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Sample records for gamma aminobutyric acid

  1. Lactic acid bacterial cell factories for gamma-aminobutyric acid.

    PubMed

    Li, Haixing; Cao, Yusheng

    2010-11-01

    Gamma-aminobutyric acid is a non-protein amino acid that is widely present in organisms. Several important physiological functions of gamma-aminobutyric acid have been characterized, such as neurotransmission, induction of hypotension, diuretic effects, and tranquilizer effects. Many microorganisms can produce gamma-aminobutyric acid including bacteria, fungi and yeasts. Among them, gamma-aminobutyric acid-producing lactic acid bacteria have been a focus of research in recent years, because lactic acid bacteria possess special physiological activities and are generally regarded as safe. They have been extensively used in food industry. The production of lactic acid bacterial gamma-aminobutyric acid is safe and eco-friendly, and this provides the possibility of production of new naturally fermented health-oriented products enriched in gamma-aminobutyric acid. The gamma-aminobutyric acid-producing species of lactic acid bacteria and their isolation sources, the methods for screening of the strains and increasing their production, the enzymatic properties of glutamate decarboxylases and the relative fundamental research are reviewed in this article. And the potential applications of gamma-aminobutyric acid-producing lactic acid bacteria were also referred to.

  2. Increased brain uptake of gamma-aminobutyric acid in a rabbit model of hepatic encephalopathy

    SciTech Connect

    Bassett, M.L.; Mullen, K.D.; Scholz, B.; Fenstermacher, J.D.; Jones, E.A. )

    1990-03-01

    Transfer of the inhibitory neurotransmitter gamma-aminobutyric acid across the normal blood-brain barrier is minimal. One prerequisite for gamma-aminobutyric acid in plasma contributing to the neural inhibition of hepatic encephalopathy would be that increased transfer of gamma-aminobutyric acid across the blood-brain barrier occurs in liver failure. The aim of the present study was to determine if brain gamma-aminobutyric acid uptake is increased in rabbits with stage II-III (precoma) hepatic encephalopathy due to galactosamine-induced fulminant hepatic failure. A modification of the Oldendorf intracarotid artery-injection technique was applied. (3H) gamma-aminobutyric acid, (14C) butanol, and 113mIn-labeled serum protein (transferrin) were injected simultaneously 4 s before decapitation. The ipsilateral brain uptake index of gamma-aminobutyric acid was determined from measurements of the 3 isotopes in 5 brain regions. Uncorrected or simple brain uptake indices of (3H) gamma-aminobutyric acid and (113mIn) transferrin were calculated using (14C) butanol as the highly extracted reference compound. The (113mIn) transferrin data were also used to correct the brain uptake index of (3H) gamma-aminobutyric acid for intravascular retention of (3H) gamma-aminobutyric acid. The methodology adopted minimized problems attributable to rapid (3H) gamma-aminobutyric acid metabolism, and slow brain washout and recirculation of the radiolabeled tracers. Both the uncorrected and corrected brain uptake indices of gamma-aminobutyric acid as well as the simple brain uptake index of transferrin were significantly increased in both stage II and III hepatic encephalopathy in all brain regions studied. Moreover, these brain uptake indices were significantly greater in stage III hepatic encephalopathy than in stage II hepatic encephalopathy.

  3. Separation of gamma-aminobutyric acid from fermented broth.

    PubMed

    Li, Haixing; Qiu, Ting; Chen, Yan; Cao, Yusheng

    2011-12-01

    Gamma-aminobutyric acid (GABA) is a non-proteinaceous amino acid that is widely distributed in nature and acts as the major inhibitory neurotransmitter in the mammalian brain. This study aimed to find a separation method for getting high-purity GABA from a fermented broth. Firstly, a fermented broth with a high content of GABA (reaching 997 ± 51 mM) was prepared by fermentation with Lactobacillus brevis NCL912. GABA purification was conducted by successive centrifugation, filtration, decoloration, desalination, ion-exchange chromatography (IEC), and crystallization. Inorganic salt (Na₂SO₄) was removed from the both by desalination with 70% ethanol solution. A ninhydrin test strip was designed for the real-time detection of GABA during IEC. The recovery rate for the whole purification process was about 50%. The purified product was characterized by thin-layer chromatography and HPLC, and its purity reached 98.66 ± 2.36%.

  4. Reduced gamma-aminobutyric acid concentration is associated with physical disability in progressive multiple sclerosis

    PubMed Central

    Solanky, Bhavana S.; Muhlert, Nils; Tur, Carmen; Edden, Richard A. E.; Wheeler-Kingshott, Claudia A. M.; Miller, David H.; Thompson, Alan J.; Ciccarelli, Olga

    2015-01-01

    Neurodegeneration is thought to be the major cause of ongoing, irreversible disability in progressive stages of multiple sclerosis. Gamma-aminobutyric acid is the principle inhibitory neurotransmitter in the brain. The aims of this study were to investigate if gamma-aminobutyric acid levels (i) are abnormal in patients with secondary progressive multiple sclerosis compared with healthy controls; and (ii) correlate with physical and cognitive performance in this patient population. Thirty patients with secondary progressive multiple sclerosis and 17 healthy control subjects underwent single-voxel MEGA-PRESS (MEscher-GArwood Point RESolved Spectroscopy) magnetic resonance spectroscopy at 3 T, to quantify gamma-aminobutyric acid levels in the prefrontal cortex, right hippocampus and left sensorimotor cortex. All subjects were assessed clinically and underwent a cognitive assessment. Multiple linear regression models were used to compare differences in gamma-aminobutyric acid concentrations between patients and controls adjusting for age, gender and tissue fractions within each spectroscopic voxel. Regression was used to examine the relationships between the cognitive function and physical disability scores specific for these regions with gamma-aminobuytric acid levels, adjusting for age, gender, and total N-acetyl-aspartate and glutamine-glutamate complex levels. When compared with controls, patients performed significantly worse on all motor and sensory tests, and were cognitively impaired in processing speed and verbal memory. Patients had significantly lower gamma-aminobutyric acid levels in the hippocampus (adjusted difference = −0.403 mM, 95% confidence intervals −0.792, −0.014, P = 0.043) and sensorimotor cortex (adjusted difference = −0.385 mM, 95% confidence intervals −0.667, −0.104, P = 0.009) compared with controls. In patients, reduced motor function in the right upper and lower limb was associated with lower gamma-aminobutyric acid

  5. Reduced gamma-aminobutyric acid concentration is associated with physical disability in progressive multiple sclerosis.

    PubMed

    Cawley, Niamh; Solanky, Bhavana S; Muhlert, Nils; Tur, Carmen; Edden, Richard A E; Wheeler-Kingshott, Claudia A M; Miller, David H; Thompson, Alan J; Ciccarelli, Olga

    2015-09-01

    Neurodegeneration is thought to be the major cause of ongoing, irreversible disability in progressive stages of multiple sclerosis. Gamma-aminobutyric acid is the principle inhibitory neurotransmitter in the brain. The aims of this study were to investigate if gamma-aminobutyric acid levels (i) are abnormal in patients with secondary progressive multiple sclerosis compared with healthy controls; and (ii) correlate with physical and cognitive performance in this patient population. Thirty patients with secondary progressive multiple sclerosis and 17 healthy control subjects underwent single-voxel MEGA-PRESS (MEscher-GArwood Point RESolved Spectroscopy) magnetic resonance spectroscopy at 3 T, to quantify gamma-aminobutyric acid levels in the prefrontal cortex, right hippocampus and left sensorimotor cortex. All subjects were assessed clinically and underwent a cognitive assessment. Multiple linear regression models were used to compare differences in gamma-aminobutyric acid concentrations between patients and controls adjusting for age, gender and tissue fractions within each spectroscopic voxel. Regression was used to examine the relationships between the cognitive function and physical disability scores specific for these regions with gamma-aminobuytric acid levels, adjusting for age, gender, and total N-acetyl-aspartate and glutamine-glutamate complex levels. When compared with controls, patients performed significantly worse on all motor and sensory tests, and were cognitively impaired in processing speed and verbal memory. Patients had significantly lower gamma-aminobutyric acid levels in the hippocampus (adjusted difference = -0.403 mM, 95% confidence intervals -0.792, -0.014, P = 0.043) and sensorimotor cortex (adjusted difference = -0.385 mM, 95% confidence intervals -0.667, -0.104, P = 0.009) compared with controls. In patients, reduced motor function in the right upper and lower limb was associated with lower gamma-aminobutyric acid concentration in the

  6. Reduced gamma-aminobutyric acid concentration is associated with physical disability in progressive multiple sclerosis.

    PubMed

    Cawley, Niamh; Solanky, Bhavana S; Muhlert, Nils; Tur, Carmen; Edden, Richard A E; Wheeler-Kingshott, Claudia A M; Miller, David H; Thompson, Alan J; Ciccarelli, Olga

    2015-09-01

    Neurodegeneration is thought to be the major cause of ongoing, irreversible disability in progressive stages of multiple sclerosis. Gamma-aminobutyric acid is the principle inhibitory neurotransmitter in the brain. The aims of this study were to investigate if gamma-aminobutyric acid levels (i) are abnormal in patients with secondary progressive multiple sclerosis compared with healthy controls; and (ii) correlate with physical and cognitive performance in this patient population. Thirty patients with secondary progressive multiple sclerosis and 17 healthy control subjects underwent single-voxel MEGA-PRESS (MEscher-GArwood Point RESolved Spectroscopy) magnetic resonance spectroscopy at 3 T, to quantify gamma-aminobutyric acid levels in the prefrontal cortex, right hippocampus and left sensorimotor cortex. All subjects were assessed clinically and underwent a cognitive assessment. Multiple linear regression models were used to compare differences in gamma-aminobutyric acid concentrations between patients and controls adjusting for age, gender and tissue fractions within each spectroscopic voxel. Regression was used to examine the relationships between the cognitive function and physical disability scores specific for these regions with gamma-aminobuytric acid levels, adjusting for age, gender, and total N-acetyl-aspartate and glutamine-glutamate complex levels. When compared with controls, patients performed significantly worse on all motor and sensory tests, and were cognitively impaired in processing speed and verbal memory. Patients had significantly lower gamma-aminobutyric acid levels in the hippocampus (adjusted difference = -0.403 mM, 95% confidence intervals -0.792, -0.014, P = 0.043) and sensorimotor cortex (adjusted difference = -0.385 mM, 95% confidence intervals -0.667, -0.104, P = 0.009) compared with controls. In patients, reduced motor function in the right upper and lower limb was associated with lower gamma-aminobutyric acid concentration in the

  7. The gamma-aminobutyric acid shunt contributes to closing the tricarboxylic acid cycle in Synechocystis sp PCC 6803

    SciTech Connect

    Xiong, W; Brune, D; Vermaas, WFJ

    2014-07-16

    A traditional 2-oxoglutarate dehydrogenase complex is missing in the cyanobacterial tricarboxylic acid cycle. To determine pathways that convert 2-oxoglutarate into succinate in the cyanobacterium Synechocystis sp. PCC 6803, a series of mutant strains, Delta sll1981, Delta slr0370, Delta slr1022 and combinations thereof, deficient in 2-oxoglutarate decarboxylase (Sll1981), succinate semialdehyde dehydrogenase (Slr0370), and/or in gamma-aminobutyrate metabolism (Slr1022) were constructed. Like in Pseudomonas aeruginosa, N-acetylornithine aminotransferase, encoded by slr1022, was shown to also function as gamma-aminobutyrate aminotransferase, catalysing gamma-aminobutyrate conversion to succinic semialdehyde. As succinic semialdehyde dehydrogenase converts succinic semialdehyde to succinate, an intact gamma-aminobutyrate shunt is present in Synechocystis. The Delta sll1981 strain, lacking 2-oxoglutarate decarboxylase, exhibited a succinate level that was 60% of that in wild type. However, the succinate level in the Delta slr1022 and Delta slr0370 strains and the Delta sll1981/Delta slr1022 and Delta sll1981/Delta slr0370 double mutants was reduced to 20-40% of that in wild type, suggesting that the gamma-aminobutyrate shunt has a larger impact on metabolite flux to succinate than the pathway via 2-oxoglutarate decarboxylase. C-13-stable isotope analysis indicated that the gamma-aminobutyrate shunt catalysed conversion of glutamate to succinate. Independent of the 2-oxoglutarate decarboxylase bypass, the gamma-aminobutyrate shunt is a major contributor to flux from 2-oxoglutarate and glutamate to succinate in Synechocystis sp. PCC 6803.

  8. A validated method for gas chromatographic analysis of gamma-aminobutyric acid in tall fescue herbage

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Gamma-Aminobutyric acid (GABA) is an inhibitory neurotransmitter in animals that is also found in plants and has been associated with plant responses to stress. A simple and relatively rapid method of GABA separation and quantification was developed from a commercially available kit for serum amino...

  9. Effects of beer and hop on ionotropic gamma-aminobutyric acid receptors.

    PubMed

    Aoshima, Hitoshi; Takeda, Katsuichi; Okita, Yoichi; Hossain, Sheikh Julfikar; Koda, Hirofumi; Kiso, Yoshinobu

    2006-04-01

    Beer induced the response of the ionotropic gamma-aminobutyric acid receptors (GABA(A) receptors) expressed in Xenopus oocytes, indicating the presence of gamma-aminobutyric acid (GABA)-like activity. Furthermore, the pentane extract of the beer, hop (Humulus lupulus L.) oil, and myrcenol potentiated the GABA(A) receptor response elicited by GABA. The GABA(A) receptor responses were also potentiated by the addition of aliphatic esters, most of which are reported to be present in beer flavor. Aliphatic esters showed the tendency to decrease in the potentiation of the GABA(A) receptor response with an increase in their carbon chain length. When myrcenol was injected to mice prior to intraperitoneal administration of pentobarbital, the pentobarbital-induced sleeping time of mice increased additionally. Therefore, the beer contained not only GABA-like activity but also the modulator(s) of the GABA(A) receptor response.

