The prolonged seizures of generalized status epilepticus (SE) in man and rodents trigger a series of molecular and cellular events that eventually culminate in the appearance of spontaneous seizures, i.e., epilepsy. These events include selective neuronal degeneration, inflammatory reactions involving reactive microglia and astroglia, selective axonal sprouting with new synapse formation, neurogenesis, and a myriad of changes in synaptic efficacy in the hippocampus. The breadth of the phenotypic consequences of SE, and their elaboration over days and weeks, raises the possibility that one or more broad regulators of gene expression could mediate some or even many of these consequences.
As described above and schematized in , REST is a transcriptional repressor that recruits histone deacetylases, demethylases and methyltransferases to cause epigenetic remodeling of chromatin architecture around the REST target genes (Ding, et al. 2008
; Garriga-Canut et al., 2006
; Huang et al., 1999
; Mulligan et al., 2008
; Roopra et al., 2004
; Roopra et al., 2000
; Tahiliani et al., 2007
). The most prominent modifications occur on H3K4 and H3K9 ()(Zheng, et al. 2009
REST is strongly induced in hippocampal pyramidal and dentate granule neurons after SE induced by kainate (Palm, et al. 1998
) or pilocarpine (). More than 1300 genes, or approximately 5% of the protein-encoding genome, are confirmed REST targets (Bruce, et al. 2004
, Johnson, et al. 2006
) including many genes known to be involved in neuronal excitability (Roopra, et al. 2001
), making REST an excellent candidate transcription factor to mediate seizure-induced widespread changes in gene expression. REST target genes are highly over-represented among the differentially expressed genes after SE (Lelutiu et al., in preparation), consistent with a wide scope of REST’s influence. Cooperation among the multiple histone modifying enzymes recruited by REST suggests that the ultimate effect on transcription can be context- and cell-dependent. For example, although REST is best known as a transcriptional repressor, in some cases there is credible evidence that REST or its truncated splice variant, REST4, might act as an activator of gene expression (Abramovitz, et al. 2008
, Kuwabara, et al. 2004
). The picture is thus emerging of a very dynamic set of REST protein complexes working together to determine cell-specific and activity-dependent gene expression profiles that, in turn, drive incredibly diverse biological processes. REST recruits at least three classes of epigenetic modifier enzymes (), some of which might have a net effect of promoting and others opposing epileptogenesis. In this respect, whether the net effect of REST-mediated changes in gene expression promotes or opposes epileptogenesis is controversial at this point.
Rapid induction of REST protein in dentate granule neurons 5 hr after pilocarpine-induced SE. Hsieh and Dingledine, unpublished.
Some ion channel genes are both epigenetically regulated and repressed after seizures including Gria2, which encodes GluA2 (Huang, Myers and Dingledine 1999
, Myers, et al. 1998
), and HCN1 (McClelland, et al. 2011
). HCN1 (hyperpolarization-activated cyclic nucleotide-regulated cation channel) channels typically function to dampen excitability in cortical neurons. Mice lacking HCN1 channels in the forebrain exhibit more seizures and higher mortality in both kindling and pilocarpine models (Santoro, et al. 2010
). This finding suggests that HCN1 downregulation following severe seizures may contribute to disease progression. McClelland et al. (2012) showed that REST binding to the RE1 element in the HCN1 promoter in the hippocampus was augmented two days after kainate-induced status epilepticus, and that intraventricular administration of oligodeoxynucleotides targeted to the HCN1-RE1 both disrupted REST binding to the HCN1 promoter and prevented downregulation of HCN1 protein. Importantly, rescue of HCN1 protein levels was accompanied by full restoration of the Ih
current amplitude in CA1 pyramidal cell dendrites and by fewer spontaneous seizures in the chronic epilepsy phase. The experimental design did not rule out roles for REST target genes other than HCN1, but this study does strongly reinforce the notion that REST induction after status epilepticus contributes to the development of epilepsy.
