Benzodiazepines are drugs used to treat anxiety disorders and epilepsy due to their hypnotic, anxiolytic and anticonvulsant functions. But due to existing adverse effects such as decreased psychomotor activity and impaired memory, new and safer drugs are sought. Therefore, this study seeks to evaluate the anxiolytic and anticonvulsant action of chalcone (E)-3-(furan-2-yl)-1-(2-hydroxyphenyl)prop-2-en-1-one against wildadult zebrafish, and the mechanisms of action involved in such effects.The acute toxicity of chalcone was analyzed during 96 hours, and the anxiolytic behavior of fish treated with chalcone was evaluated in light/dark tests and open field tests (n=6 animals/group). In the test to evaluate the anticonvulsant effect of the sample, Pentylenetetrazole (PTZ) was used to induce seizures by immersion. Next, we investigated the mechanism of action with the γ-aminobutyric acid (GABA) antagonist and performed a molecular coupling study. The results showed that chalcone at doses of 4, 20, and 40 mg/kg was non-toxic and altered fish locomotion (****p<0.0001 vs. control). All analyzed doses exhibited possible anxiolytic effects (* p < 0.05, *** p < 0.001 vs. control) in the light/dark test. This effect was blocked by the antagonist flumazenil (FMZ) at a dose of 40 mg/kg (# p < 0.05 vs. Fmz+Chalcone). In addition to exhibiting anxiolytic effects, chalcone demonstrated anticonvulsant effects via GABA, as confirmed by a binding/activity study (### p < 0.01, #### p < 0.0001 vs. Fmz+Chalcone).

This research is designed to investigate the sedative effects of naringenin (NAR) and diazepam against thiopental sodium‐induced sleeping mice. NAR (5 and 10 mg kg), diazepam (DZP) (2 mg kg), flumazenil (FLN) (0.1 mg kg), and their combinations are administered intraperitoneally to mice. After 30 min, sleep is induced with intraperitoneal thiopental sodium (20 mg kg), and sleep latency and duration are measured. In silico analysis investigates the role of GABA receptors. NAR significantly reduces sleep latency and prolongs sleep duration in a dose‐dependent manner, with the combination of NAR‐10 and DZP‐2 showing a synergistic effect. The combination of the antagonist (FLN) (NAR‐10 and FLN‐0.1) indicates that latency increases and sleep duration decreases compared to NAR‐10 alone. Furthermore, in silico docking studies corroborates these results, demonstrating a significant binding affinity of NAR (−8.3 kcal mol), which is comparable to the standard ligand DZP (−8.7 kcal mol) and FLN (−7.0 kcal mol) for the GABAA (6X3X) receptor, indicating a GABAergic mechanism. Pharmacokinetics and toxicity evaluations confirm NAR's potential as a safe therapeutic agent, with a high LD50 (2000 mg kg) and minimal toxicity. These findings highlight NAR's potential as a GABAergic sedative agent, requiring further research before being used clinically to treat sleep disorders.

Assessment of sedative activity of Chrysin: Behavioral approach with pharmacokinetics, toxicological profile and molecular docking.

The loss of synaptic vesicle glycoprotein 2A (SV2A) can lead to dysfunction of GABAergic neurons, but a direct comparison of SV2A and GABAA receptor densities in humans has not been assessed. This study evaluated SV2A and GABAA receptor abnormalities in patients with drug-resistant epilepsy (DRE) and compared the patterns to glucose hypometabolism. Eleven patients with DRE were retrospectively recruited and underwent PET imaging with [18F]fluorodeoxyglucose ([18F]FDG), [18F]Flumazenil (FMZ), and [18F]SynVesT-1. Visual assessments counted abnormal metabolic brain regions based on the Anatomical Automatic Labeling (AAL) atlas, while voxel-level analyses delineated the abnormal metabolic distributions. The relationship between hypo-metabolic distributions and the age of epilepsy onset was analyzed. The hypometabolic regions in [18F]FDG PET, identified in the AAL atlas, was significantly broader than in [18F]FMZ (p = 0.0005) and [18F]SynVesT-1 (p = 0.0010) PET, with no statistical difference observed between [18F]FMZ and [18F]SynVesT-1 PET (p > 0.05). The voxel number in [18F]FDG PET was significantly higher than that of the [18F]FMZ and [18F]SynVesT-1 PET in both hypo-intensity area and severe hypo-intensity area. The ratio of the voxel number between these two area was higher for [18F]SynVesT-1 PET compared to [18F]FDG PET (p = 0.0195) and [18F]FMZ PET (p = 0.0237), and positively correlated with the age of epilepsy onset (r = 0.7397, p = 0.0145). [18F]FMZ and [18F]SynVesT-1 PET images revealed a more restricted pattern of reduced uptake compared to [18F]FDG PET in DRE patients. The age of epilepsy onset correlated with a reduction in [18F]SynVesT-1 uptake but not in [18F]FMZ or [18F]FDG uptake.

