Anticonvulsant effects of Paeonia daurica subsp. macrophylla root extracts in pentylenetetrazol-induced seizure models in mice.

INTRODUCTION: In the present study, anticonvulsant effects of aqueous extract (AE), hydro-alcoholic crude extract (HE), and its fractions (F-CHCl3, F-EtOAc, F-MeOH) of Paeonia daurica subsp. macrophylla (P. daurica ssp. macrophylla) root examined by using a pentylenetetrazol-induced model (PTZ) on mice. METHODS: HE and its fractions as well as AE, in concentrations of (100, 200 and 400mg/kg), valproate (Val) (100 and 200mg/kg), and saline (negative control) (10mg/kg) were injected intraperitoneally (i.p.) 30min before PTZ (80mg/kg, i.p.). The time taken before the onset of myoclonic convulsions (MC), MC duration, time taken before the onset of generalized tonic-clonic seizures (GTCS), the duration of GTCS, and the percentage of GTCS and mortality protection recorded. The plant's anticonvulsant mechanisms were assessed using flumazenil (5mg/kg, i.p.) before AE (100, 200, and 400mg/kg, i.p.) injection. GraphPad Prism software was used to compare the differences between various treatment groups with one-way analysis of variance (ANOVA) followed by Tukey-Krammer multiple comparison tests. RESULTS: All the plant samples except F-EtOAc significantly delayed the onset and decreased the duration of PTZ-induced MCS and GTCS, and significantly reduced the GTCS and mortality rate. Pretreatment with flumazenil diminished the significant anticonvulsant effects of AE against PTZ-induced seizures. CONCLUSIONS: It can report that extract of P. daurica ssp. macrophylla might be a helpful guide for future studies in the treatment of epilepsy.

Caffeine Decreases Topiramate Levels in Zebrafish Larvae in a Pentylenetetrazol-Induced Seizure Model.

Epilepsy ranks as the second-most prevalent neurological disease, and is characterized by seizures resulting in neurobiological and behavioral impairment. Naturally occurring in coffee beans or tea leaves, the alkaloid caffeine (CAF) is the most prevalent global stimulant. Caffeine has been observed to influence epileptic seizures and the efficacy of antiepileptic medications, with a notable impact on topiramate (TPM). This study aimed to explore the influence of CAF on TPM's anticonvulsant effects in zebrafish larvae within a PTZ-induced seizure model, concurrently determining TPM concentrations through a sophisticated analytical approach based on ultrahigh-performance liquid chromatography and subsequent mass spectrometric detection. Zebrafish larvae four days post-fertilization were incubated for 18 h with varying doses of TPM or combinations of CAF + TPM, and locomotor activity was then assessed. Seizures were induced by introducing a PTZ solution to achieve a final concentration of 20 mM. Utilizing liquid chromatography-mass spectrometry (LC-MS/MS), TPM levels in the larvae were quantified. CAF co-administration (especially in higher doses) with TPM caused a decrease in the average locomotor activity in the larvae compared to TPM alone. Moreover, CAF decreased TPM levels in the larvae at all investigated doses. In conclusion, these findings offer a novel perspective on the interplay between CAF and TPM, shedding light on previously unexplored facets. The potential impact of CAF consumption in assisting with epileptic seizure control, unless proven otherwise, suggests a noteworthy consideration for future research and clinical practices.

Therapeutic potential of CB1R activation by Qingyangshen glycoside M1 for seizure relief.

