Non-epileptic Control Research Articles

Adenylyl Cyclases: Expression in the Developing Rat Thalamus and Their Role in Absence Epilepsy

Adenylyl cyclases (ACs) synthesize the second messenger cyclic AMP (cAMP) which influences the function of multiple ion channels. Former studies point to a malfunction of cAMP-dependent ion channel regulation in thalamocortical relay neurons that contribute to the development of the absence epileptic phenotype of a rat genetic model (WAG/Rij). Here, we provide detailed information about the thalamic gene and protein expression of Ca(2+)/calmodulin-activated AC isoforms in rat thalamus. Data from WAG/Rij were compared to those from non-epileptic controls (August-Copenhagen Irish rats) to elucidate whether differential expression of ACs contributes to the dysregulation of thalamocortical activity. At one postnatal stage (P21), we found the gene expression of two specific Ca(2+)-activated AC isoforms (AC-1 and AC-3) to be significantly down-regulated in epileptic tissue, and we identified the isoform AC-1 to be the most prominent one in both strains. However, Western blot data and analysis of enzymatic AC activity revealed no differences between the two strains. While basal AC activity was low, cAMP production was boosted by application of a forskolin derivative up to sevenfold. Despite previous hints pointing to a major contribution of ACs, the presented data show that there is no apparent causality between AC activity and the occurrence of the epileptic phenotype.

Metoprine induced behavioral modifications and brain regional histamine increase in WAG/Rij and Wistar rats

The effects of metoprine, an inhibitor of histamine N-methyltransferase, on open field activity and brain regional histamine (HA) content were examined in rats with mixed, absence and audiogenic, epilepsy (WAG/Rij-AGS), rats with audiogenic epilepsy (Wistar-AGS) and in non-epileptic control rats (Wistar-nAGS). HA content was increased by metoprine (20mg/kg, i.p.) in the cortex, striatum, thalamus, hypothalamus and hippocampus of the rats from all three tested groups. However, WAG/Rij rats showed a lower rate of metoprine-induced HA accumulation in the striatum and thalamus than Wistar rats. For the open field test, the main effect of metoprine (20mg/kg, i.p.) was a general increase of locomotor activity although distinctive features, such as hyperlocomotion and exaggerated sniffing, were characteristic for the epileptic rats (WAG/Rij-AGS and Wistar-AGS, respectively). Individual rats from all the groups showed stereotyped behavior of shuttle type and head bobbing. Electroencephalographic data obtained in WAG/Rij-AGS rats confirmed that metoprine-induced behavioral activation was accompanied by suppression of spike-wave discharges, the main hallmark of absence seizures. Taken together, these results show that inhibition of the histamine catabolism may induce motor activation of particular patterns in epileptic rats and provoke stereotyped behavior.

Determinants of active convulsive epilepsy in rural Cameroon: a population based case–control study

Purpose: The aim of this study was to investigate the determinants of active convulsive epilepsy in rural Cameroon, in order to inform prevention and control strategies at the local and regional levels.Methods: This case–control study took place in the rural health district of Bafut, North-West Region of Cameroon. People with epilepsy receiving care in local health facilities or identified in the community were matched with non-epileptic controls for age and sex. The Limoges questionnaire was used for case definitions and screening for risk factors for epilepsy. Potential determinants were investigated through logistic regression models.Results: In all 170 participants (85 cases, with 85 controls) were included. Fewer cases than controls were married (27% vs. 58%; P<0·001) and more cases than control were living with parents (56% vs. 28%; P<0·001). In final multivariable regression analyses, a family history of epilepsy (odd ratio 3·11; 95% confidence interval: 1·61–6·01; P<0·001) and a poor obstetrical history (odd ratio: 4·34; 95% confidence interval: 1·29–14·58; P = 0·018) were significant determinants of active convulsive epilepsy.Conclusion: In this rural area of Cameroon, the previously reported high prevalence of active convulsive epilepsy could be explained by a positive family history and a poor obstetrical history. The later being a potentially modifiable determinant, there is a need for additional efforts into maternal and child health, and for integrated disease control programs to mitigate the social impact of epilepsy

