Severe epilepsy resulting from genetic interaction between Scn2a and Kcnq2

A mutation in the voltage-gated sodium-channel Scn2a results in moderate epilepsy in transgenic Scn2a(Q54) mice maintained on a C57BL/6J strain background. The onset of progressive epilepsy begins in adults with short-duration partial seizures that originate in the hippocampus. The underlying abnormality is an increase in persistent sodium current in hippocampal neurons. The voltage-gated potassium channel Kcnq2 is responsible for generating M current (I(KM)) that is thought to control excitability and limit repetitive firing of hippocampal neurons. To determine whether impaired M current would exacerbate the seizure phenotype of Scn2a(Q54) mice, we carried out genetic crosses with two mutant alleles of Kcnq2. Szt1 mice carry a spontaneous deletion that removes the C-terminal domain of Kcnq2. A novel Kcnq2 missense mutation V182M was identified by screening the offspring of ENU-treated males for reduced threshold to electrically evoked minimal clonic seizures. Double mutant mice carrying the Scn2a(Q54) transgene together with either of the Kcnq2 mutations exhibited severe epilepsy with early onset, generalized tonic-clonic seizures and juvenile lethality by 3 weeks of age. This dramatic exacerbation of the sodium-channel mutant phenotype indicates that M current plays a critical role in preventing seizure initiation and spreading in this animal model. The genetic interaction between Scn2a and Kcnq2 demonstrates that combinations of mild alleles of monogenic epilepsy genes can result in severe disease and provides a model for complex inheritance of human epilepsy. The data suggest that interaction between these genes might contribute to the variable expressivity observed in human families with sodium-channel mutations. In a screen of 23 SMEI patients with missense mutations of SCN1A, no second-site mutations in KCNQ2 were identified.     

Na+ channel expression and neuronal function in the Na+/H+ exchanger 1 null mutant mouse

Mice lacking Na(+)/H(+) exchanger 1 (NHE1) suffer from recurrent seizures and die early postnatally. Although the mechanisms for seizures are not well established, our previous electrophysiological work has shown that neuronal excitability and Na(+) current density are increased in hippocampal CA1 neurons of these mutant mice. However, it is unknown whether this increased density is related to altered expression or functional regulation of Na(+) channels. In this work, we asked three questions: is the increased excitability limited to CA1 neurons, is the increased Na(+) current density related to an increased Na(+) channel expression, and, if so, which Na(+) channel subtype(s) is upregulated? Using neurophysiological, autoradiographic, and immunoblotting techniques, we showed that both CA1 and cortical neurons have an increase in membrane excitability and Na(+) current density; Na(+) channel density is selectively upregulated in the hippocampus and cortex (P < 0.05); and Na(+) channel subtype I is significantly increased in the hippocampus and Na(+) channel subtype II is increased in the cortex. Our results demonstrate that mice lacking NHE1 upregulate their Na(+) channel expression in the hippocampal and cortical regions selectively; this leads to an increase in Na(+) current density and membrane excitability. We speculate that neuronal overexcitability due to Na(+) channel upregulation in the hippocampus and cortex forms the basis of epileptic seizures in NHE1 mutant mice.     

Hippocampal seizure resistance and reduced neuronal excitotoxicity in mice lacking the Cav2.3 E/R-type voltage-gated calcium channel

