Injection Of Kainic Acid Research Articles

Phenotypic differences based on lateralization of intrahippocampal kainic acid injection in female mice

Clinical evidence indicates that patients with temporal lobe epilepsy (TLE) often show differential outcomes of comorbid conditions in relation to the lateralization of the seizure focus. A particularly strong relationship exists between the side of seizure focus and the propensity for distinct reproductive endocrine comorbidities in women with TLE. Therefore, here we evaluated whether targeting of left or right dorsal hippocampus for intrahippocampal kainic acid (IHKA) injection, a model of TLE, produces different outcomes in hippocampal granule cell dispersion, body weight gain, and multiple measures of reproductive endocrine dysfunction in female mice. One, two, and four months after IHKA or saline injection, in vivo measurements of estrous cycles and weight were followed by ex vivo examination of hippocampal dentate granule cell dispersion, circulating ovarian hormone and corticosterone levels, ovarian morphology, and pituitary gene expression. IHKA mice with right-targeted injection (IHKA-R) showed greater granule cell dispersion and pituitary Fshb expression compared to mice with left-targeted injection (IHKA-L). By contrast, pituitary expression of Lhb and Gnrhr were higher in IHKA-L mice compared to IHKA-R, but these values were not different from respective saline-injected controls. IHKA-L mice also showed an increased rate of weight gain compared to IHKA-R mice. Increases in estrous cycle length, however, were similar in both IHKA-L and IHKA-R mice. These findings indicate that although major reproductive endocrine dysfunction phenotypes present similarly after targeting left or right dorsal hippocampus for IHKA injection, distinct underlying mechanisms based on lateralization of epileptogenic insult may contribute to produce similar emergent reproductive endocrine outcomes.

Temporal and spatial changes in synaptic vesicle glycoprotein 2A (SV2A) under kainic acid induced epileptogenesis: An autoradiographic study

Synaptic Vesicle Glycoprotein 2A (SV2A) has been proposed as a presynaptic marker in several neurological disorders. Not only is SV2A the target for the antiepileptic drug levetiracetam, but also considered a marker of mature pre-synapses. In this study, we aimed to assess the binding of [3H]UCB-J as a selective radioligand for SV2A to visualize and determine changes during different stages of epileptogenesis by in-vitro autoradiography in rat models of temporal lobe epilepsy. Two different kainic acid (KA) injection routes were used to model temporal lobe epilepsy in the rat; a systemic (10 mg/kg KA injected intraperitoneally) and a local model (1.875 mM KA injected intrahippocampally). Brain tissue was sampled at different time points after the initial status epilepticus and semi-quantitative [3H]UCB-J autoradiography was performed to determine temporal and spatial changes under the progression of epileptogenesis. A decrease in [3H]UCB-J binding was observed in many brain areas in the acute phases after both types of kainic acid administration. Peak reductions occurred slightly before in systemic-treated animals (within 3–10 days) than after local-treated animals (within 5–15 days). Interestingly in the systemic model, we observed a full restoration in the binding level 30 days after the treatment in most areas probably reflecting neuronal reorganization. However, after the local injection in the hippocampus, the binding in the hippocampus, and in temporal and piriform cortices did not return to basal levels. The time-course profile displayed lateralization in the local model. These results demonstrate changes in the amount of a presynaptic SV2A binding site after seizures and suggest that SV2A may have importance in eliciting spontaneous seizures and/or be a biomarker for epileptogenesis. The present study shows that SV2A is a biomarker of acute phase epileptogenesis in specific brain regions.

Piperine Provides Neuroprotection against Kainic Acid-Induced Neurotoxicity via Maintaining NGF Signalling Pathway.