Cytokines and Absence Seizures in a Genetic Rat Model

We investigated the role of two cytokines, IL-1β and TNF-α, in the development of absence seizures using a genetic model of absence epilepsy in WAG/Rij rats. We administered these cytokines to animals systemically and measured the number of spike-wave discharges (SWDs) in the EEG. We also coadministered IL-1β with the GABA reuptake inhibitor tiagabine and measured the levels of IL-1β and TNF-α in the brain and blood plasma of 2-, 4-, and 6-month-old WAG/Rij rats and animals that served as a non-epileptic control (ACI). We found that IL-1β induced a significant increase in SWDs 2-5 h after administration, while TNF-α enhanced SWDs much later. Both cytokines enhanced passive behavior; body temperature was elevated only after TNF-α. The action of tiagabine was potentiated by earlier IL-1β injection, even when IL-1β was no longer active. Young WAG/Rij rats showed higher levels of TNF-α in blood serum than young ACI rats; the effects in the brain tended to be opposite. The marked differences in timing of the increase in SWDs suggest different time scales for the action of both cytokines tested. It is proposed that the results found after TNF-α are due to the de novo synthesis of IL-1β. TNF-α may possess neuroprotective effects. IL-1β might increase GABA-ergic neurotransmission. The changes in the efficacy of antiepileptic drugs related to changes in the cytokine systems may have some clinical relevance.

Vagus nerve stimulation has antidepressant potential in the kainic acid model for temporal lobe epilepsy

Aim : Depression represents one of the most common and disabling comorbidities in epilepsy patients. There is a growing consensus that both disorders have a common neurobiological cause. Vagus nerve stimulation (VNS) is an add-on treatment for patients with refractory epilepsy or depression. Both the mechanism of this comorbidity and the mechanism of action of VNS are poorly understood. In this study, we investigated the antidepressant potential of VNS in the kainic acid (KA) model for temporal lobe epilepsy (TLE). Methods : Male Sprague Dawley rats (n=29) were implanted with a cuff-electrode around the left vagus nerve. One week after surgery, half of the animals were subjected to a kainic acid KA-induced status epilepticus (SE). The other half of the animals were injected with saline (Sal). Both groups were subdivided in a VNS-treated group and a SHAM-treated group, yielding 4 experimental groups: Sal-SHAM (n=8), Sal-VNS (n=7), KA-SHAM (n=7), KA-VNS (n=7). Five weeks post-SE, the behavioral equivalent of anhedonia (i.e. inability to experience pleasure), was assessed using the saccharin preference test (SPT). To control for loss of taste due to KA-induced SE and subsequent neuronal loss, a quinine preference (of aversion) test (QPT) was performed. Both VNS groups (Sal-VNS and KA-VNS) received 2 weeks of therapeutic VNS, both SHAM groups received SHAM treatment. After this period, SPT and QPT were performed again in the same animals. Results : KA rats showed a significant reduction of taste preference towards saccharin compared to Sal rats (25% vs. 95 %, p < 0.001). No difference in aversion towards quinine was found. Two weeks of VNS significantly increased saccharin preference (SP) in the KA-VNS group (14% to 60% p < 0.001), while no difference could be demonstrated in SHAM-treated animals of the KA group. VNS had no influence on the QPT. There were no significant changes of SP in Sal-VNS and Sal-SHAM group. Conclusion : KA-injected epileptic rats display signs of anhedonia based on the SPT compared to non-epileptic control rats. Two weeks of VNS significantly reduced this anhedonia, suggesting an antidepressant effect of VNS in this rodent model for TLE.

