><pub-id pub-id-type="doi">10.1111/j.1365-2796.2011.02431.x</pub-id></element-citation></ref><ref id="bib51"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>McClelland</surname><given-names>S</given-names></name><name><surname>Flynn</surname><given-names>C</given-names></name><name><surname>Dubé</surname><given-names>C</given-names></name><name><surname>Richichi</surname><given-names>C</given-names></name><name><surname>Zha</surname><given-names>Q</given-names></name><name><surname>Ghestem</surname><given-names>A</given-names></name><name><surname>Esclapez</surname><given-names>M</given-names></name><name><surname>Bernard</surname><given-names>C</given-names></name><name><surname>Baram</surname><given-names>TZ</given-names></name></person-group><year>2011a</year><article-title>Neuron-restrictive silencer factor-mediated hyperpolarization-activated cyclic nucleotide gated channelopathy in experimental temporal lobe epilepsy</article-title><source>Annals of Neurology</source><volume>70</volume><fpage>454</fpage><lpage>464</lpage><pub-id pub-id-type="doi">10.1002/ana.22479</pub-id></element-citation></ref><ref id="bib52"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>McClelland</surname><given-names>S</given-names></name><name><surname>Korosi</surname><given-names>A</given-names></name><name><surname>Cope</surname><given-names>J</given-names></name><name><surname>Ivy</surname><given-names>A</given-names></name><name><surname>Baram</surname><given-names>TZ</given-names></name></person-group><year>2011b</year><article-title>Emerging roles of epigenetic mechanisms in the enduring effects of early-life stress and experience on learning and memory</article-title><source>Neurobiology of Learning and Memory</source><volume>96</volume><fpage>79</fpage><lpage>88</lpage><pub-id pub-id-type="doi">10.1016/j.nlm.2011.02.008</pub-id></element-citation></ref><ref id="bib53"><element-citation publication-type="book"><person-group person-group-type="author"><name><surname>Meisler</surname><given-names>MH</given-names></name><name><surname>O'Brien</surname><given-names>JE</given-names></name></person-group><year>2012</year><article-title>Gene interactions and modifiers in epilepsy</article-title><person-group person-group-type="editor"><name><surname>Noebel</surname><given-names>JL</given-names></name><name><surname>Avoli</surname><given-names>M</given-names></name><name><surname>Rogawski</surname><given-names>MA</given-names></name><name><surname>Olsen</surname><given-names>RW</given-names></name><name><surname>Delgado-Escueta</surname><given-names>AV</given-names></name></person-group><source>Jasper's basic mechanisms of the epilepsies</source><edition>4th edition</edition><publisher-loc>New York</publisher-loc><publisher-name>Oxford University Press</publisher-name><fpage>763</fpage><lpage>770</lpage></element-citation></ref><ref id="bib54"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Mootha</surname><given-names>VK</given-names></name><name><surname>Lindgren</surname><given-names>CM</given-names></name><name><surname>Eriksson</surname><given-names>KF</given-names></name><name><surname>Subramanian</surname><given-names>A</given-names></name><name><surname>Sihag</surname><given-names>S</given-names></name><name><surname>Lehar</surname><given-names>J</given-names></name><name><surname>Puigserver</surname><given-names>P</given-names></name><name><surname>Carlsson</surname><given-names>E</given-names></name><name><surname>Ridderstråle</surname><given-names>M</given-names></name><name><surname>Laurila</surname><given-names>E</given-names></name><name><surname>Houstis</surname><given-names>N</given-names></name><name><surname>Daly</surname><given-names>MJ</given-names></name><name><surname>Patterson</surname><given-names>N</given-names></name><name><surname>Mesirov</surname><given-names>JP</given-names></name><name><surname>Golub</surname><given-names>TR</given-names></name><name><surname>Tamayo</surname><given-names>P</given-names></name><name><surname>Spiegelman</surname><given-names>B</given-names></name><name><surname>Lander</surname><given-names>ES</given-names></name><name><surname>Hirschhorn</surname><given-names>JN</given-names></name><name><surname>Altshuler</surname><given-names>D</given-names></name><name><surname>Groop</surname><given-names>LC</given-names></name></person-group><year>2003</year><article-title>PGC-1alpha-responsive genes involved in oxidative phosphorylation are coordinately downregulated in human diabetes</article-title><source>Nature Genetics</source><volume>34</volume><fpage>267</fpage><lpage>273</lpage><pub-id pub-id-type="doi">10.1038/ng1180</pub-id></element-citation></ref><ref id="bib55"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Mortazavi</surname><given-names>A</given-names></name><name><surname>Leeper Thompson</surname><given-names>EC</given-names></name><name><surname>Garcia</surname><given-names>ST</given-names></name><name><surname>Myers</surname><given-names>RM</given-names></name><name><surname>Wold</surname><given-names>B</given-names></name></person-group><year>2006</year><article-title>Comparative genomics modeling of the NRSF/REST repressor network: from single conserved sites to genome-wide repertoire</article-title><source>Genome Research</source><volume>16</volume><fpage>1208</fpage><lpage>1221</lpage><pub-id pub-id-type="doi">10.1101/gr.4997306</pub-id></element-citation></ref><ref