The neuroprotective properties of piperine, the major alkaloid extracted from black pepper, have been under investigation, but its mechanism of action in excitotoxicity is still poorly understood. This study aimed to evaluate the protective effects of piperine with a focus on nerve growth factor (NGF) signalling in a kainic acid (KA) rat model of excitotoxicity. Rats were administered intraperitoneally (i.p.) piperine (10 or 50 mg/kg) before KA injection (15 mg/kg, i.p.). Our results show that KA exposure in rats caused seizure behaviour, intrinsic neuronal hyperactivity, glutamate elevation, hippocampal neuronal damage, and cognitive impairment. These KA-induced alterations could be restored to the normal state by piperine treatment. In addition, piperine decreased the expression of the NGF precursor proNGF and NGF-degrading protease matrix metalloproteinase 9, whereas it increased the expression of proNGF processing enzyme matrix metalloproteinase 7, NGF, and NGF-activated receptor TrkA in the hippocampus of KA-treated rats. Furthermore, KA decreased phosphorylation of the protein kinase B (Akt) and glycogen synthase kinase 3β (GSK3β) in the hippocampus, and piperine reversed these changes. Our data suggest that piperine protects hippocampal neurons against KA-induced excitotoxicity by upregulating the NGF/TrkA/Akt/GSK3β signalling pathways.

Antiseizure Effects of Cannabidiol Leading to Increased Peroxisome Proliferator-Activated Receptor Gamma Levels in the Hippocampal CA3 Subfield of Epileptic Rats.

We evaluated the effects of cannabidiol (CBD) on seizures and peroxisome proliferator-activated receptor gamma (PPARγ) levels in an animal model of temporal lobe epilepsy (TLE). Adult male Sprague-Dawley rats were continuously monitored by video-electrocorticography up to 10 weeks after an intraperitoneal kainic acid (15 mg/kg) injection. Sixty-seven days after the induction of status epilepticus and the appearance of spontaneous recurrent seizures in all rats, CBD was dissolved in medium-chain triglyceride (MCT) oil and administered subcutaneously at 120 mg/kg (n = 10) or 12 mg/kg (n = 10), twice a day for three days. Similarly, the vehicle was administered to ten epileptic rats. Brain levels of PPARγ immunoreactivity were compared to those of six healthy controls. CBD at 120 mg/kg abolished the seizures in 50% of rats (p = 0.033 vs. pre-treatment, Fisher’s exact test) and reduced total seizure duration (p < 0.05, Tukey Test) and occurrence (p < 0.05). PPARγ levels increased with CBD in the hippocampal CA1 subfield and subiculum (p < 0.05 vs. controls, Holm–Šidák test), but only the highest dose increased the immunoreactivity in the hippocampal CA3 subfield (p < 0.001), perirhinal cortex, and amygdala (p < 0.05). Overall, these results suggest that the antiseizure effects of CBD are associated with upregulation of PPARγ in the hippocampal CA3 region.

Neferine, an Alkaloid from Lotus Seed Embryos, Exerts Antiseizure and Neuroprotective Effects in a Kainic Acid-Induced Seizure Model in Rats.

Current anti-seizure drugs fail to control approximately 30% of epilepsies. Therefore, there is a need to develop more effective anti-seizure drugs, and medicinal plants provide an attractive source for new compounds. This study aimed to evaluate the possible anti-seizure and neuroprotective effects of neferine, an alkaloid from the lotus seed embryos of Nelumbo nucifera, in a kainic acid (KA)-induced seizure rat model and its underlying mechanisms. Rats were intraperitoneally (i.p.) administrated neferine (10 and 50 mg/kg) 30 min before KA injection (15 mg/kg, i.p.). Neferine pretreatment increased seizure latency and reduced seizure scores, prevented glutamate elevation and neuronal loss, and increased presynaptic protein synaptophysin and postsynaptic density protein 95 expression in the hippocampi of rats with KA. Neferine pretreatment also decreased glial cell activation and proinflammatory cytokine (interleukin-1β, interleukin-6, tumor necrosis factor-α) expression in the hippocampi of rats with KA. In addition, NOD-like receptor 3 (NLRP3) inflammasome, caspase-1, and interleukin-18 expression levels were decreased in the hippocampi of seizure rats pretreated with neferine. These results indicated that neferine reduced seizure severity, exerted neuroprotective effects, and ameliorated neuroinflammation in the hippocampi of KA-treated rats, possibly by inhibiting NLRP3 inflammasome activation and decreasing inflammatory cytokine secretion. Our findings highlight the potential of neferine as a therapeutic option in the treatment of epilepsy.