Eicosanoid levels in the neocortex of drug-resistant epileptic patients submitted to epilepsy surgery

There is an increasing body of evidence implicating eicosanoids (arachidonic acid metabolites) in the experimental generation of epileptic seizures and the development of epilepsy. Our purpose was to measure the synthesis of eicosanoids from the cyclooxygenase and lipoxygenase pathways in human brain neocortex tissue samples obtained from epileptic patients, and to compare them with non-epileptic control subjects. Epileptic neocortex specimens demonstrated a significant increase (P<0.001) in the levels of three eicosanoids derived from the cyclooxygenase pathway: Prostaglandin E(2) (PGE(2)), Thromboxane A(2) (TXA(2)), and Prostacyclin (PGI(2)), compared to controls. In the epileptic samples the level of TXA(2) was twice as much the levels of PGI(2), while in the control samples the levels of PGI(2) were slightly higher than TXA(2). Conversely, there were no detectable levels of eicosanoids derived from the lipoxygenase pathway: Leukotriene B(4) (LTB(4)) and Leukotriene C(4) (LTC(4)). The lack of leukotrienes synthesis illustrates that COX pathway is dominant in neocortex of epileptic patients. Our human data are consistent with the results obtained in experimental animal models of epilepsy. The important increase in PGE(2) and TXA(2) suggests that selective inhibition of prostanoid synthesis or blockage of prostanoid receptors might provide novel antiepileptic strategies in human epilepsy.

Increased expression of placental growth factor in patients with temporal lobe epilepsy and a rat model

Placental growth factor (PIGF) plays a role in angiogenesis and neuroprotection. It has been suggested that angiogenesis and blood–brain barrier damage are involved in the pathophysiology of epilepsy. In this study, we investigated the PIGF expression in the temporal neocortices of 11 patients with pharmaco-resistant temporal lobe epilepsy (TLE) and 6 non-epileptic controls, using double immunofluorescence labeling, immunohistochemistry and Western blotting. We also assessed PIGF expression pattern in a rat model of TLE induced by lithium chloride-pilocarpine. We found that PIGF expression was significantly elevated in patients with TLE than in control. TLE patients with initial injuries had significantly higher PIGF level than those without initial injuries. In the TLE rat model, PIGF upregulation started at 6h after status epilepticus and maintained at significant high level for up to 60days. These results suggest that the augmentation of brain PIGF is associated with development of epilepsy.

Ictal SPECT of Thalamocortical Coupling in a Patient with Frontal Absence

Petit mal absence has been reported with 3-Hz generalized spike-and-wave discharges induced by secondary bilateral synchrony. Absence seizure may be present in patients with frontal lobe epilepsy. The thalamic rhythmogenic mechanisms responsible for spike-and-wave discharges have been investigated, providing a better understanding of the underlying anatomico-physiological mechanisms. We report the thalamocortical coupling in a patient with frontal absence by performing synchronous ictal single photon emission computed tomography (SPECT) analysis. Ictal SPECT revealed thalamic hyperperfusion combined with ipsilateral frontal cortical hyperperfusion in the patient. Moreover, lateral indexes of cerebral blood flow in the frontal region and thalamus were higher than those from non-epileptic control subjects. Thalamocortical coupling was thus revealed by ictal SPECT. Frontal absences should be considered as a secondarily generalized epilepsy syndrome originating from the frontal regions. The thalamus may play a crucial role as a pacemaker of rhythmic electroencephalographic activities such as secondary bilateral synchronous discharges in patients with frontal absences.

Association study of functional polymorphisms in serotonin transporter gene with temporal lobe epilepsy in Han Chinese population

Serotoninergic dysfunction was reported to be involved in aetiology of temporal lobe epilepsy (TLE). Serotonin (5-HT) is actively cleared from synaptic cleft by serotonin transporter (5-HTT). We investigated the association between three common polymorphisms of 5-HTT gene, which may influence gene expression or function, and risk for TLE. Three hundred and thirty-four patients with TLE and four hundred and eighty-seven non-epileptic control subjects from Han Chinese origin were enrolled for the present study. Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method was used for genotyping. 10-repeat allele frequency of 17 bp variable number of tandem repeats in the second intron (5-HTTVNTR) was moderately higher in patients with TLE than in controls (9.1% vs. 6.1%, P=0.0187, OR=1.55, 95%CI=1.07-2.26). Our study suggested 10-repeat allele of 5-HTTVNTR may be associated with TLE susceptibility.