id="bib56"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Naruse</surname><given-names>Y</given-names></name><name><surname>Aoki</surname><given-names>T</given-names></name><name><surname>Kojima</surname><given-names>T</given-names></name><name><surname>Mori</surname><given-names>N</given-names></name></person-group><year>1999</year><article-title>Neural restrictive silencer factor recruits mSin3 and histone deacetylase complex to repress neuron-specific traget genes</article-title><source>Proceedings of the National Academy of Sciences of USA</source><volume>96</volume><fpage>13691</fpage><lpage>13696</lpage><pub-id pub-id-type="doi">10.1073/pnas.96.24.13691</pub-id></element-citation></ref><ref id="bib57"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Nichols</surname><given-names>M</given-names></name><name><surname>Weih</surname><given-names>F</given-names></name><name><surname>Schmid</surname><given-names>W</given-names></name><name><surname>DeVack</surname><given-names>C</given-names></name><name><surname>Kowenz-Leutz</surname><given-names>E</given-names></name><name><surname>Luckow</surname><given-names>B</given-names></name><name><surname>Boshart</surname><given-names>M</given-names></name><name><surname>Schütz</surname><given-names>G</given-names></name></person-group><year>1992</year><article-title>Phosphorylation of CREB affects its binding to high and low affinity sites; implications for cAMP induced gene transcription</article-title><source>The EMBO Journal</source><volume>11</volume><fpage>3337</fpage><lpage>3346</lpage></element-citation></ref><ref id="bib58"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Nikitidou</surname><given-names>L</given-names></name><name><surname>Kanter-Schlifke</surname><given-names>I</given-names></name><name><surname>Dhondt</surname><given-names>J</given-names></name><name><surname>Carmeliet</surname><given-names>P</given-names></name><name><surname>Lambrechts</surname><given-names>D</given-names></name><name><surname>Kokaia</surname><given-names>M</given-names></name></person-group><year>2012</year><article-title>VEGF receptor-2 (FLK-1) overexpression in mice counteracts focal epileptic seizures</article-title><source>PLOS ONE</source><volume>7</volume><fpage>e40535</fpage><pub-id pub-id-type="doi">10.1371/journal.pone.0040535</pub-id></element-citation></ref><ref id="bib59"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Noam</surname><given-names>Y</given-names></name><name><surname>Bernard</surname><given-names>C</given-names></name><name><surname>Baram</surname><given-names>TZ</given-names></name></person-group><year>2011</year><article-title>Towards an integrated view of HCN channel role in epilepsy</article-title><source>Current Opinion in Neurobiology</source><volume>21</volume><fpage>873</fpage><lpage>879</lpage><pub-id pub-id-type="doi">10.1016/j.conb.2011.06.013</pub-id></element-citation></ref><ref id="bib60"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Noh</surname><given-names>KM</given-names></name><name><surname>Hwang</surname><given-names>JY</given-names></name><name><surname>Follenzi</surname><given-names>A</given-names></name><name><surname>Athanasiadou</surname><given-names>R</given-names></name><name><surname>Miyawaki</surname><given-names>T</given-names></name><name><surname>Greally</surname><given-names>JM</given-names></name><name><surname>Bennett</surname><given-names>MV</given-names></name><name><surname>Zukin</surname><given-names>RS</given-names></name></person-group><year>2012</year><article-title>Repressor element-1 silencing transcription factor (REST)-dependent epigenetic remodelling is critical to ischemia-induced neuronal death</article-title><source>Proceedings of the National Academy of Sciences of USA</source><volume>109</volume><fpage>E962</fpage><lpage>E971</lpage><pub-id pub-id-type="doi">10.1073/pnas.1121568109</pub-id></element-citation></ref><ref id="bib61"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Oberbauer</surname><given-names>AM</given-names></name><name><surname>Belanger</surname><given-names>JM</given-names></name><name><surname>Grossman</surname><given-names>DI</given-names></name><name><surname>Regan</surname><given-names>KR</given-names></name><name><surname>Famula</surname><given-names>TR</given-names></name></person-group><year>2010</year><article-title>Genome-wide linkage scan for loci associated with epilepsy in Belgian shepherd dogs</article-title><source>BMC Genetics</source><volume>11</volume><fpage>35</fpage><pub-id pub-id-type="doi">10.1186/1471-2156-11-35</pub-id></element-citation></ref><ref id="bib62"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Oliva</surname><given-names>M</given-names></name><name><surname>Berkovic</surname><given-names>SF</given-names></name><name><surname>Petrou</surname><given-names>S</given-names></name></person-group><year>2012</year><article-title>Sodium channels and the neurobiology of epilepsy</article-title><source>Epilepsia</source><volume>53</volume><fpage>1849</fpage><lpage>1859</lpage><pub-id pub-id-type="doi">10.1111/j.1528-1167.2012.03631.x</pub-id></element-citation></ref><ref id="bib63"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Palm</surname><given-names>K</given-names></name><name><surname>Belluardo</surname><given-names>N</given-names></name><name><surname>Metsis</surname><given-names>M</given-names></name><name><surname>Timmusk</surname><given-names>T</given-names></name></person-group><year>1998</year><article-title>Neuronal expression of zinc finger transcription factor REST/NRSF/XBR gene</article-title><source>The