Seizure activity triggers tau hyperphosphorylation and amyloidogenic pathways.

Although epilepsies and neurodegenerative disorders show pathophysiological similarities, their direct functional associations are unclear. Here, we tested the hypothesis that experimental seizures can induce tau hyperphosphorylation and amyloidogenic modifications over time, with intersections with neuroinflammation. We used a model of mesial temporal lobe epilepsy (MTLE) where unilateral intrahippocampal injection of kainic acid (KA) in C57BL/6mice elicits epileptogenesis and spontaneous focal seizures. We used a model of generalized status epilepticus (SE) obtained by intraperitoneal KA injection in C57BL/6mice. We performed analyses and cross-comparisons according to a schedule of 72 h, 1 week, and 8 weeks after KA injection. In experimental MTLE, we show AT100, PHF1, and CP13 tau hyperphosphorylation during epileptogenesis (72h-1week) and long-term (8weeks) during spontaneous seizures in the ipsilateral hippocampi, the epileptogenic zone. These pathological modifications extended to the contralateral hippocampus, a seizure propagating zone with no histological lesion or sclerosis. Two kinases, Cdk5 and GSK3β, implicated in the pathological phosphorylation of tau, were activated. In this MTLE model, the induction of the amyloidogenic pathway (APP, C99, BACE1) was prominent and long-lasting in the epileptogenic zone. These Alzheimer's disease (AD)-relevant markers, established during seizure progression and recurrence, reciprocated an enduring glial (GFAP, Iba1) inflammation and the inadequate activation of the endogenous, anti-inflammatory, glucocorticoid receptor system. By contrast, a generalized SE episode provoked a predominantly transient induction of tau hyperphosphorylation and amyloidogenic markers in the hippocampus, along with resolving inflammation. Finally, we identified overlapping profiles of long-term hippocampal tau hyperphosphorylation by comparing MTLE to J20mice, the latter a model relevant to AD. MTLE and a generalized SE prompt persistent and varying tau hyperphosphorylation or amyloidogenic modifications in the hippocampus. In MTLE, an AD-relevant molecular trajectory intertwines with neuroinflammation, spatiotemporally involving epileptogenic and nonlesional seizure propagating zones.

Fibroblast growth factor 13 is involved in the pathogenesis of temporal lobe epilepsy.

Temporal lobe epilepsy (TLE) is the most common drug-resistant epilepsy in adults, with pathological mechanisms remaining to be fully elucidated. Fibroblast Growth Factor 13 (FGF13) encodes an intracellular protein involved in microtubule stabilization and regulation of voltage-gated sodium channels (VGSCs) function. FGF13 mutation has been identified in patients with inherent seizure, suggesting a potential association between FGF13 and the etiology of TLE. Here, we set to explore the pathological role of FGF13 in the etiology of TLE. We found that the expression of FGF13 was increased in the cortical lesions and CA1 region of sclerotic hippocampus and correlated with the seizure frequency in TLE patients. Also, Fgf13 expression was increased in the hippocampus of chronic TLE mice generated by kainic acid (KA) injection. Furthermore, Fgf13 knockdown or overexpression was respectively found to attenuate or potentiate the effects of KA on axonal length, somatic area and the VGSCs-mediated current in the hippocampal neurons. Taken together, these findings suggest that FGF13 is involved in the pathogenesis of TLE by modulating microtubule activity and neuronal excitability.

Pathological Human Tau Induces Alterations in the Brain Insulin Signaling Cascade.

