Gene Expression Underlying Changes in Network Excitability

Katarzyna Lukasiuk

Raymond Dingledine

Daniel H. Lowenstein

Asla Pitkänen

Introduction

Remarkable progress in the fields of molecular biology and molecular genetics has added a new dimension to epilepsy research. Just few years ago, studies on the basic mechanisms of epilepsy were limited primarily to anatomic, electrophysiologic, and pharmacologic analyses. However, molecular approaches have provided a more direct and specific means for studying biochemical events in the brain, leading to the recognition that seizures and other forms of injury can induce complex changes in gene expression. These changes are likely to regulate the molecular architecture of neuronal cells and circuits that contribute to longer-term process of epileptogenesis as well as seizure initiation, spread, and cessation. The application of molecular biology to investigate epilepsy can be divided into two broad categories: (a) studies on the genetic or inherited basis of epilepsies (which will not be discussed here) and (b) studies on the role of alterations in expression levels of specific genes or ensembles of genes during epileptogenesis and in epilepsy. This chapter focuses mainly on recent results of global analyses of gene expression in humans and in experimental models of epileptogenesis and epilepsy that provide new understanding of cellular and metabolic processes occurring in the brain affected by the disease.

Basic Principles of Gene Expression

This section provides a brief review of the main steps involved in the conversion of genetic information encoded by DNA into proteins (for a detailed review, see Drlica⁴⁴).

DNA Replication

The genetic code for all cellular proteins is contained in DNA. In its native state, DNA exists as a double helix of deoxyribonucleic acids bound together by phosphate and hydrogen bonds. The hydrogen bonding between DNA strands ensures that the base adenosine (A) will pair with thymidine (T), whereas guanine (G) pairs with cytosine (C). Replication (or duplication) of DNA requires unraveling and separation of the complementary strands of DNA, attachment of DNA-binding proteins, annealing of small primer fragments, and creation of new complementary strands via DNA polymerase enzymes. Because each strand is replicated once, the process is termed semiconservative. DNA replication occurs prior to cell division or mitosis, but not in nondividing cells. The combination of highly specialized enzymes and the ability to “proofread” during the synthesis of a new strand of DNA makes this a remarkably reliable process. Even nondividing cells must constantly maintain the integrity of the DNA by using a host of scanning, editing, and repair enzymes. The double-stranded nature of DNA helps to ensure that a mutation in one strand will be rapidly detected and repaired by the cell.

Transcription

During transcription, the enzyme RNA polymerase recognizes and attaches to a specific region at the starting point of a gene. RNA polymerase synthesizes a single-stranded RNA molecule that is complementary to the DNA template, with the exception that ribonucleic acids are substituted for deoxyribonucleic acids and the base uracil (U) is substituted for thymidine in the coding scheme. Transcription continues until the enzyme reaches a termination sequence in the gene. Once the full-length gene is transcribed, segments that do not code for a specific portion of a protein (termed introns) are excised, and only the exons, which contain specific information for the coding of proteins, remain. In addition, a long tail of adenosine bases is attached to the RNA, thereby identifying it as “messenger” RNA (mRNA). Messenger RNA is then used to translate the genetic code from ribonucleic acids into amino acids and proteins.

The mechanisms controlling the recognition and attachment of RNA polymerase to the starting point of a gene are critical determinants of the pattern of gene expression in a cell. Specific DNA-binding proteins known as transcription factors can attach to a regulatory element upstream from the starting point of the gene. Depending on the type of transcription factor, this may either promote or inhibit binding of the RNA polymerase and subsequent transcription. Similarly, other upstream and downstream regions known as enhancer or repressor sites can also influence transcriptional activity. These transcription factors are critical links in the process of stimulus-induced regulation of gene expression.

Translation

Transcribed mRNA is translated into protein by an interpretive process involving ribosomes, transfer RNA (tRNA), and a variety of other proteins that facilitate the process, such as initiation factors and elongation factors. Protein synthesis begins with the attachment of the mRNA to a ribosome near the “start codon.” A codon is a triplet of RNA bases that codes for either a specific amino acid or a start or stop to the translation procedure. A tRNA, which carries a particular amino acid, then joins the complex based on the complementary nucleotide sequence of the mRNA (codon) and tRNA (anticodon). The polypeptide chain is created by sequentially bringing pairs of
tRNAs together on the ribosome using the mRNA as a template. A peptidyl transferase reaction forms a new peptide bond between the two amino acids on the tRNAs. This process continues until the end of the mRNA encoding sequence is reached, and the newly formed polypeptide is released in the cell with the aid of specific release factors.