Voltage-gated calcium channels are key components in the etiology and pathogenesis of epilepsies. Former studies mainly focused on P/Q-type Ca(v)2.1 and T-type Ca(v)3.2 Ca(2+) channels involved in absence epileptogenesis, but recent findings also point to an intriguing role of the Ca(v)2.3 E/R-type Ca(2+) channel in ictogenesis and seizure propagation. Based on the observation that Ca(v)2.3 is thought to induce plateau potentials in CA1 pyramidal cells, which can trigger epileptiform activity, our recent investigation revealed reduced PTZ-seizure susceptibility and altered seizure architecture in Ca(v)2.3(-/-) mice compared with controls. In the present study we tested hippocampal seizure susceptibility in Ca(v)2.3-deficient mice using surface and deep intrahippocampal telemetric EEG recordings as well as phenotypic seizure video analysis. Administration of kainic acid (30 mg/kg ip) revealed clear alteration in behavioral seizure architecture and dramatic resistance to limbic seizures in Ca(v)2.3(-/-) mice compared with controls, whereas no difference in hippocampal EEG seizure activity between both genotypes could be detected at this suprathreshold dosage. The same tendency was observed for NMDA seizure susceptibility (150 mg/kg ip) approaching the level of significance. In addition, histochemical analysis within the hippocampus revealed that excitotoxic effects after kainic acid administration are absent in Ca(v)2.3(-/-) mice, whereas Ca(v)2.3(+/+) animals exhibited clear and typical signs of excitotoxic cell death. These findings clearly indicate that the Ca(v)2.3 voltage-gated calcium channel plays a crucial role in both hippocampal ictogenesis and seizure generalization and is of central importance in neuronal degeneration after excitotoxic events.     

GIRK1、2在慢性颞叶癫痫大鼠海马内的表达变化

目的:探讨G蛋白偶联内向整流钾通道(GIRK)在海人酸诱导慢性颞叶癫痫大鼠海马内的表达变化及其在癫痫中的作用。方法:采用海人酸颞叶癫痫模型,运用原位杂交、免疫细胞化学检测海马齿状回、CA1、CA3区GIRK1、2mRNA和蛋白表达;Westernblotting检测海马整体GIRK1、2蛋白变化。结果:GIRK1、2mRNA和蛋白在大鼠海马内分布广泛;慢性颞叶癫痫海马DG区GIRK2mRNA和蛋白表达增高有显著差异(P＜0.01);GIRK1mRNA在海马DG区表达有差异(P＜0.05);Westernblotting检测海马GIRK1、2未有明显变化。结论:GIRK1、2在慢性癫痫大鼠海马内的表达增高,特别在齿状回,提示是机体对神经元网络过度兴奋的代偿或适应性反应;其合成增加将降低神经元的兴奋性,阻止海马内过度兴奋的扩散(DG-CA3-CA1)。     

Seizure susceptibility and epileptogenesis are decreased in transgenic rats overexpressing neuropeptide Y

Functional studies in epileptic tissue indicate that neuropeptide Y and some of its peptide analogs potently inhibit seizure activity. We investigated seizure susceptibility in transgenic rats overexpressing the rat neuropeptide Y gene under the control of its natural promoter. Seizures were induced in adult transgenic male rats and their wild-type littermates by i.c.v. injection of 0.3 microg kainic acid or by electrical kindling of the dorsal hippocampus. Transgenic rats showed a significant reduction in the number and duration of electroencephalographic seizures induced by kainate by 30% and 55% respectively (P<0.05 and 0.01). Transgenic rats were also less susceptible to epileptogenesis than wild-type littermates as demonstrated by a 65% increase in the number of electrical stimuli required to induce stage 5 seizures (P<0.01). This phenotype was associated with a strong and specific expression of neuropeptide Y mRNA in area CA1, a brain area involved in the seizure network. We conclude that endogenous neuropeptide Y overexpression in the rat hippocampus is associated with inhibition of seizures and epileptogenesis suggesting that this system may be a valuable target for developing novel antiepileptic treatments.     