The process of neurodegeneration in Alzheimer’s disease has been associated with a disruption of insulin signaling cascade in neurons, and to insulin resistance. T2DM correlates with Alzheimer’s disease, but mechanisms of interaction are unknown. We have developed a mouse model of tau induced neurodegeneration expressing pseudo-phosphorylated tau [Pathological Human Tau (PH-Tau)] in neurons. This model (PH-Tau-Tg) recapitulated cognitive decline and neurodegeneration observed in AD. In this study we examined if expression of PH-Tau could affect neuronal excitability and insulin receptor signaling. Neuronal excitability was investigated using intracerebral recordings of extracellular field potentials from prefrontal cortex after insulin and kainic acid (KA) injection. Analysis of baseline recordings indicated an increased excitability of PH-Tau-Tg as evidenced by higher spectrum densities (PSDs) of high frequencies brain waves. Injection of insulin (1IU, s.c) led to a decrease of fast ripples PSDs, more pronounced in PH-Tau-Tg mice than controls. Subsequent injection of kainic acid (KA, 5 mg/kg, s.c) led to significant increase in firing rate, amplitude of extracellular field potentials and PSDs of high frequency brain waves in control mice only. To further investigate the role of insulin in PH-Tau-Tg mice, we subjected mice to a glucose tolerance test. We found that PH-Tau-Tg mice were significantly hyperglycemic prior to glucose injection. Interestingly, the PH-Tau-Tg mice showed a moderate increase at 30 min due to the higher baseline, indicating a low sensitivity of insulin receptor in these mice. This is consistent with increased levels of activated insulin receptors in the brain and the inhibitory effect of insulin on ictal activity post KA injection in PH-Tau-Tg mice. We suggest that these mice have reduced insulin sensitivity (hyperglycemia) and as a compensatory mechanism there is overactivation/expression of insulin receptor in the brain rendering neuronal circuits resistant to seizure induction after injection of insulin. These data indicate that insulin signal transduction pathway is altered in PH-Tau-Tg mice, and that injection of exogenous insulin reduces hypersynchronous bursting activity of field potentials recorded from cortical neuronal circuits. We propose that the appearance of abnormal tau might potentiate the toxic environment by interfering with the insulin signaling cascade in the brain of patients with Alzheimer’s disease.

18F]DPA-714 PET imaging for the quantitative evaluation of early spatiotemporal changes of neuroinflammation in rat brain following status epilepticus.

Most antiepileptic drug therapies are symptomatic and adversely suppress normal brain function by nonspecific inhibition of neuronal activity. In recent times, growing evidence has suggested that neuroinflammation triggered by epileptic seizures might be involved in the pathogenesis of epilepsy. Although the potential effectiveness of anti-inflammatory treatment for curing epilepsy has been extensively discussed, the limited quantitative data regarding spatiotemporal characteristics of neuroinflammation after epileptic seizures makes it difficult to be realized. We quantitatively analyzed the spatiotemporal changes in neuroinflammation in the early phase after status epilepticus in rats, using translocator protein (TSPO) positron emission tomography (PET) imaging, which has been widely used for the quantitative evaluation of neuroinflammation in several animal models of CNS disease. The second-generation TSPO PET probe, [18F]DPA-714, was used for brain-wide quantitative analysis of neuroinflammation in the brains of rats, when the status epilepticus was induced by subcutaneous injection of kainic acid (KA, 15 mg/kg) into those rats. A series of [18F]DPA-714 PET scans were performed at 1, 3, 7, and 15 days after status epilepticus, and the corresponding histological changes, including activation of microglia and astrocytes, were confirmed by immunohistochemistry. Apparent accumulation of [18F]DPA-714 was observed in several KA-induced epileptogenic regions, such as the amygdala, piriform cortex, ventral hippocampus, mediodorsal thalamus, and cortical regions 3 days after status epilepticus, and was reversibly displaced by unlabeled PK11195 (1 mg/kg). Consecutive [18F]DPA-714 PET scans revealed that accumulation of [18F]DPA-714 was focused in the KA-induced epileptogenic regions from 3 days after status epilepticus and was further maintained in the amygdala and piriform cortex until 7 days after status epilepticus. Immunohistochemical analysis revealed that activated microglia but not reactive astrocytes were correlated with [18F]DPA-714 accumulation in the KA-induced epileptogenic regions for at least 1 week after status epilepticus. These results indicate that the early spatiotemporal characteristics of neuroinflammation quantitatively evaluated by [18F]DPA-714 PET imaging provide valuable evidence for developing new anti-inflammatory therapies for epilepsy. The predominant activation of microglia around epileptogenic regions in the early phase after status epilepticus could be a crucial therapeutic target for curing epilepsy.