Journal of Neuroscience</source><volume>18</volume><fpage>1280</fpage><lpage>1296</lpage></element-citation></ref><ref id="bib64"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Palm</surname><given-names>K</given-names></name><name><surname>Metsis</surname><given-names>M</given-names></name><name><surname>Timmusk</surname><given-names>T</given-names></name></person-group><year>1999</year><article-title>Neuron-specific splicing of zinc finger transcription factor REST/NRSF/XBR is frequent in neuroblastomas and conserved in human, mouse and rat</article-title><source>Brain Research Molecular Brain Research</source><volume>72</volume><fpage>30</fpage><lpage>39</lpage><pub-id pub-id-type="doi">10.1016/S0169-328X(99)00196-5</pub-id></element-citation></ref><ref id="bib65"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Papale</surname><given-names>LA</given-names></name><name><surname>Makinson</surname><given-names>CD</given-names></name><name><surname>Christopher Ehlen</surname><given-names>J</given-names></name><name><surname>Tufik</surname><given-names>S</given-names></name><name><surname>Decker</surname><given-names>MJ</given-names></name><name><surname>Paul</surname><given-names>KN</given-names></name><name><surname>Escayg</surname><given-names>A</given-names></name></person-group><year>2013</year><article-title>Altered sleep regulation in a mouse model of SCN1A-derived genetic epilepsy with febrile seizures plus (GEFS+)</article-title><source>Epilepsia</source><volume>54</volume><fpage>625</fpage><lpage>634</lpage><pub-id pub-id-type="doi">10.1111/epi.12060</pub-id></element-citation></ref><ref id="bib66"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Pathak</surname><given-names>HR</given-names></name><name><surname>Weissinger</surname><given-names>F</given-names></name><name><surname>Terunuma</surname><given-names>M</given-names></name><name><surname>Carlson</surname><given-names>GC</given-names></name><name><surname>Hsu</surname><given-names>FC</given-names></name><name><surname>Moss</surname><given-names>SJ</given-names></name><name><surname>Coulter</surname><given-names>DA</given-names></name></person-group><year>2007</year><article-title>Disrupted dentate granule cell chloride regulation enhances excitability during development of temporal lobe epilepsy</article-title><source>The Journal of Neuroscience</source><volume>27</volume><fpage>14012</fpage><lpage>14022</lpage><pub-id pub-id-type="doi">10.1523/JNEUROSCI.4390-07.2007</pub-id></element-citation></ref><ref id="bib67"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Paxinos</surname><given-names>G</given-names></name><name><surname>Watson</surname><given-names>C</given-names></name><name><surname>Pennisi</surname><given-names>M</given-names></name><name><surname>Topple</surname><given-names>A</given-names></name></person-group><year>1985</year><article-title>Bregma, lambda and the interaural midpoint in stereotaxic surgery with rats of different sex, strain and weight</article-title><source>Journal of Neuroscience Methods</source><volume>13</volume><fpage>139</fpage><lpage>143</lpage><pub-id pub-id-type="doi">10.1016/0165-0270(85)90026-3</pub-id></element-citation></ref><ref id="bib68"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Pernhorst</surname><given-names>K</given-names></name><name><surname>Raabe</surname><given-names>A</given-names></name><name><surname>Niehusmann</surname><given-names>P</given-names></name><name><surname>van Loo</surname><given-names>KM</given-names></name><name><surname>Grote</surname><given-names>A</given-names></name><name><surname>Hoffmann</surname><given-names>P</given-names></name><name><surname>Cichon</surname><given-names>S</given-names></name><name><surname>Sander</surname><given-names>T</given-names></name><name><surname>Schoch</surname><given-names>S</given-names></name><name><surname>Becker</surname><given-names>AJ</given-names></name></person-group><year>2011</year><article-title>Promoter variants determine gamma-aminobutyric acid homeostasis-related gene transcription in human epileptic hippocampi</article-title><source>Journal of Neuropathology and Experimental Neurology</source><volume>70</volume><fpage>1080</fpage><lpage>1088</lpage><pub-id pub-id-type="doi">10.1097/NEN.0b013e318238b9af</pub-id></element-citation></ref><ref id="bib69"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Pernhorst</surname><given-names>K</given-names></name><name><surname>van Loo</surname><given-names>KM</given-names></name><name><surname>von Lehe</surname><given-names>M</given-names></name><name><surname>Priebe</surname><given-names>L</given-names></name><name><surname>Cichon</surname><given-names>S</given-names></name><name><surname>Herms</surname><given-names>S</given-names></name><name><surname>Hoffmann</surname><given-names>P</given-names></name><name><surname>Helmstaedter</surname><given-names>C</given-names></name><name><surname>Sander</surname><given-names>T</given-names></name><name><surname>Schoch</surname><given-names>S</given-names></name><name><surname>Becker</surname><given-names>AJ</given-names></name></person-group><year>2013</year><article-title>Rs6295 promoter variants of the serotonin type 1A receptor are differentially activated by c-Jun in vitro and correlate to transcript levels in human epileptic brain tissue</article-title><source>Brain Research</source><volume>1499</volume><fpage>136</fpage><lpage>144</lpage><pub-id pub-id-type="doi">10.1016/j.brainres.2012.12.045</pub-id></element-citation></ref><ref