GABA-transaminase Inhibitor Research Articles

Sustained Inhibition of GABA-AT by OV329 Enhances Neuronal Inhibition and Prevents Development of Benzodiazepine Refractory Seizures.

γ-Aminobutyric acid (GABA) is the principal inhibitory neurotransmitter in the adult brain which mediates its rapid effects on neuronal excitability via ionotropic GABAA receptors. GABA levels in the brain are critically dependent upon GABA-aminotransferase (GABA-AT) which promotes its degradation. Vigabatrin, a low-affinity GABA-AT inhibitor, exhibits anticonvulsant efficacy, but its use is limited due to cumulative ocular toxicity. OV329 is a rationally designed, next-generation GABA-AT inhibitor with enhanced potency. We demonstrate that sustained exposure to OV329 in mice reduces GABA-AT activity and subsequently elevates GABA levels in the brain. Parallel increases in the efficacy of GABAergic inhibition were evident, together with elevations in electroencephalographic delta power. Consistent with this, OV329 exposure reduced the severity of status epilepticus and the development of benzodiazepine refractory seizures. Thus, OV329 may be of utility in treating seizure disorders and associated pathologies that result from neuronal hyperexcitability.

Is hepatic GABA transaminase a promising target for obesity and epilepsy treatments?

γ-Aminobutyric acid (GABA) transaminase (GABA-T) is a GABA-degrading enzyme that plays an essential role in regulating GABA levels and maintaining supplies of GABA. Although GABA in the mammalian brain was discovered 70 years ago, research on GABA and GABA-T has predominantly focused on the brain. Notwithstanding the high activity and expression of GABA-T in the liver, the exact functions of GABA-T in the liver remain unknown. This article reviews the up-to-date information on GABA-T in the liver. It presents recent findings on the role of liver GABA-T in food intake suppression and appetite regulation. Finally, the potential functions of liver GABA-T in other neurological diseases, natural GABA-T inhibitors, and future perspectives in this research area are discussed.

VIGAB-STAT—A Phase IIa Feasibility Trial of Irreversible GABA-transaminase Inhibition as Adjunct Treatment of Status Epilepticus After Cardiac Arrest (P4-2.003)

VIGAB-STAT—A Phase IIa Feasibility Trial of Irreversible GABA-transaminase Inhibition as Adjunct Treatment of Status Epilepticus After Cardiac Arrest (P4-2.003)

GABA-transaminase: A Key Player and Potential Therapeutic Target for Neurological Disorders.

Neurological disorders such as epilepsy, autism, Huntington's disease, multiple sclerosis, and Alzheimer's disease alter brain functions like cognition, mood, movements, and language, severely compromising the well-being of persons, suffering from their negative effects. The neurotransmitters (GABA, glutamate, norepinephrine, dopamine) are found to be involved in neuronal signaling and neurotransmission. GABA, a "commanding neurotransmitter" is directly or indirectly associated with various neurological disorders. GABA is metabolized to succinic semialdehyde by a mitochondrial gamma-aminobutyric acid-transaminase (GABA-T) enzyme. Therefore, the alterations in the GABA performance in the distinct regions of the brain via GABA-T overstimulation or inhibition would play a vital role in the pathogenesis of various neurological disorders. This review emphasizes the leading participation of GABA-T in neurological disorders like Huntington's disease, epilepsy, autism, Alzheimer's disease, and multiple sclerosis. In Huntington's disease, epilepsy, and multiple sclerosis, the surfeited performance of GABA-T results in diminished levels of GABA, whereas in autism, the subsidence of GABA-T activity causes the elevation in GABA contents, which is responsible for behavioral changes in these disorders. Therefore, GABA-T inhibitors (in Huntington's disease, epilepsy, and multiple sclerosis) or agonists (in autism) can be used therapeutically. In the context of Alzheimer's disease, some researchers favor the stimulation of GABA-T activity whereas some disagree with it. Therefore, the activity of GABA-T concerning Alzheimer's disease is still unclear. In this way, studies of GABA-T enzymatic activity in contrast to neurological disorders could be undertaken to understand and be considered a therapeutic target for several GABA-ergic CNS diseases.

