Succinic semialdehyde dehydrogenase deficiency: a metabolic and genomic approach to diagnosis.

Genomic sequencing offers an untargeted, data-driven approach to genetic diagnosis; however, variants of uncertain significance often hinder the diagnostic process. The discovery of rare genomic variants without previously known functional evidence of pathogenicity often results in variants being overlooked as potentially causative, particularly in individuals with undifferentiated phenotypes. Consequently, many neurometabolic conditions, including those in the GABA (gamma-aminobutyric acid) catabolism pathway, are underdiagnosed. Succinic semialdehyde dehydrogenase deficiency (SSADHD, OMIM #271980) is a neurometabolic disorder in the GABA catabolism pathway. The disorder is due to bi-allelic pathogenic variants in ALDH5A1 and is usually characterized by moderate-to-severe developmental delays, hypotonia, intellectual disability, ataxia, seizures, hyperkinetic behavior, aggression, psychiatric disorders, and sleep disturbances. In this study, we utilized an integrated approach to diagnosis of SSADHD by examining molecular, clinical, and metabolomic data from a single large commercial laboratory. Our analysis led to the identification of 16 patients with likely SSADHD along with three novel variants. We also showed that patients with this disorder have a clear metabolomic signature that, along with molecular and clinical findings, may allow for more rapid and efficient diagnosis. We further surveyed all available pathogenic/likely pathogenic variants and used this information to estimate the global prevalence of this disease. Taken together, our comprehensive analysis allows for a global approach to the diagnosis of SSADHD and provides a pathway to improved diagnosis and potential incorporation into newborn screening programs. Furthermore, early diagnosis facilitates referral to genetic counseling, family support, and access to targeted treatments-taken together, these provide the best outcomes for individuals living with either GABA-TD or SSADHD, as well as other rare conditions.

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.

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.

γ-Aminobutyric acid (GABA) improves pesticide detoxification in plants.

It is important to ensure food safety to study the technology and mechanism of pesticide residues degradation in crops. Though γ-aminobutyric acid (GABA) has been widely reported to involve in plant stress resistance, whether exogenous application or endogenous regulation of GABA by gene-editing technology can promote the pesticide detoxification is not clear in plants. Using tomato and chlorothalonil (CHT) as research models, we discovered that exogenous application of GABA or endogenous elevation of GABA by knockout of pyruvate-dependent GABA transaminase promoted both CHT metabolism and plant stress tolerance to CHT. This is closely related to the active adaptation of GABA to CHT stress by regulating the plant GABA shunt pathway and polyamine pathway. The transcriptome data revealed 17 target genes that may be closely related to the involvement of GABA in CHT metabolism, including 4 peroxidases, 5 glycosyltransferases, 4 glutathione S-transferases, and 4 ABC transporters. In addition, the glutathione detoxification pathway and antioxidative enzyme also actively participated in the GABA-induced CHT detoxification process, which played an important role in relieving CHT stress. As a result, GABA significantly increased the photosynthetic capacity of tomato leaves under CHT stress. While studying photosynthesis, we unexpectedly found that GABA promotes stomatal closure in terms of decreased stomatal conductance and stomatal diameter. This result implies that GABA can reduce CHT absorption by regulating stomatal movement in leaves. Together, we provided a novel viewpoint that foliar application of GABA or metabolic engineering of GABA is an effective approach to reduce the risk of pesticide contamination in crop production.

Characterization of GABA-Transaminase Gene from Mulberry (Morus multicaulis) and Its Role in Salt Stress Tolerance.

Gamma-aminobutyric acid (GABA) has been reported to accumulate in plants when subjected to salt stress, and GABA-transaminase (GABA-T) is the main GABA-degrading enzyme in the GABA shunt pathway. So far, the salt tolerance mechanism of the GABA-T gene behind the GABA metabolism remains unclear. In this study, the cDNA (designated MuGABA-T) of GABA-T gene was cloned from mulberry, and our data showed that MuGABA-T protein shares some conserved characteristics with its homologs from several plant species. MuGABA-T gene was constitutively expressed at different levels in mulberry tissues, and was induced substantially by NaCl, ABA and SA. In addition, our results demonstrated that exogenous application of GABA significantly reduced the salt damage index and increased plant resistance to NaCl stress. We further performed a functional analysis of MuGABA-T gene and demonstrated that the content of GABA was reduced in the transgenic MuGABA-T Arabidopsis plants, which accumulated more ROS and exhibited more sensitivity to salt stress than wild-type plants. However, exogenous application of GABA significantly increased the activities of antioxidant enzymes and alleviated the active oxygen-related injury of the transgenic plants under NaCl stress. Moreover, the MuGABA-T gene was overexpressed in the mulberry hairy roots, and similar results were obtained for sensitivity to salt stress in the transgenic mulberry plants. Our results suggest that the MuGABA-T gene plays a pivotal role in GABA catabolism and is responsible for a decrease in salt tolerance, and it may be involved in the ROS pathway in the response to salt stress. Taken together, the information provided here is helpful for further analysis of the function of GABA-T genes, and may promote mulberry resistance breeding in the future.