id="bib70"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Pitkänen</surname><given-names>A</given-names></name><name><surname>Bolkvadze</surname><given-names>T</given-names></name><name><surname>Immonen</surname><given-names>R</given-names></name></person-group><year>2011</year><article-title>Anti-epileptogenesis in rodent post-traumatic epilepsy models</article-title><source>Neuroscience Letters</source><volume>497</volume><fpage>163</fpage><lpage>171</lpage><pub-id pub-id-type="doi">10.1016/j.neulet.2011.02.033</pub-id></element-citation></ref><ref id="bib71"><element-citation publication-type="book"><person-group person-group-type="author"><name><surname>Poolos</surname><given-names>NP</given-names></name></person-group><year>2012</year><article-title>Hyperpolarization-activated cyclic nucleotide-gated (HCN) Ion channelopathy in epilepsy</article-title><person-group person-group-type="editor"><name><surname>Noebel</surname><given-names>JL</given-names></name><name><surname>Avoli</surname><given-names>M</given-names></name><name><surname>Rogawski</surname><given-names>MA</given-names></name><name><surname>Olsen</surname><given-names>RW</given-names></name><name><surname>Delgado-Escueta</surname><given-names>AV</given-names></name></person-group><source>Jasper's basic mechanisms of the epilepsies</source><edition>4th edition</edition><publisher-loc>New York</publisher-loc><publisher-name>Oxford University Press</publisher-name><fpage>85</fpage><lpage>95</lpage></element-citation></ref><ref id="bib72"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Pozzi</surname><given-names>D</given-names></name><name><surname>Lignani</surname><given-names>G</given-names></name><name><surname>Ferrea</surname><given-names>E</given-names></name><name><surname>Contestabile</surname><given-names>A</given-names></name><name><surname>Paonessa</surname><given-names>F</given-names></name><name><surname>D'Alessandro</surname><given-names>R</given-names></name><name><surname>Lippiello</surname><given-names>P</given-names></name><name><surname>Boido</surname><given-names>D</given-names></name><name><surname>Fassio</surname><given-names>A</given-names></name><name><surname>Meldolesi</surname><given-names>J</given-names></name><name><surname>Valtorta</surname><given-names>F</given-names></name><name><surname>Benfenati</surname><given-names>F</given-names></name><name><surname>Baldelli</surname><given-names>P</given-names></name></person-group><year>2013</year><article-title>REST/NRSF-mediated intrinsic homeostasis protects neuronal networks from hyperexcitability</article-title><source>The EMBO Journal</source><volume>32</volume><fpage>2994</fpage><lpage>3007</lpage><pub-id pub-id-type="doi">10.1038/emboj.2013.231</pub-id></element-citation></ref><ref id="bib73"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Racine</surname><given-names>RJ</given-names></name></person-group><year>1972</year><article-title>Modification of seizure activity by electrical stimulation. I. After-discharge threshold</article-title><source>Electroencephalography and Clinical Neurophysiology</source><volume>32</volume><fpage>269</fpage><lpage>279</lpage><pub-id pub-id-type="doi">10.1016/0013-4694(72)90176-9</pub-id></element-citation></ref><ref id="bib74"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Richichi</surname><given-names>C</given-names></name><name><surname>Brewster</surname><given-names>AL</given-names></name><name><surname>Bender</surname><given-names>RA</given-names></name><name><surname>Simeone</surname><given-names>TA</given-names></name><name><surname>Zha</surname><given-names>Q</given-names></name><name><surname>Yin</surname><given-names>HZ</given-names></name><name><surname>Weiss</surname><given-names>JH</given-names></name><name><surname>Baram</surname><given-names>TZ</given-names></name></person-group><year>2008</year><article-title>Mechanisms of seizure-induced 'transcriptional channelopathy' of hyperpolarization-activated cyclic nucleotide gated (HCN) channels</article-title><source>Neurobiology of Disease</source><volume>29</volume><fpage>297</fpage><lpage>305</lpage><pub-id pub-id-type="doi">10.1016/j.nbd.2007.09.003</pub-id></element-citation></ref><ref id="bib75"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Robison</surname><given-names>AJ</given-names></name><name><surname>Nestler</surname><given-names>EJ</given-names></name></person-group><year>2011</year><article-title>Transcriptional and epigenetic mechanisms of addiction</article-title><source>Nature Reviews Neuroscience</source><volume>12</volume><fpage>623</fpage><lpage>637</lpage><pub-id pub-id-type="doi">10.1038/nrn3111</pub-id></element-citation></ref><ref id="bib76"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Rodenas-Ruano</surname><given-names>A</given-names></name><name><surname>Chavez</surname><given-names>AE</given-names></name><name><surname>Cossi</surname><given-names>MJ</given-names></name><name><surname>Castillo</surname><given-names>PE</given-names></name><name><surname>Zukin</surname><given-names>RS</given-names></name></person-group><year>2012</year><article-title>REST-dependent epigenetic remodeling promotes the developmental switch in synaptic NMDA receptors</article-title><source>Nature Neuroscience</source><volume>15</volume><fpage>1382</fpage><lpage>1390</lpage><pub-id pub-id-type="doi">10.1038/nn.3214</pub-id></element-citation></ref><ref