Exploring Ocimum basilicum's Secondary Metabolites: Inhibition and Molecular Docking against Rhynchophorus ferrugineus for Optimal Action.

The objective of our work is to create a practical procedure to produce in vitro cell suspensions of O. basilicum and to ascertain the factors that encourage enhanced secondary metabolite production. We investigated the impact of these metabolites on Rhynchophorus ferrugineus's adult and larval target enzymes. The explants were cultivated on Murashige and Skoog (MS) media with 0.1 to 1 mg/L plant growth regulators (PGRs) to create calluses. 2,4-Dichlorophenoxyacetic acid (2,4-D), kinetin, 1-naphthylacetic acid (NAA), and indole-3-butryic acid (IBA) at 0.5, 0.5, 0.1, and 1 mg/L, respectively, with 3% sucrose led to the highest biomass accumulation. In cell suspensions, the total phenolic content (TPC) and total flavonoid content (TFC) were 39.68 and 5.49 mg/g DW, respectively, with abiotic Verticillium dahliae as an activator. Rosmarinic acid, ursolic acid, nepetoidin A and B, salvigenin, and quercetin-3-O-rutinoside as flavonoids and phenolics were analyzed using UPLC-I TQD MS, with the highest concentrations reached after 40 days. The extract demonstrates insecticidal activity against the fourth-instar larvae of R. ferrugineus, with adults at 1197 µg/mL and 12.5 µg/larvae as LC50 and LD50 values. The extract inhibited acetylcholine esterase (AChE), acid phosphatases (ACPs), alkaline phosphatases (ALPs), and gamma-aminobutyric acid-transaminase (GABA-T) in larval tissue in vitro, with IC50 values of 124.2, 149.3, 157.8, and 204.8 µg/mL, and in vivo, with IC50 values of 157.2, 179.4, 185.3, and 241.6 µg/mL, after 24 h. Pure compounds identified the activity of the extract, showing the inhibition of AChE, ACPs, ALPs, and GABA-T with IC50 values ˂ 200 µg/mL (in vitro). The ABMET examination revealed good oral permeability, and docking tests showed that the compounds bind AChE, ACPs, ALPs, and GABA-T. These findings show that a green bioprocessing method such as an O. basilicum cell suspension is a quick and straightforward technique for producing phenolic compounds, and it may be used to develop sustainable bio-insecticides and new green procedures.

Chronic intermittent convection-enhanced delivery of vigabatrin to the bilateral subthalamic nucleus in an acute rat seizure model

Targeted intracerebral drug delivery is an attractive experimental approach for the treatment of drug-resistant epilepsies. In this regard, the subthalamic nucleus (STN) represents a focus-independent target involved in the remote modulation and propagation of seizure activity. Indeed, acute and chronic pharmacological inhibition of the STN with vigabatrin (VGB), an irreversible inhibitor of GABA transaminase, has been shown to produce antiseizure effects. This effect, however, is lost over time as tolerance develops with chronic, continuous intracerebral pharmacotherapy. Here we investigated the antiseizure effects of chronic intermittent intra-STN convection-enhanced delivery of VGB in an acute rat seizure model focusing on circumventing tolerance development and preventing adverse effects. Timed intravenous pentylenetetrazol (PTZ) seizure threshold testing was conducted before and after implantation of subcutaneous drug pumps and bilateral intra-STN cannulas. Drug pumps infused vehicle or VGB twice daily (0.4 µg) or once weekly (2.5 µg, 5 µg) over three weeks. Putative adverse effects were evaluated and found to be prevented by intermittent compared to previous continuous VGB delivery. Clonic seizure thresholds were more clearly raised by intra-STN VGB compared to myoclonic twitch. Both twice daily and once weekly intra-STN VGB significantly elevated clonic seizure thresholds depending on dose and time point, with responder rates of up to 100% observed at tolerable doses. However, tolerance could not be completely avoided, as tolerance rates of 40–75% were observed with chronic VGB treatment. Results indicate that the extent of tolerance development after intermittent intra-STN VGB delivery varies depending on infusion dose and regimen.