Loss of efferent projections of the hippocampal formation in the mouse intrahippocampal kainic acid model

Unilateral intrahippocampal injection of kainic acid is used as a model of medial temporal lobe epilepsy and provides a platform to study the mechanisms of epilepsy. Here, we used an AAV-9 EYFP-tagged viral vector as an anterograde tracer, injected into the dorsal and ventral hippocampus after kainic acid injection, to map out the efferent hippocampal projections after the development of spontaneous seizures in this model. The purpose of the study was to identify the extent of changes in hippocampal efferent system in several brain regions that receive significant inputs from the hippocampus. Loss of efferent hippocampal fibers was greatest in the retrosplenial cortex where neuronal loss was also observed. Loss of fibers was also observed in the fornix without any specific effect in the lateral mammillary nuclei. Although expected, these observations provide further evidence of the broader network effects as a result of hippocampal cell loss.

Neuroprotective effects of tannic acid against kainic acid-induced seizures in mice.

Epileptic seizures are associated with the release of potentially neurotoxic amount of glutamate, which results in the over-production of free radicals and inflammatory factors, and induction of neuronal cell death. Current study evaluated the effect of tannic acid (TA) on Kainic acid (KA)-induced seizures in mice. Mice were divided into the six groups. Group I was administrated with normal saline (NS; 1mL/kg, intraperitoneally (i.p.)), Group II was injected with KA (15mg/kg, i.p.), Groups III was treated with diazepam (DZ; 20mg/kg, i.p.) and KA (15mg/kg, i.p.), Groups IV-VI were treated with TA (25, 50 and 100mg/kg, i.p.) and KA (15mg/kg, i.p.). Animals received all treatments 30min before injection of KA. After the injection of KA, mice were observed for seizure (latency, activity and duration) and mortality for 2h. In the brain tissue, oxidative stress, apoptosis, and inflammatory markers were evaluated in addition to the determination of histological alterations in the CA1 molecular layer of hippocampus. Treatment with TA significantly increased seizure latency and decreased seizure duration and activity, but could not significantly decrease mice mortality. This effect was associated with the reduction of oxidative stress, inflammation, and apoptosis. Furthermore, treatment with TA significantly improved KA-induced pyramidal cell loss and change in the arrangement of CA1 molecular layer. Tannic acid may be useful in the control of epileptic seizures through regulating oxidative stress, inflammation and apoptosis.

Electrophysiological Evidence for Anti-epileptic Property of Taurine.

Taurine, 2-aminoethanesulfonic acid, is one of the most abundant free amino acids especially in excitable tissues, with wide physiological actions. Several lines of evidence suggest that taurine may function as a potent inhibitory neuromodulator that regulate neuronal activity in many cerebral areas. Parenteral injection of kainic acid (KA), a glutamate receptor agonist, causes severe and stereotyped behavioral convulsions in mice and is used as a rodent model for human temporal lobe epilepsy. In the adult brain, inhibitory GABAergic interneurons modulate the activity of principal excitatory cells via their GABAA receptors and thus adjust excitatory output of neuronal circuits. The goal of this study was to examine the potential anti-convulsive effects of the neuro-active amino acid taurine, in the mouse model of limbic seizures. We used the glutamic acid decarboxylase (GAD) inhibitor isoniazid (100mg.kg-1, s.c.) which induces seizures by interfering with GABA synthesis through inhibition of GAD activity followed by kainic acid (5mg.kg-1, s.c.) a glutamate receptor agonist which is commonly used to induce limbic seizures.Using intracerebral recordings of field potentials found that taurine (43mg.kg-1, s.c.) had a significant anti-epileptic effect when injected prior to isoniazid and KA. Furthermore, injection of taurine to a mouse undergoing limbic seizure completely stopped burst population spikes and restored neuronal firing to its baseline. Therefore, taurine is potentially capable of treating seizure-associated brain damage.