Aspects of Gene Expression Unique to the Central Nervous System

When the techniques of modern molecular biology are applied to the study of epilepsy and neural injury, certain special aspects of the central nervous system (CNS) must be taken into consideration. Although these unique characteristics do not necessarily limit the use of molecular methods in neurobiology, they do provide insight into the complexities of gene regulation that are unique to the signal processing and cellular repair mechanisms of the CNS. This section discusses several of these aspects.

One of the most notable aspects of neurons is that they are postmitotic.⁹² Most mature neurons, therefore, are required to maintain their functional integrity for, ideally, the lifetime of the organism. To do this, neurons use many “housekeeping” functions. These functions include replacing and repairing cytoskeletal structures, replenishing membrane and membrane-associated proteins, synthesizing and packaging neurotransmitters, and maintaining integrity of synaptic contacts. If cell injury occurs, genes are activated in an attempt to restore function and to protect from further injury.¹⁷^,²⁷^,⁵⁴^,¹⁰⁹^,¹¹¹^,¹²⁴^,¹⁴⁵ If cell death occurs, in certain systems new synaptic contacts are made by the surviving presynaptic neurons, presumably in response to the lost functional connectivity of the dead neuron.¹¹⁰

In addition to being postmitotic, neurons have a considerable amount of phenotypic diversity and plasticity. Phenotypic diversity refers to the unique characteristics of various types of neurons, such as differences in neuron size and shape, classes of neurotransmitters and peptides released, firing conditions and frequencies, and responses to stress and injury.¹⁰⁸^,¹⁷⁴ Phenotypic plasticity refers to the ability of an individual neuron to alter its function. There are numerous levels of control of phenotypic plasticity. Neurons can transiently modify the quantity of neurotransmitters they release in response to both external stimuli and intracellular signaling mechanisms, as has been suggested in certain forms of long-term potentiation.¹¹^,¹¹⁶ Phenotypic plasticity is a dynamic process that is highly dependent on the molecular aspects of cellular physiology. Neurons can modulate the relative amounts and properties of proteins they express through changes in gene expression, editing of mRNA transcripts, and a variety of posttranslational modifications. For example, in response to repeated stimulation, neurons can synthesize new ion channels and membrane receptors that may contribute to long-lasting changes in membrane excitability.⁴²^,⁵⁷^,⁹⁸^,¹¹⁸^,¹²⁶ Similarly, following stimulation or seizures, neurons may modify the response characteristics of specific voltage-dependent ion channels and membrane receptors and alter expression of mRNAs coding for various synapse-modifying proteins.⁴¹^,¹¹⁹ Phenotypic plasticity is an ongoing and constant component of neuronal and glial behavior and is an essential ingredient in the ability of the CNS to interact with the environment.

An especially interesting example of phenotypic diversity in neurons comes from the observation that protein synthesis may be highly localized in specific regions of the cell that are distant from the nucleus and regulated independently from translation occurring in the cell body. In most cells, translation of mRNA into protein occurs within the cell body, and newly synthesized proteins are then transported to their appropriate site based on intracellular signaling mechanisms. Most neurons are polarized cells with three fairly distinct compartments: (a) the dendritic processes, (b) a central regulatory soma, and (c) axonal projections ending with presynaptic terminals. Each of these regions has specific functions and interactions with its microenvironment.³³ mRNA can be differentially distributed and translated in dendritic processes.¹³^,³³ Furthermore, ribosomes necessary for carrying out protein translation have also been located in dendrites. This localized distribution of mRNA and ribosomes suggests that dendrites have the capacity to support highly localized and site-specific protein synthesis. This additional level of phenotypic plasticity may prove to be an important element in the synaptic modifications implicated in activity-dependent learning. It is interesting that there is no evidence that a similar phenomenon of localized protein synthesis occurs in axons.³³ This implies that the neuron may rely on intracellular transport mechanisms, such as axoplasmic flow and microtubule systems, to deliver newly synthesized protein to the axon terminal.