Protective effects of mossy fiber lesions against kainic acid-induced seizures and neuronal degeneration

The effects of a hippocampal mossy fiber lesion have been determined on neuronal degeneration and limbic seizures provoked by the subsequent intracerebroventricular administration of kainic acid to unanesthetized rats. Mossy fiber lesions were made either by transecting this pathway unilaterally or by destroying the dentate granule cells unilaterally or bilaterally with colchicine. All control rats eventually developed status epilepticus and each temporally discrete seizure that preceded status epilepticus was recorded from the hippocampus ipsilateral to the kainic acid infusion before the contralateral hippocampus. A mossy fiber lesion of the ipsilateral hippocampus prevented the development of status epilepticus in 26% of subjects and in 52% of subjects seizures were recorded from the contralateral hippocampus before the ipsilateral hippocampus. Unlike electrographic records from other treatment groups, those from rats which had received a bilateral colchicine lesion exhibited no consistent pattern indicative of seizure propagation from one limbic region to another. A bilateral, but not a unilateral, mossy fiber lesion also dramatically attenuated the behavioral expression of the seizures. Regardless of its effects on kainic acid-induced electrographic and behavioral seizures, a mossy fiber lesion always substantially reduced or completely prevented the degeneration of ipsilateral hippocampal CA3-CA4 neurons. This protective effect was specific for those hippocampal neurons deprived of mossy fiber innervation. Neurons in other regions of the brain were protected from degeneration only when the mossy fiber lesion also prevented the development of electrographic status epilepticus. These results suggest that the hippocampal mossy fibers constitute an important, though probably not an obligatory, link in the circuit responsible for the spread of kainic acid seizures. Degeneration of CA3-CA4 neurons appears to depend upon (1) the duration of hippocampal seizure activity and (2) an as yet undefined influence of or interaction with the mossy fiber projection which enhances the neurodegenerative effect of the seizures.     

Clinicopathologic findings in mesial temporal sclerosis treated with gamma knife radiotherapy

Mesial temporal sclerosis (MTS) is the most common cause of medically intractable temporal lobe epilepsy. Histologic findings include hippocampal atrophy with neuronal loss in the dentate, CA1, and CA3/CA4 regions with gliosis. The conventional treatment of patients with intractable epilepsy secondary to MTS has been surgical excision. Gamma knife radiotherapy (GKR) has recently been suggested as a less invasive alternative to surgery. To date, the histologic changes that occur in this setting after GKR have not been well described. The clinicopathologic features of 4 patients with MTS who received GKR and underwent subsequent surgical resection or autopsy were retrospectively reviewed. The study group is composed of 4 patients (3 women, 1 men) with ages 55, 48, 22, and 20 years, respectively, at the time of GKR. There were 2 patients who had a history of infantile febrile seizures, and 2 who had a central nervous infection during infancy. All 4 patients had a long-standing (13-36 years) history of temporal lobe seizures resistant to medical management. Imaging studies, electroencephalogram, and surgical specimens all confirmed the diagnosis of MTS. The oldest of the 4 patients died 1 month after receiving GKR, presumably because of post-gamma knife persistent seizure complications. The postmortem neuropathology on this patient was unremarkable for any radiation effect changes but showed evidence of MTS. The remaining 3 patients underwent surgical resection for persistent seizures at 18, 22, and 20 months, respectively, post-gamma knife. These 3 surgical specimens showed variable degrees of radiation effect changes in the temporal lobe, hippocampus, and amygdala, including chronic (lymphocytes and macrophages) perivascular inflammation (3/3), vascular sclerosis (3/3), foci of edema with necrosis (3/3; extensive in 2 patients), reactive astrocytosis (3/3), microglial proliferation (1/3), and microcalcifications (1/3). Patients with MTS who underwent GKR can develop typical radiation changes over time. Treatment of individuals with MTS via GKR may not always be adequate in controlling seizures. Radiation therapy effect may contribute to persistent seizures after GKR in some patients with MTS.     