  10. Synthesis and proton NMR spectroscopy of intra-vesicular gamma-aminobutyric acid (GABA).

    PubMed

    Wang, Luke Y-J; Tong, Rong; Kohane, Daniel S

    2013-01-01

    We report the synthesis of vesicles containing gamma-aminobutyric acid (GABA), and their proton nuclear magnetic resonance ((1)H NMR) spectra. These vesicles were constructed to more closely mimic the intracellular environment wherein GABA exists. For this study, these GABA-containing vesicles were examined under (1)H NMR as a potential platform for future studies on the differences between aqueous phantoms, ex vivo brain extracts, and in vivo magnetic resonance spectroscopy results. We found that intra-vesicular GABA faithfully yielded the chemical shifts and J-coupling constants of free aqueous GABA, alongside the chemical shift signals of the vesicle wall.

  11. New insights into {gamma}-aminobutyric acid catabolism: Evidence for {gamma}-hydroxybutyric acid and polyhydroxybutyrate synthesis in Saccharomyces cerevisiae.

    PubMed

    Bach, Benoît; Meudec, Emmanuelle; Lepoutre, Jean-Paul; Rossignol, Tristan; Blondin, Bruno; Dequin, Sylvie; Camarasa, Carole

    2009-07-01

    The gamma-aminobutyrate (GABA) shunt, an alternative route for the conversion of alpha-ketoglutarate to succinate, involves the glutamate decarboxylase Gad1p, the GABA transaminase Uga1p and the succinate semialdehyde dehydrogenase Uga2p. This pathway has been extensively described in plants and animals, but its function in yeast remains unclear. We show that the flux through Gad1p is insignificant during fermentation in rich sugar-containing medium, excluding a role for this pathway in redox homeostasis under anaerobic conditions or sugar stress. However, we found that up to 4 g of exogenous GABA/liter was efficiently consumed by yeast. We studied the fate of this consumed GABA. Most was converted into succinate, with a reaction yield of 0.7 mol/mol. We also showed that a large proportion of GABA was stored within cells, indicating a possible role for this molecule in stress tolerance mechanisms or nitrogen storage. Furthermore, based on enzymatic and metabolic evidence, we identified an alternative route for GABA catabolism, involving the reduction of succinate-semialdehyde into gamma-hydroxybutyric acid and the polymerization of gamma-hydroxybutyric acid to form poly-(3-hydroxybutyric acid-co-4-hydroxybutyric acid). This study provides the first demonstration of a native route for the formation of this polymer in yeast. Our findings shed new light on the GABA pathway and open up new opportunities for industrial applications.

  12. Pre-staining paper chromatography method for quantification of gamma-aminobutyric acid.

    PubMed

    Li, Haixing; Qiu, Ting; Cao, Yusheng; Yang, Jiyan; Huang, Zhibing

    2009-06-19

    The routine method of paper chromatography includes five steps: spotting, separating, drying, spraying/immersing and color development. In this paper, a pre-staining paper chromatography which only consisted of spotting, separating and color development was developed for quantitative analysis of gamma-aminobutyric acid. Compared to the routine paper chromatography, the improved method is clean, rapid, inexpensive and reproducible. The effects of ninhydrin concentration, color temperature, color time and Cu(2+) concentration on the color yield in the ninhydrin reaction were optimized. And then the pre-staining paper chromatography coupled with vis spectrophotometry was applied to gamma-aminobutyric acid quantification. The results indicated that the limit of detection was 0.05 mg mL(-1) and the linear range was from 0.5 to 20.0 mg mL(-1). Furthermore, an excellent correlation coefficient was observed with an R(2)=0.998. The method is accurate (RSD<2.64%), and has good recoveries (102.7-103.9%). The validation of the modified technique was verified by a HPLC method.

  13. Isolation and characterization of a gamma-aminobutyric acid producing strain Lactobacillus buchneri WPZ001 that could efficiently utilize xylose and corncob hydrolysate.

    PubMed

    Zhao, Anqi; Hu, Xiaoqing; Pan, Lu; Wang, Xiaoyuan

    2015-04-01

    Lactobacillus buchneri strain WPZ001 that could efficiently produce gamma-aminobutyric acid was isolated from Chinese fermented sausages. Optimal cultivation conditions for gamma-aminobutyric acid production in L. buchneri WPZ001 were determined, and xylose was found to be the best carbon source. Using xylose as the sole carbon source, 70 g/L gamma-aminobutyric acid was produced by flask fermentation of L. buchneri WPZ001 for 48 h, and the harvested cells could continue to convert monosodium glutamate to gamma-aminobutyric acid in buffer and produce 59 g gamma-aminobutyric acid after eight runs of biotransformation; the total yield of gamma-aminobutyric acid reached 129 g/L. This combination strategy also worked well when the low-cost corncob hydrolysate was used as the sole carbon source, and the yield of gamma-aminobutyric acid reached 117 g/L. The results indicate that L. buchneri WPZ001 has great potential for industrial production of gamma-aminobutyric acid. PMID:25524701

  14. Succinic semialdehyde as a substrate for the formation of gamma-aminobutyric acid.

    PubMed

    van Bemmelen, F J; Schouten, M J; Fekkes, D; Bruinvels, J

    1985-11-01

    The conversion of succinic semialdehyde into gamma-aminobutyric acid (GABA) by GABA-transaminase was measured in rat brain homogenate in the presence of different concentrations of the cosubstrate glutamate. The calculated kinetic parameters of succinic semialdehyde for GABA-transaminase were a limiting Km value of 168 microM and a limiting Vmax value of 38 mumol g-1 h-1. Combination with previously obtained data for the conversion of GABA into succinic semialdehyde revealed a kEq value of 0.04, indicating that equilibrium of GABA-transaminase is biased toward the formation of GABA. The increased formation of GABA in the presence of succinic semialdehyde was not due to an increased conversion of glutamate into GABA by glutamic acid decarboxylase. Therefore these results indicate that succinic semialdehyde can act as a precursor for GABA synthesis.

  15. Benzodiazepine receptor agonists affect both binding and gating of recombinant alpha1beta2gamma2 gamma-aminobutyric acid-A receptors.

    PubMed

    Mercik, Katarzyna; Piast, Michał; Mozrzymas, Jerzy W

    2007-05-28

    Benzodiazepines are known to act by enhancing the effect of gamma-aminobutyric acid-A receptor agonists. Positive modulation by benzodiazepines is typically ascribed to upregulation of agonist binding affinity but their effect on gamma-aminobutyric acid-A receptor gating remain unclear. In this work, we have used the ultrafast application system to examine the impact of flurazepam and zolpidem on recombinant alpha1beta2gamma2 gamma-aminobutyric acid-A receptors. As expected, both drugs strongly enhanced currents evoked by low [gamma-aminobutyric acid]. These compounds, however, also affected currents elicited by saturating agonist concentration. In particular, flurazepam and zolpidem reduced current amplitudes and slowed down the recovery process in paired-pulse experiments. Moreover, flurazepam accelerated the current rise time and zolpidem enhanced the rate and extent of desensitization. We propose that flurazepam and zolpidem modulate gamma-aminobutyric acid-A receptors by strong enhancement of agonist binding with a superimposed limited effect on the receptor gating.

  16. Genome Sequence of Lactobacillus curieae CCTCC M 2011381T, a Novel Producer of Gamma-aminobutyric Acid

    PubMed Central

    Wang, Ying; Wang, Yu; Lang, Chong; Wei, Dongzhi; Xu, Ping

    2015-01-01

    Lactobacillus curieae CCTCC M 2011381T is a novel species of the genus Lactobacillus and a gamma-aminobutyric acid producer that was isolated from stinky tofu brine. Here, we present a 2.19-Mb assembly of its genome, which may provide further insights into the molecular mechanisms underlying its beneficial properties. PMID:26021929

  17. Effect of diphenylhydantoin on gamma aminobutyric acid (GABA) and succinate activity in rat Purkinje cells.

    PubMed Central

    Hitchcock, E; Gabra-Sanders, T

    1977-01-01

    A study has been made of the effect of diphenylhydantoin (DPH) upon the levels of gamma aminobutyric acid (GABA) and succinic dehydrogenase in rat Purkinje cells. DPH was administered over 26 days in chronic experiments using controls receiving the same injection vehicle without DPH. Animals in this group received daily 1.25 mg/kg body weight, 12.5 mg/kg body weight, and 50 mg/kg body weight DPH. Acute experiments were carried out over the course of not more than four days, three groups of animals receiving 75 mg/kg body weight, 87.5 mg/kg body weight, and 100 mg/kg body weight DPH. No effect upon succinic dehydrogenase could be demonstrated at any dose level. There was a significant progressive loss of GABA with increasing dosage of DPH. Images PMID:903771

  18. Neurons and glia in cat superior colliculus accumulate [3H]gamma-aminobutyric acid (GABA).

    PubMed

    Mize, R R; Spencer, R F; Sterling, P

    1981-11-01

    We have examined by autoradiography the labeling pattern in the cat superior colliculus following injection of tritiated gamma-aminobutyric acid (GABA). Silver grains were heavily distributed within the zonal layer and the upper 200 micrometer of the superficial gray. Fewer grains were observed deeper within the superficial gray, and still fewer were found within the optic and intermediate gray layers. The accumulation of label was restricted to certain classes of neuron and glia. Densely labeled neurons were small (8-12 micrometer in diameter) and located primarily within the upper 200 micrometer. Dark oligodendrocytes and astrocytes showed a moderate accumulation of label while pale oligodendrocytes and microglia were unlabeled. Label was also selectively accumulated over several other types of profile within the neuropil, including presynaptic dendrites, axons, and axon terminals.

  19. Serum hormonal concentrations following transcendental meditation--potential role of gamma aminobutyric acid.

    PubMed

    Elias, A N; Wilson, A F

    1995-04-01

    Transcendental mediation (TM) is a stylized form of physical and mental relaxation which is associated with changes in the secretion and release of several pituitary hormones. The hormonal changes induced by TM mimic the effects of the inhibitory neurotransmitter gamma aminobutyric acid (GABA). It is hypothesized that TM produces changes in pituitary hormone secretion by enhancing hypothalamic GABAergic tone, and its anxiolytic effects by promoting GABAergic tone in specific areas of the brain. This mechanism is similar to the effects of synthetic anxiolytic and tranquilizing agents such as benzodiazepines that bind to components of the GABA-A (GABAA) receptor. TM, therefore, may produce relaxation by enhancing the effects of an endogenous neurotransmitter analogous to the effects of endorphins in runners who reportedly experience a 'runner's high'.

  20. Prolonged treatment with gamma-aminobutyric acid (GABA)-mimetic substances in prepubertal male rats.

    PubMed

    Debeljuk, L; Díaz, M D; Maines, V M; Seilicovich, A

    1983-06-01

    The effect of chronic treatment with a gamma-aminobutyric acid (GABA)-mimetic compound, progabide, and an inhibitor of GABA-transaminase, gamma-acetylenic GABA (GAG), was tested in prepubertal male rats. The effect of gamma-butyrolactone (GBL), given orally, was also tested. The rats treated with progabide did not show any difference in body, testicular, or seminal vesicle weights or serum prolactin levels, as compared with control rats. Treatment with GAG, at both dose levels used, did not significantly affect body weight. Testicular weight was significantly lower in the group of rats treated with the low dosage of GAG (5 mg/kg), and serum prolactin was significantly lower in the rats treated with the high dosage of GAG (20 mg/kg) as compared with control rats. In the first experiment performed with GBL, the rats given this compound had significantly lower body and testicular weights as compared with control rats. In the second experiment, GBL-treated rats had body weights similar to those of control rats, but testicular weights were significantly decreased. Prolonged treatment with GABA mimetics may affect the hypothalamic-pituitary-testicular axis.

  1. Amiloride inhibition of gamma-aminobutyric acid(A) receptors depends upon the alpha subunit subtype.

    PubMed

    Fisher, Janet L

    2002-06-01

    gamma-Aminobutyric acid(A) (GABA(A)) receptors (GABARs) are responsible for most fast inhibitory neurotransmission in the mammalian brain. The GABARs contain several allosteric modulatory sites, many of which are useful clinically. The activity of most of these modulators depends upon the subunit composition of the receptor. The diuretic amiloride was previously reported to inhibit GABARs in frog sensory neurons. We measured its effects on recombinant GABARs to determine its mechanism of action at mammalian receptors and to examine the effect of subunit composition. Amiloride acted primarily as a competitive antagonist, reducing the sensitivity of the receptor to GABA without affecting the maximal current amplitude. Receptors containing an alpha6 subunit were about 10-fold more sensitive to amiloride than those containing other alpha subunits. In contrast, the identity of the beta or gamma subtype had little effect on amiloride sensitivity. Although several other modulators have specific effects at alpha6-containing receptors, amiloride is the first inhibitor to be reported with no additional dependence on the identity of the beta or gamma subunit. Therefore, it probably represents a unique modulatory site on the GABAR, which could be useful for developing drugs targeting these receptors. The selective activity of amiloride could also be helpful for isolating the contribution of receptors composed of alpha6 subtypes in heterogeneous native GABAR populations.

  2. Active transport of. gamma. -aminobutyric acid and glycine into synaptic vesicles

    SciTech Connect

    Kish, P.E.; Fischer-Bovenkerk, C.; Ueda, T. )

    1989-05-01

    Although {gamma}-aminobutyric acid (GABA) and glycine are recognized as major amino acid inhibitory neurotransmitters in the central nervous system, their storage is poorly understood. In this study the authors have characterized vesicular GABA and glycine uptakes in the cerebrum and spinal cord, respectively. They present evidence that GABA and glycine are each taken up into isolated synaptic vesicles in an ATP-dependent manner and that the uptake is driven by an electrochemical proton gradient. Uptake for both amino acids exhibited kinetics with low affinity similar to a vesicular glutamate uptake. The ATP-dependent GABA uptake was not inhibited by the putative amino acid neurotransmitters glycine, taurine, glutamate, or aspartate or by GABA analogs, agonists, and antagonists. Similarly, ATP-dependent glycine uptake was hardly affected by GABA, taurine, glutamate, or aspartate or by glycine analogs or antagonists. The GABA uptake was not affected by chloride, which is in contrast to the uptake of the excitatory neurotransmitter glutamate, whereas the glycine uptake was slightly stimulated by low concentrations of chloride. Tissue distribution studies indicate that the vesicular uptake systems for GABA, glycine, and glutamate are distributed in different proportions in the cerebrum and spinal cord. These results suggest that the vesicular uptake systems for GABA, glycine, and glutamate are distinct from each other.