ETHNOPHARMACOLOGICAL RELEVANCE: Cynanchum otophyllum C.K.Schneid.PI.Wilson, commonly referred as ''Qingyangshen'' (QYS), is a traditional folk medicine from Yunnan, renowned for its efficacy in neurological and psychiatric disorders. Glycosides isolated from QYS have shown promise in alleviating epilepsy, however, mechanisms of action and specific molecular targets remain to be elucidated. AIM OF THE STUDY: The study aimed to evaluate the anticonvulsant effects of Qingyangshen glycosides M1 (M1), a C21 steroidal glycoside from QYS, on pentylenetetrazol (PTZ)-induced convulsions in zebrafish (Danio rerio), and its neuroprotective effect on Glutamate (Glu)-induced damage to PC12 cells, and importantly to identify its potential molecular targets. MATERIALS AND METHODS: To evaluate anticonvulsant activity of M1, 7 days-post-fertilization (7-dpf) animals were pretreated (by immersion) and then exposed to PTZ (10 mM) solution. Furthermore, Glu-induced PC12 cell damage was employed to investigate the neuroprotective and anti-apoptotic capacity. Cells were pretreated with various concentrations of M1 (0-10 µM) for 12 h and then co-treated with Glu (15 mM) for an additional 24 h. The cell viability, apoptosis rate and apoptosis-related proteins (p-PI3K, PI3K, Akt, p-Akt, CREB, p-CREB, BDNF, Bax and Bcl-2) were measured using CCK-8, annexin V/PI and Western blot assays. To model the expected interaction between M1 and candidate cannabinoid receptor type 1 (CB1R), ERK phosphorylation, molecular docking, and drug affinity responsive target stability (DARTS) techniques were employed. Finally, CB1R antagonist Rimonabant (Rim) was validated by co-administration in both zebrafish and cells to confirm the requirement of CB1R for M1 efficacy. RESULTS: At a concentration of 400 µM, M1 dramatically reversed PTZ-induced convulsive-like behaviors in zebrafish, as evidenced by a significant reduction in locomotor activity. In the context of Glu-induced cytotoxicity, M1 (10 µM) demonstrated a notable increase in cell viability and suppressed apoptosis through modulation of the Bax/Bcl-2 ratio and activation of the PI3K/Akt/CREB/BDNF signaling axis. These effects were facilitated through CB1R activation. In contrast, Rim dampened the beneficial activities of M1 as a cannabinoid agonist. CONCLUSIONS: These results demonstrated that M1 as a potential CB1R activator, exhibiting anticonvulsive effects in a PTZ-induced zebrafish model and neuroprotective properties via the PI3K/Akt/CREB/BDNF signaling axis in a Glu-induced PC12 cell injury model. Notably, the observed seizure relief attenuated by CB1R chemical antagonism.

Behavioral and neurotransmitter changes on antiepileptic drugs treatment in the zebrafish pentylenetetrazol-induced seizure model.

Epilepsy, a recurrent neurological disorder involving abnormal neurotransmitter kinetics in the brain, has emerged as a global health concern. The mechanism of epileptic seizures is thought to involve a relative imbalance between excitatory and inhibitory neurotransmitters. Despite the recent advances in clinical and basic research on the pathogenesis of epilepsy, the complex relationship between the neurotransmitter changes and behavior with and without antiepileptic drugs (AEDs) during seizures remains unclear. To investigate the effects of AEDs such as levetiracetam (LEV), carbamazepine (CBZ), and fenfluramine (FFR) on key neurotransmitters in the pentylenetetrazol (PTZ)-induced seizures in adult zebrafish, we examined the changes in glutamic acid, gamma-aminobutyric acid (GABA), serotonin (5-HT), 5-hydroxyindoleacetic acid (5-HIAA), choline, acetylcholine, norepinephrine, dopamine (DA), 3,4-dihydroxyphenylacetic acid (DOPAC), and adenosine. In this study, we observed that 5-HT and DA levels in the brain increased immediately after PTZ-induced seizures. Behavioral tests clearly showed that all of these AEDs suppressed the PTZ-induced seizures. Upon treatment of PTZ-induced seizures with these AEDs, CBZ decreased the glutamic acid and FFR increased the GABA levels; however, no neurotransmitter changes were observed in the brain after LEV administration. Thus, we demonstrated a series of neurotransmitter changes linked to behavioral changes during PTZ-induced epileptic seizures when LEV, CBZ, or FFR were administered. These findings will lead to a more detailed understanding of the pathogenesis of epilepsy associated with behavioral and neurotransmitter changes under AED treatment.

Immature Status Epilepticus Alters the Temporal Relationship between Hippocampal Interictal Epileptiform Discharges and High-frequency Oscillations.