Epilepsy and Alterations in Gene Expression

An “Intelligent Guess” = Candidate Gene Approach

Changes in gene expression leading to network reorganization and neuronal hyperexcitability are crucial to understanding epilepsy and have been studied with traditional methods. Several candidate genes encoding proteins having a role in brain excitability have been tested. Obviously, much attention has been given to genes coding for proteins directly influencing electrical properties of neurons, such as receptors and channels. The other group of genes has been those coding for proteins involved in remodeling of neuronal networks, such as growth factors and their receptors, synaptic proteins, or other structural proteins. However, such studies have concentrated mostly on seizure-induced alteration and only within hours after seizures. These data were reviewed recently⁴⁹^,¹⁷⁵ and are only briefly mentioned here. Less information is available on alteration in expression of selected genes during late epileptogenesis and in epileptic tissue. In this chapter, we briefly review only data on changes in the expression of genes that are hypothesized to influence brain excitability directly or indirectly. Data from both human tissue and experimental models of epileptogenesis and epilepsy are included, but data on immediate effect of seizures or status epilepticus, reflecting influence of increased neuronal activity on gene expression, are omitted.

Traditional Methods Most Commonly Used for Studies of Gene Expression Levels

The reverse transcription polymerase chain reaction (RT-PCR) is a powerful technique for making many copies of DNA from minute quantities of RNA targets used as starting material. For gene expression analysis, the first step is synthesis of complementary DNA (cDNA) on the basis of mRNA extracted from the studied tissue using the enzyme reverse transcriptase. The PCR is then used to amplify cDNA representing the gene of interest. PCR uses an enzymatic reaction to repeatedly copy the original DNA fragment present in the reaction. The PCR reaction mixture includes four basic components: (a) the target DNA, (b) oligonucleotide “primers,” which are designed to hybridize the sequence of DNA to be amplified, (c) an excess of individual nucleotides (deoxyadenosine 5′-triphosphate [dATP], deoxycytidine 5′-triphosphate
[dCTP], thymidine 5′-triphosphate [dTTP], deoxyguanosine 5′-triphosphate [dGTP]), and (d) a DNA polymerase that is extremely stable at a high temperatures. With the use of an automated heating block, this mixture is cycled through a series of temperature changes that are designed to allow the polymerase to repeatedly duplicate the target DNA. Each cycle begins with a high temperature (e.g., 94°C) to separate the strands of target DNA (“strand separation phase”). The temperature is then dropped (i.e., 42°C–60°C, depending on reaction conditions) to allow the oligonucleotides to anneal (“primer annealing phase”). Then the temperature is increased to 72°C, which is the optimal temperature for Taq polymerase activity, so that the DNA replication occurs using the oligonucleotides as the primers (“primer extension phase”). Repeated cycling through these steps results in amplification of the DNA in a highly efficient fashion.

A Northern blot is a method for detecting the presence of a particular mRNA transcript in a sample. RNA is isolated from tissue or cells using standard extraction procedures and separated by size via gel electrophoresis. The size-separated RNA is transferred to a membrane, and the membrane is then exposed to a radiolabeled probe. If the mRNA of interest is in the sample, the probe will specifically hybridize to the mRNA and produce a signal at the predicted size on the autoradiogram.

In situ hybridization provides a means of determining the pattern of mRNA expression at the cellular level. Whereas Northern analysis uses mRNA derived from homogenized tissue, in situ hybridization uses fixed tissue sections so that the cellular anatomy is preserved. Otherwise, the technique uses the same concepts described for Northern analysis. The tissue sections are mounted on slides and prepared in a way that allows the labeled probe to have access to the cellular compartments and to hybridize with the mRNA. Radioactively labeled probe is then detected with X-ray film. To obtain cellular detail, the slides are coated with photographic emulsion, such that the radiolabeled probe will expose the emulsion directly overlying the cells. The slides are then “developed” in the same manner as film, and silver grains in the emulsion seen as black specks reflect the presence of this signal, signifying the presence of the specific mRNA transcript. In case of fluorescently labeled probe, the results of hybridization are analyzed using standard fluorescent microscopy techniques.

Immediate-Early Genes

In recent decades, a class of genes termed immediate-early genes (IEGs) has been identified that appears to function as a vital link between acute activity-dependent events and long-term changes in gene expression.¹⁵^,³¹^,⁶³^,⁸⁸^,⁸⁹^,¹⁰²^,¹⁵⁰^,¹⁵⁴^,¹⁷⁵ By definition, induction of expression of IEG mRNAs is rapid and does not depend on protein synthesis. Protein products of IEGs often serve as transcription factors regulating the transcription of other genes (e.g., c-fos, c-jun, c-myc, jun-B, krox-20, krox-24, fra-1, zif/268, etc.).