Evaluation of the pharmacological profile of GABA aminotransferase inhibitor diels-alder adducts with potential anticonvulsant activity

Evaluation of the pharmacological profile of GABA aminotransferase inhibitor diels-alder adducts with potential anticonvulsant activity

Abstract 257: VIGAB-STAT- A Phase IIa Feasibility Trial of Irreversible GABA-Transaminase Inhibition as Adjunct Treatment of Status Epilepticus After Cardiac Arrest

Background: Effective therapeutic interventions for post-anoxic status epilepticus (PASE) are lacking. Vigabatrin (VGB) could be an attractive adjunct drug in PASE by increasing GABA availability in inhibitory synapses; however, enteral malabsorption in the post-cardiac arrest (CA) period and difficulty enrolling in the FDA mandatory registry could hinder its use in this setting. We hypothesize that CA survivors absorb VGB despite vasopressor use, concurrent enteral feeding, and regardless of gastric or post-pyloric drug delivery. Goals: Prove feasibility and study pharmacokinetics of a single enteric VGB load within 48 hours of PASE onset. Methods: We administered a single load 1125-4500 mg VGB (based on creatinine clearance; CrCl) in adults with non-traumatic CA, with electrographic SE and gastric or duodenal access. Plasma samples were collected at 0, 0.5, 1, 2, 3, 6, 12, 24, 48, 72 and 168 hours. Results: Interim results from a cohort of 6 subjects comprised of 67% male (4 of 6), 83% White (5 of 6), with a median age 62 years 22-68), median BMI 30 Kg/m 2 (20-53), and median CrCl 44cc/min (22-186) are presented. All subjects received VGB within 48h window; median VGB dose was 2250 mg (1125-4500). The median time to peak (Tmax) was 2 h (1-3); median peak concentration (Cmax) was 38.1 mcg/ml (16.7 -91.8). Median 24h VGB concentration (C24) was 9.4 mcg/ml (3.4-14.2) and half-life (T1/2) was 16 h (10-24). The AUC 0-24 value was 318 mcg*h/ml (262-1012). No significant association was found between sex or BMI and Cmax. Three out of six patients achieved EEG response following VGB load; no association with dose or concentration was noted. Conclusions: All patients achieved detectable VGB levels with peak occurring between 1-3 hours post-administration. Enteric administration of VGB does not preclude phase IIb clinical trials of VGB in PASE. Registration: URL: http://www.clinicaltrials.gov ; Unique identifier: NCT04772547

Sex-related exacerbation of injury-induced mechanical hypersensitivity in GAD67 haplodeficient mice.

Decreased GABA levels in injury-induced loss of spinal inhibition are still under intense interest and debate. Here, we show that GAD67 haplodeficient mice exhibited a prolonged injury-induced mechanical hypersensitivity in postoperative, inflammatory, and neuropathic pain models. In line with this, we found that loss of 1 copy of the GAD67-encoding gene Gad1 causes a significant decrease in GABA contents in spinal GABAergic neuronal profiles. Consequently, GAD67 haplodeficient males and females were unresponsive to the analgesic effect of diazepam. Remarkably, all these phenotypes were more pronounced in GAD67 haplodeficient females. These mice had significantly much lower amount of spinal GABA content, exhibited an exacerbated pain phenotype during the second phase of the formalin test, developed a longer lasting mechanical hypersensitivity in the chronic constriction injury of the sciatic nerve model, and were unresponsive to the pain relief effect of the GABA-transaminase inhibitor phenylethylidenehydrazine. Our study provides strong evidence for a role of GABA levels in the modulation of injury-induced mechanical pain and suggests a potential role of the GABAergic system in the prevalence of some painful diseases among females.

Exogenous γ-aminobutyric acid (GABA) alleviates nitrogen deficiency by mediating nitrate uptake and assimilation in Andrographis paniculata seedlings