  3. Effects of traditionally used anxiolytic botanicals on enzymes of the gamma-aminobutyric acid (GABA) system.

    PubMed

    Awad, R; Levac, D; Cybulska, P; Merali, Z; Trudeau, V L; Arnason, J T

    2007-09-01

    In Canada, the use of botanical natural health products (NHPs) for anxiety disorders is on the rise, and a critical evaluation of their safety and efficacy is required. The purpose of this study was to determine whether commercially available botanicals directly affect the primary brain enzymes responsible for gamma-aminobutyric acid (GABA) metabolism. Anxiolytic plants may interact with either glutamic acid decarboxylase (GAD) or GABA transaminase (GABA-T) and ultimately influence brain GABA levels and neurotransmission. Two in vitro rat brain homogenate assays were developed to determine the inhibitory concentrations (IC50) of aqueous and ethanolic plant extracts. Approximately 70% of all extracts that were tested showed little or no inhibitory effect (IC50 values greater than 1 mg/mL) and are therefore unlikely to affect GABA metabolism as tested. The aqueous extract of Melissa officinalis (lemon balm) exhibited the greatest inhibition of GABA-T activity (IC50 = 0.35 mg/mL). Extracts from Centella asiatica (gotu kola) and Valeriana officinalis (valerian) stimulated GAD activity by over 40% at a dose of 1 mg/mL. On the other hand, both Matricaria recutita (German chamomile) and Humulus lupulus (hops) showed significant inhibition of GAD activity (0.11-0.65 mg/mL). Several of these species may therefore warrant further pharmacological investigation. The relation between enzyme activity and possible in vivo mode of action is discussed. PMID:18066140

  4. [Neuropharmacological regulation of pituitary hormones. Role of acetylcholine, gamma-aminobutyric acid, histamine and endorphins (author's transl)].

    PubMed

    Martin-du Pan, R; Gomez, F

    1981-01-01

    The secretion of pituitary hormones is controlled by hypothalamic hormones which are synthetized by neurosecreting cells whose activity is modulated by different neurotransmitters as dopamine and serotonin. Centrally acting drugs interfere with the activity of these neurotransmitters. Thus they may influence the secretion of pituitary hormones. Acetylcholine, gamma-aminobutyric acid, histamine and endorphins seem also to influence the pituitary secretion. The endocrine effects of drugs (opiates, antiparkinson, antiepileptic, and antihistaminic agents) acting on these neurotransmitters is reviewed.

  5. Effects of gamma-aminobutyric acid agonist and antagonist drugs on local cerebral glucose utilization

    SciTech Connect

    Palacios, J.M.; Kuhar, M.J.; Rapoport, S.I.; London, E.D.

    1982-07-01

    The (/sup 14/C)2-deoxy-D-glucose method of Sokoloff et al. was used to study local cerebral glucose utilization (LCGU) in rats treated with gamma-aminobutyric acid (GABA) agonist (muscimol and 4,5,6,7-tetrahydroisoxazolo(5,4-C)pyridin-3-ol, THIP) and antagonist (bicuculline) drugs. It was of interest to determine if the pattern of LCGU responses to GABA agonists and antagonists administered systemically in vivo would reflect the known distributions of markers for central GABAergic synapses. The patterns of LCGU responses to muscimol and THIP generally were similar. Most brain regions showed dose-dependent decreases in LCGU; others showed no effects; but the red nucleus showed an increase. The GABA antagonist bicuculline produced convulsions and variable LCGU responses, depending on the time of administration. Bicuculline also partially antagonized the depressant effects of muscimol of LCGU. The magnitudes and distribution of in vivo cerebral metabolic responses to specific GABA agonists were not correlated simply with markers for GABAergic synapses. This lack of correlation indicates that additional factors, such as neural circuitry, regulate the LCGU responses to GABAergic drugs.

  6. Identification of gamma-aminobutyric acid and its binding sites in Caenorhabditis elegans

    SciTech Connect

    Schaeffer, J.M.; Bergstrom, A.R.

    1988-01-01

    Gamma-aminobutyric acid (GABA), glutamate decarboxylase and GABA-transaminase were identified in the nematode Caenorhabditis elegans. The concentration of GABA in C. elegans is approximately 10-fold lower than the concentration of GABA in rat brain. Glutamate decarboxylase and GABA-transaminase, the GABA anabolic and catabolic enzymes, are also present in C. elegans. Crude membrane fractions were prepared from C. elegans and used to study specific (/sup 3/H) GABA binding sites. GABA binds to C. elegans membranes with high affinity and low capacity. Muscimol is a competitive inhibitor of specific GABA binding with a K/sub I/ value of 120 nM. None of the other GABA agonists or antagonists inhibited greater than 40% of the specific GABA binding at concentrations up to 10/sup -4/M. Thirteen spider venoms were examined as possible GABA agonists or antagonists, the venom from Calilena agelenidae inhibits specific GABA binding with a K/sub I/ value of 6 nl/ml. These results suggest that GABA has a physiological role as a neurotransmitter in C. elegans.

  7. Gamma-aminobutyric acid circuits shape response properties of auditory cortex neurons.

    PubMed

    Wang, Jian; McFadden, Sandra L; Caspary, Donald; Salvi, Richard

    2002-07-19

    Neurons containing gamma aminobutyric acid (GABA) are widely distributed throughout the primary auditory cortex (AI). We investigated the effects of endogenous GABA by comparing response properties of 110 neurons in chinchilla AI before and after iontophoresis of bicuculline, a GABA(A) receptor antagonist, and/or CGP35348, a GABA(B) receptor antagonist. GABA(A) receptor blockade significantly increased spontaneous and driven discharge rates, dramatically decreased the thresholds of many neurons, and constricted the range of thresholds across the neural population. Some neurons with 'non-onset' temporal discharge patterns developed an onset pattern that was followed by a long pause. Interestingly, the excitatory response area typically expanded on both sides of the characteristic frequency; this expansion exceeded one octave in a third of the sample. Although GABA(B) receptor blockade had little effect alone, the combination of CGP35348 and bicuculline produced greater increases in driven rate and expansion of the frequency response area than GABA(A) receptor blockade alone, suggesting a modulatory role of local GABA(B) receptors. The results suggest that local GABA inhibition contributes significantly to intensity and frequency coding by controlling the range of intensities over which cortical neurons operate and the range of frequencies to which they respond. The inhibitory circuits that generate nonmonotonic rate-level functions are separate from those that influence other response properties of AI neurons. PMID:12106684

  8. Production of gamma-aminobutyric acid by Streptococcus salivarius subsp. thermophilus Y2 under submerged fermentation.

    PubMed

    Yang, S-Y; Lü, F-X; Lu, Z-X; Bie, X-M; Jiao, Y; Sun, L-J; Yu, B

    2008-04-01

    Gamma-aminobutyric acid (GABA), a major inhibitory neurotransmitter in the central nervous system, has several well-known physiological functions and has been applied to the production of many drugs and functional foods. The technology of GABA production via submerged fermentation by Streptococcus salivarius subsp. thermophilus Y2 was investigated in this paper. It indicated that the GABA production was related to the biochemical characteristics of glutamate decarboxylase (GAD) of S. salivarius subsp. thermophilus Y2. After 24 h of fermentation at 37 degrees C, which is the suitable culture conditions for GAD-production, then the culture condition were adjusted to the optimal temperature (40 degrees C) and pH (4.5) for the GAD reaction activity in biotransformation of cells and pyridoxal 5'-phosphate (0.02 mmol/l) were added to the broth at the 48 h, the GABA production was increased up to 1.76-fold, reaching 7984.75 +/- 293.33 mg/l. The strain shows great potential use as a starter for GABA-containing yoghurt, cheese and other functional fermented food productions. PMID:17514494

  9. Effects of white rice containing enriched gamma-aminobutyric acid on blood pressure

    PubMed Central

    Nishimura, Mie; Yoshida, Shin-ichi; Haramoto, Masafumi; Mizuno, Hidenori; Fukuda, Tomohiko; Kagami-Katsuyama, Hiroyo; Tanaka, Aiko; Ohkawara, Tatsuya; Sato, Yuji; Nishihira, Jun

    2015-01-01

    Gamma-aminobutyric acid (GABA) is an inhibitory neurotransmitter with beneficial effects including antihypertension and antistress properties. In this study, we examined the effects of GABA-enriched white rice (GABA rice) on blood pressure (BP) in 39 mildly hypertensive adults in a randomized, double-blind, placebo-controlled study. The participants were divided into a test group (n = 22) who consumed rice with 11.2 mg GABA/100 g of rice and a placebo group (n = 17) who consumed rice with 2.7 mg GABA/100 g of rice. For 8 weeks, the participants took 150 g of either the GABA rice or the placebo rice. Hematological examinations were performed on both groups at 0, 4, and 8 weeks after the start of rice consumption. Home BP was self-measured two times daily, morning and evening, from 1 weeks before to 2 weeks after the intervention. Although the hospital BP and evening BP measurements of the participants showed no significant change, consumption of the GABA rice improved the morning BP compared with the placebo rice after the 1st week and during the 6th and 8th weeks. These results showed the possibility that the GABA rice improves morning hypertension. PMID:26870683

  10. Gamma-aminobutyric acid (GABA) and neuropeptides in neural areas mediating motion-induced emesis

    NASA Technical Reports Server (NTRS)

    Damelio, F.; Daunton, Nancy G.; Fox, Robert A.

    1991-01-01

    Immunocytochemical methods were employed to localize the neurotransmitter amino acid gamma-aminobutyric acid and the neuropeptides substance P and Met-enkephalin in the area postrema (AP), area subpostrema (ASP), nucleus of the tractus solitarius (NTS), dorsal motor nucleus of the vagus nerve (DMNV), and lateral vestibular nucleus (LVN). Glutamic acid decarboxylase immunoreactive (GAD-IR) terminals and fibers were observed in the AP and particularly in the ASP. A gradual decrease in the density of terminals was seen towards the solitary complex. The DMNV revealed irregularly scattered GAD-IR terminals within the neuropil or closely surrounding neuronal cell bodies. The LVN, particularly the dorsal division, showed numerous axon terminals which were mostly localize around large neurons and their proximal dendrites. Substance P immunoreactive (SP-IR) terminals and fibers showed high density in the solitary complex, in particular within the lateral division. The ASP showed medium to low density of SP-IR fibers and terminals. The AP exhibited a small number of fibers and terminals irregularly distributed. The DMNV revealed a high density of SP-IR terminals and fibers that were mainly concentrated in the periphery. Very few terminals were detected in the LVN. Met-enkephalin immunoreactive (Met-Enk-IR) fibers and terminals showed high density and uniform distribution in the DMNV. Scattered terminals and fibers were observed in the AP, ASP, and NTS (particularly the lateral division). The very few fibers were observed in the LVN surrounded the neuronal cell bodies. The present report is part of a study designed to investigate the interaction between neuropeptides and conventional neurotransmitters under conditions producing motion sickness and in the process of sensory-motor adaptation.

  11. Up-regulation of gamma-aminobutyric acid transporter I mediates ethanol sensitivity in mice.

    PubMed

    Hu, J-H; Ma, Y-H; Yang, N; Mei, Z-T; Zhang, M-H; Fei, J; Guo, L-H

    2004-01-01

    Ethanol is among the most widely abused drugs in the world. Chronic ethanol consumption leads to ethanol tolerance and addiction, and impairs learning and memory. Na+/Cl- dependent GABA transporters play an important role in controlling the concentration of GABA in the synaptic cleft, and thus they control the intensity and duration of synaptic transmission of GABA. It has been suggested that GABAergic system is involved in ethanol consumption, tolerance and addiction, because chronic ethanol consumption alters the expression of GABAA receptors and drugs on GABA receptors affect ethanol actions. The results of the present study reveal that that activity of GABA transporters in mouse brain after 15-min acute ethanol injection or after chronic ethanol consumption is increased. Moreover, mice pre-injected with a competitive or a noncompetitive antagonist of gamma-aminobutyric acid transporter subtype 1 (GAT1) showed high sensitivity to the sedative/hypnotic effects of ethanol. In contrast, transgenic mice overexpressing GAT1 displayed low sensitivity to ethanol, as shown by the righting reflex test. Mice overexpressing GAT1 survived a lethal dose of ethanol (9 g/kg, i.p.) longer, maintained locomotor activity longer after a sub-lethal dose (1.75 g/kg, i.p.) and exhibited a higher median lethal dose than wild-type littermates. These results suggest that GAT1 plays an important role in sensitivity to ethanol, and might be a therapeutic target for alcoholism prevention and treatment. Acute and chronic ethanol administration resulted in the increase of GABA transporter function. Use of GAT1 selective inhibitors and GAT1 overexpressing mice thus demonstrate that GAT1 should be an important protein mediating sensitivity to ethanol in mice.

  12. Functional reconstitution of the. gamma. -aminobutyric acid transporter from synaptic vesicles using artificial ion gradients

    SciTech Connect

    Hell, J.W.; Edelmann, L.; Hartinger, J.; Jahn, R. )

    1991-12-24

    The {gamma}-aminobutyric acid transporter of rat brain synaptic vesicles was reconstituted in proteoliposomes, and its activity was studied in response to artificially created membrane potentials or proton gradients. Changes of the membrane potential were monitored using the dyes oxonol VI and 3,3{prime}-diisopropylthiodicarbocyanine iodide, and changes of the H{sup +} gradient were followed using acridine orange. An inside positive membrane potential was generated by the creation of an inwardly directed K{sup +} gradient and the subsequent addition of valinomycin. Under these conditions, valinomycin evoked uptake of ({sup 3}H)GABA which was saturable. Similarly, ({sup 3}H)glutamate uptake was stimulated by valinomycin, indicating that both transporters can be driven by the membrane potential. Proton gradients were generated by the incubation of K{sup +}-loaded proteoliposomes in a buffer free of K{sup +} or Na{sup +} ions and the subsequent addition of nigericin. Proton gradients were also generated via the endogenous H{sup +} ATPase by incubation of K{sup +}-loaded proteoliposomes in equimolar K{sup +} buffer in the presence of valinomycin. These proton gradients evoked nonspecific, nonsaturable uptake of GABA and {beta}-alanine but not of glycine in proteoliposomes as well as protein-free liposomes. Therefore, transporter activity was monitored using glycine as an alternative substrate. Proton gradients generated by both methods elicited saturable glycine uptake in proteoliposomes. Together, these data confirm that the vesicular GABA transporter can be energized by both the membrane potential and the pH gradient and show that transport can be achieved by artificial gradients independently of the endogenous proton ATPase.