Wortmannin通过抑制癫痫大鼠海马自噬活性发挥神经保护作用

目的:探讨癫痫大鼠海马自噬活性的变化及自噬抑制剂wortmannin(WM)对致痫大鼠海马神经元的保护作用。方法:实验大鼠分为对照组、致痫2 h、8 h、16 h、24 h、72 h组和WM干预组。应用HE染色和Nissl染色观察癫痫大鼠海马神经元损伤的变化。免疫印迹检测海马组织微管相关蛋白1轻链3(LC3)Ⅱ/LC3Ⅰ的比值作为自噬活性的指标。结果:LC3Ⅱ/LC3Ⅰ比值在大鼠癫痫发作2 h时开始升高,24 h达到高峰,持续升高至少72 h。致痫24 h时海马CA1区出现明显神经元损伤和丢失。WM干预组CA1区存活神经元数目(100.88±18.73)显著高于癫痫组(70.16±5.09)(P0.05),LC3Ⅱ/LC3Ⅰ的比值低于癫痫组(P0.05)。结论:癫痫发作导致的海马损伤时存在自噬激活现象;WM通过抑制自噬活性减轻癫痫发作所致的海马损伤,具有神经保护作用。     

Mouse models of human KCNQ2 and KCNQ3 mutations for benign familial neonatal convulsions show seizures and neuronal plasticity without synaptic reorganization

The childhood epilepsy syndrome of benign familial neonatal convulsions (BFNC) exhibits the remarkable feature of clinical remission within a few weeks of onset and a favourable prognosis, sparing cognitive abilities despite persistent expression of the mutant KCNQ2 or KCNQ3 potassium channels throughout adulthood. To better understand such dynamic neuroprotective plasticity within the developing brain, we introduced missense mutations that underlie human BFNC into the orthologous murine Kcnq2 (Kv7.2) and Kcnq3 (Kv7.3) genes. Mutant mice were examined for altered thresholds to induced seizures, spontaneous seizure characteristics, hippocampal histology, and M-current properties of CA1 hippocampal pyramidal neurons. Adult Kcnq2 ^A306T/+ and Kcnq3 ^G311V/+ heterozygous knock-in mice exhibited reduced thresholds to electrically induced seizures compared to wild-type littermate mice. Both Kcnq2 ^A306T/A306T and Kcnq3 ^G311V/G311V homozygous mutant mice exhibited early onset spontaneous generalized tonic-clonic seizures concurrent with a significant reduction in amplitude and increased deactivation kinetics of the neuronal M-current. Mice had recurrent seizures into adulthood that triggered molecular plasticity including ectopic neuropeptide Y (NPY) expression in granule cells, but without hippocampal mossy fibre sprouting or neuronal loss. These novel knockin mice recapitulate proconvulsant features of the human disorder yet show that inherited M-current defects spare granule cells from reactive changes in adult hippocampal networks. The absence of seizure-induced pathology found in these epileptic mouse models parallels the benign neurodevelopmental cognitive profile exhibited by the majority of BFNC patients.     

Epileptic tolerance is associated with enduring neuroprotection and uncoupling of the relationship between CA3 damage,neuropeptide Y rearrangement and spontaneous seizures following intra-amygdala kainic acid-induced status epilepticus in mice

Brief, non-harmful seizures can activate endogenous protective programmes which render the brain resistant to damage caused by prolonged seizure episodes. Whether protection in epileptic tolerance is long-lasting or influences the subsequent development of epilepsy is uncertain. Presently, we investigated the relationship between hippocampal pathology, neuropeptide Y rearrangement and spontaneous seizures in sham- and seizure-preconditioned mice after status epilepticus induced by intra-amygdala kainate. Seizure-induced neuronal death at 24 h was significantly reduced in the ipsilateral hippocampal CA3 and hilus of tolerance mice compared to sham-preconditioned animals subject to status epilepticus. Damage to the CA3-hilus remained reduced in tolerance mice 21 days post-status. In sham-preconditioned mice subject to status epilepticus correlative statistics showed there was a strong inverse relationship between CA3, but not hilar, neuron counts and the number of spontaneous seizures. A strong positive association was also found between neuropeptide Y score and spontaneous seizure count in these mice. In contrast, there was no significant association between spontaneous seizure count and CA3 neuron loss or neuropeptide Y rearrangement in the tolerance mice. These data show that tolerance-conferred neuroprotection is long-lasting and that tolerance disrupts the normal association between CA3 damage, synaptic rearrangement and occurrence of spontaneous seizures in this model     