Altered Expression of Par3, aPKC-λ, and Lgl1 in Hippocampus in Kainic Acid-Induced Status Epilepticus Rat Model

Introduction: Mossy fiber sprouting (MFS) is a frequent histopathological finding in temporal lobe epilepsy (TLE) and is involved in the pathology of TLE. However, molecular signals underlying MFS remain unclear. Partitioning defective 3(Par3), atypical protein kinase C-λ(aPKC-λ), and lethal giant larvae 1(Lgl1) were involved in the neuronal polarity and axon growth. The potential roles of those proteins in MFS and epileptogenesis of TLE were investigated.Material and Methods: The epileptic rat models were established by intracerebroventricular injection of kainic acid (KA). The degree of MFS was measured by using Timm staining, Neuronal loss and the expression aPKC-λ, Par3, and Lgl1 in hippocampus were measured by using immunohistochemistry and western blot analysis.Results: The neuronal loss in CA3 region was observed from 3 days to 8 weeks, while the neuronal loss in the hilar region was observed from 1 to 8 weeks in experimental group. The Timm score in the CA3 region in experimental group was significantly higher than that in the control group from 2 to 8 weeks. Compared with control group, the expressions of Par3 and Lgl1 were upregulated and the expression of aPKC-λ was downregulated in the experimental groups. Positive correlation between the Par3 expression and Timm scores, and the negative correlation between the aPKC-λ expression and Timm scores in CA3 region were discovered in experimental group.Conclusion: The findings of the present study indicated that aPKC-λ, Par3, and Lgl1 may be involved in MFS and in the epileptogenesis of temporal lobe epilepsy.

TIGAR plays neuroprotective roles in KA-induced excitotoxicity through reducing neuroinflammation and improving mitochondrial function

Excitotoxicity refers to the ability of excessive extracellular excitatory amino acids to damage neurons via receptor activation. It is a crucial pathogenetic process in neurodegenerative diseases. TP53 is confirmed to be involved in excitotoxicity. It is demonstrated that TP53 induced glycolysis and apoptotic regulator (TIGAR)-regulated metabolic pathway can protect against neuronal injury. However, the role of TIGAR in excitotoxicity and specific mechanisms is still unknown. In this study, an in vivo excitotoxicity model was constructed via stereotypical kainic acid (KA) injection into the striatum of mice. KA reduced TIGAR expression levels, neuroinflammatory responses and mitochondrial dysfunction. TIGAR overexpression could reverse KA-induced neuronal injury by reducing neuroinflammation and improving mitochondrial function, thereby exerting neuroprotective effects. Therefore, this study could provide a potential therapeutic target for neurodegenerative diseases.

Intravenous kainic acid induces status epilepticus and late onset seizures in mice

We set out to establish a novel model of temporal lobe epilepsy (TLE) in a mouse. We sought to induce TLE through the injection of kainic acid (KA) into the tail vein with subsequent development of status epilepticus (SE). Using C57BL/6 mice, we implanted hippocampal EEG recording electrodes before or after injection of KA or phosphate buffered saline (PBS). Video and EEG analysis were conducted to evaluate for SE and development of recurrent seizures, the hallmark of TLE. All mice injected with KA developed SE while those who were injected with PBS did not. Of the animals injected with KA monitored for recurrent seizures following SE, 33% developed spontaneous recurrent seizures while those injected with PBS did not. Injection of KA through the tail vein of a mouse reliably and rapidly induces SE which remits spontaneously and leads to the development of TLE in a subset of mice.