Expression of IEGs in neurons has been shown to be induced by a wide range of stimuli, including membrane depolarization due to both physiologic¹⁴^,¹⁵^,⁵⁵^,⁶⁷^,⁷⁰^,⁸³^,⁸⁴^,⁸⁵^,⁸⁶ and nonphysiologic stimulation,³⁷^,⁴¹^,⁴²^,⁸²^,⁸⁸^,¹⁰⁵^,¹¹³^,¹²⁷^,¹³³^,¹⁴⁷^,¹⁵²^,¹⁶¹ mechanical trauma,⁴³ and ischemia.¹⁴⁰ Studies using seizures induced by kindling, electroconvulsive seizures, electrolytic lesions, and chemoconvulsants have also shown that IEGs are markedly induced in specific CNS regions such as the hippocampus and cerebral cortex. Much effort has been invested in understanding molecular pathways leading to induction of IEGs expression following seizures, and the results of these studies are reviewed in detail elsewhere.¹³⁹^,¹⁷⁵

For the purposes of this review, an important question is, what are the target genes for transcription factors (IEGs and other) participating in the development of epilepsy? Several target genes that can influence neuronal excitability and their relation to transcription factors governing their expression have been described. For example, expression of the metabotropic γ-aminobutyric acid receptor GABA_BR1a and GABA_BR1b isoforms in hippocampal neurons is mediated by the cAMP response element–binding protein (CREB), which binds to unique cAMP response elements in the alternative promoter regions. CREB is then critical for transcriptional mechanisms that control GABA_BR1 subunit levels in vivo.¹⁵³ In addition, the activating transcription factor-4 (ATF4) differentially regulates GABA_BR1a and GABA_BR1b promoter activity.¹⁵³

The DREAM transcription factor that is expressed widely in the nervous system can be induced by seizures.²⁹^,¹²³ Its target genes include genes related to neuronal plasticity such as c-fos and preprodynorphin.²⁸^,²⁹

The AP-1 transcription factor (which is composed of proteins belonging to the Fos and Jun families) has been shown to regulate expression of nerve growth factor (NGF) after hilus lesion–induced seizures⁴⁸ and proenkephalin and prodynorphin after kainic acid–induced seizures.⁹⁶^,¹⁴³

There are also IEGs upregulated during epileptogenesis that lack transcription factor functions and can have lasting effect on neuronal excitability. One of these proteins is homer-1, which codes for a scaffold protein anchoring metabotropic glutamate receptors to the cytoskeleton and regulates pyramidal neuron excitability.²⁴^,¹⁴¹^,¹⁴⁸

Neurotransmitter Receptors and Ion Channels

Alterations in the expression of mRNAs encoding neurotransmitter receptors and ion channels are likely candidates for affecting network excitability. Table 1 gives a summary of such changes in experimental and human epilepsy.

In the mature nervous system, GABA_A receptors function as ligand-gated chloride channels that confer fast inhibitory synaptic transmission. Several studies have demonstrated abnormal GABA_A receptor function in epileptic tissue.²²^,⁶⁴^,⁸⁰ Because receptor properties depend on subunit composition,¹¹³^,¹⁶⁹ altered expression of GABA_A-receptor subunits can explain functional abnormalities. In fact, changes in mRNA expression for selected GABA_A-receptor subunits have been reported in epilepsy models induced by kainic acid¹⁶² and pilocarpine²³^,¹⁴⁶ and electrically induced status epilepticus (SE),¹⁰³^,¹³⁴ as well as following hippocampal kindling.¹³⁴

Metabotropic GABA_B-receptor expression has been studied in epileptic animals. Changes in the expression of mRNA encoding GABA_B receptors occur in the hippocampus of patients with temporal lobe epilepsy (TLE)⁶⁰ as well as in kainic acid–induced epilepsy,⁵⁹ hippocampal kindling,¹³⁴ and epilepsy following electrically-induced SE.¹³⁴