Epilepsy, a globally prevalent neurologic disorder, necessitates precise identification of the epileptogenic zone (EZ) for effective surgical management. While the individual utilities of FDG-PET and flumazenil (FMZ)-PET have been demonstrated, their combined efficacy in localizing the epileptogenic zone remains underexplored. We aim to improve the noninvasive prediction of EZ in temporal lobe epilepsy (TLE) by combining FDG-PET and FMZ-PET with statistical feature extraction and machine learning. This study included 20 drug-resistant patients with unilateral TLE (14 mesial TLE, 6 lateral TLE) and 2 control groups (n = 29 for FDG, n = 20 for FMZ). EZ of each patient was confirmed by postsurgical pathology and 1-year follow-up, while propagation zone (PZ) and noninvolved zone (NIZ) were derived from the epileptogenicity index based on presurgical stereo-encephalography (SEEG) monitoring. Whole brain PET scans were obtained with dual tracers [18F]FDG and [18F]FMZ on separate days, from which standard uptake value ratio (SUVR) was calculated by global mean scaling. Low-order statistical parameters of SUVRs and t-maps derived against control groups were extracted. Additionally, fused FDG and FMZ features were created by using arithmetic operations. Spearman correlation was used to investigate the associations between FDG and FMZ, while multiple linear regression analyses were used to explore the interaction effects of imaging features in predicting epileptogenicity. Crafted imaging features were used to train logistic regression models to predict EZ, whose performance was evaluated by using 10-fold cross-validation at ROI level, and leave-1-patient-out cross-validation at patient level. FDG SUVR significantly decreased in EZ and PZ compared with NIZ, while FMZ SUVR in EZ significantly differed from PZ. Interaction effects were found between FDG and FMZ in their prediction of epileptogenicity. Fusion of FDG and FMZ provided the best prediction model with an area under the curve (AUC) of 0.86 [0.84-0.87] for EZ versus NIZ and an AUC of 0.79 [0.77-0.81] for EZ versus PZ, eliminating 100% false-positives in 50% of patients, and ≥80% FPs in 90% of patients at patient level. Combined FDG and FMZ offer a promising avenue for noninvasive localization of the epileptogenic zone in TLE, potentially refining surgical planning.

This study investigated the anxiolytic, anticonvulsant and memory preservation effects of the flavonoid robinin. The compound, administered at doses of 4, 20 and 40mg/kg, did not show toxicity after 96 h of monitoring. In behavioural experiments with zebrafish, robinin did not cause significant changes in motor functions, but it impairs locomotor activity and demonstrates anxiolytic properties, evidenced by the increase in the time spent in the clean zone of the protector. A minimum effective dose (4mg/kg) was blocked by flumazenil (FMZ), providing interaction with GABAA receptors and decreasing an anxiolytic profile similar to that of diazepam, without causing sedation. In addition, a dose of 40mg/kg was able to reverse seizures, increasing the latency to enter the seizure stages, an effect that was also blocked by FMZ. Robinin (40mg/kg) also prevented memory variation in an inhibitory avoidance test. In silico absorption, distribution, metabolism and excretion tests indicated that robinine presents gradual intestinal absorption and low distribution in the central nervous system. In molecular docking, the compound was exposed in the layer with CAII and GABAA receptors, corroborating the anxiolytic and anticonvulsant effects. The results suggest that robinine has therapeutic potential in the treatment of anxiety and seizures, in addition to offering memory protection, representing an advantageous alternative to benzodiazepines, with a promising neuroprotective potential for the pharmaceutical industry.