Age-related amyloid beta deposition in transgenic mice overexpressing both Alzheimer mutant presenilin 1 and amyloid beta precursor protein Swedish mutant is not associated with global neuronal loss

To analyze the relationship between the deposition of amyloid beta peptides (Abeta) and neuronal loss in transgenic models of Alzheimer's disease (AD), we examined the frontal neocortex (Fc) and CA1 portion of hippocampus (CA1) in PSAPP mice doubly expressing AD-associated mutant presenilin 1 (PS1) and Swedish-type mutant beta amyloid precursor protein (APPsw) by morphometry of Abeta burden and neuronal counts. Deposition of Abeta was detected as early as 3 months of age in the Fc and CA1 of PSAPP mice and progressed to cover 28.3% of the superior frontal cortex and 18.4% of CA1 at 12 months: approximately 20- (Fc) and approximately 40- (CA1) fold greater deposition than in APPsw mice. There was no significant difference in neuronal counts in either CA1 or the frontal cortex between nontransgenic (non-tg), PS1 transgenic, APPsw, and PSAPP mice at 3 to 12 months of age. In the PSAPP mice, there was disorganization of the neuronal architecture by compact amyloid plaques, and the average number of neurons was 8 to 10% fewer than the other groups (NS, P > 0.10) in CA1 and 2 to 20% fewer in frontal cortex (NS, P = 0.31). There was no loss of total synaptophysin immunoreactivity in the Fc or dentate gyrus molecular layer of the 12-month-old PSAPP mice. Thus, although co-expression of mutant PS1 with Swedish mutant betaAPP leads to marked cortical and limbic Abeta deposition in an age-dependent manner, it does not result in the dramatic neuronal loss in hippocampus and association cortex characteristic of AD.     

Conditional transgenic suppression of M channels in mouse brain reveals functions in neuronal excitability, resonance and behavior

In humans, mutations in the KCNQ2 or KCNQ3 potassium-channel genes are associated with an inherited epilepsy syndrome. We have studied the contribution of KCNQ/M-channels to the control of neuronal excitability by using transgenic mice that conditionally express dominant-negative KCNQ2 subunits in brain. We show that suppression of the neuronal M current in mice is associated with spontaneous seizures, behavioral hyperactivity and morphological changes in the hippocampus. Restriction of transgene expression to defined developmental periods revealed that M-channel activity is critical to the development of normal hippocampal morphology during the first postnatal weeks. Suppression of the M current after this critical period resulted in mice with signs of increased neuronal excitability and deficits in hippocampus-dependent spatial memory. M-current-deficient hippocampal CA1 pyramidal neurons showed increased excitability, reduced spike-frequency adaptation, attenuated medium afterhyperpolarization and reduced intrinsic subthreshold theta resonance. M channels are thus critical determinants of cellular and neuronal network excitability, postnatal brain development and cognitive performance.     

Persistent regional increases in brain-derived neurotrophic factor in the flurothyl model of epileptogenesis are dependent upon the kindling status of the animal