id="bib77"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Rojas</surname><given-names>A</given-names></name><name><surname>Dingledine</surname><given-names>R</given-names></name></person-group><year>2013</year><article-title>Ionotropic glutamate receptors: regulation by G-protein-coupled receptors</article-title><source>Molecular Pharmacology</source><volume>83</volume><fpage>746</fpage><lpage>752</lpage><pub-id pub-id-type="doi">10.1124/mol.112.083352</pub-id></element-citation></ref><ref id="bib78"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Roopra</surname><given-names>A</given-names></name><name><surname>Dingledine</surname><given-names>R</given-names></name><name><surname>Hsieh</surname><given-names>J</given-names></name></person-group><year>2012</year><article-title>Epigenetics and epilepsy</article-title><source>Epilepsia</source><volume>53</volume><supplement>Suppl 9</supplement><fpage>2</fpage><lpage>10</lpage><pub-id pub-id-type="doi">10.1111/epi.12030</pub-id></element-citation></ref><ref id="bib79"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Roopra</surname><given-names>A</given-names></name><name><surname>Huang</surname><given-names>Y</given-names></name><name><surname>Dingledine</surname><given-names>R</given-names></name></person-group><year>2001</year><article-title>Neurological disease: listening to gene silencers</article-title><source>Molecular Interventions</source><volume>1</volume><fpage>219</fpage><lpage>228</lpage></element-citation></ref><ref id="bib80"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Rowley</surname><given-names>S</given-names></name><name><surname>Patel</surname><given-names>M</given-names></name></person-group><year>2013</year><article-title>Mitochondrial involvement and oxidative stress in temporal lobe epilepsy</article-title><source>Free Radical Biology & Medicine</source><volume>62</volume><fpage>121</fpage><lpage>131</lpage><pub-id pub-id-type="doi">10.1016/j.freeradbiomed.2013.02.002</pub-id></element-citation></ref><ref id="bib81"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Saeed</surname><given-names>AI</given-names></name><name><surname>Bhagabati</surname><given-names>NK</given-names></name><name><surname>Braisted</surname><given-names>JC</given-names></name><name><surname>Liang</surname><given-names>W</given-names></name><name><surname>Sharov</surname><given-names>V</given-names></name><name><surname>Howe</surname><given-names>EA</given-names></name><name><surname>Li</surname><given-names>J</given-names></name><name><surname>Thiagarajan</surname><given-names>M</given-names></name><name><surname>White</surname><given-names>JA</given-names></name><name><surname>Quackenbush</surname><given-names>J</given-names></name></person-group><year>2006</year><article-title>TM4 microarray software suite</article-title><source>Methods in Enzymology</source><volume>411</volume><fpage>134</fpage><lpage>193</lpage><pub-id pub-id-type="doi">10.1016/S0076-6879(06)11009-5</pub-id></element-citation></ref><ref id="bib82"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Saeed</surname><given-names>AI</given-names></name><name><surname>Sharov</surname><given-names>V</given-names></name><name><surname>White</surname><given-names>J</given-names></name><name><surname>Li</surname><given-names>J</given-names></name><name><surname>Liang</surname><given-names>W</given-names></name><name><surname>Bhagabati</surname><given-names>N</given-names></name><name><surname>Braisted</surname><given-names>J</given-names></name><name><surname>Klapa</surname><given-names>M</given-names></name><name><surname>Currier</surname><given-names>T</given-names></name><name><surname>Thiagarajan</surname><given-names>M</given-names></name><name><surname>Sturn</surname><given-names>A</given-names></name><name><surname>Snuffin</surname><given-names>M</given-names></name><name><surname>Rezantsev</surname><given-names>A</given-names></name><name><surname>Popov</surname><given-names>D</given-names></name><name><surname>Ryltsov</surname><given-names>A</given-names></name><name><surname>Kostukovich</surname><given-names>E</given-names></name><name><surname>Borisovsky</surname><given-names>I</given-names></name><name><surname>Liu</surname><given-names>Z</given-names></name><name><surname>Vinsavich</surname><given-names>A</given-names></name><name><surname>Trush</surname><given-names>V</given-names></name><name><surname>Quackenbush</surname><given-names>J</given-names></name></person-group><year>2003</year><article-title>TM4: a free, open-source system for microarray data management and analysis</article-title><source>Biotechniques</source><volume>34</volume><fpage>374</fpage><lpage>378</lpage></element-citation></ref><ref id="bib83"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Schoenherr</surname><given-names>CJ</given-names></name><name><surname>Anderson</surname><given-names>DJ</given-names></name></person-group><year>1995</year><article-title>The neuron restrictive silencer factor (NRSF): a coordinate repressor of multiple neuron-specific genes</article-title><source>Science</source><volume>267</volume><fpage>1360</fpage><lpage>1363</lpage><pub-id pub-id-type="doi">10.1126/science.7871435</pub-id></element-citation></ref><ref id="bib84"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Sedaghat</surname><given-names>Y</given-names></name><name><surname>Bui</surname><given-names>HH</given-names></name><name><surname>Mazur</surname><given-names>C</given-names></name><name><surname>Monia</surname><given-names>BP</given-names></name></person-group><year>2013</year><article-title>Identification