This study investigated the anxiolytic and anticonvulsant effects and safety profile of limonene enantiomers and their oxidized derivatives. The toxicity test was performed by monitoring the animals for 96 hours, with no deaths or significant toxicity observed up to the highest dose, which allowed the determination of the LD50. Doses of 4, 20 and 40 mg/kg were tested, with no toxicity observed up to 96 h (LD50>40 mg/kg). Anxiolytic activity was measured in a preference test for light and dark areas, and the effect of the compounds was evaluated in the presence of serotonergic antagonists. The (S)-(-)-LIM and (R)-(+)-LIM enantiomers showed anxiolytic effects, with (S)-(-)-LIM being effective at all doses. In the anticonvulsant test, the oxidized derivatives, such as perilyl acid (PAC), significantly delayed PTZ-induced seizures, an effect blocked by flumazenil (FMZ). The oxidized derivatives, especially perilyl acid (PAC), showed anxiolytic effects at all doses and significantly delayed the three PTZ-induced seizure events. This effect was blocked by FMZ, suggesting a relationship between PAC and the GABAergic pathway. PAC, being the most oxidized derivative, was the most effective for both anxiety and delaying seizure progression, suggesting that oxidation of limonene compounds may increase their therapeutic efficacy.

The soy isoflavone daidzin (DZN) has been considered a hopeful bioactive compound having diverse biological activities, including anxiolytic, memory-enhancing, and antiepileptic effects, in experimental animals. However, its sedative and hypnotic effects are yet to be discovered. This study aimed to evaluate its sedative/hypnotic effect on Swiss mice. Additionally, in silico studies were also performed to see the possible molecular mechanisms behind the tested neurological effect. For this, male Swiss albino mice were treated with DZN (5, 10, or 20mg/kg) intraperitoneally (i.p.) with or without the standard GABAergic medication diazepam(DZP) and/or flumazenil(FLU) and checked for the onset and duration of sleeping time using thiopental sodium-induced as well as DZP-induced sleeping tests. A molecular docking study was also performed to check its interaction capacity with the α1 and β2 subunits of the GABAA receptor. Findings suggest that DZN dose-dependently and significantly reduced the latency while increasing the duration of sleep in animals. In combination therapy, DZN shows synergistic effects with the DZP-2 and DZP-2 + FLU-0.01 groups, resulting in significantly (p < 0.05) reduced latency and increased sleep duration. Further, molecular docking studies demonstrate that DZN has a strong binding affinity of - 7.2kcal/mol, which is closer to the standard ligand DZP (- 8.3kcal/mol) against the GABAA (6X3X) receptor. Molecular dynamic simulations indicated stability and similar binding locations for DZP and DZN with 6X3X. In conclusion, DZN shows sedative effects on Swiss mice, possibly through the GABAA receptor interaction pathway.