γ-Aminobutyric acid (GABA) plays significant metabolic and signaling roles in plant stress responses. Recent studies have proposed that GABA alleviates plant nitrogen (N) deficient stress; however, the mechanism by which GABA mediates plant N deficiency adaptation remains not yet well understood. Herein we found in a medicinal plant Andrographis paniculata that 5 mmol L−1 exogenous GABA promoted plant growth under N deficient (1 mmol L−1 NO3−) condition, with remarkably increments in total N and NO3− concentrations in plants. GABA increased N assimilation and protein synthesis by up-regulating the activities and expression of N metabolic enzymes. GABA also increased the accumulation of α-ketoglutarate and malate, which could facilitate the assimilation of NO3−. Inhibition of NR by Na2WO4 counteracted the promoting effects of GABA on plant growth, and the effects of GABA were not affected by L-DABA and 3-MP, the inhibitors of GABA transaminase (GABA-T) and glutamate decarboxylase (GAD), respectively. These results suggested that the nutritional role of GABA was excluded in promoting plant growth under low N condition. The results of 15N isotopic tracing and NRTs transcription indicated that exogenous GABA could up-regulate NRT2.4 and NRT3.2 to increase plant NO3− uptake under N deficient condition. Interestingly, primidone, an inhibitor of GABA receptor, impeded the effects of GABA on plant growth and N accumulation. Thus, our results revealed that exogenous GABA acted as a signal to up-regulate NRTs via its receptor to increase NO3− uptake, and subsequently promoted NO3− assimilation to alleviate N deficiency in A. paniculata.

GABAergic Signaling during Spinal Cord Stimulation Reduces Cardiac Arrhythmias in a Porcine Model.

Neuraxial modulation, including spinal cord stimulation, reduces cardiac sympathoexcitation and ventricular arrhythmogenesis. There is an incomplete understanding of the molecular mechanisms through which spinal cord stimulation modulates cardiospinal neural pathways. The authors hypothesize that spinal cord stimulation reduces myocardial ischemia-reperfusion-induced sympathetic excitation and ventricular arrhythmias through γ-aminobutyric acid (GABA)-mediated pathways in the thoracic spinal cord. Yorkshire pigs were randomized to control (n = 11), ischemia-reperfusion (n = 16), ischemia-reperfusion plus spinal cord stimulation (n = 17), ischemia-reperfusion plus spinal cord stimulation plus γ-aminobutyric acid type A (GABAA) or γ-aminobutyric acid type B (GABAB) receptor antagonist (GABAA, n = 8; GABAB, n = 8), and ischemia-reperfusion plus GABA transaminase inhibitor (GABAculine, n = 8). A four-pole spinal cord stimulation lead was placed epidurally (T1 to T4). GABA modulating pharmacologic agents were administered intrathecally. Spinal cord stimulation at 50 Hz was applied 30 min before ischemia. A 56-electrode epicardial mesh was used for high-resolution electrophysiologic recordings, including activation recovery intervals and ventricular arrhythmia scores. Immunohistochemistry and Western blots were performed to measure GABA receptor expression in the thoracic spinal cord. Cardiac ischemia led to myocardial sympathoexcitation with reduction in activation recovery interval (mean ± SD, -42 ± 11%), which was attenuated by spinal cord stimulation (-21 ± 17%, P = 0.001). GABAA and GABAB receptor antagonists abolished spinal cord stimulation attenuation of sympathoexcitation (GABAA, -9.7 ± 9.7%, P = 0.043 vs. ischemia-reperfusion plus spinal cord stimulation; GABAB, -13 ± 14%, P = 0.012 vs. ischemia-reperfusion plus spinal cord stimulation), while GABAculine alone caused a therapeutic effect similar to spinal cord stimulation (-4.1 ± 3.7%, P = 0.038 vs. ischemia-reperfusion). The ventricular arrhythmia score supported these findings. Spinal cord stimulation during ischemia-reperfusion increased GABAA receptor expression with no change in GABAB receptor expression. Thoracic spinal cord stimulation reduces ischemia-reperfusion-induced sympathoexcitation and ventricular arrhythmias through activation of GABA signaling pathways. These data support the hypothesis that spinal cord stimulation-induced release of GABA activates inhibitory interneurons to decrease primary afferent signaling from superficial dorsal horn to sympathetic output neurons in the intermediolateral nucleus.