Not Just a Cycle: Three gab Genes Enable the Non-Cyclic Flux Toward Succinate via GABA Shunt in 'Candidatus Liberibacter asiaticus'-Infected Citrus.

Although the mitochondria retain all required enzymes for an intact tricarboxylic acid (TCA) cycle, plants might shift the cyclic flux from the TCA cycle to an alternative noncyclic pathway via γ-aminobutyric acid (GABA) shunt under specific physiological conditions. We hypothesize that several genes may ease this noncyclic flux and contribute to the citrus response to the phytopathogenic bacterium 'Candidatus Liberibacter asiaticus', the causal agent of Huanglongbing in citrus. To test this hypothesis, we used multiomics techniques (metabolomics, fluxomics, and transcriptomics) to investigate the potential roles of putative gab homologies from Valencia sweet orange (Citrus sinensis). Our findings showed that 'Ca. L. asiaticus' significantly increased the endogenous GABA and succinate content but decreased ketoglutarate in infected citrus plants. Citrus genome harbors three putative gab genes, including amino-acid permease (also known as GABA permease; CsgabP), GABA transaminase (CsgabT), and succinate-semialdehyde dehydrogenase (also known as GABA dehydrogenase; CsgabD). The transcript levels of CsgabP, CsgabT, and CsgabD were upregulated in citrus leaves upon the infection with 'Ca. L. asiaticus' and after the exogenous application of GABA or deuterium-labeled GABA isotope (GABA-D6). Moreover, our finding showed that exogenously applied GABA is quickly converted to succinate and fed into the TCA cycle. Likewise, the fluxomics study showed that GABA-D6 is rapidly metabolized to succinate-D4. Our work proved that GABA shunt and three predicated gab genes from citrus, support the upstream noncyclic flux toward succinate rather than an intact TCA cycle and contribute to citrus defense responses to 'Ca. L. asiaticus'.[Formula: see text] Copyright © 2022 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

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) 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, homeostatic model assessment for insulin resistance (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.

Hepatocyte membrane potential regulates serum insulin and insulin sensitivity by altering hepatic GABA release.

Hepatic lipid accumulation in obesity correlates with the severity of hyperinsulinemia and systemic insulin resistance. Obesity-induced hepatocellular lipid accumulation results in hepatocyte depolarization. We have established that hepatocyte depolarization depresses hepatic afferent vagal nerve firing, increases GABA release from liver slices, and causes hyperinsulinemia. Preventing hepatic GABA release or eliminating the ability of the liver to communicate to the hepatic vagal nerve ameliorates the hyperinsulinemia and insulin resistance associated with diet-induced obesity. In people with obesity, hepatic expression of GABA transporters is associated with glucose infusion and disposal rates during a hyperinsulinemic euglycemic clamp. Single-nucleotide polymorphisms in hepatic GABA re-uptake transporters are associated with an increased incidence of type 2 diabetes mellitus. Herein, we identify GABA as a neuro-hepatokine that is dysregulated in obesity and whose release can be manipulated to mute or exacerbate the glucoregulatory dysfunction common to obesity.

Effects of two kinds of fishery drugs on the expressions of GAD and GABA-T mRNA in crucian carp (Carassius auratus gibelio).