The aim was to investigate the long-term effects of a single episode of immature Status Epilepticus (SE) on the excitability of the septal and temporal hippocampus in vitro, by studying the relationship between interictal-like epileptiform discharges (IEDs) and high-frequency oscillations (HFOs; Ripples, Rs and Fast Ripples, FRs). A pentylenetetrazol-induced Status Epilepticus-(SE)-like generalized seizure was induced at postnatal day 20 in 22 male and female juvenile rats, sacrificed >40 days later to prepare hippocampal slices. Spontaneous IEDs induced by Mg2+-free ACSF were recorded from the CA3 area of temporal (T) or septal (S) slices. Recordings were band-pass filtered off-line revealing Rs and FRs and a series of measurements were conducted, with mean values compared with those obtained from age-matched controls (CTRs). In CTR S (vs T) slices, we recorded longer R & FR durations, a longer HFO-IED temporal overlap, higher FR peak power and more frequent FR initiation preceding IEDs (% events). Post-SE, in T slices all types of events duration (IED, R, FR) and the time lag between their onsets (R-IED, FR-IED, R-FR) increased, while FR/R peak power decreased; in S slices, the IED 1st population spike and the FR amplitudes, the R and FR peak power and the (percent) events where Rs or FRs preceded IEDs all decreased. The CA3 IED-HFO relationship offers insights to the septal-to-temporal synchronization patterns; its post-juvenile-SE changes indicate permanent modifications in the septotemporal excitability gradient. Moreover, these findings are in line to region-specific regulation of various currents post-SE, as reported in literature.

Luteolin ameliorates pentetrazole-induced seizures through the inhibition of the TLR4/NF-κB signaling pathway.

Epilepsy represents a prevalent neurological disorder in the population, and the existing antiepileptic drugs (AEDs) often fail to adequately control seizures. Inflammation is recognized as a pivotal factor in the pathophysiology of epilepsy. Luteolin, a natural flavonoid extract, possesses anti-inflammatory properties and exhibits promising neuroprotective activity. Nevertheless, the precise molecular mechanisms underlying the antiepileptic effects of luteolin remain elusive. In this study, we established a rat model of epilepsy using pentylenetetrazole (PTZ) to induce seizures. A series of behavioral experiments were conducted to assess behavioral abilities and cognitive function. Histological techniques, including HE staining, Nissl staining, and TUNEL staining, were employed to assess hippocampal neuronal damage. Additionally, Western blotting, RT-qPCR, and ELISA were utilized to analyze the expression levels of proteins involved in the TLR4/IκBα/NF-κB signaling pathway, transcription levels of apoptotic factors, and levels of inflammatory cytokines, respectively. Luteolin exhibited a dose-dependent reduction in seizure severity, prolonged the latency period of seizures, and shortened seizure duration. Furthermore, luteolin prevented hippocampal neuronal damage in PTZ-induced epileptic rats and partially restored behavioral function and learning and memory abilities. Lastly, PTZ kindling activated the TLR4/IκBα/NF-κB pathway, leading to elevated levels of the cytokines TNF-α, IL-6 and IL-1ß, which were attenuated by luteolin. Luteolin exerted anticonvulsant and neuroprotective activities in the PTZ-induced epileptic model. Its mechanism was associated with the inhibition of the TLR4/IκBα/NF-κB pathway, alleviating the immune-inflammatory response in the post-epileptic hippocampus.

Respiratory activity during seizures induced by pentylenetetrazole.

This study investigated the respiratory activity in adult Wistar rats across different behavioral seizure severity induced by pentylenetetrazole (PTZ). Animals underwent surgery for electrodes implantation, allowing simultaneous EEG and diaphragm EMG (DIAEMG) recordings and the respiratory frequency and DIAEMG amplitude were measured. Seizures were acutely induced through PTZ injection and classified based on a pre-established score, with absence-like seizures (spike wave discharge (SWD) events on EEG) representing the lowest score. The respiratory activity was grouped into the different seizure severities. During absence-like and myoclonic jerk seizures, the breathing frequency decreased significantly (∼50% decrease) compared to pre- and post-ictal periods. Pronounced changes occurred with more severe seizures (clonic and tonic) with periods of apnea, especially during tonic seizures. Apnea duration was significantly higher in tonic compared to clonic seizures. Notably, during PTZ-induced tonic seizures the apnea events were marked by tonic DIAEMG contraction (tonic-phase apnea). In the majority of animals (5 out of 7) this was a fatal event in which the seizure-induced respiratory arrest preceded the asystole. In conclusion, we provide an assessment of the respiratory activity in the PTZ-induced acute seizures and showed that breathing dysfunction is more pronounced in seizures with higher severity.