Glutamate, the major excitatory neurotransmitter in the brain, acts on ionotropic and metabotropic receptors. Sixteen genes encoding for ionotropic receptors that belong to three functional families (α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid [AMPA], kainate, and N-methyl-D-aspartic acid [NMDA] receptors) have been identified.³⁸ Alterations in the expression of AMPA receptor subunits have been demonstrated in human epilepsy⁶¹^,¹²¹ as well as in electrically-⁹⁰^,¹⁷³ and pentylenetetrazole-induced kindling.⁴⁶ Changes in kainate receptor subunits occur in human epileptic tissue⁶⁸ and the kainate model of epilepsy¹⁶³ as well as in amygdala and hippocampal kindling.⁷⁵^,⁹¹ Changes in the expression of NMDA subunits were observed in patients with TLE,⁷^,¹²¹^,¹²²^,¹³⁰ in rats kindled with amygdala⁹⁴ or hippocampal stimulation,¹⁰⁰^,¹⁰¹ and following intra-amygdala injection of kainic acid.¹⁴⁴

Table 1 Summary of studies using conventional methods to demonstrate alterations in the expression of genes encoding neurotransmitter receptors, transporters, and ion channels

Authors	Model/disorder	Structure	Gene
*Ionotropic GABA receptors*
Rice et al. (1996)¹⁴⁶	Pilocarpine-induced SE	CA1–CA3	↓α2, α5 at 1–2 mo
Tsunashima et al. (1997)¹⁶²	Kainic acid–induced SE	Hippocampus	↓α2, α3, α5, β1, β3, γ2, δ; ↑ α1, α4, β2 at 6–24 hr ↓α5, δ; ↑α1, α3, β1, β2 at 7–30 d
Brooks-Kayal et al (1998)²⁶	Pilocaprine-induced SE	Dentate gyrus	↓α1, β1; ↑ α4, β3 chronically
Lauren et al. (2003)¹⁰³	Amygdala stimulation model of TLE	CA3 CA1	↓α2; ↑β3 in epileptic rats ↓α4; ↑β3 in epileptic rats
Nishimura et al. (2005)¹³⁴	Hippocampal kindling Electrically induced SE	Dentate gyrus CA3 Dentate gyrus	↓δ; ↑α4, β1–3, γ2 at 7–30 d ↓α2 and δ1–3 ↓α5 at 24 hr ↓δ, α5; ↑α4, β1–3, γ2 at 7–30 d
*Metabotropic GABA receptors*
Furtinger et al. (2003)⁵⁹	Kainic acid–induced SE	Hippocampus	↓ GABA_B-1, GABA_B-2 acutely
Furtinger et al. (2003)⁶⁰	Human TLE	Dentate gyrus	↑ GABA_B-1 GABA_B-2
Nishimura et al. (2005)¹³⁴	Hippocampal kinding and electrically induced SE	Dentate gyrus	↑ GABA_B-2 acutely and chronically
*Ionotropic glutamate receptors*
Wong et al. (1993)¹⁷³	Seizures		↓↑GluR1
Garcia-Ladona et al. (1994)⁶¹	Human TLE	Hippocampus	↑ GluR1
Kamphuis et al. (1994)⁹⁰	Kindling	Hippocampus	↑ GluR1 flip up to 4 wk
Kamphuis et al. (1995)⁹¹	Hippocampal kindling	Dentate gyrus	↓ GluR7 at 28 d
Bayer et al. (1995)⁷	Human TLE	Dentate gyrus	↑ NR2
Kraus et al. (1994, 1996)¹⁰⁰^,¹⁰¹	Hippocampal kindling	Hippocampus	↓ NR1 splice variant at 28 d
Grigorenko et al. (1997)⁶⁸	Human TLE	Hippocampus	↑ GluR6
Mathern et al. (1997)¹²¹	Human TLE	Dentate gyrus	↑ NR2
Rafiki et al. (1998)¹⁴⁴	Intra-amygdalar kainic injection	Hippocampus	↑ NR1-2a, NR-2b, NR1-3a, NR1-3b, NR1-3b at 21 d–4 mo
Mathern et al. (1997)¹²¹	Human TLE	Hippocampus	↑ GluR1
Mathern et al. (1999)¹²²	Human TLE	Dentate gyrus CA2/3	↑ NR2A, NR2B ↑ NR2B, NR1; ↓ NR2A