Altered expression of brain monocarboxylate transporter 1 in models of temporal lobe epilepsy

Monocarboxylate transporter 1 (MCT1) facilitates the transport of monocarboxylate fuels (lactate, pyruvate and ketone bodies) and acidic drugs, such as valproic acid, across cell membranes. We recently reported that MCT1 is deficient on microvessels in the epileptogenic hippocampal formation in patients with medication-refractory temporal lobe epilepsy (TLE). To further define the role of MCT1 in the pathophysiology of TLE, we used immunohistochemistry and stereological analysis to localize and quantify the transporter in the hippocampal formation in three novel and highly relevant rat models of TLE and in nonepileptic control animals. One model utilizes methionine sulfoximine to induce brain glutamine synthetase deficiency and recurrent limbic seizures, while two models employ an episode of perforant pathway stimulation to cause epilepsy. MCT1 was lost on microvessels and upregulated on astrocytes in the hippocampal formation in all models of TLE. Notably, the loss of MCT1 on microvessels was not due to a reduction in microvessel density. The similarities in MCT1 expression among human subjects with TLE and several animal models of the disease strongly suggest a critical role of this molecule in the pathogenesis of TLE. We hypothesize that the downregulation of MCT1 may promote seizures via impaired uptake of ketone bodies and antiepileptic drugs by the epileptogenic brain. We also propose that the overexpression of MCT1 on astrocytes may lead to increased uptake or release of monocarboxylates by these cells, with important implications for brain metabolism and excitability. These hypotheses can now be rigorously tested in several animal models that replicate key features of human TLE.

Differential aberrant sprouting in temporal lobe epilepsy with psychiatric co-morbidities

Psychiatric co-morbidities in epilepsy are common in patients with temporal lobe epilepsy (TLE). Pathological alterations in TLE are well characterised; however, neuropathologic data are relatively scale regarding the association between psychiatric diseases and epilepsy. Our objective was to evaluate the clinical data of 46 adult TLE patients with and without psychiatric co-morbidities and to correlate the data with hippocampal neuronal density and mossy fiber sprouting. Accordingly, patients were grouped as follows: TLE patients without history of psychiatric disorder (TLE, n=16), TLE patients with interictal psychosis (TLE+P, n=14), and TLE patients with major depression (TLE+D, n=16). Hippocampi from autopsies served as non-epileptic controls (n=10). TLE+P exhibited significantly diminished mossy fiber sprouting and decreased neuronal density in the entorhinal cortex when compared with TLE. TLE+P showed significantly poorer results in verbal memory tasks. TLE+D exhibited significantly increased mossy fiber sprouting length when compared with TLE and TLE+P. Further, a higher proportion of TLE+D and TLE+P presented secondarily generalised seizures than did TLE. Our results indicate that TLE patients with psychiatric disorders have distinct features when compared with TLE patients without psychiatric co-morbidities and that these changes may be involved in either the manifestation or the maintenance of psychiatric co-morbidities in epilepsy.

Cellular neuropathology of absence epilepsy in the neocortex: a population of glial cells rather than neurons is impaired in genetic rat model.