  13. Identification of a lithium interaction site in the gamma-aminobutyric acid (GABA) transporter GAT-1.

    PubMed

    Zhou, Yonggang; Zomot, Elia; Kanner, Baruch I

    2006-08-01

    The sodium- and chloride-dependent electrogenic gamma-aminobutyric acid (GABA) transporter GAT-1, which transports two sodium ions together with GABA, is essential for synaptic transmission by this neurotransmitter. Although lithium by itself does not support GABA transport, it has been proposed that lithium can replace sodium at one of the binding sites but not at the other. To identify putative lithium selectivity determinants, we have mutated the five GAT-1 residues corresponding to those whose side chains participate in the sodium binding sites Na1 and Na2 of the bacterial leucine-transporting homologue LeuT(Aa). In GAT-1 and in most other neurotransmitter transporter family members, four of these residues are conserved, but aspartate 395 replaces the Na2 residue threonine 354. At varying extracellular sodium, lithium stimulated sodium-dependent transport currents as well as [3H]GABA uptake in wild type GAT-1. The extent of this stimulation was dependent on the GABA concentration. In mutants in which aspartate 395 was replaced by threonine or serine, the stimulation of transport by lithium was abolished. Moreover, these mutants were unable to mediate the lithium leak currents. This phenotype was not observed in mutants at the four other positions, although their transport properties were severely impacted. Thus at saturating GABA, the site corresponding to Na2 behaves as a low affinity sodium binding site where lithium can replace sodium. We propose that GABA participates in the other sodium binding site, just like leucine does in the Na1 site, and that at limiting GABA, this site determines the apparent sodium affinity of GABA transport.

  14. Gamma-aminobutyric acid aggravates nephrotoxicity induced by cisplatin in female rats

    PubMed Central

    Peysepar, Elham; Soltani, Nepton; Nematbakhsh, Mehdi; Eshraghi-Jazi, Fatemeh; Talebi, Ardeshir

    2016-01-01

    Introduction: Cisplatin (CP) is a major antineoplastic drug for treatment of solid tumors. CP-induced nephrotoxicity may be gender-related. This is while gamma-aminobutyric acid (GABA) is an inhibitory neurotransmitter in the central nervous system that has renoprotective impacts on acute renal injury. Objectives: This study was designed to investigate the protective role of GABA against CP-induced nephrotoxicity in male and female rats. Materials and Methods: Sixty Wistar male and female rats were used in eight experimental groups. Both genders received GABA (50 μg/kg/day; i. p.) for 14 days and CP (2.5 mg/kg/day; i. p.) was added from day 8 to the end of the study, and they were compared with the control groups. At the end of the study, all animals were sacrificed and the serum levels of blood urea nitrogen (BUN), creatinine (Cr), nitrite, malondialdehyde (MDA), and magnesium (Mg) were measured. The kidney tissue damage was also determined via staining. Results: CP significantly increased the serum levels of Cr and BUN, kidney weight, and kidney tissue damage score in both genders (P<0.05). GABA did not attenuate these markers in males; even these biomarkers were intensified in females. Serum level of Mg, and testis and uterus weights did not alter in the groups. However, the groups were significantly different in terms of nitrite and MDA levels. Conclusion: It seems that GABA did not improve nephrotoxicity induced by CP-treated rats, and it exacerbated renal damage in female rats. PMID:27689121

  15. Vertical organization of gamma-aminobutyric acid-accumulating intrinsic neuronal systems in monkey cerebral cortex

    SciTech Connect

    DeFelipe, J.; Jones, E.G.

    1985-12-01

    Light and electron microscopic methods were used to examine the neurons in the monkey cerebral cortex labeled autoradiographically following the uptake and transport of (/sup 3/H)-gamma-aminobutyric acid (GABA). Nonpyramidal cell somata in the sensory-motor areas and primary visual area (area 17) were labeled close to the injection site and at distances of 1 to 1.5 mm beyond the injection site, indicating labeling by retrograde axoplasmic transport. This labeling occurred preferentially in the vertical dimension of the cortex. Prior injections of colchicine, an inhibitor of axoplasmic transport, abolished all labeling of somata except those within the injection site. In each area, injections of superficial layers (I to III) produced labeling of clusters of cell somata in layer V, and injections of the deep layers (V and VI) produced labeling of clusters of cell somata in layers II and III. In area 17, injections of the superficial layers produced dense retrograde cell labeling in three bands: in layers IVC, VA, and VI. Vertically oriented chains of silver grains linked the injection sites with the resulting labeled cell clusters. In all areas, the labeling of cells in the horizontal dimension was insignificant. Electron microscopic examination of labeled neurons confirms that the neurons labeled at a distance from an injection site are nonpyramidal neurons, many with somata so small that they would be mistaken for neuroglial cells light microscopically. They receive few axosomatic synapses, most of which have symmetric membrane thickenings. The vertical chains of silver grains overlie neuronal processes identifiable as both dendrites and myelinated axons, but unmyelinated axons may also be included. The clusters of (/sup 3/H)GABA-labeled cells are joined to one another and to adjacent unlabeled cells by junctional complexes, including puncta adherentia and multi-lamellar cisternal complexes.

  16. Further characterization of [3H]gamma-aminobutyric acid release from isolated neuronal growth cones: role of intracellular Ca2+ stores.

    PubMed

    Lockerbie, R O; Gordon-Weeks, P R

    1986-04-01

    We have recently shown that growth cones isolated from neonatal rat forebrain possess uptake and release mechanisms for the neurotransmitter gamma-aminobutyric acid. About half of the K+-induced release of [3H]gamma-aminobutyric acid from isolated growth cones is dependent on extracellular Ca2+. The remaining component of the [3H]gamma-aminobutyric acid release is unaffected by removal of extracellular Ca2+ and is resistant to blockade by the voltage-sensitive Ca2+-channel blocker methoxyverapamil. In the present series of experiments we have used caffeine to assess the possible role of intracellular stores of Ca2+ in supporting that component of the K+-induced release of [3H]gamma-aminobutyric acid from isolated growth cones that is independent of extracellular Ca2+. We have chosen caffeine because of its well established effect of releasing Ca2+ from smooth endoplasmic reticulum in muscle. We found that caffeine can release [3H]gamma-aminobutyric acid from isolated growth cones. This effect persists in Ca2+-free medium, in the presence of methoxyverapamil and in the absence of Na+. Furthermore, isobutylmethylxanthine could not substitute for caffeine suggesting that the caffeine effect is not due to phosphodiesterase inhibition and the subsequent rise in intracellular cyclic nucleotides. A combination of the mitochondrial poisons, Antimycin A and sodium azide had no effect on the release of [3H]gamma-aminobutyric acid induced either by caffeine or by high K+. We conclude that caffeine causes the release of Ca2+ from a non-mitochondrial store within the growth cone and that this Ca2+ store supports that component of the K+-induced release of [3H]gamma-aminobutyric acid that is independent of extracellular Ca2+.

  17. Effects of free fatty acids, ethanol and development on gamma-aminobutyric acid and glutamate fluxes in rat nerve endings.

    PubMed

    Hitzemann, R; Mark, C; Panini, A

    1982-12-15

    The effects of type A (cis-unsaturated) and type B (trans-unsaturated and saturated) fatty acids, 1% and 3% ethanol (v/v), and development (7 days) on the thermodynamics of glutamate and gamma-aminobutyric acid (GABA) transport into cortical rat brain nerve endings were examined. The effects of the various manipulations, which are known to affect membrane fluidity, may be summarized. Three percent ethanol and oleic acid increased delta S degrees and delta S+ for glutamate transport and decreased delta H degrees and delta H+. Type B fatty acids had the opposite effects. In comparison to glutamate transport, GABA transport was less affected by the various manipulations and showed less specificity in terms of the fatty acid effects. Similarly, the effects of development on the thermodynamic parameters for glutamate and GABA transport were not consistent. Glutamate transport into 7-day nerve endings showed thermodynamic behavior similar to that seen when type A fatty acids were incorporated into adult nerve endings. In contrast, GABA transport into 7-day nerve endings had the character of adult nerve endings into which type B fatty acids were incorporated.

  18. Comparison of gamma-aminobutyric acid effects in different parts of the cat ileum.

    PubMed

    Pencheva, N; Itzev, D; Milanov, P

    1999-02-26

    The effects of gamma-aminobutyric acid (GABA) and those of a GABA(A) (muscimol) and a GABA(B) (baclofen) receptor agonists were determined on the spontaneous activity of longitudinally or circularly oriented preparations (segments) isolated from terminal, proximal and distal parts of the cat ileum. GABA applied at 1 microM to 2 mM caused dose-dependent biphasic changes (relaxation and contraction) in spontaneous activity of the longitudinal and circular layers in the terminal and distal parts of the cat ileum and monophasic changes (contraction) in the proximal part. The potency of GABA to elicit relaxant and/or contractile effects in different parts of the ileum showed a proximal-to-terminal increasing pattern. In the longitudinal layer of the distal and terminal ileum, muscimol (100 microM) mimicked the relaxation phase of the GABA effect, while baclofen (100 microM) simulated the contractile phase. Bicuculline, atropine and tetrodotoxin abolished GABA- and muscimol-induced relaxation and suppressed, but failed to prevent GABA- and baclofen-induced contractions. In addition, 2-hydroxysaclofen antagonized the baclofen-induced contractile effect, reduced the GABA-induced contractile phase but failed to prevent GABA- and muscimol-induced relaxation. In the circular layer of the same regions, muscimol mimicked the biphasic GABA effects, while baclofen was without effect. Bicuculline, atropine and tetrodotoxin completely prevented the GABA- and muscimol effects, while 2-hydroxysaclofen failed to antagonize them. In the longitudinal and circular layers of the proximal ileum, muscimol (100 microM) exerted a 'GABA-like' transient contractile effect, while baclofen (100 microM) did not elicit any response. Bicuculline, atropine and tetrodotoxin antagonized the GABA- and muscimol-induced contractile responses of longitudinal and circular layers, while 2-hydroxysaclofen was ineffective. The results suggested that the inhibitory and/or excitatory action of GABA on

  19. Antiepileptic potential of matrine via regulation the levels of gamma-aminobutyric acid and glutamic acid in the brain.

    PubMed

    Xiang, Jun; Jiang, Yugang

    2013-12-05

    Our present study aimed to determine the antiepileptic activity of matrine, and explore the possible molecular mechanism. To evaluate the antiepileptic activity of matrine, seizures in mice induced by PTZ and MES were established, then the pentobarbital sodium-induced anaesthetizing time and locomotor activity tests in mice were also carried out. For the molecular mechanism investigations, contents of aspartic acid (Asp), gamma-aminobutyric acid (GABA), glutamic acid (Glu), glycine (Gly) in seizures mice were determined; then, the chronic seizures rats induced by PTZ were prepared, and western blotting was used to determine the expressions of GAD 65, GABAA and GABAB in the brains. In the results, matrine showed significant antiepileptic effects on seizures mice induced by MES and PTZ. Moreover, the pentobarbital sodium-induced anaesthetizing time and locomotor activity tests were also demonstrated that matrine had obvious antiepileptic effects. Additionally, our results revealed that after treatment with matrine, contents of GABA can be elevated, and the contents of Glu were obviously decreased. Furthermore, western blotting revealed that the mechanism regarding the antiepileptic effect of may be related to the up-regulations of GAD 65 and GABAA in the brain. Collectively, we suggested that matrine can be developed as an effective antiseptic drug.

  20. Antiepileptic Potential of Matrine via Regulation the Levels of Gamma-Aminobutyric Acid and Glutamic Acid in the Brain

    PubMed Central

    Xiang, Jun; Jiang, Yugang

    2013-01-01

    Our present study aimed to determine the antiepileptic activity of matrine, and explore the possible molecular mechanism. To evaluate the antiepileptic activity of matrine, seizures in mice induced by PTZ and MES were established, then the pentobarbital sodium-induced anaesthetizing time and locomotor activity tests in mice were also carried out. For the molecular mechanism investigations, contents of aspartic acid (Asp), gamma-aminobutyric acid (GABA), glutamic acid (Glu), glycine (Gly) in seizures mice were determined; then, the chronic seizures rats induced by PTZ were prepared, and western blotting was used to determine the expressions of GAD 65, GABAA and GABAB in the brains. In the results, matrine showed significant antiepileptic effects on seizures mice induced by MES and PTZ. Moreover, the pentobarbital sodium-induced anaesthetizing time and locomotor activity tests were also demonstrated that matrine had obvious antiepileptic effects. Additionally, our results revealed that after treatment with matrine, contents of GABA can be elevated, and the contents of Glu were obviously decreased. Furthermore, western blotting revealed that the mechanism regarding the antiepileptic effect of may be related to the up-regulations of GAD 65 and GABAA in the brain. Collectively, we suggested that matrine can be developed as an effective antiseptic drug. PMID:24317434

  1. The effect of antivitamin B6 administration on gamma-aminobutyric acid metabolism in retina and electroretinogram.

    PubMed

    Mizuno, A; Kamada, Y; Kunita, M; Matsuda, M

    1980-01-01

    The effect of several antivitamin B6 on gamma-aminobutyric acid (GABA) metabolism was studied in the rat retina. The rat electroretinogram (ERG) was also recorded after administration of these drugs. Aminooxyacetic acid (AOAA) and hydrazine administration increased the GABA content and inhibited the GABA degrading enzyme, GABA transaminase in retina. In addition, there drugs elongated the peak latency of the oscillatory potential in the rat ERG. In contrast, 4-deoxypyridoxine (DOP) or isonicotinic acid hydrazide (INAH) administration decreased the GABA content and inhibited the GABA synthesizing enzyme, glutamic acid decarboxylase in retina, and administration of these drugs together with AOAA lessened the degrees of elevation of GABA content and of the elongation of the peak latency produced as compared with AOAA alone, though neither of the former drugs had a significant effect on ERG. The retinal GABA seems to play an important role in relation to the oscillatory potential of ERG.