Inhibition of PTEN-induced kinase 1 autophosphorylation may assist in preventing epileptogenesis induced by pentylenetetrazol.

PTEN-induced kinase 1 (PINK1) autophosphorylation-triggered mitophagy is the main mitophagic pathway in the nervous system. Moreover, multiple studies have confirmed that mitophagy is closely related to the occurrence and development of epilepsy. Therefore, we speculated that the PINK1 autophosphorylation may be involved in epileptogenesis by mediating mitophagic pathway. This study aimed to explore the contribution of activated PINK1 to epileptogenesis induced by pentylenetetrazol (PTZ) in Sprague‒Dawley rats. During PTZ-induced epileptogenesis, the levels of phosphorylated PINK1 were increased, accompanied by elevated mitophagy, mitochondria oxidative stress and neuronal damage. After microRNA intervention targeting translocase outer mitochondrial membrane 7 (TOM7) or overlapping with the m-AAA protease 1 homolog (OMA1), the levels of PINK1 phosphorylation, mitophagy, mitochondrial oxidative stress, neuronal injury were observed in the rats with induced epileptogenesis. Furthermore, inhibiting of the expression of TOM7, a positive regulator of PINK1 autophosphorylation, reversed the increase in PINK1 phosphorylation and alleviated mitophagy, neuronal injury, thereby preventing epileptogenesis. In contrast, reducing the levels of OMA1, a negative regulator of PINK1 autophosphorylation, led to increased phosphorylation of PINK1, accompanied by aggravated neuronal injury and ultimately, epileptogenesis. This study confirmed the contribution of activated PINK1 to PTZ-induced epileptogenesis and suggested that the inhibition of PINK1 autophosphorylation may assist in preventing epileptogenesis.

Exploring the neuroprotective potential of antimicrobial peptides from Dinoponera quadriceps venom against pentylenetetrazole-induced seizures in vivo.

Epilepsy affects around 50 million people worldwide and 30% of patients have difficulty controlling the disease. The search for substances that can fill the existing gaps in the treatment of epilepsy is of great importance. Arthropod venoms are promising sources for this purpose due to the presence of small peptides that modulate the activity of ion channels and neuron receptors. The aim of this study was to investigate dinoponeratoxins from the Dinoponera quadriceps ant venom (M-PONTX-Dq3a, M-PONTX-Dq3b and M-PONTX-Dq3c) as potential anticonvulsants. We evaluated them in a seizure model induced by pentylenetetrazole (PTZ) in male swiss mice. Interestingly, intraperitoneal treatment with each peptide increased the time until the first seizure and the percentage of survival, with M-PONTX-Dq3b showing the best results. M-PONTX-Dq3a was discarded due to the appearance of some signs of toxicity with the increase in malondialdehyde (MDA) levels in the striatum. Both, M-PONTX-Dq3b and M-PONTX-Dq3c decreased iNOS and TNF-α in the hippocampus. Notably, M-PONTX-Dq3c treatment decreased the levels of MDA and nitrite in the cortex and hippocampus. Our results indicate that, M-PONTX-Dq3b and M-PONTX-Dq3c have anticonvulsant activity and exhibit anti-inflammatory effects in epilepsy, offering new perspectives for biopharmaceutical development.

Integrated non-targeted metabolomics and network pharmacology to reveal the mechanisms of berberine in the long-term treatment of PTZ-induced epilepsy.