The chronic neuronal burden of traumatic brain injury (TBI) is not fully characterized by routine imaging, limiting understanding of the role of neuronal substrates in adverse outcomes. To determine whether tissues that appear healthy on routine imaging can be investigated for selective neuronal loss using [11C]flumazenil (FMZ) positron emission tomography (PET) and to examine whether this neuronal loss is associated with long-term outcomes. In this cross-sectional study, data were collected prospectively from 2 centers (University of Cambridge in the UK and Weill Cornell Medicine in the US) between September 1, 2004, and May 31, 2021. Patients with TBI (>6 months postinjury) were compared with healthy control participants (all aged >18 years). Individuals with neurological disease, benzodiazepine use, or contraindication to magnetic resonance imaging were excluded. Data were retrospectively collated with nonconsecutive recruitment, owing to convenience and scanner or PET ligand availability. Data were analyzed between February 1 and September 30, 2023. Flumazenil voxelwise binding potential relative to nondisplaceable binding potential (BPND). Selective neuronal loss identified with FMZ PET was compared between groups on voxelwise and regional scales, and its association with functional, cognitive, and psychological outcomes was examined using Glasgow Outcome Scale (GOS) scores, measures of sustained executive attention (animal and sustained fluency), and 36-Item Short Form Health Survey (SF-36) scores. Diffusion tensor imaging was used to assess structural connectivity of regions of cortical damage, and its association with thalamic selective neuronal loss. In this study, 24 patients with chronic TBI (mean [SD] age, 39.2 [12.3] years; 18 men [75.0%]) and 33 healthy control participants (mean [SD] age, 47.6 [20.5] years; 23 men [69.7%]) underwent FMZ PET. Patients with TBI had a median time of 29 (range, 7-95) months from injury to scan. They displayed selective neuronal loss in thalamic nuclei, over and above gross volume loss in the left thalamus, and bilateral central, mediodorsal, ventral-lateral dorsal, anterior, and ventral anterior thalamic nuclei, across a wide range of injury severities. Neuronal loss was associated with worse functional outcome using GOS scores (left thalamus, left ventral anterior, and bilateral central, mediodorsal, and anterior nuclei), worse cognitive outcome on measures of sustained executive attention (left thalamus, bilateral central, and right mediodorsal nuclei), and worse emotional outcome using SF-36 scores (right central thalamic nucleus). Chronic thalamic neuronal loss partially mirrored the location of primary cortical contusions, which may indicate secondary injury mechanisms of transneuronal degeneration. The findings of this study suggest that selective thalamic vulnerability may have chronic neuronal consequences with relevance to long-term outcome, suggesting the evolving and potentially lifelong thalamic neuronal consequences of TBI. FMZ PET is a more sensitive marker of the burden of neuronal injury than routine imaging; therefore, it could inform outcome prognostication and may lead to the development of individualized precision medicine approaches.

Fluorine-18 (18F) flumazenil (FMZ) has been synthesized using various precursors, and its role has been explored in imaging Gamma-aminobutyric acid-A receptors. The main objective was to synthesize (18F) FMZ using isotopic substitution. Around 18 ± 2 GBq was added to the module, dried, and radiolabeling was standardized with 3.0 mg of the FMZ precursor at various temperatures (110°C -160°C) for 10-30 min. The product was finally eluted with 20% ethanol (in phosphate buffer). The final product was characterized by high-performance liquid chromatography (HPLC). The stability was evaluated in water, saline, and phosphate-buffered saline for 4 h. The radiolabelling efficiency of cartridge-based purification was 16 ± 4% (n = 10) with a radiochemical purity of 96.5 ± 1.8%, whereas in HPLC-based purification, the yield was 10 ± 4% (n = 5) with a radiochemical purity of 97.3 ± 1.4%. The specific activity was 120 ± 20 GBq/μmol. (18F) FMZ was successfully synthesized using an isotopic approach and could be used as an alternative cheaper option for the synthesis.

Mefenamic acid (MFA), a common analgesic, causes central nervous system (CNS) toxicity at high doses with a proposed activity on the Gamma-aminobutyric acid (GABA) system. However, it remains unknown whether flumazenil (FMZ), a GABA type A receptor (GABAAR) antagonist, can reverse MFA toxicity. The behavioral and neurophysiological effects of MFA were investigated in mice with and without FMZ pre-treatment. The elevated zero maze (EZM) and marble burying tests were used to assess anxiety-like behaviors and burying activities, respectively. The standard bar test was used to evaluate catalepsy, while the actophotometer test was used to measure locomotor activity. Seizure intensity was scored, and fatalities were counted. Without FMZ pre-treatment, MFA induced behavioral and neurophysiological effects in a dose-dependent manner as follows: At a dose of 20 mg/kg, i.p, MFA-treated mice exhibited anxiety-like behaviors, which was determined by a significant increase in the time spent in the closed areas and a significant decrease in the number of entries to the open areas of the EZM apparatus. These mice also showed a significant decrease in the burying activity, manifested as a significant decrease in the number of buried marbles. At 40 mg/kg, i.p., MFA-treated mice showed catalepsy that was associated with a significant decrease in locomotor activity. At a dose of 80 mg/kg, i.p., mice developed fatal tonic-clonic seizures (seizure score = 4). Pre-treatment with FMZ (5 mg/kg, i.p.) significantly reversed the anxiety-like behaviors and restored marble-burying activity. Additionally, FMZ prevented catalepsy, significantly restored locomotor activity, reduced seizure intensity (seizure score = 0.3) and significantly reduced mortalities. The present study's findings indicate that activation of the GABAAR is involved in the CNS toxicity of MFA, and FMZ reverses MFA toxicity by interfering with this receptor.