It is well accepted that absence epilepsy is not accompanied by structural brain abnormalities. In the present report, we challenged this view based on microscopic analysis of neocortex in a genetic model of absence epilepsy, WAG/Rij rats. Density of neurons and glial cells was measured in the motor, somatosensory and cingular cortical areas in epileptic WAG/Rij rats and in non-epileptic control ACI rats. More extensive and significant differences between two strains were found in a population of glial cells and less significant - in neurons. In contract to ACI rats, WAG/Rij rats showed (1) a deficit of glial cells and a lower glia-neuron index in the somatosensory and cingulate areas (deep layers); (2) a reduced number of neurons locally in the motor cortex. The somatosensory cortex (deep layers) is known to play a key role in triggering of epileptic discharges, and an impairment of glia-neuron interactions in this area might underlie pathological processes in a primary epileptic focus. In the motor cortex, epileptiform activity is known to reach the highest amplitude, and this may cause or result from a deficit of neurons. Our data suggest the critical role of glial cells and glia-neuron interactions in pathogenesis of absence epilepsy.

Perirhinal cortex hyperexcitability in pilocarpine‐treated epileptic rats

The perirhinal cortex (PC), which is heavily connected with several epileptogenic regions of the limbic system such as the entorhinal cortex and amygdala, is involved in the generation and spread of seizures. However, the functional alterations occurring within an epileptic PC network are unknown. Here, we analyzed this issue by using in vitro electrophysiology and immunohistochemistry in brain tissue obtained from pilocarpine-treated epileptic rats and age-matched, nonepileptic controls (NECs). Neurons recorded intracellularly from the PC deep layers in the two experimental groups had similar intrinsic and firing properties and generated spontaneous depolarizing and hyperpolarizing postsynaptic potentials with comparable duration and amplitude. However, spontaneous and stimulus-induced epileptiform discharges were seen with field potential recordings in over one-fifth of pilocarpine-treated slices but never in NEC tissue. These network events were reduced in duration by antagonizing NMDA receptors and abolished by NMDA + non-NMDA glutamatergic receptor antagonists. Pharmacologically isolated isolated inhibitory postsynaptic potentials had reversal potentials for the early GABA(A) receptor-mediated component that were significantly more depolarized in pilocarpine-treated cells. Experiments with a potassium-chloride cotransporter 2 antibody identified, in pilocarpine-treated PC, a significant immunostaining decrease that could not be explained by neuronal loss. However, interneurons expressing parvalbumin and neuropeptide Y were found to be decreased throughout the PC, whereas cholecystokinin-positive cells were diminished in superficial layers. These findings demonstrate synaptic hyperexcitability that is contributed by attenuated inhibition in the PC of pilocarpine-treated epileptic rats and underscore the role of PC networks in temporal lobe epilepsy.

Anomalous levels of Cl− transporters cause a decrease of GABAergic inhibition in human peritumoral epileptic cortex

Several factors contribute to epileptogenesis in patients with brain tumors, including reduced γ-aminobutyric acid (GABA)ergic inhibition. In particular, changes in Cl(-) homeostasis in peritumoral microenvironment, together with alterations of metabolism, are key processes leading to epileptogenesis in patients afflicted by glioma. It has been recently proposed that alterations of Cl(-) homeostasis could be involved in tumor cell migration and metastasis formation. In neurons, the regulation of intracellular Cl(-) concentration ([Cl(-) ](i) ) is mediated by NKCC1 and KCC2 transporters: NKCC1 increases while KCC2 decreases [Cl(-) ](i) . Experiments were thus designed to investigate whether, in human epileptic peritumoral cortex, alterations in the balance of NKCC1 and KCC2 activity may decrease the hyperpolarizing effects of GABA, thereby contributing to epileptogenesis in human brain tumors. Membranes from peritumoral cortical tissues of epileptic patients afflicted by gliomas (from II to IV WHO grade) and from cortical tissues of nonepileptic patients were injected into Xenopus oocytes leading to the incorporation of functional GABA(A) receptors. The GABA-evoked currents were recorded using standard two-microelectrode voltage-clamp technique. In addition, immunoblot analysis and immunohistochemical staining were carried out on membranes and tissues from the same patients. We found that in oocytes injected with epileptic peritumoral cerebral cortex, the GABA-evoked currents had a more depolarized reversal potential (E(GABA) ) compared to those from nonepileptic healthy cortex. This difference of E(GABA) was abolished by the NKCC1 blocker bumetanide or unblocking of KCC2 with the Zn(2+) chelator TPEN. Moreover, Western blot analysis revealed an increased expression of NKCC1, and more modestly, of KCC2 transporters in epileptic peritumoral tissues compared to nonepileptic control tissues. In addition, NKCC1 immunoreactivity was strongly increased in peritumoral cortex with respect to nonepileptic cortex, with a prominent expression in neuronal cells. We report that the positive shift of E(GABA) in epileptic peritumoral human cortex is due to an altered expression of NKCC1 and KCC2, perturbing Cl(-) homeostasis, which might lead to a consequent reduction in GABAergic inhibition. These findings point to a key role of Cl(-) transporters KCC2 and NKCC1 in tumor-related epilepsy, suggesting a more specific drug therapy and surgical approaches for the epileptic patients afflicted by brain tumors.