Kikuchi et al. (2000)⁹³	Amygdala kindling	Frontal and temporal cortex Piriform cortex	↑ NR1 at 4 wk ↓ NR1 at 4 wk
Ekonomou et al. (2001)⁴⁶	Pentylenetetrazole-induced kindling	CA1 and dentate gyrus cortex	↓ GluR2, GluR3 at 1 wk ↑ GluR2 at 1 mo
Neder et al. (2002)¹³⁰	Human TLE	Sclerotic hippocampus Nonsclerotic hippocampus	↓ NR1 ↑ NR1
Hikiji et al. (1993)⁷⁵	Amygdala kindling	CA3	↑ KA1 at 28 d
Ullal et al. (2005)¹⁶³	Kainic acid–induced SE	Hippocampus	↓ GluR7 at 3 mo; ↑ GluR5 at 9 mo
*Metabotropic glutamate receptors*
Akbar et al (1996)¹	Amygdala kindling	Hippocampus	↓ mGluR5; ↑ mGluR1 at 24 hr
Blumcke et al. (2000)¹⁹	Kindling Kainic acid–induced SE	Hippocampus	↑ mGluR1
*Glutamate transporters*
Akbar et al. (1998)²	Genetically epilepsy-prone rats	Cortex, striatum, and hippocampi	↓ GLT-1
Ingram et al. (2000)⁷⁸	GAERS rats	Thalamus Cortex	↑ GLT-1 ↑ GLAST
Doi et al. (2000)⁴⁰	Fe(3+)-induced epileptogenesis		↑ EAAC-1 up to 30 d ↓ GLAST in epileptic animals
Ingram et al. (2001)⁷⁹	EL mice	CA3, parietal cortex	↓ GLT-1
Crino et al. (2002)³⁵	Pilocarpine-induced SE	Dentate gyrus	↑ EAAC1 in epileptic animals
Proper et al. (2002)¹⁴²	Human TLE	Hippocampus	↑ GLT-1, EAAC1
*Voltage-gated sodium channels*
Aronica et al. (2001)⁴	Hippocampal stimulation	Hippocampus	↑ NaCh II and III up to 3 mo
Whitaker et al. (2001)¹⁷²	Human TLE	CA3 CA4	↓ NaCh II ↑ NaCh III
Ellerkmann et al. (2003)⁴⁷	Pilocarpine-induced SE	Dentate gyrus	↓ Nav 1.2, Nav 1.6 up to 30 d
Klein et al. (2004)⁹⁵	WAG/Rij absence epileptic rats	Cortex	↑ Nav 1.1 and Nav 1.6
*Calcium channels*
Hendriksen et al. (1997)⁷²	Hippocampal kindling	Hippocampus CA3	↑ VDCC-α1A, -α1D, -α1E; ↓ -α1E during initial kindling ↑ VDCC-α1B in fully kindled animals
Vigues et al. (1999)¹⁶⁸	Kainic acid–induced SE	Hippocampus	↑ Q-type α1A up to 7 d
Talley et al. (2000)¹⁵⁸	GAERS rats	Thalamus	↑ T-type α1H, α1G
Blalock et al. (2001)¹⁸	Kindling	CA1	↓ L-type α 1D
*Hyperpolarization-activated cyclic nucleotide–gated cation channels (HCNs)*
Brewster et al. (2002)²⁵	Rat febrile seizures	CA1 CA3	↓ HCN1; ↑ HCN2 at 1 wk ↑ HCN2 at 1 wk
Bender et al. (2003)¹²	Human TLE	Dentate gyrus	↑ HCN1
	Pilocarpine-induced SE
CA1, CA1 subfield of the hippocampus proper; CA3, CA3 subfield of the hippocampus proper; CA4, CA4 subfield of the hippocampus proper; EAAC, excitatory amino acid carrier; EAAT, excitatory amino acid transporter; GABAR, γ-aminobutyric acid receptor; GAERS, genetic absence epilepsy rats from Strasbourg; GLAST, glutamate/aspartate transporter; GluR, glutamate receptor; GLT, glutamate transporter; HCN, hyperpolarization-activated cyclic nucleotide–gated cation channel; KA1, kainic acid receptor subunit 1; mGluR, metabotropic glutamate receptor; NR, N-methyl-D-aspartate receptor subunit; SE, status epilepticus; TLE, temporal lobe epilepsy; VDCC, voltage-dependent calcium channel.