Anticonvulsant appraisal of benzylideneacetophenone analogues in Swiss mice

Background: Diketones and α,β-unsaturated ketone are useful intermediates for the synthesis of antiepileptic moieties. Epilepsy is a neurological disorder that affects about fififty million people worldwide. The aim of the study was to assess benzylideneacetophenone analogues for their potential antiepileptic effects in mice. Methods: The compounds used in the pharmacological assay which include; 4-Dimethylaminochalcone (A), 1(4-Nitrophenyl) – 3 (ϒ -benzopyranoyl)-2-propen – 1-one (B), 1(4' –Nitrophenyl) – 3 (1,3-benzodioxolyl) 2 – propen – 1 – one (C), 1-Phenyl-3- (benzodioxolyl) – 2 –propen – 1-one (D), 4-Chlorochalcone (E), 4'-Nitro-4-dimethylaminochalcone (F) 4'-Nitro-4-methoxychalcone (G), were assayed using three models; Strychnine, Pentylenetetrazole (PTZ) and Maximal Electroshock (MES). The samples were given at 500, 1000 and 1500 mg/kg while standard drug diazepam and distilled water were given at 5 mg and 10 ml/kg intraperitoneal respectively. Convulsion inducing agents (Strychnine 2 mg/kg and PTZ, 100 mg/kg) were administered intraperitoneal 1hour after the administration of test samples while MES model electroconvulsive shock was applied using electroconvulsive machine.  Results: The samples (A-G) showed significant (P<0.03, 0.003, 0.0007 and 0.0001) effect in delaying onset of seizure compared to control in strychnine model, while in PTZ model samples (A-F) evaluated at 500,1000 and 1500 mg/kg are equipotent at all the doses by signifificantly (P<0.0001) and sample G (P<0.0007) reduced the onset of seizure compared to control. However in MES model samples (A-F and G) evaluated at 500, 1000 and 1500 mg/kg are equipotent at all the doses by significantly (P<0.0001; 0.0007), reduced the duration of seizure compared to control respectively.  Conclusion: Benzylideneacetophenone analogues showed various effects in control of seizure in all the models used.This effect is due to delayed onset and reduction of seizure duration. The samples could act by stimulating the glycine, Gamma Aminobutyric acid (GABA) receptor, inhibition of GABAAminotransferase and prolong inactivation of sodium ion channel.

Early Developmental PMCA2b Expression Protects From Ketamine-Induced Apoptosis and GABA Impairments in Differentiating Hippocampal Progenitor Cells.

PMCA2 is not expressed until the late embryonic state when the control of subtle Ca2+ fluxes becomes important for neuronal specialization. During this period, immature neurons are especially vulnerable to degenerative insults induced by the N-methyl-D-aspartate (NMDA) receptor blocker, ketamine. As H19-7 hippocampal progenitor cells isolated from E17 do not express the PMCA2 isoform, they constitute a valuable model for studying its role in neuronal development. In this study, we demonstrated that heterologous expression of PMCA2b enhanced the differentiation of H19-7 cells and protected from ketamine-induced death. PMCA2b did not affect resting [Ca2+]c in the presence or absence of ketamine and had no effect on the rate of Ca2+ clearance following membrane depolarization in the presence of the drug. The upregulation of endogenous PMCA1 demonstrated in response to PMCA2b expression as well as ketamine-induced PMCA4 depletion were indifferent to the rate of Ca2+ clearance in the presence of ketamine. Yet, co-expression of PMCA4b and PMCA2b was able to partially restore Ca2+ extrusion diminished by ketamine. The profiling of NMDA receptor expression showed upregulation of the NMDAR1 subunit in PMCA2b-expressing cells and increased co-immunoprecipitation of both proteins following ketamine treatment. Further microarray screening demonstrated a significant influence of PMCA2b on GABA signaling in differentiating progenitor cells, manifested by the unique regulation of several genes key to the GABAergic transmission. The overall activity of glutamate decarboxylase remained unchanged, but Ca2+-induced GABA release was inhibited in the presence of ketamine. Interestingly, PMCA2b expression was able to reverse this effect. The mechanism of GABA secretion normalization in the presence of ketamine may involve PMCA2b-mediated inhibition of GABA transaminase, thus shifting GABA utilization from energetic purposes to neurosecretion. In this study, we show for the first time that developmentally controlled PMCA expression may dictate the pattern of differentiation of hippocampal progenitor cells. Moreover, the appearance of PMCA2 early in development has long-standing consequences for GABA metabolism with yet an unpredictable influence on GABAergic neurotransmission during later stages of brain maturation. In contrast, the presence of PMCA2b seems to be protective for differentiating progenitor cells from ketamine-induced apoptotic death.