Hippocampal injury and learning deficits following non-convulsive status epilepticus in periadolescent rats

The effects of non-convulsive status epilepticus (NCSE) on the developing brain remain largely elusive. Here we investigated potential hippocampal injury and learning deficits following one or two episodes of NCSE in periadolescent rats. Non-convulsive status epilepticus was induced with subconvulsive doses of intrahippocampal kainic acid (KA) under continuous EEG monitoring in postnatal day 43 (P43) rats. The RKA group (repeated KA) received intrahippocampal KA at P43 and P44, the SKA group (single KA injection) received KA at P43 and an intrahippocampal saline injection at P44. Controls were sham-treated with saline. The modified two-way active avoidance (MAAV) test was conducted between P45 and P52 to assess learning of context-cued and tone-signaled electrical foot-shock avoidance. Histological analyses were performed at P52 to assess hippocampal neuronal densities, as well as potential reactive astrocytosis and synaptic dysfunction with GFAP (glial fibrillary acidic protein) and synaptophysin (Syp) staining, respectively. Kainic acid injections resulted in electroclinical seizures characterized by behavioral arrest, oromotor automatisms and salivation, without tonic-clonic activity. Compared to controls, both the SKA and RKA groups had lower rates of tone-signaled shock avoidance (p < 0.05). In contextual testing, SKA rats were comparable to controls (p > 0.05), but the RKA group had learning deficits (p < 0.05). Hippocampal neuronal densities were comparable in all groups. Compared to controls, both the SKA and RKA groups had higher hippocampal GFAP levels (p < 0.05). The RKA group also had lower hippocampal Syp levels compared to the SKA and control groups (p < 0.05), which were comparable (p > 0.05). We show that hippocampal NCSE in periadolescent rats results in a seizure burden-dependent hippocampal injury accompanied by cognitive deficits. Our data suggest that the diagnosis and treatment of NCSE should be prompt.

Pericardial Injection of Kainic Acid Induces a Chronic Epileptic State in Larval Zebrafish.

Epilepsy is a common disorder of the brain characterized by spontaneous recurrent seizures, which develop gradually during a process called epileptogenesis. The mechanistic processes underlying the changes of brain tissue and networks toward increased seizure susceptibility are not fully understood. In rodents, injection of kainic acid (KA) ultimately leads to the development of spontaneous epileptic seizures, reflecting similar neuropathological characteristics as seen in patients with temporal lobe epilepsy (TLE). Although this model has significantly contributed to increased knowledge of epileptogenesis, it is technically demanding, costly to operate and hence not suitable for high-throughput screening of anti-epileptic drugs (AEDs). Zebrafish, a vertebrate with complementary advantages to rodents, is an established animal model for epilepsy research. Here, we generated a novel KA-induced epilepsy model in zebrafish larvae that we functionally and pharmacologically validated. KA was administered by pericardial injection at an early zebrafish larval stage. The epileptic phenotype induced was examined by quantification of seizure-like behavior using automated video recording, and of epileptiform brain activity measured via local field potential (LFP) recordings. We also assessed GFP-labeled GABAergic and RFP-labeled glutamatergic neurons in double transgenic KA-injected zebrafish larvae, and examined the GABA and glutamate levels in the larval heads by liquid chromatography with tandem mass spectrometry detection (LC-MS/MS). Finally, KA-injected larvae were exposed to five commonly used AEDs by immersion for pharmacological characterization of the model. Shortly after injection, KA induced a massive damage and inflammation in the zebrafish brain and seizure-like locomotor behavior. An abnormal reorganization of brain circuits was observed, a decrease in both GABAergic and glutamatergic neuronal population and their associated neurotransmitters. Importantly, these changes were accompanied by spontaneous and continuous epileptiform brain discharges starting after a short latency period, as seen in KA rodent models and reminiscent of human pathology. Three out of five AEDs tested rescued LFP abnormalities but did not affect the seizure-like behavior. Taken together, for the first time we describe a chemically-induced larval zebrafish epilepsy model offering unique insights into studying epileptogenic processes in vivo and suitable for high-throughput AED screening purposes and rapid genetic investigations.

TWIK-1 BAC-GFP Transgenic Mice, an Animal Model for TWIK-1 Expression.