IL-1β is induced in reactive astrocytes in the somatosensory cortex of rats with genetic absence epilepsy at the onset of spike-and-wave discharges, and contributes to their occurrence

Interleukin (IL)-1β plays a crucial role in the mechanisms of limbic seizures in rodent models of temporal lobe epilepsy. We addressed whether activation of the IL-1β signaling occurs in rats with genetic absence epilepsy (GAERS) during the development of spike-and-wave discharges (SWDs). Moreover, we studied whether inhibition of IL-1β biosynthesis in GAERS could affect SWD activity. IL-1β expression and glia activation were studied by immunocytochemistry in the forebrain of GAERS at postnatal days (PN)14, PN20, and PN90 and in age-matched non-epileptic control Wistar rats. In PN14 GAERS, when no SWDs have developed yet, IL-1β immunostaining was undetectable, and astrocytes and microglia showed a resting phenotype similar to control Wistar rats. In 3 out of 9 PN20 GAERS, IL-1β was observed in activated astrocytes of the somatosensory cortex; the cytokine expression was associated with the occurrence of immature-type of SWDs. In all adult PN90 GAERS, when mature SWDs are established, IL-1β was observed in reactive astrocytes of the somatosensory cortex but not in adjacent cortical areas or in extra-cortical regions. An age-dependent c-fos activation was found in the somatosensory cortex of GAERS with maximal levels reached in PN90 rats; c-fos was also induced in some thalamic nuclei in PN20 and PN90 GAERS. Inhibition of IL-1β biosynthesis in PN90 GAERS by 4-day systemic administration of a specific ICE/Caspase-1 blocker, significantly reduced both SWD number and duration. These results show that IL-1β is induced in reactive astrocytes of the somatosensory cortex of GAERS at the onset of SWDs. IL-1β has pro-ictogenic properties in this model, and thus it may play a contributing role in the mechanisms underlying the occurrence of absence seizures.

Electrical stimulation of the epileptic focus in absence epileptic WAG/Rij rats: assessment of local and network excitability

The study aims to investigate whether there is a higher excitability in the deep cortical layers of the peri-oral region of the somatosensory cortex as compared to other cortical regions in absence epileptic WAG/Rij rats and whether this is unique for this type of epileptic rats, as would be predicted by the cortical focus theory of absence epilepsy. Excitability of cortical structures was assessed in a double pulse paradigm (inter-pulse interval 400 ms, 400 μs pulse duration, varying stimulation intensities (20-100 μA)). Electrical stimulation was applied to the subgranular layers of the somatosensory and motor cortex of freely moving WAG/Rij and control Wistar rats. Electrical evoked potentials (EEPs) and afterdischarges (ADs) were recorded during wakefulness, drowsiness and non-REM sleep. WAG/Rij rats, stimulated in the somatosensory cortex, showed higher amplitudes for the N1 and N3 components of the EEPs as compared to WAG/Rij rats stimulated in the motor cortex. This effect was present in all states of alertness and at all tested intensities. In addition, this effect was not (N1) or to much less extent (N3) present in nonepileptic control rats. Stimulation-induced 8 Hz ADs were predominantly found in WAG/Rij rats. ADs were longer after stimulation in the somatosensory than in the motor cortex and preferentially occurred during drowsiness. There is a heightened excitability in the deep layer neurons of the perioral region of somatosensory cortex, which is unique for WAG/Rij rats. Moreover, the presence of 8 Hz ADs might point toward additional changes in the cortico-thalamo-cortical network. Drowsiness is an excellent state for 8 Hz ADs, mimicking spike and wave discharges (SWDs). The results are in good agreement with the cortical-focus theory of absence epilepsy.