  2. The role of glutamate and gamma-aminobutyric acid in fear extinction: clinical implications for exposure therapy.

    PubMed

    Davis, Michael; Myers, Karyn M

    2002-11-15

    Although much is now known about the neural basis of fear acquisition, the mechanisms of fear inhibition or suppression remain largely obscure. Fear inhibition is studied in the laboratory through the use of an extinction procedure, in which an animal (typically a rat) is exposed to nonreinforced presentations of a conditioned stimulus (CS; e.g., a light or tone) that had previously been paired with a fear-inducing unconditioned stimulus (US; e.g., a mild footshock). Over the course of such training, the conditioned fear response exhibited by the rat in the presence of the CS is reduced in amplitude and frequency. This procedure is analogous to those employed in the treatment of fear dysregulation in humans, which typically involve exposure to the feared object in the absence of any overt danger. Recent work on the neural basis of extinction indicates that the neurotransmitters gamma-aminobutyric acid (GABA) and glutamate are critically involved. Gamma-aminobutyric acid may act to inhibit brain areas involved in fear learning (e.g., the amygdala), and glutamate, acting at N-methyl-D-aspartate receptors, may play a role in the neural plasticity that permits this GABA-mediated inhibition to be exerted appropriately. These insights have significant implications for the conduct of extinction-based clinical interventions for fear disorders.

  3. Production of gamma-aminobutyric acid in black raspberry juice during fermentation by Lactobacillus brevis GABA100.

    PubMed

    Kim, Ja Young; Lee, Moo Young; Ji, Geun Eog; Lee, Yeon Sook; Hwang, Keum Taek

    2009-03-15

    Black raspberry juice was fermented to produce gamma-aminobutyric acid (GABA) using lactic acid bacteria (Lactobacillus brevis GABA 100) at different temperatures (25, 30, or 37 degrees C) and pHs (3.5, 4, 4.5, 5, 5.5, or 6) for 15 days. Concentrations of GABA in the juices were determined during fermentation using HPLC. GABA was produced continuously even if the viable bacterial counts markedly decreased. The fermentation at 30 degrees C generally showed higher production of GABA in the juices than those at 25 and 37 degrees C. The GABA in the juices fermented at 30 degrees C reached the maximum levels on the 12th day. The juices fermented at lower pH and lower temperature showed a lower degradation of monomeric anthocyanins. The results suggest that black raspberry juice can be GABA enriched using lactic acid bacteria.

  4. Effects of glutamate decarboxylase and gamma-aminobutyric acid (GABA) transporter on the bioconversion of GABA in engineered Escherichia coli.

    PubMed

    Le Vo, Tam Dinh; Kim, Tae Wan; Hong, Soon Ho

    2012-05-01

    Gamma-aminobutyric acid (GABA) is a non-essential amino acid and a precursor of pyrrolidone, a monomer of nylon 4. GABA can be biosynthesized through the decarboxylation of L: -glutamate by glutamate decarboxylase. In this study, the effects of glutamate decarboxylase (gadA, gadB), glutamate/GABA antiporter (gadC) and GABA aminotransferase (gabT) on GABA production were investigated in Escherichia coli. Glutamate decarboxylase was overexpressed alone or with the glutamate/GABA antiporter to enhance GABA synthesis. GABA aminotransferase, which redirects GABA into the TCA cycle, was knock-out mutated. When gadB and gadC were co-overexpressed in the gabT mutant strain, a final GABA concentration of 5.46 g/l was obtained from 10 g/l of monosodium glutamate (MSG), which corresponded to a GABA yield of 89.5%.

  5. A GC-ECD method for estimation of free and bound amino acids, gamma-aminobutyric acid, salicylic acid, and acetyl salicylic acid from Solanum lycopersicum (L.).

    PubMed

    Meher, Hari Charan; Gajbhiye, Vijay T; Singh, Ghanendra

    2011-01-01

    A gas chromatograph with electron capture detection method for estimation of selected metabolites--amino acids (free and bound), gamma-aminobutyric acid (GABA), salicylic acid (SA), and acetyl salicylic acid (ASA) from tomato--is reported. The method is based on nitrophenylation of the metabolites by 1-fluoro-2, 4-dinitrobenzene under aqueous alkaline conditions to form dinitophenyl derivatives. The derivatives were stable under the operating conditions of GC. Analysis of bound amino acids comprised perchloric acid precipitation of protein, alkylation (carboxymethylation) with iodoacetic acid, vapor-phase hydrolysis, and derivatization with 1-fluoro-2,4-dinitrobenzene in that order. The metabolites were resolved in 35 min, using a temperature-programmed run. The method is rapid, sensitive, and precise. It easily measured the typical amino acids (aspartate, asparagine, glutamate, glutamine, alanine, leucine, lysine, and phenylalanine) used for identification and quantification of a protein, resolved amino acids of the same mass (leucine and isoleucine), satisfactorily measured sulfur amino acid (methionine, cystine, and cysteine), and quantified GABA, SA, and ASA, as well. The developed method was validated for specificity, linearity, and precision. It has been applied and recommended for estimation of 25 metabolites from Solanum lycopersicum (L.).

  6. A GC-ECD method for estimation of free and bound amino acids, gamma-aminobutyric acid, salicylic acid, and acetyl salicylic acid from Solanum lycopersicum (L.).

    PubMed

    Meher, Hari Charan; Gajbhiye, Vijay T; Singh, Ghanendra

    2011-01-01

    A gas chromatograph with electron capture detection method for estimation of selected metabolites--amino acids (free and bound), gamma-aminobutyric acid (GABA), salicylic acid (SA), and acetyl salicylic acid (ASA) from tomato--is reported. The method is based on nitrophenylation of the metabolites by 1-fluoro-2, 4-dinitrobenzene under aqueous alkaline conditions to form dinitophenyl derivatives. The derivatives were stable under the operating conditions of GC. Analysis of bound amino acids comprised perchloric acid precipitation of protein, alkylation (carboxymethylation) with iodoacetic acid, vapor-phase hydrolysis, and derivatization with 1-fluoro-2,4-dinitrobenzene in that order. The metabolites were resolved in 35 min, using a temperature-programmed run. The method is rapid, sensitive, and precise. It easily measured the typical amino acids (aspartate, asparagine, glutamate, glutamine, alanine, leucine, lysine, and phenylalanine) used for identification and quantification of a protein, resolved amino acids of the same mass (leucine and isoleucine), satisfactorily measured sulfur amino acid (methionine, cystine, and cysteine), and quantified GABA, SA, and ASA, as well. The developed method was validated for specificity, linearity, and precision. It has been applied and recommended for estimation of 25 metabolites from Solanum lycopersicum (L.). PMID:21391500

  7. Presence of Gamma-Aminobutyric Acid (Gaba) in the Pedal Mucus of the Critically Endangered Species Patella ferruginea.

    PubMed

    Rivera-Ingraham, G A; Espinosa, F; Krock, B

    2015-05-01

    Patella ferruginea is a giant patellid limpet endemic to the Mediterranean Sea. It presently is in danger of extinction, and many have called for developing conservation measures including the mass production of spats for re-introduction projects. However, so far all attempts have been relatively unsuccessful. Previous work analyzing the effects of gamma-aminobutyric acid (GABA) on the recruitment of patellid limpets has shown that larvae respond to the presence of this signaling molecule. This response could explain the gregarious distribution typical of this species. In the present study, we demonstrated that GABA is naturally secreted by P. ferruginea in the pedal mucus. GABA is preferentially secreted during the summer, coinciding with the reproductive resting period of the species. Further research should aim to analyze the effects of GABA on larval development and metamorphosis in order to assess its potential use to improve conservation efforts.

  8. Cannabinoid receptor agonist WIN 55,212-2 inhibits rat cortical dialysate gamma-aminobutyric acid levels.

    PubMed

    Ferraro, L; Tomasini, M C; Cassano, T; Bebe, B W; Siniscalchi, A; O'Connor, W T; Magee, P; Tanganelli, S; Cuomo, V; Antonelli, T

    2001-10-15

    The effects of the cannabinoid receptor agonist WIN 55,212-2 (0.1-5 mg/kg i.p.) on endogenous extracellular gamma-aminobutyric acid (GABA) levels in the cerebral cortex of the awake rat was investigated by using microdialysis. WIN 55,212-2 (1 and 5 mg/kg i.p.) was associated with a concentration-dependent decrease in dialysate GABA levels (-16% +/- 4% and -26% +/- 4% of basal values, respectively). The WIN 55,212-2 (5 mg/kg i.p.) induced-inhibition was counteracted by a dose (0.1 mg/kg i.p.) of the CB(1) receptor antagonist SR141716A, which by itself was without effect on cortical GABA levels. These findings suggest that cannabinoids decrease cortical GABA levels in vivo, an action that might underlie some of the cognitive and behavioral effects of acute exposure to marijuana.

  9. Gamma-aminobutyric acid depletion affects stomata closure and drought tolerance of Arabidopsis thaliana.

    PubMed

    Mekonnen, Dereje Worku; Flügge, Ulf-Ingo; Ludewig, Frank

    2016-04-01

    A rapid accumulation of γ-aminobutyric acid (GABA) during biotic and abiotic stresses is well documented. However, the specificity of the response and the primary role of GABA under such stress conditions are hardly understood. To address these questions, we investigated the response of the GABA-depleted gad1/2 mutant to drought stress. GABA is primarily synthesized from the decarboxylation of glutamate by glutamate decarboxylase (GAD) which exists in five copies in the genome of Arabidopsis thaliana. However, only GAD1 and GAD2 are abundantly expressed, and knockout of these two copies dramatically reduced the GABA content. Phenotypic analysis revealed a reduced shoot growth of the gad1/2 mutant. Furthermore, the gad1/2 mutant was wilted earlier than the wild type following a prolonged drought stress treatment. The early-wilting phenotype was due to an increase in stomata aperture and a defect in stomata closure. The increase in stomata aperture contributed to higher stomatal conductance. The drought oversensitive phenotype of the gad1/2 mutant was reversed by functional complementation that increases GABA level in leaves. The functionally complemented gad1/2 x pop2 triple mutant contained more GABA than the wild type. Our findings suggest that GABA accumulation during drought is a stress-specific response and its accumulation induces the regulation of stomatal opening thereby prevents loss of water.

  10. Non-neuronal release of gamma-aminobutyric Acid by embryonic pluripotent stem cells.

    PubMed

    Teng, Lin; Tang, Ya-Bin; Sun, Fan; An, Shi-Min; Zhang, Chun; Yang, Xin-Jie; Lv, Hao-Yu; Lu, Qin; Cui, Yong-Yao; Hu, Jin-Jia; Zhu, Liang; Chen, Hong-Zhuan

    2013-11-15

    γ-Aminobutyric acid (GABA), the principle inhibitory transmitter in the mature central nervous system, is also involved in activities outside the nervous system. Recent studies have shown that functional GABA receptors are expressed in embryonic stem (ES) cells and these receptors control ES cell proliferation. However, it is not clear whether ES cells have their own GABAergic transmission output machinery that can fulfill GABA release or whether the cells merely process the GABA receptors by receiving and responding to the diffused GABA released elsewhere. To get further insight into this unresolved problem, we detected the repertoire of components for GABA synthesis, storage, reaction, and termination in ES and embryonal carcinoma stem cells by biological assays, and then directly quantified released GABA in the intercellular milieu from these pluripotent stem (PS) cells by an analytical chemical assay based on high-performance liquid chromatography coupled with electrospray ionization tandem mass spectrometry (HPLC-ESI-MS/MS). We found that embryonic PS cells processed a GABAergic circuit machinery and spontaneously released GABA, which suggests the potential that embryonic PS cells could autonomously establish a GABA niche via release of the transmitter.

  11. Potentiation of Gamma Aminobutyric Acid Receptors (GABAAR) by Ethanol: How Are Inhibitory Receptors Affected?

    PubMed Central

    Förstera, Benjamin; Castro, Patricio A.; Moraga-Cid, Gustavo; Aguayo, Luis G.

    2016-01-01

    In recent years there has been an increase in the understanding of ethanol actions on the type A γ-aminobutyric acid chloride channel (GABAAR), a member of the pentameric ligand gated ion channels (pLGICs). However, the mechanism by which ethanol potentiates the complex is still not fully understood and a number of publications have shown contradictory results. Thus many questions still remain unresolved requiring further studies for a better comprehension of this effect. The present review concentrates on the involvement of GABAAR in the acute actions of ethanol and specifically focuses on the immediate, direct or indirect, synaptic and extra-synaptic modulatory effects. To elaborate on the immediate, direct modulation of GABAAR by acute ethanol exposure, electrophysiological studies investigating the importance of different subunits, and data from receptor mutants will be examined. We will also discuss the nature of the putative binding sites for ethanol based on structural data obtained from other members of the pLGICs family. Finally, we will briefly highlight the glycine gated chloride channel (GlyR), another member of the pLGIC family, as a suitable target for the development of new pharmacological tools. PMID:27199667

  12. Gamma-aminobutyric acid depletion affects stomata closure and drought tolerance of Arabidopsis thaliana.

    PubMed

    Mekonnen, Dereje Worku; Flügge, Ulf-Ingo; Ludewig, Frank

    2016-04-01

    A rapid accumulation of γ-aminobutyric acid (GABA) during biotic and abiotic stresses is well documented. However, the specificity of the response and the primary role of GABA under such stress conditions are hardly understood. To address these questions, we investigated the response of the GABA-depleted gad1/2 mutant to drought stress. GABA is primarily synthesized from the decarboxylation of glutamate by glutamate decarboxylase (GAD) which exists in five copies in the genome of Arabidopsis thaliana. However, only GAD1 and GAD2 are abundantly expressed, and knockout of these two copies dramatically reduced the GABA content. Phenotypic analysis revealed a reduced shoot growth of the gad1/2 mutant. Furthermore, the gad1/2 mutant was wilted earlier than the wild type following a prolonged drought stress treatment. The early-wilting phenotype was due to an increase in stomata aperture and a defect in stomata closure. The increase in stomata aperture contributed to higher stomatal conductance. The drought oversensitive phenotype of the gad1/2 mutant was reversed by functional complementation that increases GABA level in leaves. The functionally complemented gad1/2 x pop2 triple mutant contained more GABA than the wild type. Our findings suggest that GABA accumulation during drought is a stress-specific response and its accumulation induces the regulation of stomatal opening thereby prevents loss of water. PMID:26940489

  13. The importance of glutamate, glycine, and {gamma}-aminobutyric acid transport and regulation in manganese, mercury and lead neurotoxicity

    SciTech Connect

    Fitsanakis, Vanessa A.; Aschner, Michael . E-mail: michael.aschner@vanderbilt.edu

    2005-05-01

    Historically, amino acids were studied in the context of their importance in protein synthesis. In the 1950s, the focus of research shifted as amino acids were recognized as putative neurotransmitters. Today, many amino acids are considered important neurochemicals. Although many amino acids play a role in neurotransmission, glutamate (Glu), glycine (Gly), and {gamma}-aminobutyric acid (GABA) are among the more prevalent and better understood. Glu, the major excitatory neurotransmitter, and Gly and GABA, the major inhibitory neurotransmitters, in the central nervous system, are known to be tightly regulated. Prolonged exposure to environmental toxicants, such as manganese (Mn), mercury (Hg), or lead (Pb), however, can lead to dysregulation of these neurochemicals and subsequent neurotoxicity. While the ability of these metals to disrupt the regulation of Glu, Gly and GABA have been studied, few articles have examined the collective role of these amino acids in the respective metal's mechanism of toxicity. For each of the neurotransmitters above, we will provide a brief synopsis of their regulatory function, including the importance of transport and re-uptake in maintaining their optimal function. Additionally, the review will address the hypothesis that aberrant homeostasis of any of these amino acids, or a combination of the three, plays a role in the neurotoxicity of Mn, Hg, or Pb.