Brain-derived neurotrophic factor (BDNF) appears to be both regulated by and a regulator of epileptogenesis. In the flurothyl (HFE) model of kindling mice exposed to successive flurothyl trials over 8 days express a rapid, long-lasting reduction in generalized seizure threshold and a more slowly evolving change in seizure phenotype in response to subsequent flurothyl exposure. The BDNF genotype of particular mouse strains appears to influence the epileptogenic progression in this model. Thus, we hypothesized that BDNF signaling pathways are altered by flurothyl-induced seizures. Following HFE kindling, fully kindled (eight seizures) adult male C57BI/6J mice had significantly elevated whole brain BDNF levels through at least 28 days after their final seizure. Mice that received only four HFE seizures (not kindled) had elevated BDNF levels, but only at 1 day post-seizure (DPSz), while BDNF levels were not significantly altered in mice receiving just one HFE seizure at any time point studied. Regional expression patterns of BDNF in the hippocampus, hypothalamus, and frontal cortex were also elevated by one DPSz and returned to control values by 14 DPSz in mice that received four HFE seizures. No changes were seen in the cerebellum, striatum, or piriform cortex. In contrast, fully kindled mice had significantly elevated BDNF levels within the hippocampus, hypothalamus, neocortex, and striatum that remained elevated through at least 14 DPSz, while levels were unchanged in the cerebellum and piriform cortex. Regional results were confirmed using anti-BDNF immunohistochemistry (IHC). Despite changes in BDNF levels following HFE kindling, we were unable to demonstrate alterations either in full-length tyrosine kinase receptor B (TrkB) expression (Western blot and IHC) or in truncated TrkB (IHC) expression levels. Together, these data suggest a model of a positive feedback loop involving seizure activity and seizure number and persistently modified BDNF signaling pathways that influences seizure phenotypes within the HFE kindling paradigm. Thus, long-term elevations in BDNF may be responsible in part for epileptogenic processes and the development of human refractory epilepsies.     

Acupuncture inhibits kainic Acid-induced hippocampal cell death in mice

We examined whether acupuncture can reduce both the incidence of seizures and hippocampal cell death using a mouse model of kainic acid (KA)-induced epilepsy. ICR mice were given acupuncture once a day at acupoint HT8 (sobu) bilaterally during 2 days before KA injection. After an intracerebroventricular injection of 0.1 microg of KA, acupuncture treatment was subsequently administered once more (total 3 times), and the degree of seizure was observed for 20 min. Three hours after injection, the survival of neuronal cells and the expressions of c-Fos, c-Jun, and glutamate decarboxylase (GAD)-67 in the CA1 and CA3 were determined using immunohistochemistry and Western blotting techniques. Acupuncture reduced the severity of the KA-induced epileptic seizure and the rate of neural cell death, and it also decreased the expressions of c-Fos and c-Jun induced by KA in the hippocampus. Furthermore, acupuncture increased GAD-67 expressions in the same areas. These results demonstrated that it could inhibit the KA-induced epileptic seizure and hippocampal cell death by increasing GAD-67 expressions.     

Sodium currents in isolated rat CA1 pyramidal and dentate granule neurones in the post-status epilepticus model of epilepsy

Status epilepticus (SE) was induced in the rat by long-lasting electrical stimulation of the hippocampus. After a latent period of 1 week, spontaneous seizures occurred which increased in frequency and severity in the following weeks, finally culminating after 3 months in a chronic epileptic state. In these animals we determined the properties of voltage-dependent sodium currents in acutely isolated CA1 pyramidal neurones and dentate granule cells using the whole-cell voltage-clamp technique. The conductance of the fast transient sodium current was larger in SE rats (84+/-7 nS versus 56+/-6 nS) but related to a difference in cell size so that the neurones had a similar specific sodium conductance (control: 7.8+/-0.8 nS/pF, SE: 6.7+/-0.8 nS/pF). Current activation and inactivation were characterised by a Boltzmann function. After SE the voltage dependence of activation was shifted to more negative potentials (control: -45.1+/-1.4 mV, SE: -51.5+/-2.9 mV, P<0.05). In combination with a small shift in the voltage dependence of inactivation to more depolarised potentials (control: -68.8+/-2.3 mV, SE: -66.3+/-2.3 mV), it resulted in a window current that was much increased in the SE neurones (median: 64 pA in control, 217 pA in SE, P<0.05). The peak of this window current shifted to more hyperpolarised potentials (control: -44 mV, SE: -50 mV, P<0.05). No differences were found in the sodium currents analysed in dentate granule cells of control and SE animals. The changes observed in CA1 neurones after SE contribute to enhanced excitability in particular when membrane potential is near firing threshold. They can, at least partly, explain the lower threshold for epileptic activity in SE animals. The comparison of CA1 with DG neurones in the same rats demonstrates a differential response in the two cell types that participated in very similar seizure activity.     