Changes in intracellular protein expression in cortex, thalamus and hippocampus in a genetic rat model of absence epilepsy

Epilepsy is a chronic disorder characterized by repeated seizures resulting from abnormal activation of neurons in the brain. Although mutations in genes related to Na +, K +, Ca 2+ channels have been defined, few studies show intracellular protein changes. We have used proteomics to investigate the expression of soluble proteins in a genetic rat model of absence epilepsy “Genetic Absence Epilepsy Rats from Strasbourg (GAERS)”. The advantage of this technique is its high throughput quantitative and qualitative detection of all proteins with their post-translational modifications at a given time. The parietal cortex and thalamus, which are the regions responsible for the generation of absence seizures, and the hippocampus, which is not involved in this activity, were dissected from GAERS and from non-epileptic control rat brains. Proteins from each tissue sample were isolated and separated by two-dimensional gel electrophoresis. Spots that showed significantly different levels of expression between controls and GAERS were identified by nano LC–ESI-MS/MS. Identified proteins were: ATP synthase subunit delta and the 14-3-3 zeta isoform in parietal cortex; myelin basic protein and macrophage migration inhibitory factor in thalamus; and macrophage migration inhibitory factor and 0-beta 2 globulin in hippocampus. All protein expressions were up-regulated in GAERS except 0-beta globulin. These soluble proteins are related to energy generation, signal transduction, inflammatory processes and membrane conductance. These results indicate that not only membrane proteins but also cytoplasmic proteins may take place in the pathophysiology and can be therapeutic targets in absence epilepsy.

Protective role for type-1 metabotropic glutamate receptors against spike and wave discharges in the WAG/Rij rat model of absence epilepsy

Eight-month old WAG/Rij rats, which developed spontaneous occurring absence seizures, showed a reduced function of mGlu1 metabotropic glutamate receptors in the thalamus, as assessed by in vivo measurements of DHPG-stimulated polyphosphoinositide hydrolysis, in the presence of the mGlu5 antagonist MPEP as compared to age-matched non-epileptic control rats. These symptomatic 8-month old WAG/Rij rats also showed lower levels of thalamic mGlu1α receptors than age-matched controls and 2-month old (pre-symptomatic) WAG/Rij rats, as detected by immunoblotting. Immunohistochemical and in situ hybridization analysis indicated that the reduced expression of mGlu1 receptors found in symptomatic WAG/Rij rats was confined to an area of the thalamus that excluded the ventroposterolateral nucleus. No mGlu1 receptor mRNA was detected in the reticular thalamic nucleus. Pharmacological manipulation of mGlu1 receptors had a strong impact on absence seizures in WAG/Rij rats. Systemic treatment with the mGlu1 receptor enhancer SYN119, corresponding to compound RO0711401, reduced spontaneous spike and wave discharges spike-wave discharges (SWDs) in epileptic rats. Subcutaneous doses of 10 mg/kg of SYN119 only reduced the incidence of SWDs, whereas higher doses (30 mg/kg) also reduced the mean duration of SWDs. In contrast, treatment with the non-competitive mGlu1 receptor antagonist, JNJ16259685 (2.5 and 5 mg/kg, i.p.) increased the incidence of SWDs. These data suggest that absence epilepsy might be associated with a reduction of mGlu1 receptors in the thalamus, and that compounds that amplify the activity of mGlu1 receptors might be developed as novel anti-absence drugs. This article is part of a Special Issue entitled ‘Trends in Neuropharmacology: In Memory of Erminio Costa’.