GABAA receptor-positive modulators are well-known to induce sedation, sleep, and general anesthesia. Conversely, GABAA receptor negative allosteric modulators (GABAARNAMs) can increase arousal and induce seizures. Motivated by our studies with patients with hypersomnia, and our discovery that two GABAARNAMs can restore the Excitation/Inhibition (E/I) balance in vitro and arousal in vivo, we chose to screen 11 compounds that have been reported to modulate arousal, to see if they shared a GABA-related mechanism. We determined modulation with both conventional and microfluidic patch clamp methods. We found that receptor activation was variably modulated by all 11 compounds: Rifampicin (RIF), Metronidazole (MET), Minocycline (MIN), Erythromycin (ERY), Ofloxacin (OFX), Chloroquine (CQ), Hydroxychloroquine sulfate (HCQ), Flumazenil (FLZ), Pentylenetetrazol (PTZ), (-)-Epigallocatechin Gallate (EGCG), and clarithromycin (CLR). The computational modeling of modulator-receptor interactions predicted drug action at canonical binding sites and novel orphan sites on the receptor. Our findings suggest that multiple avenues of investigation are now open to investigate large and brain-penetrant molecules for the treatment of patients with diminished CNS E/I balance.

Anxiety disorders are highly prevalent and chronic disorders with treatment resistance to current pharmacotherapies occurring in approximately one in three patients. It has been postulated that flumazenil (FMZ) is efficacious in the management of anxiety disorders via the removal of α4β2δ gamma-aminobutyric acid A receptors. To assess the safety and feasibility of continuous low-dose FMZ infusions for the management of generalised anxiety disorder (GAD) and collect preliminary efficacy data. Uncontrolled, open-label pilot study. Participants had a primary diagnosis of generalised anxiety disorder (GAD) and received two consecutive subcutaneous continuous low-dose FMZ infusions. Each infusion contained 16 mg of FMZ and was delivered over 96 ± 19.2 h. The total dose of FMZ delivered was 32 mg over approximately 8 days. Sodium valproate was given to participants at risk of seizure. The primary outcome was the change in stress and anxiety subscale scores on the Depression Anxiety Stress Scale-21 between baseline, day 8, and day 28. Nine participants with a primary diagnosis of GAD were treated with subcutaneous continuous low-dose FMZ infusions; seven participants met the criteria for treatment resistance. There was a significant decrease in anxiety and stress between baseline and day 8 and baseline and day 28. There was also a significant improvement in subjective sleep quality from baseline to day 28 measured by the Jenkins Sleep Scale. No serious adverse events occurred. This study presents preliminary results for subcutaneous continuous low-dose FMZ's effectiveness and safety in GAD. The findings suggest that it is a safe, well-tolerated, and feasible treatment option in this group of patients. Future randomised control trials are needed in this field to determine the efficacy of this treatment.

Parkinson's disease (PD) is a progressive neurodegenerative disease associated with loss of dopaminergic neurons in the substantia nigra pars compacta. Although aging is the primary cause, environmental and genetic factors have also been implicated in its etiology. In fact, the sporadic nature of PD (i.e., unknown etiology) renders the uncovering of the exact pathogenic mechanism(s) or development of effective pharmacotherapies challenging. In search of novel neuroprotectants, we showed that butyrate (BUT), a short-chain fatty acid, protects against salsolinol (SALS)-induced toxicity in human neuroblastoma-derived SH-SY5Y cells, which are considered an in-vitro model of PD. Dihydromyricetin (DHM), a flavonoid derived from Asian medicinal plant, has also shown effectiveness against oxidative damage and neuroinflammation, hallmarks of neurodegenerative diseases. Here we show that pretreatment of SH-SY5Y cells with DHM concentration-dependently prevented SALS-induced toxicity and that a combination of DHM and BUT resulted in a synergistic protection. The effects of both DHM and BUT in turn could be completely blocked by flumazenil (FLU), a GABAA antagonist acting at benzodiazepine receptor site, and by bicuculline (BIC), a GABAA antagonist acting at orthosteric site. Beta-hydroxybutyrate (BHB), a free fatty acid 3 (FA3) receptor antagonist, also fully blocked the protective effect of DHM. BHB was shown previously to only partially block the protective effect of BUT. Thus, there are some overlaps and some distinct differences in protective mechanisms of DHM and BUT against SALS-induced toxicity. It is suggested that a combination of DHM and BUT may have therapeutic potential in PD. However, further in-vivo verifications are necessary.