The objective of this study was to investigate the effects of difloxacin (DIF) and avermectin (AVM) on glutamate decarboxylase (GAD) and GABA-transaminase (GABA-T) in different tissues of crucian carp (Carassius auratus gibelio). After the treatments of DIF and AVM, the mRNA expressions of GAD and GABA-T in different tissues were detected by quantitative real-time PCR (qPCR). The results showed that the mRNA expressions of GAD65, GAD67, and GABA-T in the telencephalon (Tel), mesencephalon (Mes), cerebella (Cer), and medulla oblongata (Med) were downregulated significantly with the safe dose (SD, 20 mg/kg) of DIF (P < 0.05 or P < 0.01). While the expressions of GAD65 and GAD67 in the kidney at 12 h had strikingly upregulated to 13.81 ± 1.06** and 150.67 ± 12.85** times. Treated with the lethal dose of 50% (LD50, 2840 mg/kg b. W.) of DIF, the mRNA expressions of GAD65, GAD67, and GABA-T in all tissues were increased significantly (P < 0.01). The results of AVM group showed that the mRNA expressions of GAD65, GAD67, and GABA-T both in the central and peripheral tissues were all remarkably downregulated at the safe concentration (SC, 0.0039 mg/L) and the lethal concentration of 50% (LC50, 0.039 mg/L), except for the mRNA inhibitions of GAD65, GAD67, and GABA-T in the muscle at 2 h which sharply downregulated to 0.20 ± 0.02ΔΔ × 10-2, 0.57 ± 0.06ΔΔ × 10-1 and 0.44 ± 0.02ΔΔ × 10-1, respectively (P < 0.01).

Role of pyridoxine in GABA synthesis and degradation in the hippocampus.

Pyridoxal-5'-phosphate, the active form of vitamin B6, is associated with activities of several enzymes and the treatment of various neurological disorders. Here, we investigated the effects of pyridoxine on the immunoreactivity and protein levels of γ-aminobutyric acid (GABA)-synthesizing and degradation enzymes such as glutamic acid decarboxylase (GAD), GABA transaminase (GABA-T), and succinic semialdehyde dehydrogenase (SSADH), in the hippocampus of mice. The mice intraperitonially received physiological saline and 350 mg/kg pyridoxine, twice a day for 21 days, and were euthanized 2 h after the final dose. In the vehicle-treated group, we observed GAD67 immunoreactivity in the stratum pyramidale of the CA1 and CA3 region, Schaffer collateral, polymorphic layer, and outer granule cell layer of the dentate gyrus. Pyridoxine administration significantly increased GAD67 immunoreactivity, while significantly decreasing GABA-T immunoreactivity in pyridoxine-treated mouse hippocampi (CA1 region and dentate gyrus). In the stratum lacunosum-moleculare of CA1 region, GABA-T immunoreactivity was significantly increased in the pyridoxine-treated group compared to that in the vehicle-treated group, although GAD67 immunoreactivity was similarly observed in these groups. Alternatively, there were no significant differences in SSADH immunoreactivity in any regions of the hippocampus between the vehicle- and pyridoxine-treated groups. Western blot analysis showed significant increases in GAD67 and GABA-T protein levels in the pyridoxine-treated group compared with those in the vehicle-treated group. Therefore, pyridoxine administration facilitates GABA turnover in mouse hippocampus by modulating the GABA-synthesizing and degradation enzymes.

Super-Agrobacterium ver. 4: Improving the Transformation Frequencies and Genetic Engineering Possibilities for Crop Plants.

Agrobacterium tumefaciens has been utilized for both transient and stable transformations of plants. These transformation methods have been used in fields such as breeding GM crops, protein production in plant cells, and the functional analysis of genes. However, some plants have significantly lower transient gene transfer and stable transformation rates, creating a technical barrier that needs to be resolved. In this study, Super-Agrobacterium was updated to ver. 4 by introducing both the ACC deaminase (acdS) and GABA transaminase (gabT) genes, whose resultant enzymes degrade ACC, the ethylene precursor, and GABA, respectively. A. tumefaciens strain GV2260, which is similar to other major strains (EHA105, GV3101, LBA4404, and MP90), was used in this study. The abilities of the Super-Agrobacterium ver. 4 were evaluated in Erianthus ravennae, Solanum lycopersicum "Micro-Tom," Nicotiana benthamiana, and S. torvum. Super-Agrobacterium ver. 4 showed the highest T-DNA transfer (transient transformation) frequencies in E. ravennae and S. lycopersicum, but not in N. benthamiana and S. torvum. In tomato, Super-Agrobacterium ver. 4 increased the stable transformation rate by 3.6-fold compared to the original GV2260 strain. Super-Agrobacterium ver. 4 enables reduction of the amount of time and labor required for transformations by approximately 72%, and is therefore a more effective and powerful tool for plant genetic engineering and functional analysis, than the previously developed strains. As our system has a plasmid containing the acdS and gabT genes, it could be used in combination with other major strains such as EHA105, EHA101, LBA4404, MP90, and AGL1. Super-Agrobacterium ver. 4, could thus possibly be a breakthrough application for improving basic plant science research methods.