  14. Effects of Frequency Drift on the Quantification of Gamma-Aminobutyric Acid Using MEGA-PRESS

    PubMed Central

    Tsai, Shang-Yueh; Fang, Chun-Hao; Wu, Thai-Yu; Lin, Yi-Ru

    2016-01-01

    The MEGA-PRESS method is the most common method used to measure γ-aminobutyric acid (GABA) in the brain at 3T. It has been shown that the underestimation of the GABA signal due to B0 drift up to 1.22 Hz/min can be reduced by post-frequency alignment. In this study, we show that the underestimation of GABA can still occur even with post frequency alignment when the B0 drift is up to 3.93 Hz/min. The underestimation can be reduced by applying a frequency shift threshold. A total of 23 subjects were scanned twice to assess the short-term reproducibility, and 14 of them were scanned again after 2–8 weeks to evaluate the long-term reproducibility. A linear regression analysis of the quantified GABA versus the frequency shift showed a negative correlation (P < 0.01). Underestimation of the GABA signal was found. When a frequency shift threshold of 0.125 ppm (15.5 Hz or 1.79 Hz/min) was applied, the linear regression showed no statistically significant difference (P > 0.05). Therefore, a frequency shift threshold at 0.125 ppm (15.5 Hz) can be used to reduce underestimation during GABA quantification. For data with a B0 drift up to 3.93 Hz/min, the coefficients of variance of short-term and long-term reproducibility for the GABA quantification were less than 10% when the frequency threshold was applied. PMID:27079873

  15. Effects of Frequency Drift on the Quantification of Gamma-Aminobutyric Acid Using MEGA-PRESS

    NASA Astrophysics Data System (ADS)

    Tsai, Shang-Yueh; Fang, Chun-Hao; Wu, Thai-Yu; Lin, Yi-Ru

    2016-04-01

    The MEGA-PRESS method is the most common method used to measure γ-aminobutyric acid (GABA) in the brain at 3T. It has been shown that the underestimation of the GABA signal due to B0 drift up to 1.22 Hz/min can be reduced by post-frequency alignment. In this study, we show that the underestimation of GABA can still occur even with post frequency alignment when the B0 drift is up to 3.93 Hz/min. The underestimation can be reduced by applying a frequency shift threshold. A total of 23 subjects were scanned twice to assess the short-term reproducibility, and 14 of them were scanned again after 2–8 weeks to evaluate the long-term reproducibility. A linear regression analysis of the quantified GABA versus the frequency shift showed a negative correlation (P < 0.01). Underestimation of the GABA signal was found. When a frequency shift threshold of 0.125 ppm (15.5 Hz or 1.79 Hz/min) was applied, the linear regression showed no statistically significant difference (P > 0.05). Therefore, a frequency shift threshold at 0.125 ppm (15.5 Hz) can be used to reduce underestimation during GABA quantification. For data with a B0 drift up to 3.93 Hz/min, the coefficients of variance of short-term and long-term reproducibility for the GABA quantification were less than 10% when the frequency threshold was applied.

  16. Dynamic changes in gamma-aminobutyric acid and glutamate decarboxylase activity in oats (Avena nuda L.) during steeping and germination.

    PubMed

    Xu, Jian Guo; Hu, Qing Ping; Duan, Jiang Lian; Tian, Cheng Rui

    2010-09-01

    Gamma-aminobutyric acid (GABA) is the principal inhibitory neurotransmitter in the central nervous system and provides beneficial effects for human and other animals health. To accumulate GABA, samples from two different naked oat cultivars, Baiyan II and Bayou I, were steeped and germinated in an incubator. The content of GABA and glutamic acid as well as the activity of the glutamate decarboxylase (GAD) in oats during steeping and germination were investigated with an amino acid automatic analyzer. Compared with raw groats, an increase in GABA content of oat groats during steeping and germination was continuously observed for two oat cultivars. The activity of GAD increased greatly at the end of steeping and the second stage of germination for Baiyan II and Bayou I, respectively. Glutamic acid content of treated oat groats was significantly lower than that in raw groats until the later period of germination. GABA was correlated (p<0.01) significantly and positively with the glutamic acid rather than GAD activity in the current study. The results indicates that steeping and germination process under highly controlled conditions can effectively accumulate the GABA in oat groats for Baiyan II and Bayou I, which would greatly facilitate production of nutraceuticals or food ingredients that enable consumers to gain greater access to the health benefits of oats. However, more assays need to be further performed with more oat cultivars.

  17. Cellular synthesis and axonal transport of gamma-aminobutyric acid in a photoreceptor cell of the barnacle.

    PubMed Central

    Koike, H; Tsuda, K

    1980-01-01

    1. [3H]glutamate or [3H]gamma-aminobutyric acid (GABA) was injected into the photoreceptor cell of the lateral ocellus of Balanus eburneus, in order to study the transmitter substance of the cell. 2. The photoreceptor cell synthesized [3H]GABA from injected [3H]glutamate. 3. The newly formed [3H]GABA moved inside the photoreceptor axon towards the axon terminal with a velocity of about 0.9 mm/hr. Injected [3H]GABA moved at 0.9 mm/hr and also at 0.4 mm/hr. 4. Axonally transported [3H]GABA reached the axon terminal within several hours following the injection. It did not accumulate at the terminal, but gradually disappeared. 5. Light-microscope and electron-microscope autoradiography following the injection of [3H]GABA revealed that [3H]-reacted silver grains were present in a certain type of axon terminal. The terminal thus identified as that of a photoreceptor cell contains many clear, polymorphic synaptic vesicles about 300-500 A in diameter, some dense-cored vesicles 700-1300 A in diameter, and glycogen granules. The terminal forms many synapses, and each synapse has a synaptic dense body. The terminal always faces two post-synaptic elements at the synapse, forming a triad with a gap distance of about 160-200 A. 6. A GABA analogue, [3H]di-aminobutyric acid, was selectively taken up into the terminals previously identified as those of photoreceptors. 7. These results support the notion that the transmitter substance of the photoreceptor cell of the barnacle is GABA. Images Plate 1 Plate 2 PMID:6160239

  18. Electrophoretic method for the determination of the proportion of gamma-aminobutyric acid in a mixture of labeled neurotransmitter and its catabolites

    SciTech Connect

    Cupello, A.; Rapallino, M.V.; Besio, G.; Mainardi, P.

    1987-01-01

    An electrophoretic method for the separation of gamma-aminobutyric acid (GABA) from its metabolites after GABA-transaminase attack is presented. The method is based on the fact that at neutral pH GABA has no net electrical charge, whereas its major metabolites, succinic acid and Krebs cycle intermediates, are negatively charged. The method appears to be especially suitable for evaluation of true-labeled neurotransmitter within the radioactivity which is found in synaptosomes after labeled GABA-uptake studies.

  19. Direct protein-protein coupling enables cross-talk between dopamine D5 and gamma-aminobutyric acid A receptors.

    PubMed

    Liu, F; Wan, Q; Pristupa, Z B; Yu, X M; Wang, Y T; Niznik, H B

    2000-01-20

    GABA(A) (gamma-aminobutyric-acid A) and dopamine D1 and D5 receptors represent two structurally and functionally divergent families of neurotransmitter receptors. The former comprises a class of multi-subunit ligand-gated channels mediating fast interneuronal synaptic transmission, whereas the latter belongs to the seven-transmembrane-domain single-polypeptide receptor superfamily that exerts its biological effects, including the modulation of GABA(A) receptor function, through the activation of second-messenger signalling cascades by G proteins. Here we show that GABA(A)-ligand-gated channels complex selectively with D5 receptors through the direct binding of the D5 carboxy-terminal domain with the second intracellular loop of the GABA(A) gamma2(short) receptor subunit. This physical association enables mutually inhibitory functional interactions between these receptor systems. The data highlight a previously unknown signal transduction mechanism whereby subtype-selective G-protein-coupled receptors dynamically regulate synaptic strength independently of classically defined second-messenger systems, and provide a heuristic framework in which to view these receptor systems in the maintenance of psychomotor disease states.

  20. Two-step production of gamma-aminobutyric acid from cassava powder using Corynebacterium glutamicum and Lactobacillus plantarum.

    PubMed

    Yang, Taowei; Rao, Zhiming; Kimani, Bernard Gitura; Xu, Meijuan; Zhang, Xian; Yang, Shang-Tian

    2015-08-01

    Production of gamma-aminobutyric acid (GABA) from crop biomass such as cassava in high concentration is desirable, but difficult to achieve. A safe biotechnological route was investigated to produce GABA from cassava powder by C. glutamicum G01 and L. plantarum GB01-21. Liquefied cassava powder was first transformed to glutamic acid by simultaneous saccharification and fermentation with C. glutamicum G01, followed by biotransformation of glutamic acid to GABA with resting cells of L. plantarum GB01-21 in the reaction medium. After optimizing the reaction conditions, the maximum concentration of GABA reached 80.5 g/L with a GABA productivity of 2.68 g/L/h. This is the highest yield ever reported of GABA production from cassava-derived glucose. The bioprocess provides the added advantage of employing nonpathogenic microorganisms, C. glutamicum and L. plantarum, in microbial production of GABA from cassava biomass, which can be used in the food and pharmaceutical industries.

  1. Gamma-aminobutyric acid production using immobilized glutamate decarboxylase followed by downstream processing with cation exchange chromatography.

    PubMed

    Lee, Seungwoon; Ahn, Jungoh; Kim, Yeon-Gu; Jung, Joon-Ki; Lee, Hongweon; Lee, Eun Gyo

    2013-01-15

    We have developed a gamma-aminobutyric acid (GABA) production technique using his-tag mediated immobilization of Escherichia coli-derived glutamate decarboxylase (GAD), an enzyme that catalyzes the conversion of glutamate to GABA. The GAD was obtained at 1.43 g/L from GAD-overexpressed E. coli fermentation and consisted of 59.7% monomer, 29.2% dimer and 2.3% tetramer with a 97.6% soluble form of the total GAD. The harvested GAD was immobilized to metal affinity gel with an immobilization yield of 92%. Based on an investigation of specific enzyme activity and reaction characteristics, glutamic acid (GA) was chosen over monosodium glutamate (MSG) as a substrate for immobilized GAD, resulting in conversion of 2.17 M GABA in a 1 L reactor within 100 min. The immobilized enzymes retained 58.1% of their initial activities after ten consecutive uses. By using cation exchange chromatography followed by enzymatic conversion, GABA was separated from the residual substrate and leached GAD. As a consequence, the glutamic acid was mostly removed with no detectable GAD, while 91.2% of GABA was yielded in the purification step.

  2. Determination of glutamic acid and gamma-aminobutyric acid in Ringer's solution without desalination at the femtomole level by gas chromatography chemical ionization mass spectrometry.

    PubMed

    Murayama, K; Shindo, N; Mineki, R; Ohta, K

    1981-04-01

    For the quantification of glutamic acid in Ringer's solution, pentafluoropropionic methyl ester was the most sensitive derivative. The detectable concentration was 0.01 microM glutamic acid in Ringer's solution; the amount of the preparation was 1 pmol and the injection into a gas chromatograph mass spectrometer was 10 fmol. For the quantification of gamma-aminobutyric acid in Ringer's solution, the trifluoroacetal-hexafluoropropionyl ester was quantification of gamma-aminobutyric acid in Ringer's solution, the trifluoroacetal-hexafluoropropionyl ester was detectable at a concentration of 0.01 microM. Ringer's salts facilitated acylation in the order heptafluorobutyric anhydride greater than pentafluoropropionic anhydride greater than trifluoroacetic anhydride. The effect depended on esterification of carboxy groups in the order methyl ester greater than hexafluoropropionyl ester greater than butyl ester. Sodium carbonate, sodium acetate and sodium citrate also facilitated acylation with pentafluoroproionic anhydride, while sodium phosphate inhibited the acylation and sodium sulfate inhibited it slightly. The pentafluoropropionic methyl ester of glutamic acid was stable for up to 10 days, when it was dissolved in acetone and stored at -18 degrees C.

  3. Monitoring Technology for Gamma-Aminobutyric acid Production in Polished Mochi Barley Grains using a Carbon Dioxide Sensor.

    PubMed

    Watanabe, Yasuo; Kawata, Kohki; Watanabe, Seiya

    2015-06-01

    Gamma-aminobutyric acid (GABA) has many biological functions, including the inhibition of blood pressure increases and acceleration of growth hormone secretion. In this study, we discovered the utility of measuring the concentration of carbon dioxide (CO2 ) dissolved in the reaction solution, for development of a real-time and convenient technique to estimate GABA production. In addition to mochi barley bran, we examined the polished grains of three species: mochi barley (a variant of hulless barley), barley, and Japanese millet, all soaked in l-glutamic acid (l-Glu) solution at pH 4.5. We found a positive correlation between GABA and CO2 concentrations, and the production of CO2 was suppressed in the absence of l-Glu at pH 4.5. These results suggest that GABA content can be easily predicted by measuring the aqueous CO2 content using a CO2 sensor, during the process of GABA production in polished mochi barley grains and bran.