Structure-guided discovery of food-derived GABA-T inhibitors as hunters for anti-anxiety compounds.

With the acceleration of the pace of life, people may face all kinds of pressure, and anxiety has become a common mental issue that is seriously affecting human life. Safe and effective food-derived compounds may be used as anti-anxiety compounds. In this study, anti-anxiety compounds were collected and curated for database construction. Quantitative structure-activity relationship (QSAR) models were developed using a combination of various machine-learning approaches and chemical descriptors to predict natural compounds in food with anti-anxiety effects. High-throughput molecular docking was used to screen out compounds that could function as anti-anxiety molecules by inhibiting γ-aminobutyrate transaminase (GABA-T) enzyme, and 7 compounds were screened for in vitro activity verification. Pharmacokinetic analysis revealed three compounds (quercetin, lithocholic acid, and ferulic acid) that met Lipinski's Rule of Five and inhibited the GABA-T enzyme to alleviate anxiety in vitro. The established QSAR model combined with molecular docking and molecular dynamics was proved by the synthesis and discovery of novel food-derived anti-anxiety compounds.

GABAergic effects of some foods extracts via inhibition of GABA transaminase (GABA-T)

Objective: GABAergic system is a target for various groups of medications including sedatives, anxiolytics, muscle relaxants, antidepressants and antiepileptics. Several foods or food ingredients are able to affect the GABAergic system by the inhibition of the γ-aminobutyric acid (GABA) degrading enzymes including the GABA-transaminase and succinate semialdehyde dehydrogenase. The purpose of this study to investigate the inhibitory effects of tea (Camellia sinensis), coffee (Coffea arabica L.), peppermint (Mentha piperita L.), thyme (Thymus vulgaris L.), and cinnamon (Cinnamomum zeylanicum) on GABA degrading enzymes.Methods: The inhibition of the GABA-T by aqueous extracts of tea (Camellia sinensis), coffee (Coffea arabica L.), peppermint (Mentha piperita L.), cinnamon (Cinnamomum zeylanicum), and thyme (Thymus vulgaris L.) was investigated using a fluorometric microplate enzyme assay. Dose-dependent inhibition of the GABA-degrading enzymes was attained by all the food extracts tested. For determination of the IC50 values of the extracts (± 95 % CI), a linear regression was performed using Origin® (Origin® 2015G von Origin Lab Corporation, Northampton, MA 01060 USA).Results: The aqueous extract of black tea presented the strongest inhibitory activity with an IC50-value (half maximal inhibitory concentration) of 13.0 (11.0-15.3) µg/mL. The tested food extracts were successful in inhibiting the GABA-degrading enzymes even at low concentrations. Conclusion: In conclusion, the selected food extracts could serve as natural inhibitors for GABA-degrading enzymes thus, they could increase the GABA concentration in the brain.

Inhibition of GABA-Transaminase and GABA-Transporters in the Brain by Vigabatrin and Tiagabine Prevents Seizure Development in Rats Breathing Hyperbaric Oxygen

Inhibition of GABA-Transaminase and GABA-Transporters in the Brain by Vigabatrin and Tiagabine Prevents Seizure Development in Rats Breathing Hyperbaric Oxygen

Dietary GABA and its combination with vigabatrin mimic calorie restriction and induce antiobesity-like effects in lean mice

The anti-obesity and anti-diabetic effects of γ-aminobutyric acid (GABA) are mainly expressed via glucose and insulin regulation in mice. It is unknown whether dietary GABA exerts anti-obesity effects via other pathways. Herein, we report that a high dietary GABA intake (5%) significantly suppressed food intake (−30%), body-weight gain, and fat accumulation, induced ketogenesis, improved glucose metabolism, and elevated the levels of circulating GABA and β-aminoisobutyric acid. These changes suggest that 5% GABA intake possibly induces calorie restriction. Interestingly, a combination of low-dose dietary GABA (0.5% and 2%) and vigabatrin (inhibitor of GABA transaminase (GABA-T)) markedly increased the levels of circulating GABA and strongly exerted antiobesity-like effects. Brain GABA levels increased slightly upon 5% GABA intake, but significantly upon intake of the low-dose GABA–vigabatrin combination. Therefore, the manipulation of peripheral and brain GABA metabolism by targeting GABA-T may lead to the development of novel interventions for overeating and obesity.