2-Pyrrolidinone and Succinimide as Clinical Screening Biomarkers for GABA-Transaminase Deficiency: Anti-seizure Medications Impact Accurate Diagnosis.

Broad-scale untargeted biochemical phenotyping is a technology that supplements widely accepted assays, such as organic acid, amino acid, and acylcarnitine analyses typically utilized for the diagnosis of inborn errors of metabolism. In this study, we investigate the analyte changes associated with 4-aminobutyrate aminotransferase (ABAT, GABA transaminase) deficiency and treatments that affect GABA metabolism. GABA-transaminase deficiency is a rare neurodevelopmental and neurometabolic disorder caused by mutations in ABAT and resulting in accumulation of GABA in the cerebrospinal fluid (CSF). For that reason, measurement of GABA in CSF is currently the primary approach to diagnosis. GABA-transaminase deficiency results in severe developmental delay with intellectual disability, seizures, and movement disorder, and is often associated with death in childhood. Using an untargeted metabolomics platform, we analyzed EDTA plasma, urine, and CSF specimens from four individuals with GABA-transaminase deficiency to identify biomarkers by comparing the biochemical profile of individual patient samples to a pediatric-centric population cohort. Metabolomic analyses of over 1,000 clinical plasma samples revealed a rich source of biochemical information. Three out of four patients showed significantly elevated levels of the molecule 2-pyrrolidinone (Z-score ≥2) in plasma, and whole exome sequencing revealed variants of uncertain significance in ABAT. Additionally, these same patients also had elevated levels of succinimide in plasma, urine, and CSF and/or homocarnosine in urine and CSF. In the analysis of clinical EDTA plasma samples, the levels of succinimide and 2-pyrrolidinone showed a high level of correlation (R = 0.73), indicating impairment in GABA metabolism and further supporting the association with GABA-transaminase deficiency and the pathogenicity of the ABAT variants. Further analysis of metabolomic data across our patient population revealed the association of elevated levels of 2-pyrrolidinone with administration of vigabatrin, a commonly used anti-seizure medication and a known inhibitor of GABA-transaminase. These data indicate that anti-seizure medications may alter the biochemical and metabolomic data, potentially impacting the interpretation and diagnosis for the patient. Further, these data demonstrate the power of combining broad scale genotyping and phenotyping technologies to diagnose inherited neurometabolic disorders and support the use of metabolic phenotyping of plasma to screen for GABA-transaminase deficiency.

Attenuation of the effects of oxidative stress by the MAO-inhibiting antidepressant and carbonyl scavenger phenelzine.

Phenelzine (ß-phenylethylhydrazine) is a monoamine oxidase (MAO)-inhibiting antidepressant with anxiolytic properties. It possesses a number of important pharmacological properties which may alter the effects of oxidative stress. After conducting a comprehensive literature search, the authors of this review paper aim to provide an overview and discussion of the mechanisms by which phenelzine may attenuate oxidative stress. It inhibits γ-aminobutyric acid (GABA) transaminase, resulting in elevated brain GABA levels, inhibits both MAO and primary amine oxidase and, due to its hydrazine-containing structure, reacts chemically to sequester a number of reactive aldehydes (e.g. acrolein and 4-hydroxy-2-nonenal) proposed to be implicated in oxidative stress in a number of neurodegenerative disorders. Phenelzine is unusual in that it is both an inhibitor of and a substrate for MAO, the latter action producing at least one active metabolite, ß-phenylethylidenehydrazine (PEH). This metabolite inhibits GABA transaminase, is a very weak inhibitor of MAO but a strong inhibitor of primary amine oxidase, and sequesters aldehydes. Phenelzine may ameliorate the effects of oxidative stress by reducing formation of reactive metabolites (aldehydes, hydrogen peroxide, ammonia/ammonia derivatives) produced by the interaction of MAO with biogenic amines, by sequestering various other reactive aldehydes and by inhibiting primary amine oxidase. In PC12 cells treated with the neurotoxin MPP+, phenelzine has been reported to reduce several adverse effects of MPP+. It has also been reported to reduce lipid peroxidative damage induced in plasma and platelet proteins by peroxynitrite. In animal models, phenelzine has a neuroprotective effect in global ischemia and in cortical impact traumatic brain injury. Recent studies reported in the literature on the possible involvement of acrolein in spinal cord injury and multiple sclerosis indicate that phenelzine can attenuate adverse effects of acrolein in these models. Results from studies in our laboratories on effects of phenelzine and PEH on primary amine oxidase (which catalyzes formation of toxic aldehydes and is overexpressed in Alzheimer's disease), on sequestration of the toxic aldehyde acrolein, and on reduction of acrolein-induced toxicity in mouse cortical neurons are also reported.