  4. The cardiovascular and subjective effects of methamphetamine combined with gamma-vinyl-gamma-aminobutyric acid (GVG) in non-treatment seeking methamphetamine-dependent volunteers.

    PubMed

    De La Garza, Richard; Zorick, Todd; Heinzerling, Keith G; Nusinowitz, Steve; London, Edythe D; Shoptaw, Steven; Moody, David E; Newton, Thomas F

    2009-11-01

    Gamma-vinyl-gamma-aminobutyric acid (GVG) elevates central nervous system gamma-aminobutyric acid (GABA) levels by irreversibly inhibiting GABA transaminase. An open-label clinical trial in humans suggested that GVG may reduce cocaine and methamphetamine use. To test safety and to obtain preliminary data on efficacy of GVG for treating methamphetamine dependence, we conducted a double-blind, placebo-controlled, parallel group study of GVG interaction with the cardiovascular and subjective effects produced by methamphetamine. Non-treatment seeking methamphetamine-dependent volunteers received either GVG (N=8) or placebo (N=9) by random assignment. GVG treatment was initiated at 1 g/day and increased to 5 g/day. After reaching the target dose of 5 g/day, participants received methamphetamine (15+30 mg, IV), and cardiovascular and subjective effects were assessed. No serious adverse events were noted, and the total number of adverse events was similar between the treatment groups. Considering the full time course and peak effects independently, no significant differences were detected between the groups for systolic or diastolic blood pressures, or heart rate, following methamphetamine exposure. Some methamphetamine-induced cardiovascular changes approached significance (p<0.10) and may warrant attention in future trials. Methamphetamine-induced subjective effects ("any drug effect", "high", "crave methamphetamine") were statistically similar between GVG and placebo treatment groups. Pharmacokinetic data indicate that GVG treatment did not alter methamphetamine or amphetamine plasma levels, and there was no association between methamphetamine or amphetamine plasma levels and peak cardiovascular effects. Taken together, the data indicate that GVG treatment is generally well tolerated but not efficacious in attenuating the positive subjective effects of methamphetamine in the laboratory.

  5. Dual action of isoflurane on the gamma-aminobutyric acid (GABA)-mediated currents through recombinant alpha(1)beta(2)gamma(2L)-GABA(A)-receptor channels.

    PubMed

    Neumahr, S; Hapfelmeier, G; Scheller, M; Schneck, H; Franke, C; Kochs, E

    2000-05-01

    Isoflurane (ISO) increased the agonist-induced chloride flux through the gamma-aminobutyric acid A receptor (GABA(A)R). This may reflect an anesthetic-induced increase in the apparent agonist affinity. A dual effect of anesthetics was postulated for both the nicotinic acetylcholine receptor (nAChR) and the GABA(A)R. We tested the hypothesis that, in addition to a blocking effect, ISO increases gamma-aminobutyric acid (GABA)-gated currents through recombinant GABA(A)R channels. HEK293 cells were transfected with rat cDNA for alpha(1),beta(2),gamma(2L) subunits. Currents elicited by 1 mM or 0. 01 mM GABA, respectively, alone, or with increasing concentrations of ISO, were recorded by using standard patch clamp techniques. ISO reduced the peak current elicited by 1 mM GABA. Currents induced by 0.01 mM GABA were potentiated by small ISO (twofold at 0.5 mM ISO) and inhibited by larger concentrations. Withdrawal of ISO and GABA induced rebound currents, suggesting an open-channel block by ISO. These currents increased with increasing concentrations of ISO. At large concentrations of ISO, the inhibitory effect predominated and was caused by, at least partly, an open-channel block. At small concentrations of ISO, potentiation of the GABA-gated currents was more prominent. This dual action of ISO indicates different binding sites at the GABA(A)R. The balance between potentiation and block depends on the concentrations of both ISO and GABA.

  6. Gamma aminobutyric acid B and 5-hydroxy tryptamine 2A receptors functional regulation during enhanced liver cell proliferation by GABA and 5-HT chitosan nanoparticles treatment.

    PubMed

    Shilpa, Joy; Pretty, Mary Abraham; Anitha, Malat; Paulose, Cheramadathikudyil Skaria

    2013-09-01

    Liver is one of the major organs in vertebrates and hepatocytes are damaged by many factors. The liver cell maintenance and multiplication after injury and treatment gained immense interest. The present study investigated the role of Gamma aminobutyric acid (GABA) and serotonin or 5-hydroxytryptamine (5-HT) coupled with chitosan nanoparticles in the functional regulation of Gamma aminobutyric acid B and 5-hydroxy tryptamine 2A receptors mediated cell signaling mechanisms, extend of DNA methylation and superoxide dismutase activity during enhanced liver cell proliferation. Liver injury was achieved by partial hepatectomy of male Wistar rats and the GABA and 5-HT chitosan nanoparticles treatments were given intraperitoneally. The experimental groups were sham operated control (C), partially hepatectomised rats with no treatment (PHNT), partially hepatectomised rats with GABA chitosan nanoparticle (GCNP), 5-HT chitosan nanoparticle (SCNP) and a combination of GABA and 5-HT chitosan nanoparticle (GSCNP) treatments. In GABA and 5-HT chitosan nanoparticle treated group there was a significant decrease (P<0.001) in the receptor expression of Gamma aminobutyric acid B and a significant increase (P<0.001) in the receptor expression of 5-hydroxy tryptamine 2A when compared to PHNT. The cyclic adenosine monophosphate content and its regulatory protein, presence of methylated DNA and superoxide dismutase activity were decreased in GCNP, SCNP and GSCNP when compared to PHNT. The Gamma aminobutyric acid B and 5-hydroxy tryptamine 2A receptors coupled signaling elements played an important role in GABA and 5-HT chitosan nanoparticles induced liver cell proliferation which has therapeutic significance in liver disease management.

  7. Visualization of spatial distribution of gamma-aminobutyric acid in eggplant (Solanum melongena) by matrix-assisted laser desorption/ionization imaging mass spectrometry.

    PubMed

    Goto-Inoue, Naoko; Setou, Mitsutoshi; Zaima, Nobuhiro

    2010-01-01

    We applied imaging mass spectrometry (IMS) to determine the spatial distribution of gamma-aminobutyric acid (GABA). We found that GABA had a specific localization in seeds. We also visualized various biomolecules as well as GABA with higher spatial resolution than in the previous report. Our work suggests that IMS might be a powerful tool for exploring functional food factors, investigating the specific distribution of nutrients in unused natural resources, and evaluating the quality of functional foods.

  8. Gamma-Aminobutyric acid and benzodiazepine receptors in the kindling model of epilepsy: a quantitative radiohistochemical study

    SciTech Connect

    Shin, C.; Pedersen, H.B.; McNamara, J.O.

    1985-10-01

    Quantitative radiohistochemistry was utilized to study alterations of gamma-aminobutyric acid (GABA) and benzodiazepine receptors in the kindling model of epilepsy. The radioligands used for GABA and benzodiazepine receptors were (TH) muscimol and (TH)flunitrazepam, respectively. GABA receptor binding was increased by 22% in fascia dentata of the hippocampal formation but not in neocortex or substantia nigra of kindled rats. Within fascia dentata, GABA receptor binding was increased to an equivalent extent in stratum granulosum and throughout stratum moleculare; no increase was found in dentate hilus or stratum lacunosummoleculare or stratum radiatum of CA1. The increased binding was present at 24 hr but not at 28 days after the last kindled seizure. The direction, anatomic distribution, and time course of the increased GABA receptor binding were paralleled by increased benzodiazepine receptor binding. The anatomic distribution of the increased GABA receptor binding is consistent with a localization to somata and dendritic trees of dentate granule cells. The authors suggest that increased GABA and benzodiazepine receptor binding may contribute to enhanced inhibition of dentate granule cells demonstrated electrophysiologically in kindled animals.

  9. Correlation Between Hepatocyte Growth Factor (HGF) and Gamma-Aminobutyric Acid (GABA) Plasma Levels in Autistic Children.

    PubMed

    Russo, Anthony J

    2013-01-01

    There is much support for the role of Gamma-Aminobutyric acid (GABA) in the etiology of autism. Recent research has shown that hepatocyte growth factor (HGF) modulates GABAergic inhibition and seizure susceptibility. This study was designed to determine and correlate plasma levels of HGF, GABA, as well as symptom severity, in autistic children and neurotypical controls. Plasma from 48 autistic children and 29 neurotypical controls was assessed for HGF and GABA concentration using ELISAs. Symptom severity was assessed in these autistic individuals and compared to HGF and GABA concentrations. We previously reported that autistic children had significantly decreased levels of HGF. In this study, the same autistic children had significantly increased plasma levels of GABA (P = 0.002) and decreased HGF levels correlated with these increased GABA levels (r = 0.3; P = 0.05). High GABA levels correlated with increasing hyperactivity (r = 0.6; P = 0.0007) and impulsivity severity (r = 0.5; P = 0.007), tip toeing severity (r = 0.35; P = 0.03), light sensitivity (r = 0.4; P = 0.02), and tactile sensitivity (r = 0.4; P = 0.01). HGF levels did not correlate significantly with any symptom severity. These results suggest an association between HGF and GABA levels and suggest that plasma GABA levels are related to symptom severity in autistic children.

  10. Gamma-aminobutyric acid induces tumor cells apoptosis via GABABR1·β-arrestins·JNKs signaling module.

    PubMed

    Tian, Hui; Wu, Jin-Xia; Shan, Feng-Xiao; Zhang, Shang-Nuan; Cheng, Qian; Zheng, Jun-Nian; Pei, Dong-Sheng

    2015-03-01

    Gamma-aminobutyric acid (GABA), an inhibitory neurotransmitter in central nervous system, has yet been found to widely exist in tumor tissues to regulate tumor cells growth. However, the function of GABA on inducing tumor cells apoptosis and the potential mechanism are still unclear. In order to detect whether GABA via GABAB receptor GABABR1 would activate c-Jun N-terminal kinases (JNKs) to promote tumor cells apoptosis, co-immunoprecipitation assay was used to investigate the association of β-arrestins with GABABR1 and JNKs in the different four cancer cell lines. Our observation demonstrated that β-arrestins, in addition to their role in G protein-coupled receptors desensitization, had an additional function as adapter proteins to recruit JNKs to GABABR1, thereby conferring distinct enzymatic activities upon the receptor, which may trigger JNKs signal pathway involved in the regulation of cellular growth. Activated JNKs subsequently phosphorylated downstream c-Jun to transcribe a wide variety of pro-apoptotic genes. Additionally, GABA up-regulated the ratio of pro-apoptotic protein Bax to anti-apoptotic protein Bcl-2, and thus facilitated caspase-3 cleavage, leading to tumor cells apoptosis in a mitochondrial-dependent pathway. In contrast, GABABR antagonist CGP35348 reversed GABA-induced JNKs phosphorylation and its downstream proteins activation, which consequently restrained tumor cells apoptosis. Taken together, our study suggested that GABA via its receptor GABABR1 recruited β-arrestins to facilitate the activation of JNKs cascade, resulting in tumor cells growth inhibition.

  11. Analysis of subcomponents of the gamma-aminobutyric acid/benzodiazepine receptor macromolecular complex in mammalian central nervous system

    SciTech Connect

    McCabe, R.T.

    1987-01-01

    Since the presence of endogenous gamma-aminobutyric acid (GABA) may affect benzodiazepine binding to tissue sections in autoradiographic studies, a protocol designed to check for this influence has been investigated. (/sup 3/H)Flunitrazepam (1 nM) was used to label benzodiazepine receptors for autoradiographic localization. Bicuculline was added to the incubation medium of an additional set of tissue sections to antagonize any potential effect of endogenous GABA. Binding in these sections was compared to that occurring in another set in which excess GABA was added to create further GABA enhancement. Binding was also compared to adjacent sections which were treated similarly but also preincubated in distilled-deionized water to burst the cells by osmotic shock and eliminate endogenous GABA, thereby preventing any effect on benzodiazepine binding. The results indicated that endogenous GABA is indeed present in the slide-mounted tissue sections and is affecting benzodiazepine receptor binding differentially in various regions of the brain depending on the density of GABAergic innervation. Scatchard analysis of saturation data demonstrated that the alteration in BZ binding due to GABA was a result of a change in the affinity rather than number of receptors present.

  12. [Effects of gamma-aminobutyric acid on the photosynthesis and chlorophyll fluorescence parameters of muskmelon seedlings under hypoxia stress].

    PubMed

    Xia, Qing-ping; Gao, Hong-bo; Li, Jing-rui

    2011-04-01

    By the method of hydroponic culture, this paper studied the effects of exogenous gamma-aminobutyric acid (GABA) on the photosynthetic pigment contents, photosynthesis, and chlorophyll fluorescence parameters of muskmelon seedlings under hypoxia stress. Hypoxia stress induced a significant decrease of photosynthetic pigment contents, resulting in the decrease of photosynthesis. Applying GABA could significantly increase the photosynthetic pigment contents, net photosynthetic rate (P(n)), stomatal conductance (G(s)), intercellular CO2 concentration (C(i)), carboxylation efficiency (CE), maximal photochemical efficiency of PS II (F(v)/F(m)), photochemical quenching (q(P)), apparent photosynthetic electron transfer rate (ETR), and quantum yield of PS II electron transport (phi(PS II)), and decrease the stomatal limitation value (L(s)), minimal fluorescence (F(o)), and non-photochemical quenching (NPQ) under both hypoxic and normal conditions. The alleviation effect of GABA on photosynthetic characteristics was more obvious under hypoxia stress. However, simultaneously applying GABA and VGB could significantly decrease the alleviation effect of GABA under hypoxia stress.

  13. Effects of vegetable containing gamma-aminobutyric acid on the cardiac autonomic nervous system in healthy young people.