Ингибирование ГАМК-трансаминазы и ГАМК-транспортеров в головном мозге с помощью вигабатрина и тиагабина ослабляет развитие судорог у крыс при дыхании гипербарическим кислородом

Hyperbaric oxygen (HBO2) inhibits the GABAergic function in the brain, which leads to the development of convulsive reactions in the form of paroxysmal discharges on the EEG and movement disorders, similar to a generalized epileptic seizure. We hypothesize that the deficiency of GABAergic neurotransmission in HBO2, which occurs due to suppression of GABA synthesis, can be overcome by blocking the clearance of GABA from the synaptic cleft and slowing down its enzymatic degradation. The present study is aimed at studying the development of seizure reactions in rats under oxygen pressure of 5 ATA after blockade of GABA transporters with tiagabine and inhibition of GABA-transaminase with vigabatrin. New data obtained in this study are: (1) inhibition of GABA transaminase by vigabatrin or GABA transporters by tiagabine prevented the development of seizures in rats under oxygen pressure of 5 ATA; (2) oxygen convulsions in HBO2 appeared with a decrease in the brain GABA by 30-40% from the initial level; (3) inhibition of GABA transaminase by vigabatrin prevented a decrease in GABA in the striatum of rats under 5 ATA pressure. Consequently, a slowdown in synaptic clearance of GABA or a weakening of the enzymatic destruction of a mediator can compensate reduced GABA synthesis in HBO2 and increase its concentration to a level sufficient for the restoration of GABAergic function preventing the development of convulsive syndrome.

High-yield synthesis and purification of recombinant human GABA transaminase for high-throughput screening assays

Many studies have focussed on modulating the activity of γ-aminobutyric acid transaminase (GABA-T), a GABA-catabolizing enzyme, for treating neurological diseases, such as epilepsy and drug addiction. Nevertheless, human GABA-T synthesis and purification have not been established. Thus, biochemical and drug design studies on GABA-T have been performed by using porcine GABA-T mostly and even bacterial GABA-T. Here we report an optimised protocol for overexpression of 6xHis-tagged human GABA-T in human cells followed by a two-step protein purification. Then, we established an optimised human GABA-T (0.5 U/mg) activity assay. Finally, we compared the difference between human and bacterial GABA-T in sensitivity to two irreversible GABA-T inhibitors, gabaculine and vigabatrin. Human GABA-T in homodimeric form showed 70-fold higher sensitivity to vigabatrin than bacterial GABA-T in multimeric form, indicating the importance of using human GABA-T. In summary, our newly developed protocol can be an important first step in developing more effective human GABA-T modulators.

A Critical Role of Hepatic GABA in the Metabolic Dysfunction and Hyperphagia of Obesity

Hepatic lipid accumulation is a hallmark of type II diabetes (T2D) and associated with hyperinsulinemia, insulin resistance, and hyperphagia. Hepatic synthesis of GABA, catalyzed by GABA-transaminase (GABA-T), is upregulated in obese mice. To assess the role of hepatic GABA production in obesity-induced metabolic and energy dysregulation, we treated mice with two pharmacologic GABA-T inhibitors and knocked down hepatic GABA-T expression using an antisense oligonucleotide. Hepatic GABA-T inhibition and knockdown decreased basal hyperinsulinemia and hyperglycemia, and improved glucose intolerance. GABA-T knockdown improved insulin sensitivity assessed by hyperinsulinemic-euglycemic clamps in obese mice. Hepatic GABA-T knockdown also decreased food intake and induced weight loss without altering energy expenditure in obese mice. Data from people with obesity support the notion that hepatic GABA production and transport are associated with serum insulin, HOMA-IR, T2D, and BMI. These results support a key role for hepatocyte GABA production in the dysfunctional glucoregulation and feeding behavior associated with obesity.