Metabolomic analyses of vigabatrin (VGB)-treated mice: GABA-transaminase inhibition significantly alters amino acid profiles in murine neural and non-neural tissues.

The anticonvulsant vigabatrin (VGB; SabrilR) irreversibly inhibits GABA transaminase to increase neural GABA, yet its mechanism of retinal toxicity remains unclear. VGB is suggested to alter several amino acids, including homocarnosine, ß-alanine, ornithine, glycine, taurine, and 2-aminoadipic acid (AADA), the latter a homologue of glutamic acid. Here, we evaluate the effect of VGB on amino acid concentrations in mice, employing a continuous VGB infusion (subcutaneously implanted osmotic minipumps), dose-escalation paradigm (35-140 mg/kg/d, 12 days), and amino acid quantitation in eye, visual and prefrontal cortex, total brain, liver and plasma. We hypothesized that continuous VGB dosing would reveal numerous hitherto undescribed amino acid disturbances. Consistent amino acid elevations across tissues included GABA, ß-alanine, carnosine, ornithine and AADA, as well as neuroactive aspartic and glutamic acids, serine and glycine. Maximal increase of AADA in eye occurred at 35 mg/kg/d (41 ±â€¯2 nmol/g (n = 21, vehicle) to 60 ±â€¯8.5 (n = 8)), and at 70 mg/kg/d for brain (97 ±â€¯6 (n = 21) to 145 ±â€¯6 (n = 6)), visual cortex (128 ±â€¯6 to 215 ±â€¯19) and prefrontal cortex (124 ±â€¯11 to 200 ±â€¯13; mean ±â€¯SEM; p < 0.05), the first demonstration of tissue AADA accumulation with VGB in mammal. VGB effects on basic amino acids, including guanidino-species, suggested the capacity of VGB to alter urea cycle function and nitrogen disposal. The known toxicity of AADA in retinal glial cells highlights new avenues for assessing VGB retinal toxicity and other off-target effects.

GABA Transaminase Deficiency With Survival Into Adulthood.

γ-Aminobutyric acid (GABA)-transaminase deficiency is an ultra-rare disorder of GABA metabolism that was described for decades as an early-onset epileptic encephalopathy plus movement disorder and hypersomnolence with mortality in early childhood. We report 2 affected siblings in adolescence and adulthood, both with profound developmental impairment, intractable epilepsy, movement disorder, and behavioral fluctuations. This considerably expands the phenotype and longevity of this inherited neurotransmitter disease.

Corrigendum: 2-Pyrrolidinone and Succinimide as Clinical Screening Biomarkers for GABA-Transaminase Deficiency: Anti-seizure Medications Impact Accurate Diagnosis.

[This corrects the article DOI: 10.3389/fnins.2019.00394.].

White matter spongiosis with vigabatrin therapy for infantile spasms.

The histopathology, "white matter spongiosis," defined by electron microscopy (EM) as "intramyelinic edema," has been associated with vigabatrin therapy in various animal models, but its role or significance in clinical studies is unknown. We conducted a neuropathological examination on a 27-month-old boy with bilateral polymicrogyria and epilepsy after sudden unexpected death in epilepsy (SUDEP). The patient was initiated on vigabatrin at 4 months of age, which controlled infantile spasms, and was continued as maintenance therapy. Autopsy showed a combination of developmental and acquired lesions: (1) bilateral gyral malformations of the frontal, parietal, temporal, and insular cortex; (2) agenesis of the olfactory tracts and bulbs; (3) hippocampal abnormalities: dentate gyrus bilamination and granule cell dispersion; and (4) areas of microscopic bilateral, symmetric white matter spongiosis in the brainstem central tegmental tract, amiculum and hilum of the inferior olive, medial longitudinal fasciculus, paragigantocellularis lateralis, optic nerves and chiasm, and hypothalamus. The white matter spongiosis was identical to the histopathologic lesions (which by EM exhibited intramyelinic edema) that were demonstrated in animal models on vigabatrin therapy, indicating that vigabatrin toxicity is not restricted to animal models.