Recently, more direct evidence for the roles of neuronal REST expression in epilepsy has been achieved following the creation of mice in which the REST gene had been conditionally deleted in glutamatergic forebrain neurons (Hu, et al. 2011a
) or all neurons (Liu, et al. 2012
). In the initial studies a conditional REST knockout (nREST cKO-CAMKII) was created by crossing a mouse line bearing a floxed REST gene with a mouse line expressing cre recombinase under the CAMKII promoter. In the kindling model, nREST cKO-CAMKII mice exhibited dramatically accelerated seizure progression and prolonged after discharge duration compared with control mice (Hu, et al., 2011a
). This finding suggests that in the kindling model REST may function to oppose epileptogenesis. Quite a different result was obtained in the pentylenetetrazol (PTZ) model of acute seizures, using nREST cKO-NSE mice created using neuron-specific enolase cre, which ablates the REST gene from most if not all neurons. Although the initial clonic convulsions caused by PTZ were not different between nREST cKO-NSE and control mice, tonic convulsions and death required a higher PTZ dose in nREST cKO-NSE mice (Liu, et al. 2012
). These findings, considered alone, would suggest that REST might contribute to seizure initiation or generalization. Taken together with the findings from the nREST cKO-CAMKII mice, however, it is unclear whether the opposing conclusions are due to the animal model (kindling vs PTZ), the subset of cells from which the REST gene had been ablated, or some unknown, laboratory-specific factor.
A potential role for REST in the ketogenic diet therapy for epilepsy has been reported by two groups. Garriga-Canut et al. (2006)
reasoned that, because the ketogenic diet is high fat plus low carbohydrate, glycolytic inhibition itself by 2-deoxyglucose (2-DG) might replicate the anticonvulsant effect of the diet. They tested this hypothesis with the kindling model and found that systemic administration of 2-DG could retard the progression of seizure intensity during kindling. Moreover, they found that 2-DG treatment caused decreased expression of BDNF and TrkB, both REST target genes. These results are consistent with the observation that knockout of TrkB or BDNF showed reduced or zero epileptogenesis in the kindling model of Temporal Lobe Epilepsy (He, et al. 2004
). Reduction in BDNF and TrkB was accompanied by deacetylation and methylation of lysine-9 on histone-3 associated with the BDNF RE1 element that is the binding site for REST. The transcriptional corepressor CtBP (), which is allosterically regulated by NADH (see above), was shown to mediate the formation of the repressive chromatin environment in 2-DG treated animals. Following this study, the antiepileptic effect of 2-DG in the kindling model was found to be abolished in nREST-cKO-CAMKII mice (Hu, et al. 2011b
), indicating that REST expression in forebrain glutamatergic neurons could be required for the antiepileptic effect of 2-DG. Interestingly, the antiepileptic effect of the ketogenic diet itself was maintained in nREST-cKO-CAMKII mice, pointing to a benefit of the high fat diet that is independent of glycolytic inhibition.
In a related study, global ischemia elevated REST RNA and protein levels in CA1 pyramidal neurons and in dentate granule neurons of rats (Calderone, et al. 2003
). Notably, acute knockdown of REST expression prevented downregulation of the REST target gene, GluA2, which correlated with an attenuation of ischemia-induced cell death of CA1 neurons. In a follow-up study, Noh et al. (2012)
demonstrated that local depletion of REST by intra-hippocampal injection of lentiviruses expressing either REST RNAi or a dominant negative REST construct could virtually abolish the appearance of FluoroJade-stained, injured neurons in hippocampal CA1 measured 6 days after global ischemia (Noh, et al. 2012
). Systemic administration of the broad-spectrum HDAC inhibitor, trichostatin A, replicated the effects of REST knockdown. These studies indicate that REST expression and HDAC activity seem critical for ischemia-induced neurodegeneration. It will be important to extend these studies to epilepsy models.