AIMS: The increasing resistance to anti-seizure medications (ASMs) and the ambiguous mechanisms of epilepsy highlight the pressing demand for the discovery of pioneering lead compounds. Berberine (BBR) has received significant attention in recent years within the field of chronic metabolic disorders. However, the reports on the treatment of epilepsy with BBR are not systematic and the mechanism remains unclear. MAIN METHODS: In this study, the seizure behaviors of mice were recorded following subcutaneous injection of pentetrazol (PTZ). Non-targeted metabolomics was used to analyze the serum metabolites based on ultra-performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF/MS). Meanwhile, multivariate statistical methods were used for metabolite identification and pathway analysis. Furthermore, network pharmacology, molecular docking, and quantitative real-time PCR assay were used for the target identification. KEY FINDINGS: BBR had anti-seizure effects on PTZ-induced seizure mice after long-term treatment. Tryptophan metabolism and phenylalanine metabolism were involved in regulating the therapeutic effects of BBR. SIGNIFICANCE: This study reveals the potential mechanism of BBR for epilepsy treatment based on non-targeted metabolomics and network pharmacology, which provides evidence for uncovering the pathogenesis of epilepsy, suggesting that BBR is a potential lead compound for anti-epileptic treatment.

Effects of the uremic toxin indoxyl sulfate on seizure activity, learning and brain oxidative stress parameters in mice.

Patients with end-stage renal disease often have neurological disorders, with a higher incidence of memory impairment or epilepsy than in the general population. Patients undergoing hemodialysis are particularly exposed to the biological effects of uremic toxins. Indoxyl sulfate (IS) is one of the most potent uremic toxins; however, its possible effects on seizure susceptibility or memory functions have yet to be elucidated. In the current study, we focused on investigating the possible convulsant and amnesic effects of IS in recognized animal models. The study was performed on adult male Swiss mice. IS and scopolamine (SCO) were administered intraperitoneally (i.p.), and pentylenetetrazole (PTZ) was injected subcutaneously (s.c.). All substances were given as single injections. Acute IS administration (400 mg/kg) led to its accumulation in the brain. IS at doses of 200 and 400 mg/kg decreased the PTZ convulsive threshold, and at the same doses, it did not significantly affect the threshold for electroconvulsions. IS (200 and 400 mg/kg) did not impair learning in the passive avoidance test and did not increase the SCO-induced memory impairment in this test. IS increased lipid peroxidation, decreased the level of reduced glutathione, and reduced the activity of superoxide dismutase and catalase in mouse brains. Exposure to IS did not significantly change the activity of acetylcholinesterase in the brain tissue. This study shows that acute exposure to IS induces oxidative stress in the brain and potentiates PTZ-induced seizures in mice. Further studies are needed to find out whether IS-induced oxidative stress may affect epileptic seizures and/or epileptogenesis.

Maternal sevoflurane exposure increases the epilepsy susceptibility of adolescent offspring by interrupting interneuron development.

BACKGROUND: Exposure to general anesthesia influences neuronal functions during brain development. Recently, interneurons were found to be involved in developmental neurotoxicity by anesthetic exposure. But the underlying mechanism and long-term consequences remain elusive. METHODS: Pregnant mice received 2.5% sevoflurane for 6-h on gestational day 14.5. Pentylenetetrazole (PTZ)-induced seizure, anxiety- and depression-like behavior tests were performed in 30- and 60-day-old male offspring. Cortical interneurons were labeled using Rosa26-EYFP/-; Nkx2.1-Cre mice. Immunofluorescence and electrophysiology were performed to determine the cortical interneuron properties. Q-PCR and in situ hybridization (ISH) were performed for the potential mechanism, and the finding was further validated by in utero electroporation (IUE). RESULTS: In this study, we found that maternal sevoflurane exposure increased epilepsy susceptibility by using pentylenetetrazole (PTZ) induced-kindling models and enhanced anxiety- and depression-like behaviors in adolescent offspring. After sevoflurane exposure, the highly ordered cortical interneuron migration was disrupted in the fetal cortex. In addition, the resting membrane potentials of fast-spiking interneurons in the sevoflurane-treated group were more hyperpolarized in adolescence accompanied by an increase in inhibitory synapses. Both q-PCR and ISH indicated that CXCL12/CXCR4 signaling pathway downregulation might be a potential mechanism under sevoflurane developmental neurotoxicity which was further confirmed by IUE and behavioral tests. Although the above effects were obvious in adolescence, they did not persist into adulthood. CONCLUSIONS: Our findings demonstrate that maternal anesthesia impairs interneuron migration through the CXCL12/CXCR4 signaling pathway, and influences the interneuron properties, leading to the increased epilepsy susceptibility in adolescent offspring. Our study provides a novel perspective on the developmental neurotoxicity of the mechanistic link between maternal use of general anesthesia and increased susceptibility to epilepsy.