Propionate enters GABAergic neurons, inhibits GABA transaminase, causes GABA accumulation and lethargy in a model of propionic acidemia.

Propionic acidemia is the accumulation of propionate in blood due to dysfunction of propionyl-CoA carboxylase. The condition causes lethargy and striatal degeneration with motor impairment in humans. How propionate exerts its toxic effect is unclear. Here, we show that intravenous administration of propionate causes dose-dependent propionate accumulation in the brain and transient lethargy in mice. Propionate, an inhibitor of histone deacetylase, entered GABAergic neurons, as could be seen from increased neuronal histone H4 acetylation in the striatum and neocortex. Propionate caused an increase in GABA (γ-amino butyric acid) levels in the brain, suggesting inhibition of GABA breakdown. In vitro propionate inhibited GABA transaminase with a Ki of ∼1 mmol/l. In isolated nerve endings, propionate caused increased release of GABA to the extracellular fluid. In vivo, propionate reduced cerebral glucose metabolism in both striatum and neocortex. We conclude that propionate-induced inhibition of GABA transaminase causes accumulation of GABA in the brain, leading to increased extracellular GABA concentration, which inhibits neuronal activity and causes lethargy. Propionate-mediated inhibition of neuronal GABA transaminase, an enzyme of the inner mitochondrial membrane, indicates entry of propionate into neuronal mitochondria. However, previous work has shown that neurons are unable to metabolize propionate oxidatively, leading us to conclude that propionyl-CoA synthetase is probably absent from neuronal mitochondria. Propionate-induced inhibition of energy metabolism in GABAergic neurons may render the striatum, in which >90% of the neurons are GABAergic, particularly vulnerable to degeneration in propionic acidemia.

Role of nitric oxide synthase on brain GABA transaminase activity and GABA levels.

In an attempt to clarify the controversial role of nitric oxide (NO) in seizures, the effects of NO on brain GABA transaminase (GABA-T) activity and GABA levels were investigated. To this aim, the effects of the substrate (l-arginine) and inhibitors (Nω-nitro-l-arginine methyl ester, 7-nitroindazole) of NO synthase (NOS) on GABA-T activity and GABA levels in vitro and ex vivo were analyzed. In vitro NO diminished GABA-T activity and increased GABA. Ex vivo NO modified GABA-T activity and GABA levels biphasically. Inhibition of endothelial and neuronal NOS (eNOS and nNOS) had opposite effects on GABA-T activity and GABA levels, even during seizures induced by pentylenetetrazole. Different effects of NO on GABA-T activity and on GABA levels, depending on the NOS isoform involved, may explain its contradictory role in seizures, the endothelial NOS acting as an anticonvulsant and the neuronal NOS as a proconvulsant. nNOS inhibitors may represent a new generation of antiepileptics.

Neuroinformatics analyses reveal GABAt and SSADH as major proteins involved in anticonvulsant activity of valproic acid.

The unequivocal hypotheses about anticonvulsant activity of valproic acid (VPA) have always been a basic hurdle in designing next generation neurotherapeutics, particularly the anti-epileptic drugs. The present study reports about a comprehensive in-silico investigation into qualitative and quantitative binding of VPA and corresponding natural ligands of four major enzymes involved in neurotransmissions, namely-GABA transaminase (GABAt), α-keto glutarate dehydrogenase (α-KGDH), Succinate Semialdehyde dehydrogenase (SSADH) and Glutamate Decarboxylase (GAD), respectively. The molecular docking analyses revealed that VPA inhibits GABAt and α-KGDH through allosteric while SSADH through competitive mode of binding. There is an observed elevation in binding of glutamate over GAD in the presence of VPA. The docking inhibition constant (Ki) of VPA to all the studied enzymatic receptors were observed to be well below the therapeutic concentration of VPA in blood, except for α-KGDH, thus favouring GABAergic over glutamatergic mode of anticonvulsant activity of VPA. The report is probably the first comprehensive in-silico molecular study about VPA action.