BackgroundChanges in γ-aminobutyric acid (GABA) function are noted in patients with Parkinson's disease (PD) who have some non-motor impairments. However, dopamine-related GABA function and GABA-related cognitive changes are still unclear.MethodsThirteen drug-naive early-stage PD patients underwent a series of PET scans with [11C]flumazenil(FMZ) and [11C]CFT. The [11C]FMZ binding potential (BPND) derived from a Logan plot analysis was compared between PD patients and age-matched controls. The [11C]CFT radioactivity relative to the cerebellar counterpart was estimated as a semiquantitative value [11C]CFT SUVR. Correlations between [11C]FMZ BPND and [11C]CFT SUVR in the same region of interest were also examined.ResultsIn patients in the PD group, [11C]CFT SUVR was significantly lower in the putamen. The levels of [11C]FMZ BPND in the cerebral cortex (frontal lobe dominancy) and the affected-side putamen were also reduced. In addition, [11C]CFT SUVR was negatively correlated with the [11C]FMZ BPND level in the affected-side putamen. In patients in the PD group, the total frontal assessment battery (FAB) score was positively correlated with the [11C]FMZ BPND in the frontal region.ConclusionGABAergic dysfunction coexists with dopaminergic loss not only in the putamen but also over the extrastriatal region in patients with early PD and is related to frontal dysfunction. The negative correlation of [11C]CFT SUVR with [11C]FMZ BPND in the affected putamen suggests that a greater dopaminergic demise would decelerate GABA release (or an increase in tracer binding), resulting in persistent failure of the GABAergic system in PD patients.

Epilepsy is one of the most common neurological diseases globally, resulting from a disorder in brain activity. This condition can be triggered by birth trauma, traumatic brain injury (TBI), infections of the brain and stroke. More than 70 million people suffer seizures caused by neurological abnormalities. Approximately 80% of all epileptic patients reside in low-income conditions or in developing countries, and over 75% of patients do not receive proper treatment. Our previous study found an anticonvulsant property of an extract of Euterpe oleracea stone (EEOS) that caused myorelaxation, sedation, and cardiac and respiratory depression after intraperitoneal administration. The present study investigated through electroencephalographic (EEG) profiling the anticonvulsant protective properties of EEOS in induced convulsing rats. Male Wistar rats were treated with EEOS (300 mg/kg), diazepam (DZP) (5 mg/kg), pentylenetetrazol (PTZ) (60 mg/kg) and flumazenil (FMZ) (0.1 mg/kg) by intraperitoneal (i.p.). Electrodes implanted on the dura mater provided EEG data in which EEOS suppressed seizure deflagration caused by PTZ. In addition, EEOS presented no significant difference in comparison to DZP, which has the same mechanism of action. After FMZ injection, a GABAA receptor antagonist blocked the anticonvulsive effect in both the DZP and EEOS groups, suggesting that EEOS exerts it action on the GABAA receptor at the benzodiazepine (BDZ) subunit.

IntroductionSince the 1990s there has been evidence of the significant role Flumazenil (FLU) has in benzodiazepines (BZD) detoxes. The Verona Detox approach has been developed for high dose BZD and Z-drug detoxification via continuous subcutaneous infusion of FLU, a selective BZD receptor antagonist acting on the BZD subunit of the GABA-A receptor. Flumazenil is licensed in the United Kingdom and other countries to treat only BZD overdose although numerous studies have demonstrated its effectiveness in rapidly resetting GABA-A receptors, quickly reducing tolerance and dependence from BZD, and providing a safe and rapid detox from benzodiazepines.ObjectiveThe aim of this article is to provide all healthcare professional who are interested in BZD detoxification with an approach and clear practical information on how to administer FLU.MethodIn this article we outline the approach in detail, describing all medical and nursing procedures day by day. This detox treatment is indicated for patients abusing from at least 5 Defined Daily Dose (DDD) of BZDs or Z-drugs. The process lasts 7 days, and is conducted under medical supervision (daily reviews) and continuous nursing (24/7). During this period, 7mg of FLU is administered (1 mg/24) through an elastomeric pump, via continuous subcutaneous infusion.ConclusionTo this day, the largest database of FLU detoxification was published by our group, showing how this treatment is safe, with very little side effects even in patients with significant medical comorbidities.