Effects of N-acetylcysteine and acetyl-L-carnitine on acute PTZ-induced seizures in larval and adult zebrafish.

BACKGROUND: Epilepsy is a prevalent neurological disease, affecting approximately 1-2% of the global population. The hallmark of epilepsy is the occurrence of epileptic seizures, which are characterized by predictable behavioral changes reflecting the underlying neural mechanisms of the disease. Unfortunately, around 30% of patients do not respond to current pharmacological treatments. Consequently, exploring alternative therapeutic options for managing this condition is crucial. Two potential candidates for attenuating seizures are N-acetylcysteine (NAC) and Acetyl-L-carnitine (ALC), as they have shown promising neuroprotective effects through the modulation of glutamatergic neurotransmission. METHODS: This study aimed to assess the effects of varying concentrations (0.1, 1.0, and 10 mg/L) of NAC and ALC on acute PTZ-induced seizures in zebrafish in both adult and larval stages. The evaluation of behavioral parameters such as seizure intensity and latency to the crisis can provide insights into the efficacy of these substances. RESULTS: Our results indicate that both drugs at any of the tested concentrations were not able to reduce PTZ-induced epileptic seizures. On the other hand, the administration of diazepam demonstrated a notable reduction in seizure intensity and increased latencies to higher scores of epileptic seizures. CONCLUSION: Consequently, we conclude that, under the conditions employed in this study, NAC and ALC do not exhibit any significant effects on acute seizures in zebrafish.

Chemically-induced epileptic seizures in zebrafish: A systematic review.

The use of zebrafish as a model organism is gaining evidence in the field of epilepsy as it may help to understand the mechanisms underlying epileptic seizures. As zebrafish assays became popular, the heterogeneity between protocols increased, making it hard to choose a standard protocol to conduct research while also impairing the comparison of results between studies. We conducted a systematic review to comprehensively profile the chemically-induced seizure models in zebrafish. Literature searches were performed in PubMed, Scopus, and Web of Science, followed by a two-step screening process based on inclusion/exclusion criteria. Qualitative data were extracted, and a sample of 100 studies was randomly selected for risk of bias assessment. Out of the 1058 studies identified after removing duplicates, 201 met the inclusion criteria. We found that the most common chemoconvulsants used in the reviewed studies were pentylenetetrazole (n = 180), kainic acid (n = 11), and pilocarpine (n = 10), which increase seizure severity in a dose-dependent manner. The main outcomes assessed were seizure scores and locomotion. Significant variability between the protocols was observed for administration route, duration of exposure, and dose/concentration. Of the studies subjected to risk of bias assessment, most were rated as low risk of bias for selective reporting (94%), baseline characteristics of the animals (67%), and blinded outcome assessment (54%). Randomization procedures and incomplete data were rated unclear in 81% and 68% of the studies, respectively. None of the studies reported the sample size calculation. Overall, these findings underscore the need for improved methodological and reporting practices to enhance the reproducibility and reliability of zebrafish models for studying epilepsy. Our study offers a comprehensive overview of the current state of chemically-induced seizure models in zebrafish, highlighting the common chemoconvulsants used and the variability in protocol parameters. This may be particularly valuable to researchers interested in understanding the underlying mechanisms of epileptic seizures and screening potential drug candidates in zebrafish models.

(-)-α-bisabolol exerts neuroprotective effects against pentylenetetrazole-induced seizures in rats by targeting inflammation and oxidative stress.