of REST-regulated genes and pathways using a REST-targeted antisense approach</article-title><source>Nucleic Acid Therapeutics</source><volume>23</volume><fpage>389</fpage><lpage>400</lpage><pub-id pub-id-type="doi">10.1089/nat.2013.0445</pub-id></element-citation></ref><ref id="bib85"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Shah</surname><given-names>MM</given-names></name><name><surname>Huang</surname><given-names>Z</given-names></name><name><surname>Martinello</surname><given-names>K</given-names></name></person-group><year>2013</year><article-title>HCN and KV7 (M-) channels as targets for epilepsy treatment</article-title><source>Neuropharmacology</source><volume>69</volume><fpage>75</fpage><lpage>81</lpage><pub-id pub-id-type="doi">10.1016/j.neuropharm.2012.03.005</pub-id></element-citation></ref><ref id="bib86"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Singh</surname><given-names>IS</given-names></name><name><surname>He</surname><given-names>JR</given-names></name><name><surname>Calderwood</surname><given-names>S</given-names></name><name><surname>Hasday</surname><given-names>JD</given-names></name></person-group><year>2002</year><article-title>A high affinity HSF-1 binding site in the 5’ untranslated region of the murine tumour necrosis factor-alpha gene is a transcriptional repressor</article-title><source>The Journal of Biological Chemistry</source><volume>277</volume><fpage>4981</fpage><lpage>4988</lpage><pub-id pub-id-type="doi">10.1074/jbc.M108154200</pub-id></element-citation></ref><ref id="bib87"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Singh</surname><given-names>SK</given-names></name><name><surname>Kagalwala</surname><given-names>MN</given-names></name><name><surname>Parker-Thornburg</surname><given-names>J</given-names></name><name><surname>Adams</surname><given-names>H</given-names></name><name><surname>Majumder</surname><given-names>S</given-names></name></person-group><year>2008</year><article-title>REST maintains self-renewal and pluripotency of embryonic stem cells</article-title><source>Nature</source><volume>453</volume><fpage>223</fpage><lpage>227</lpage><pub-id pub-id-type="doi">10.1038/nature06863</pub-id></element-citation></ref><ref id="bib88"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Sloviter</surname><given-names>RS</given-names></name></person-group><year>1989</year><article-title>Calcium binding protein (Calbindin-D28k) and paravalbumin immunocytochemistry; localization in the rat hippocampus with specific reference to the selective vulnerability of hippocampal neurons to seizure activity</article-title><source>The Journal of Comparative Neurology</source><volume>280</volume><fpage>183</fpage><lpage>196</lpage><pub-id pub-id-type="doi">10.1002/cne.902800203</pub-id></element-citation></ref><ref id="bib89"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Soldati</surname><given-names>C</given-names></name><name><surname>Bithell</surname><given-names>A</given-names></name><name><surname>Conforti</surname><given-names>P</given-names></name><name><surname>Cattaneo</surname><given-names>E</given-names></name><name><surname>Buckley</surname><given-names>NJ</given-names></name></person-group><year>2011</year><article-title>Rescue of gene expression by modified REST decoy oligonucleotides in a cellular model of Huntington's disease</article-title><source>Journal of Neurochemistry</source><volume>116</volume><fpage>415</fpage><lpage>425</lpage><pub-id pub-id-type="doi">10.1111/j.1471-4159.2010.07122.x</pub-id></element-citation></ref><ref id="bib90"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Stark</surname><given-names>JM</given-names></name><name><surname>Barmada</surname><given-names>MM</given-names></name><name><surname>Winterberg</surname><given-names>AV</given-names></name><name><surname>Majumber</surname><given-names>N</given-names></name><name><surname>Gibbons</surname><given-names>WJ</given-names><suffix>Jnr</suffix></name><name><surname>Stark</surname><given-names>MA</given-names></name><name><surname>Sartor</surname><given-names>MA</given-names></name><name><surname>Medvedovic</surname><given-names>M</given-names></name><name><surname>Kolls</surname><given-names>J</given-names></name><name><surname>Bein</surname><given-names>K</given-names></name><name><surname>Mailaparambil</surname><given-names>B</given-names></name><name><surname>Krueger</surname><given-names>M</given-names></name><name><surname>Heinzmann</surname><given-names>A</given-names></name><name><surname>Leikauf</surname><given-names>GD</given-names></name><name><surname>Prows</surname><given-names>DR</given-names></name></person-group><year>2010</year><article-title>Genomewide association analysis of respiratory syncytial virus infection in mice</article-title><source>Journal of Virology</source><volume>84</volume><fpage>2257</fpage><lpage>2269</lpage><pub-id pub-id-type="doi">10.1128/JVI.00584-09</pub-id></element-citation></ref><ref id="bib91"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Stoppini</surname><given-names>L</given-names></name><name><surname>Buchs</surname><given-names>PA</given-names></name><name><surname>Muller</surname><given-names>D</given-names></name></person-group><year>1991</year><article-title>A simple method for organotypic cultures of nervous tissue</article-title><source>Journal of Neuroscience Methods</source><volume>37</volume><fpage>173</fpage><lpage>182</lpage><pub-id