TWIK-1 is the first identified member of the two-pore domain potassium (K2P) channels that are involved in neuronal excitability and astrocytic passive conductance in the brain. Despite the physiological roles of TWIK-1, there is still a lack of information on the basic expression patterns of TWIK-1 proteins in the brain. Here, using a modified bacterial artificial chromosome (BAC), we generated a transgenic mouse (Tg mouse) line expressing green fluorescent protein (GFP) under the control of the TWIK-1 promoter (TWIK-1 BAC-GFP Tg mice). We confirmed that nearly all GFP-producing cells co-expressed endogenous TWIK-1 in the brain of TWIK-1 BAC-GFP Tg mice. GFP signals were highly expressed in various brain areas, including the dentate gyrus (DG), lateral entorhinal cortex (LEC), and cerebellum (Cb). In addition, we found that GFP signals were highly expressed in immature granule cells in the DG. Finally, our TWIK-1 BAC-GFP Tg mice mimic the upregulation of TWIK-1 mRNA expression in the hippocampus following the injection of kainic acid (KA). Our data clearly showed that TWIK-1 BAC-GFP Tg mice are a useful animal model for studying the mechanisms regulating TWIK-1 gene expression and the physiological roles of TWIK-1 channels in the brain.

A New Therapy of Epilepsy by Enhancing Inhibition of Gabaergic Neurons

Epilepsy is a neural disorder related to the imbalance between inhibition and excitation of the brain. Enhancement of inhibition can be used to treat epilepsy. Here, we discovered that the inhibition of cortical GABAergic neurons was potentiated after high-frequency optogenetic activation but not low-frequency. GABAergic neurons were activated chemogenetically to fire at high frequency, suppressing spontaneous neuronal activities and auditory responses in the long term during the 2-hour recording. We produced chronic epileptic mice after the intrahippocampal injection of kainic acid. Viral vectors of hM3Dq were delivered intravenously to infect the inhibitory neurons in our epileptic mice. We developed an artificial-intelligent program to objectively detect each convulsive seizure before, during, and after the chemogenetic treatment. Administration of clozapine-N-oxide suppressed the epileptic activities during and even more the week after drug application. The results demonstrate a novel gene therapy for chronic epilepsy by enhancing the inhibition of global GABAergic neurons.

Characterization of metabolic activity induced by kainic acid in adult rat whole brain at the early stage: A 18FDG-PET study

ObjectiveBrain metabolic processes are not fully characterized in the kainic acid (KA)-induced Status Epilepticus (KASE). Thus, we evaluated the usefulness of 18F-fluorodeoxyglucose positron emission tomography (FDG-PET) as an experimental strategy to evaluate in vivo, in a non-invasive way, the glucose consumption in several brain regions, in a semi-quantitative study to compare and to correlate with data from electroencephalography and histology studies. Methods.Sixteen male Wistar rats underwent FDG-PET scans at basal state and after KA injection. FDG-PET images were normalized to an MRI-based atlas and segmented to locate regions. Standardized uptake values (SUV) were obtained at several time points. EEGs and cell viability by histological analysis, were also evaluated. ResultsFDG-PET data showed changes in regions such as: amygdala, hippocampus, accumbens, entorhinal cortex, motor cortex and hypothalamus. Remarkably, hippocampal hypermetabolism was found (mean SUV = 2.66 ± 0.057) 2 h after KA administration, while hypometabolism at 24 h (mean SUV = 1.83 ± 0.056) vs basal values (mean SUV = 2.19 ± 0.057). EEG showed increased spectral power values 2 h post-KA administration. Hippocampal viable-cell counting 24 h after KA was decreased, while Fluoro-Jade B-positive cells were increased, as compared to control rats, coinciding with the hypometabolism detected in the same region by semi-quantitative FDG-PET at 24 h after KASE. ConclusionsPET is suitable to measure metabolic brain changes in the rat model of status epilepticus induced by KA (KASE) at the first 24 h, compared to that of EEG; PET data may also be sensitive to cell viability.