Epilepsy is the most common neurological disorder which is accompanied with behavioral and psychiatric alternations. Current evidences have shown that (-)-α-bisabolol (BSB) possess anti-inflammatory and antioxidative effects in several animal studies. Here, we conducted present study to evaluate its neuroprotective effects against pentylenetetrazole (PTZ)-induced seizures in rats. We used fifty male rats and they were randomly assigned into 5 groups control, BSB100, PTZ, BSB50 + PTZ, BSB100 + PTZ. The animals intraperitoneally received PTZ (45 mg/kg) for ten consecutive days to induce epilepsy model. BSB in doses of 50 and 100 mg/kg was administrated orally one hour before PTZ administration for ten days. The elevated plus maze (EPM) test was carried out to assess anxiety-like behavior. The seizure intensity was evaluated according to modifies Racine's convulsion scale (RCS). Y-maze and passive avoidance were utilized to assess working memory and aversive memory. The expression of pro-inflammatory cytokines and oxidative stress factors were measured using the enzyme-linked immunosorbent assay (ELISA). The neuronal cell loss in the hilar region was assessed using Nissl staining. Results showed that PTZ-treated rats had more seizure intensity, anxiety-like behavior, memory deficits, higher levels of TNF-α, IL-1ß, and oxidative markers. Pre-treatment with BSB 100 significantly inhibited seizure intensity, anxiety-like behavior, and memory deficits; reduced levels of TNF-α, IL-1ß, and MDA oxidative markers. Collectively, outcome of this work shows that BSB at the dose of 100 mg/kg may exert neuroprotective effects by mitigating seizures, oxidative stress, and neuroinflammation, and ameliorates memory and anxiety disorders in the PTZ-induced seizure rats.

In Silico Analysis, Anticonvulsant Activity, and Toxicity Evaluation of Schisandrin B in Zebrafish Larvae and Mice.

The aim of this study is to evaluate the anticonvulsant potential of schisandrin B, a main ingredient of Schisandra chinensis extracts. Schisandrin B showed anticonvulsant activity in the zebrafish larva pentylenetetrazole acute seizure assay but did not alter seizure thresholds in the intravenous pentylenetetrazole test in mice. Schisandrin B crosses the blood-brain barrier, which we confirmed in our in silico and in vivo analyses; however, the low level of its unbound fraction in the mouse brain tissue may explain the observed lack of anticonvulsant activity. Molecular docking revealed that the anticonvulsant activity of the compound in larval zebrafish might have been due to its binding to a benzodiazepine site within the GABAA receptor and/or the inhibition of the glutamate NMDA receptor. Although schisandrin B showed a beneficial anticonvulsant effect, toxicological studies revealed that it caused serious developmental impairment in zebrafish larvae, underscoring its teratogenic properties. Further detailed studies are needed to precisely identify the properties, pharmacological effects, and safety of schisandrin B.

Seizure-induced increase in microglial cell population in the developing zebrafish brain.

Epilepsy is a chronic brain disorder characterized by unprovoked and recurrent seizures, of which 60% are of unknown etiology. Recent studies implicate microglia in the pathophysiology of epilepsy. However, their role in this process, in particular following early-life seizures, remains poorly understood due in part to the lack of suitable experimental models allowing the in vivo imaging of microglial activity. Given the advantage of zebrafish larvae for minimally-invasive imaging approaches, we sought for the first time to describe the microglial responses after acute seizures in two different zebrafish larval models: a chemically-induced epileptic model by the systemic injection of kainate at 3 days post-fertilization, and the didys552 genetic epilepsy model, which carries a mutation in scn1lab that leads to spontaneous epileptiform discharges. Kainate-treated larvae exhibited transient brain damage as shown by increased numbers of apoptotic nuclei as early as one day post-injection, which was followed by an increase in the number of microglia in the brain. A similar microglial phenotype was also observed in didys552-/- mutants, suggesting that microglia numbers change in response to seizure-like activity in the brain. Interestingly, kainate-treated larvae also displayed a decreased seizure threshold towards subsequent pentylenetetrazole-induced seizures, as shown by higher locomotor and encephalographic activity in comparison with vehicle-injected larvae. These results are comparable to kainate-induced rodent seizure models and suggest the suitability of these zebrafish seizure models for future studies, in particular to elucidate the links between epileptogenesis and microglial dynamic changes after seizure induction in the developing brain, and to understand how these modulate seizure susceptibility.

Herb pair of Polygala tenuifolia Willd and Acorus tatarinowii Schott decoction attenuates seizures and alleviates anxiety in mice: Evidence for modulating inflammation, alleviating oxidative stress and mediating GABA pathway.