Increased levels of HMGB1 and pro-inflammatory cytokines in children with febrile seizures

ObjectiveFebrile seizures are the most common form of childhood seizures. Fever is induced by pro-inflammatory cytokines during infection, and pro-inflammatory cytokines may trigger the development of febrile seizures. In order to determine whether active inflammation, including high mobility group box-1 (HMGB1) and pro-inflammatory cytokines, occurs in children with febrile seizures or epilepsy, we analyzed cytokine profiles of patients with febrile seizures or epilepsy.MethodsForty-one febrile seizure patients who visited the emergency department of Seoul National University Boramae Hospital from June 2008 to May 2009 were included in this study. Blood was obtained from the febrile seizure child patients within 30 minutes of the time of the seizure; subsequently, serum cytokine assays were performed. Control samples were collected from children with febrile illness without convulsion (N = 41) and similarly analyzed. Serum samples from afebrile status epilepticus attacks in intractable epilepsy children (N = 12), afebrile seizure attacks in generalized epilepsy with febrile seizure plus (GEFSP) children (N = 6), and afebrile non-epileptic controls (N = 7) were also analyzed.ResultsSerum HMGB1 and IL-1β levels were significantly higher in febrile seizure patients than in fever only controls (p < 0.05). Serum IL-6 levels were significantly higher in typical febrile seizures than in fever only controls (p < 0.05). Serum IL-1β levels were significantly higher in status epilepticus attacks in intractable epilepsy patients than in fever only controls (p < 0.05). Serum levels of IL-1β were significantly correlated with levels of HMGB1, IL-6, and TNF-α (p < 0.05).ConclusionsHMGB1 and pro-inflammatory cytokines were significantly higher in febrile seizure children. Although it is not possible to infer causality from descriptive human studies, our data suggest that HMGB1 and the cytokine network may contribute to the generation of febrile seizures in children. There may be a potential role for anti-inflammatory therapy targeting cytokines and HMGB1 in preventing or limiting febrile seizures or subsequent epileptogenesis in the vulnerable, developing nervous system of children.

Ultrastructural GABA immunocytochemistry in the mossy fiber terminals of Wistar and genetic absence epileptic rats receiving amygdaloid kindling stimulations

The existence of absence epilepsy and temporal lobe epilepsy in the same patient is not common in clinical practice. The reason why both types of seizures are rarely seen in the same patient is not well understood. Therefore, we aimed to investigate kindling in a well known model of human absence epilepsy, genetic absence epilepsy rats from Strasbourg (GAERS). In the present study, we analyzed whether the GABA content of GAERS that received kindling stimulations was altered in the hippocampal mossy fiber terminals compared to non-epileptic control (NEC) Wistar rats. For this purpose, we used an immunocytochemical technique at the ultrastructural level. Ultrathin sections were immunolabeled with anti-GABA antibody and transmission electron microscopy was used for the ultrastructural examination. The number of gold particles per nerve terminal was counted and the area of the nerve terminal was determined using NIH image analysis program. The GABA density was found to be higher in sham-operated GAERS than sham-operated Wistar rats. The density was increased in kindling Wistar group compared to sham-operated Wistar and kindling GAERS groups. No statistical difference was observed between sham-operated GAERS and kindling GAERS groups. The increase in GABA levels in stimulated Wistar rats may be a result of a protective mechanism. Furthermore, there may be strain differences between Wistar rats and GAERS and our findings addressing different epileptogenesis mechanisms in these strains might be a basis for future experimental studies.