pub-id-type="doi">10.1016/0165-0270(91)90128-M</pub-id></element-citation></ref><ref id="bib92"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Subramanian</surname><given-names>A</given-names></name><name><surname>Tamayo</surname><given-names>P</given-names></name><name><surname>Mootha</surname><given-names>VK</given-names></name><name><surname>Mukherjee</surname><given-names>S</given-names></name><name><surname>Ebert</surname><given-names>BL</given-names></name><name><surname>Gillette</surname><given-names>MA</given-names></name><name><surname>Paulovich</surname><given-names>A</given-names></name><name><surname>Pomeroy</surname><given-names>SL</given-names></name><name><surname>Golub</surname><given-names>TR</given-names></name><name><surname>Lander</surname><given-names>ES</given-names></name><name><surname>Mesirov</surname><given-names>JP</given-names></name></person-group><year>2005</year><article-title>Gene set enrichment analysis: a knowledge-based approach for interpreting genome-wide expression profiles</article-title><source>Proceedings of the National Academy of Sciences of USA</source><volume>102</volume><fpage>15545</fpage><lpage>15550</lpage><pub-id pub-id-type="doi">10.1073/pnas.0506580102</pub-id></element-citation></ref><ref id="bib93"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Sweatt</surname><given-names>JD</given-names></name></person-group><year>2013</year><article-title>The emerging field of neuroepigenetics</article-title><source>Neuron</source><volume>80</volume><fpage>624</fpage><lpage>632</lpage><pub-id pub-id-type="doi">10.1016/j.neuron.2013.10.023</pub-id></element-citation></ref><ref id="bib94"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Szklarczyk</surname><given-names>A</given-names></name><name><surname>Kaczmarek</surname><given-names>L</given-names></name></person-group><year>1995</year><article-title>Antisense oligodeoxyribonucleotides: stability and distribution after intracerebral injection into rat brain</article-title><source>Journal of Neuroscience Methods</source><volume>60</volume><fpage>181</fpage><lpage>187</lpage><pub-id pub-id-type="doi">10.1016/0165-0270(95)00010-R</pub-id></element-citation></ref><ref id="bib95"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Uchida</surname><given-names>H</given-names></name><name><surname>Ma</surname><given-names>L</given-names></name><name><surname>Ueda</surname><given-names>H</given-names></name></person-group><year>2010</year><article-title>Epigenetic gene silencing underlies C-fiber dysfunctions in neuropathic pain</article-title><source>The Journal of Neuroscience</source><volume>30</volume><fpage>4806</fpage><lpage>4814</lpage><pub-id pub-id-type="doi">10.1523/JNEUROSCI.5541-09.2010</pub-id></element-citation></ref><ref id="bib96"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>van Gassen</surname><given-names>KL</given-names></name><name><surname>de Wit</surname><given-names>M</given-names></name><name><surname>van Kempen</surname><given-names>M</given-names></name><name><surname>van der Hel</surname><given-names>WS</given-names></name><name><surname>van Rijen</surname><given-names>PC</given-names></name><name><surname>Jackson</surname><given-names>AP</given-names></name><name><surname>Lindhout</surname><given-names>D</given-names></name><name><surname>de Graan</surname><given-names>PN</given-names></name></person-group><year>2009</year><article-title>Hippocampal NaBeta3 expression in patients with temporal lobe epilepsy</article-title><source>Epilepsia</source><volume>50</volume><fpage>957</fpage><lpage>962</lpage><pub-id pub-id-type="doi">10.1111/j.1528-1167.2008.02015.x</pub-id></element-citation></ref><ref id="bib97"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>van Loo</surname><given-names>KM</given-names></name><name><surname>Schaub</surname><given-names>C</given-names></name><name><surname>Pernhorst</surname><given-names>K</given-names></name><name><surname>Yaari</surname><given-names>Y</given-names></name><name><surname>Beck</surname><given-names>H</given-names></name><name><surname>Schoch</surname><given-names>S</given-names></name><name><surname>Becker</surname><given-names>AJ</given-names></name></person-group><year>2012</year><article-title>Transcriptional regulation of T-type calcium channel CaV3.2: bi-directionality by early growth response 1 (Egr1) and repressor element 1 (RE-1) protein-silencing transcription factor (REST)</article-title><source>The Journal of Biological Chemistry</source><volume>287</volume><fpage>15489</fpage><lpage>15501</lpage><pub-id pub-id-type="doi">10.1074/jbc.M111.310763</pub-id></element-citation></ref><ref id="bib98"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Vezzani</surname><given-names>A</given-names></name><name><surname>French</surname><given-names>J</given-names></name><name><surname>Bartfai</surname><given-names>T</given-names></name><name><surname>Baram</surname><given-names>TZ</given-names></name></person-group><year>2011</year><article-title>The role of inflammation in epilepsy</article-title><source>Nature Reviews Neurology</source><volume>7</volume><fpage>31</fpage><lpage>40</lpage><pub-id pub-id-type="doi">10.1038/nrneurol.2010.178</pub-id></element-citation></ref><ref id="bib99"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Yaari</surname><given-names>Y</given-names></name><name><surname>Beck</surname><given-names>H</given-names></name></person-group><year>2002</year><article-title>“Epileptic neurons” in temporal lobe epilepsy</article-title><source>Brain