Currently, prolong use of standard anti-epileptics may cause tolerance and ineffective for about 30% of epileptic patients. Medicinal plants provide an attractive therapeutic effect in preventing and treating seizures in traditional and folk medicine. In this study, we investigate the antiepileptic effects of PTAT decoction on acute and chronic seizure models in mice and explore the potential mechanisms. PTAT decoction dose-dependently protected mice against MES and PTZ induced seizure. Meanwhile, it decreased the seizure severity and reduced seizure-caused anxious behavior in the PTZ-kindling mice, suggesting a significant antiepileptic activity and anxiolytic/anxiogenic potential. PTAT decoction dose-dependently increased the levels of GSH and the activity SOD and CAT, while decreased the level of MDA in the hippocampi of treated mice. Furthermore, a significant decrease in the proinflammatory cytokine levels, including TNF-α, IL-1ß, IL-6 and MCP-1 was found in treated mice compared with the mice in the vehicle + PTZ group. Moreover, PTAT decoction dose-dependently reversed the alterations induced by PTZ in GABA, GABA-T, L-GAD and glutamate levels in kindling mice, showing an effect on the modulation of the GABA neurotransmission. Thus, PTAT decoction has a promising anticonvulsant activity mediated via multiple mechanisms, which might be used as an up-and-coming phytotherapy strategy in the management of epilepsy and its complications.

A comparative study to optimize experimental conditions of pentylenetetrazol and pilocarpine-induced epilepsy in zebrafish larvae.

A common way to investigate epilepsy and the effect of antiepileptic pharmaceuticals is to analyze the movement patterns of zebrafish larvae treated with different convulsants like pentylenetetrazol (PTZ), pilocarpine, etc. Many articles have been written on this topic, but the research methods and exact settings are not sufficiently defined in most. Here we designed and executed a series of experiments to optimize and standardize the zebrafish epilepsy model. We found that during the light and the dark trials, the zebrafish larvae moved significantly more in the light, independent of the treatment, both in PTZ and pilocarpine-treated and the control groups. As expected, zebrafish larvae treated with convulsants moved significantly more than the ones in the control group, although this difference was higher between the individuals treated with PTZ than pilocarpine. When examining the optimal observation time, we divided the half-hour period into 5-minute time intervals, and between these, the first 5 minutes were found to be the most different from the others. There were fewer significant differences in the total movement of larvae between the other time intervals. We also performed a linear regression analysis with the cumulative values of the distance moved during the time intervals that fit the straight line. In conclusion, we recommend 30 minutes of drug pretreatment followed by a 10-minute test in light conditions with a 5-minute accommodation time. Our result paves the way toward improved experimental designs using zebrafish to develop novel pharmaceutical approaches to treat epilepsy.

Delayed treatment with ceftriaxone reverses the enhanced sensitivity of TBI mice to chemically-induced seizures.

The pathophysiological changes that occur after traumatic brain injury (TBI) can lead to the development of post-traumatic epilepsy, a life-long complication of brain trauma. The etiology of post-traumatic epilepsy remains unknown, but TBI brains exhibit an abnormal excitatory / inhibitory balance. In this study, we examine how brain injury alters susceptibility to chemically-induced seizures in C57Bl/6J mice, and if pharmacological enhancement of glutamate transporters can reduce chronic post-traumatic seizures. We found that controlled cortical impact (CCI) mice display delayed susceptibility to pentylenetetrazol (PTZ)-induced seizures. While CCI mice have no change in seizure susceptibility at 7d post-injury (dpi), at 70dpi they have reduced latency to PTZ-induced seizure onset, higher seizure frequency and longer seizure duration. Quantification of glutamate transporter mRNA showed that levels of Scl1a2 and Scl1a3 mRNA were increased at 7dpi, but significantly decreased at 70dpi. To test if increased levels of glutamate transporters can ameliorate delayed-onset seizure susceptibility in TBI mice, we exposed a new cohort of mice to CCI and administered ceftriaxone (200mg/kg/day) for 14d from 55-70dpi. We found that ceftriaxone significantly increased Scl1a2 and Scl1a3 in CCI mouse brain at 70dpi, and prevented the susceptibility of CCI mice to PTZ-induced seizures. This study demonstrates cortical impact can induce a delayed-onset seizure phenotype in mice. Delayed (55dpi) ceftriaxone treatment enhances glutamate transporter mRNA in the CCI brain, and reduces PTZ-induced seizures in CCI mice.