ObjectiveThe study investigated the effect 5-[(naphthalen-2-yloxy) methyl]-1,3,4-oxadiaszole2-thiol (B3) in animal model of acute epileptic shock.MethodsThe pharmacokinetics profile of B3 was checked through SwissADME software. The binding affinities of B3, diazepam, and flumazenil (FLZ) were obtained through Auto Dock and PyRx. Post docking analysis and interpretation of hydrogen bonds were performed through Discovery Studio Visualizer 2016. Molecular dynamics simulations of three complexes were carried out through Desmond software package. B3 was then proceeded in PTZ-induced acute seizures models. Flumazenil was used in animal studies for elucidation of possible mechanism of B3. After behavioral studies, the animals were sacrificed, and the brain samples were isolated and stored in 4% formalin for molecular investigations including H and E staining, IHC staining and Elisa etc.ResultsThe results demonstrate that B3 at 20 and 40 mg/kg prolonged the onset time of generalized seizures. B3 considerably increased the expression of protective glutathione S-transferase and glutathione reductase and reduced lipid peroxidation and inducible nitric oxide synthase (P < 0.001) in the cortex. B3 significantly suppressed (P < 0.01) the over expression of the inflammatory mediator tumor necrosis factor–α, whose up-regulation is reported in acute epileptic shocks.ConclusionHence, it is concluded from the aforementioned results that B3 provides neuroprotective effects PTZ-induced acute epileptic model. FLZ pretreatment resulted in inhibition of the anticonvulsant effect of B3. B3 possesses anticonvulsant effect which may be mediated through GABAA mediated antiepileptic pathway.

To investigate the effects of a fixed dose of atipamezole (AT), flumazenil (FL), and 4-aminopyridine (AP), both alone and in combination, on changes in arterial blood pressure and heart rate induced by medetomidine (ME), midazolam (MI), and ketamine (KE) under isoflurane anesthesia with controlled ventilation in healthy cats. Prospective experimental study. University animal research facility. Healthy adult mixed-breed cats were used for 8 investigation groups (6 cats per group), with ≥2weeks between interventions. Cats were anesthetized with an end-tidal isoflurane concentration of 2% under controlled ventilation. A catheter was inserted into the right or left femoral artery for arterial pressure monitoring and blood gas sampling, and ECG electrodes were placed. Upon completed preparations, cats were administered a mixture of ME (0.05mg/kg) and MI (0.5mg/kg), followed 10 minutes later by intramuscular KE (10mg/kg). Twenty minutes after KE injection, the cats received IV injection with either a physiological saline solution at 0.1mL/kg (control), or 1 of 7 variations of experimental drugs, alone or in combination: AT (0.2mg/kg), FL (0.1mg/kg), AP (0.5mg/kg), AT+FL, FL+AP, AT+AP, and AT+FL+AP. Arterial blood pressure and heart rate were continuously measured over 120 minutes after administration of potential antagonists. ME+MI+KE induced an increase in blood pressure and bradycardia. Potential antagonists alone or in combination did not significantly alter the bradycardia. FL, AP alone, and FL+AP did not significantly alter the changes in blood pressures induced by ME+MI+KE. Meanwhile, administration of AT alone or in combination reversed the increase in blood pressure induced by ME+MI+KE but transiently caused excessive hypotension. These results revealed that AT alone or in combination is effective for antagonizing hypertension induced by ME+MI+KE; however, attention should be paid to temporary hypotension in cats anesthetized with isoflurane.

Antinociceptive effects of flower extracts and the active fraction from Styrax japonicus