Pathology</source><volume>12</volume><fpage>234</fpage><lpage>239</lpage><pub-id pub-id-type="doi">10.1111/j.1750-3639.2002.tb00438.x</pub-id></element-citation></ref><ref id="bib100"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Westbrook</surname><given-names>TF</given-names></name><name><surname>Hu</surname><given-names>G</given-names></name><name><surname>Ang</surname><given-names>XL</given-names></name><name><surname>Mulligan</surname><given-names>P</given-names></name><name><surname>Pavlova</surname><given-names>NN</given-names></name><name><surname>Liang</surname><given-names>A</given-names></name><name><surname>Leng</surname><given-names>Y</given-names></name><name><surname>Maehr</surname><given-names>R</given-names></name><name><surname>Shi</surname><given-names>Y</given-names></name><name><surname>Harper</surname><given-names>JW</given-names></name><name><surname>Elledge</surname><given-names>SJ</given-names></name></person-group><year>2008</year><article-title>SCFbeta-TRCP controls oncogenic transformation and neural differentiation through REST degradation</article-title><source>Nature</source><volume>452</volume><fpage>370</fpage><lpage>374</lpage><pub-id pub-id-type="doi">10.1038/nature06780</pub-id></element-citation></ref><ref id="bib101"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Whyte</surname><given-names>WA</given-names></name><name><surname>Orlando</surname><given-names>DA</given-names></name><name><surname>Hnisz</surname><given-names>D</given-names></name><name><surname>Abraham</surname><given-names>BJ</given-names></name><name><surname>Lin</surname><given-names>CY</given-names></name><name><surname>Kagey</surname><given-names>MH</given-names></name><name><surname>Rahl</surname><given-names>PB</given-names></name><name><surname>Lee</surname><given-names>TI</given-names></name><name><surname>Young</surname><given-names>RA</given-names></name></person-group><year>2013</year><article-title>Master transcription factors and mediator establish super-enhancers at key cell identity genes</article-title><source>Cell</source><volume>153</volume><fpage>307</fpage><lpage>319</lpage><pub-id pub-id-type="doi">10.1016/j.cell.2013.03.035</pub-id></element-citation></ref><ref id="bib102"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Wood</surname><given-names>IC</given-names></name><name><surname>Belyaev</surname><given-names>ND</given-names></name><name><surname>Bruce</surname><given-names>AW</given-names></name><name><surname>Jones</surname><given-names>C</given-names></name><name><surname>Mistry</surname><given-names>M</given-names></name><name><surname>Roopra</surname><given-names>A</given-names></name><name><surname>Buckley</surname><given-names>NJ</given-names></name></person-group><year>2003</year><article-title>Interaction of the repressor element 1-silencing transcription factor (REST) with target genes</article-title><source>Journal of Molecular Biology</source><volume>334</volume><fpage>863</fpage><lpage>874</lpage><pub-id pub-id-type="doi">10.1016/j.jmb.2003.10.017</pub-id></element-citation></ref><ref id="bib103"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Yukhananov</surname><given-names>R</given-names></name><name><surname>Kissin</surname><given-names>I</given-names></name></person-group><year>2008</year><article-title>Persistent changes in spinal cord gene expression after recovery from inflammatory hyperalgesia: a preliminary study on pain memory</article-title><source>BMC Neuroscience</source><volume>9</volume><fpage>32</fpage><pub-id pub-id-type="doi">10.1186/1471-2202-9-32</pub-id></element-citation></ref><ref id="bib104"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Zeng</surname><given-names>LH</given-names></name><name><surname>Rensing</surname><given-names>NR</given-names></name><name><surname>Wong</surname><given-names>M</given-names></name></person-group><year>2009</year><article-title>The mammalian target of rapamycin signaling pathway mediates epileptogenesis in a model of temporal lobe epilepsy</article-title><source>The Journal of Neuroscience</source><volume>29</volume><fpage>6964</fpage><lpage>6972</lpage><pub-id pub-id-type="doi">10.1523/JNEUROSCI.0066-09.2009</pub-id></element-citation></ref><ref id="bib105"><element-citation publication-type="journal"><person-group person-group-type="author"><name><surname>Zuccato</surname><given-names>C</given-names></name><name><surname>Belyaev</surname><given-names>N</given-names></name><name><surname>Conforti</surname><given-names>P</given-names></name><name><surname>Ooi</surname><given-names>L</given-names></name><name><surname>Tartari</surname><given-names>M</given-names></name><name><surname>Papadimou</surname><given-names>E</given-names></name><name><surname>MacDonald</surname><given-names>M</given-names></name><name><surname>Fossale</surname><given-names>E</given-names></name><name><surname>Zeitlin</surname><given-names>S</given-names></name><name><surname>Buckley</surname><given-names>N</given-names></name><name><surname>Cattaneo</surname><given-names>E</given-names></name></person-group><year>2007</year><article-title>Widespread disruption of repressor element-1 silencing transcription factor/neuron restrictive silencer factor occupancy at its target genes in Huntington's disease</article-title><source>The Journal of Neuroscience</source><volume>27</volume><fpage>6972</fpage><lpage>6983</lpage><pub-id pub-id-type="doi">10.1523/JNEUROSCI.4278-06.2007</pub-id></element-citation></ref></ref-list></back></article> |