Increased Seizure Susceptibility and Up-regulation of nNOS Expression in Hippocampus Following Recurrent Early-life Seizures in Rats

This study aimed to determine the long-term change of seizure susceptibility and the role of nNOS on brain development following recurrent early-life seizures in rats. Video-EEG recordings were conducted between postnatal days 50 and 60. Alterations in seizure susceptibility were assayed on day 22 or 50 using the flurothyl method. Changes in nNOS expression were determined by quantitative immunoblotting on day 50. On average, rats had 8.4±2.7 seizures during 10 daily 1 hr behavioral monitoring sessions. As adults (days 50-60), all rats displayed interictal spikes in the hippocampus and/or overlying cortex. Brief electrographic seizures were recorded in only one of five animals. Rats appeared to progress from a period of marked seizure susceptibility (day 22) to one of lessened seizure susceptibility (day 50). Up-regulation of nNOS expression following early-life recurrent seizures was observed on day 50. In conclusion, these data suggested that recurrent early-life seizures had the long-term effects on seizure susceptibility late in life and up-regulatory nNOS expression on the hippocampus during brain development, and nNOS appeared to contribute to the persistent changes in seizure susceptibility, and epileptogenesis.     

Tetanus toxin induces long-term changes in excitation and inhibition in the rat hippocampal CA1 area

Intrahippocampal tetanus toxin induces a period of chronic recurrent limbic seizures in adult rats, associated with a failure of inhibition in the hippocampus. The rats normally gain remission from their seizures after 6-8 weeks, but show persistent cognitive impairment. In this study we assessed which changes in cellular and network properties could account for the enduring changes in this model, using intracellular and extracellular field recordings in hippocampal slices from rats injected with tetanus toxin or vehicle, 5 months previously.In CA1 pyramidal neurones from toxin-injected rats, the slope of the action potential upstroke was reduced by 32%, the fast afterhyperpolarisation by 32% and the slow afterhyperpolarisation by 54%, suggesting changes in voltage-dependent conductances. The excitatory postsynaptic potential slope was reduced by 60% and the population synaptic potential slope was reduced at all stimulus intensities, suggesting a reduced afferent input in CA1. Paired-pulse stimulation showed an increase of the excitability ratio and an increase of cellular excitability only for the second pulse, suggesting a reduced inhibition. The polysynaptic inhibitory postsynaptic potential was reduced by 34%, whereas neither the inhibitory postsynaptic potential at subthreshold stimulus intensities,nor the pharmacologically isolated monosynaptic inhibitory postsynaptic potential were different in toxin-injected rats, suggesting a reduced synaptic excitation of interneurones. Stratum radiatum stimuli in toxin-injected rats, and not in controls, evoked antidromic activation of CA1 neurones, demonstrating axonal sprouting into areas normally devoid of CA1 pyramidal cell axons.We conclude that this combination of enduring changes in cellular and network properties, both pro-epileptic (increased recurrent excitatory connectivity, reduced recurrent inhibition and reduced afterhyperpolarisations) and anti-epileptic (impaired firing and reduced excitation), reaches a balance that allows remission of seizures, perhaps at the price of persistent cognitive impairment.     

Bcl-w protects hippocampus during experimental status epilepticus

Experimentally evoked seizures can activate the intrinsic mitochondrial cell death pathway, components of which are modulated in the hippocampus of patients with temporal lobe epilepsy. Bcl-2 family proteins are critical regulators of mitochondrial dysfunction, but their significance in this setting remains primarily untested. Presently, we investigated the mitochondrial pathway and role of anti-apoptotic Bcl-2 proteins using a mouse model of seizure-induced neuronal death. Status epilepticus was evoked in mice by intra-amygdala kainic acid, causing cytochrome c release, processing of caspases 9 and 7, and death of ipsilateral hippocampal pyramidal neurons. Seizures caused a rapid decline in hippocampal Bcl-w levels not seen for either Bcl-2 or Bcl-xl. To test whether endogenous Bcl-w was functionally significant for neuronal survival, we investigated hippocampal injury after seizures in Bcl-w-deficient mice. Seizures induced significantly more hippocampal CA3 neuronal loss and DNA fragmentation in Bcl-w-deficient mice compared with wild-type mice. Quantitative electroencephalography analysis also revealed that Bcl-w-deficient mice display a neurophysiological phenotype whereby there was earlier polyspike seizure onset. Finally, we detected higher levels of Bcl-w in hippocampus from temporal lobe epilepsy patients compared with autopsy controls. These data identify Bcl-w as an endogenous neuroprotectant that may have seizure-suppressive functions.     

Changes in Na(+)-K(+)-Cl(-) cotransporter immunoreactivity in the gerbil hippocampus following spontaneous seizure

The immunoreactivity of Na(+)-K(+)-Cl(-) cotransporter (NKCC) in the gerbil hippocampus associated with various sequelae of spontaneous seizures were investigated in order to identify the roles of NKCC in the epileptogenesis and the recovery mechanisms in these animals. The NKCC immunoreactivities in the CA2-3 regions, the subiculum and the entorhinal cortex, were significantly more intensified in the pre-seizure group of seizure sensitive (SS) gerbils than in the seizure resistant (SR) gerbils. Following the on-set of seizure, the immunoreactivity of NKCC was significantly changed. In the hippocampal complex except the CA1 region, NKCC immunoreactivity in GABAergic neurons was significantly decreased 30 min after seizure on-set, versus the pre-seizure group. On the other hand, NKCC immunoreactivity was dramatically elevated in the CA1 regions, and 3 h postictal NKCC immunoreactivity increased significantly in the dentate gyrus and the dendrites of the pyramidal cells in the CA2-3 regions. These findings suggest that altered NKCC expression may be associated with seizure activity, and have an important role in the postictal recovery by regulating GABA-mediated inhibitory circuit in the hippocampal complex of the gerbil.     

Hippocampal neurons and glia in epileptic EL mice

Reactive changes in hippocampal astrocytes are frequently encountered in association with temporal lobe epilepsy in humans and with drug or kindling-induced seizures in animal models. These reactive changes generally involve increases in astrocyte size and number and often occur together with neuronal loss and synaptic rearrangements. In addition to producing astrocytic changes, seizure activity can also produce reactive changes in microglia, the resident macrophages of brain. In this study, we examined the effects of recurrent seizure activity on hippocampal neurons and glia in the epileptic EL mouse, a natural model of human multifactorial idiopathic epilepsy and complex partial seizures. Timm staining was used to evaluate infrapyramidal mossy fiber organization and the optical dissector method was used to count Nissl-stained neurons in hippocampus of adult (about one year of age) EL mice and nonepileptic C57BL/6J (B6) and DDY mice. Immunostaining forglial fibrillary acidic protein (GFAP) and Iba1, an actin cross-linking molecule restricted to macrophages and microglia, was used to evaluate astrocytes and microglia, respectively. The EL mice experienced about 25–30 complex partial seizures with secondary generalization during routine weekly cage changing. No significant differences were found among the mouse strains for Timm staining scores or for neuronal counts in the CA1 and CA3 pyramidal fields or in the hilus. However, the number of GFAP-positive astrocytes was significantly elevated in the stratum radiatum and hilus of EL mice, while microglia appeared hyper-ramified and were more intensely stained in EL mice than in the B6 or DDY mice in the hilus, parietal cortex, and pyriform cortex. The results indicate that recurrent seizure activity in EL mice is associated with abnormalities in hippocampal astrocytes and brain microglia, but is not associated with obvious neuronal loss or mossy fiber synaptic rearrangements. The EL mouse can be a useful model for evaluating neuron-glia interactions related to idiopathic epilepsy.