    PubMed

    Okita, Yoshimitsu; Nakamura, Harunobu; Kouda, Katsuyasu; Takahashi, Isao; Takaoka, Terumi; Kimura, Motohiko; Sugiura, Toshifumi

    2009-01-01

    The aim of this study was to investigate the effects of vegetable tablets containing Gamma-Aminobutyric Acid (GABA) intake on cardiovascular response and the autonomic nervous system in young adults. In a double-blind, randomized controlled trial, 7 healthy subjects were assigned to take vegetable tablets (10 g/trial) or control tablets (10 g/trial). We measured heart rate (HR), systolic and diastolic blood pressure, stroke volume, cardiac output, total peripheral resistance index, and the low- and high-frequency oscillatory components of heart rate variability (HRV). Two major spectral components were examined at low-frequency (LF: 0.04-0.15 Hz) and high-frequency (HF: 0.15-0.4 Hz) bands to indicate HRV. There were significant interactions in HR (p<0.01) and in LF/HF of HRV (p<0.05). HR increased after intake of control tablets, but not after that of vegetable tablets. LF/HF increased rapidly after intake of control tablets and rose slightly after vegetable tablet intake. There was no significant difference between the vegetable and control tablet trials in stroke volume, cardiac output, total peripheral resistance, systolic or diastolic blood pressure, HF, or LF. In conclusion, these results suggest the possibility that single administration of vegetable tablets containing GABA suppresses the sympathetic nervous activity leading to an elevation of blood pressure.

  14. Dependence of gamma-aminobutyric acid modulation of cholinergic transmission on nitric oxide and purines in cat terminal ileum.

    PubMed

    Pencheva, N

    1997-11-27

    The possible involvement of purines and/or nitric oxide (NO) in the gamma-aminobutyric acid (GABA)A receptor-mediated effects on the spontaneous activity of isolated preparations from longitudinal and circular muscles of cat terminal ileum was investigated. GABA had biphasic effects, which were neurogenic and muscarinic. ATP and adenosine dose dependently inhibited the activity of the muscles. A contractile response evoked by the nucleotide only was also observed. The effects of the purines were equipotent and resistant to Nomega-nitro-L-arginine (L-NNA), tetrodotoxin and to desensitization by alpha,beta-methylene adenosine 5'-triphosphate (alpha,beta-meATP), except for the contractile effect of ATP, which was abolished by alpha,beta-meATP. Pretreatment of the preparations with ATP or adenosine produced: (i) desensitization to the effects of the respective purinoceptor agonist only; and (ii) suppression of the GABA-induced responses of longitudinal and circular muscles. Hemoglobin and L-NNA greatly reduced or completely blocked the GABA(A)-induced relaxation and decreased the GABA(A)-induced contraction. Our results indicate that purines and NO, to a different extent, mediate the relaxant phase of the GABA effects in both layers. Interactions between muscarinic cholinoceptors and GABA-nitrergic pathway and a concomitant activation of postjunctional P1 and P2y purinoceptors are suggested to explain the prejunctional biphasic effects of GABA.

  15. Overexpression and optimization of glutamate decarboxylase in Lactobacillus plantarum Taj-Apis362 for high gamma-aminobutyric acid production.

    PubMed

    Tajabadi, Naser; Baradaran, Ali; Ebrahimpour, Afshin; Rahim, Raha A; Bakar, Fatimah A; Manap, Mohd Yazid A; Mohammed, Abdulkarim S; Saari, Nazamid

    2015-07-01

    Gamma-aminobutyric acid (GABA) is an important bioactive compound biosynthesized by microorganisms through decarboxylation of glutamate by glutamate decarboxylase (GAD). In this study, a full-length GAD gene was obtained by cloning the template deoxyribonucleic acid to pTZ57R/T vector. The open reading frame of the GAD gene showed the cloned gene was composed of 1410 nucleotides and encoded a 469 amino acids protein. To improve the GABA-production, the GAD gene was cloned into pMG36e-LbGAD, and then expressed in Lactobacillus plantarum Taj-Apis362 cells. The overexpression was confirmed by SDS-PAGE and GAD activity, showing a 53 KDa protein with the enzyme activity increased by sevenfold compared with the original GAD activity. The optimal fermentation conditions for GABA production established using response surface methodology were at glutamic acid concentration of 497.973 mM, temperature 36°C, pH 5.31 and time 60 h. Under the conditions, maximum GABA concentration obtained (11.09 mM) was comparable with the predicted value by the model at 11.23 mM. To our knowledge, this is the first report of successful cloning (clone-back) and overexpression of the LbGAD gene from L. plantarum to L. plantarum cells. The recombinant Lactobacillus could be used as a starter culture for direct incorporation into a food system during fermentation for production of GABA-rich products.

  16. Overexpression and optimization of glutamate decarboxylase in Lactobacillus plantarum Taj-Apis362 for high gamma-aminobutyric acid production.

    PubMed

    Tajabadi, Naser; Baradaran, Ali; Ebrahimpour, Afshin; Rahim, Raha A; Bakar, Fatimah A; Manap, Mohd Yazid A; Mohammed, Abdulkarim S; Saari, Nazamid

    2015-07-01

    Gamma-aminobutyric acid (GABA) is an important bioactive compound biosynthesized by microorganisms through decarboxylation of glutamate by glutamate decarboxylase (GAD). In this study, a full-length GAD gene was obtained by cloning the template deoxyribonucleic acid to pTZ57R/T vector. The open reading frame of the GAD gene showed the cloned gene was composed of 1410 nucleotides and encoded a 469 amino acids protein. To improve the GABA-production, the GAD gene was cloned into pMG36e-LbGAD, and then expressed in Lactobacillus plantarum Taj-Apis362 cells. The overexpression was confirmed by SDS-PAGE and GAD activity, showing a 53 KDa protein with the enzyme activity increased by sevenfold compared with the original GAD activity. The optimal fermentation conditions for GABA production established using response surface methodology were at glutamic acid concentration of 497.973 mM, temperature 36°C, pH 5.31 and time 60 h. Under the conditions, maximum GABA concentration obtained (11.09 mM) was comparable with the predicted value by the model at 11.23 mM. To our knowledge, this is the first report of successful cloning (clone-back) and overexpression of the LbGAD gene from L. plantarum to L. plantarum cells. The recombinant Lactobacillus could be used as a starter culture for direct incorporation into a food system during fermentation for production of GABA-rich products. PMID:25757029

  17. [Influence of exogenous gamma-aminobutyric acid (GABA) on GABA metabolism and amino acid contents in roots of melon seedling under hypoxia stress].

    PubMed

    Wang, Chun-Yan; Li, Jing-Rui; Xia, Qing-Ping; Wu, Xiao-Lei; Gao, Hong-Bo

    2014-07-01

    This paper investigated the influence of gamma-aminobutyric acid (GABA) on GABA metabolism and amino acid content under hypoxia stress by accurately controlling the level of dissolved oxygen in hydroponics, using the roots of melon 'Xiyu 1' seedlings as the test material. The results showed that compared with the control, the growth of roots was inhibited seriously under hypoxia stress. Meanwhile, the hypoxia-treated roots had significantly higher activities of glutamate decarboxylase (GAD), glutamate dehydrogenase (GDH), glutamate synthase (GOGAT), glutamine synthetase (GS), alanine aminotransferase (ALT), aspartate aminotransferase (AST) as well as the contents of GABA, pyruvic acid, alanine (Ala) and aspartic acid (Asp). But the contents of glutamic acid (Glu) and alpha-keto glutaric acid in roots under hypoxia stress was obviously lower than those of the control. Exogenous treatment with GABA alleviated the inhibition effect of hypoxia stress on root growth, which was accompanied by an increase in the contents of endogenous GABA, Glu, alpha-keto glutaric acid and Asp. Furthermore, under hypoxia stress, the activities of GAD, GDH, GOGAT, GS, ALT, AST as well as the contents of pyruvic acid and Ala significantly decreased in roots treated with GABA. However, adding GABA and viny-gamma-aminobutyric acid (VGB) reduced the alleviation effect of GABA on melon seedlings under hypoxia stress. The results suggested that absorption of GABA by roots could alleviate the injury of hypoxia stress to melon seedlings. This meant that GABA treatment allows the normal physiological metabolism under hypoxia by inhibiting the GAD activity through feedback and maintaining higher Glu content as well as the bal- ance of carbon and nitrogen.

  18. [Influence of exogenous gamma-aminobutyric acid (GABA) on GABA metabolism and amino acid contents in roots of melon seedling under hypoxia stress].

    PubMed

    Wang, Chun-Yan; Li, Jing-Rui; Xia, Qing-Ping; Wu, Xiao-Lei; Gao, Hong-Bo

    2014-07-01

    This paper investigated the influence of gamma-aminobutyric acid (GABA) on GABA metabolism and amino acid content under hypoxia stress by accurately controlling the level of dissolved oxygen in hydroponics, using the roots of melon 'Xiyu 1' seedlings as the test material. The results showed that compared with the control, the growth of roots was inhibited seriously under hypoxia stress. Meanwhile, the hypoxia-treated roots had significantly higher activities of glutamate decarboxylase (GAD), glutamate dehydrogenase (GDH), glutamate synthase (GOGAT), glutamine synthetase (GS), alanine aminotransferase (ALT), aspartate aminotransferase (AST) as well as the contents of GABA, pyruvic acid, alanine (Ala) and aspartic acid (Asp). But the contents of glutamic acid (Glu) and alpha-keto glutaric acid in roots under hypoxia stress was obviously lower than those of the control. Exogenous treatment with GABA alleviated the inhibition effect of hypoxia stress on root growth, which was accompanied by an increase in the contents of endogenous GABA, Glu, alpha-keto glutaric acid and Asp. Furthermore, under hypoxia stress, the activities of GAD, GDH, GOGAT, GS, ALT, AST as well as the contents of pyruvic acid and Ala significantly decreased in roots treated with GABA. However, adding GABA and viny-gamma-aminobutyric acid (VGB) reduced the alleviation effect of GABA on melon seedlings under hypoxia stress. The results suggested that absorption of GABA by roots could alleviate the injury of hypoxia stress to melon seedlings. This meant that GABA treatment allows the normal physiological metabolism under hypoxia by inhibiting the GAD activity through feedback and maintaining higher Glu content as well as the bal- ance of carbon and nitrogen. PMID:25345052

  19. Production of yogurt with enhanced levels of gamma-aminobutyric acid and valuable nutrients using lactic acid bacteria and germinated soybean extract.

    PubMed

    Park, Ki-Bum; Oh, Suk-Heung

    2007-05-01

    Yogurt with high levels of gamma-aminobutyric acid (GABA), free amino acids and isoflavones was developed using lactic acid bacteria (LAB) and germinated soybean extract. Fermented soya milk (GABA soya yogurt) produced with starter and substrate had the GABA concentration of 424.67 microg/gDW, whereas fermented milk produced by a conventional method had GABA less than 1.5 microg/gDW. The GABA soya yogurt also contained significantly high levels of free amino acids and isoflavones compared with other conventional yogurts. The results suggested that the Lactobacillus brevis OPY-1 and germinated soybean possessed a prospect to be applied in dairy and other health products with high nutritive values and functional properties.

  20. Benzodiazepine-insensitive mice generated by targeted disruption of the gamma 2 subunit gene of gamma-aminobutyric acid type A receptors.

    PubMed Central

    Günther, U; Benson, J; Benke, D; Fritschy, J M; Reyes, G; Knoflach, F; Crestani, F; Aguzzi, A; Arigoni, M; Lang, Y

    1995-01-01

    Vigilance, anxiety, epileptic activity, and muscle tone can be modulated by drugs acting at the benzodiazepine (BZ) site of gamma-aminobutyric acid type A (GABAA) receptors. In vivo, BZ sites are potential targets for endogenous ligands regulating the corresponding central nervous system states. To assess the physiological relevance of BZ sites, mice were generated containing GABAA receptors devoid of BZ sites. Following targeted disruption of the gamma 2 subunit gene, 94% of the BZ sites were absent in brain of neonatal mice, while the number of GABA sites was only slightly reduced. Except for the gamma 2 subunit, the level of expression and the regional and cellular distribution of the major GABAA receptor subunits were unaltered. The single channel main conductance level and the Hill coefficient were reduced to values consistent with recombinant GABAA receptors composed of alpha and beta subunits. The GABA response was potentiated by pentobarbital but not by flunitrazepam. Diazepam was inactive behaviorally. Thus, the gamma 2 subunit is dispensable for the assembly of functional GABAA receptors but is required for normal channel conductance and the formation of BZ sites in vivo. BZ sites are not essential for embryonic development, as suggested by the normal body weight and histology of newborn mice. Postnatally, however, the reduced GABAA receptor function is associated with retarded growth, sensorimotor dysfunction, and drastically reduced life-span. The lack of postnatal GABAA receptor regulation by endogenous ligands of BZ sites might contribute to this phenotype. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 PMID:7644489

  1. Picrotoxin inhibition mechanism of a gamma-aminobutyric acid A receptor investigated by a laser-pulse photolysis technique.

    PubMed

    Ramakrishnan, Latha; Hess, George P

    2005-06-14

    The gamma-aminobutyric acid(A) (GABA(A)) receptor, a major inhibitory neurotransmitter receptor, belongs to a family of membrane-bound proteins that regulate signal transmission between approximately 10(12) cells of the nervous system. It plays a major role in many neurological disorders, including epilepsy. It is the target of many pharmacological agents, including the convulsant picrotoxin. Here, we present the mechanism of inhibition by picrotoxin of the rat alpha1beta2gamma2L GABA(A) receptor investigated using rapid kinetic techniques in combination with whole-cell current recordings. The following new results were obtained by using transient kinetic techniques, the cell-flow method and the laser-pulse photolysis (LaPP) technique with a microsecond to millisecond time resolution. (i) The apparent dissociation constant of picrotoxin for the open-channel form of the receptor was approximately 5 times higher than that of the closed-channel form. (ii) Picrotoxin increased the channel-closing rate constant (k(cl)) approximately 4-fold, while the rate constant for channel opening (k(op)) remained essentially unaffected. (iii) The mechanism indicates that picrotoxin binds to an allosteric site of the receptor with higher affinity for the closed-channel form than for the open-channel form and thereby inhibits the receptor by decreasing 4-fold its channel-opening equilibrium constant [Phi(I)(-)(1) = k(op(I))/k(cl(I))]. (iv) The mechanism further indicates that compounds that bind with equal affinity to the picrotoxin-binding site on the open-channel form of the receptor and the closed-channel form will not affect the channel-opening equilibrium and can, therefore, displace picrotoxin and prevent inhibition of the GABA(A) receptor by picrotoxin. Such compounds may be therapeutically useful in counteracting the effects of compounds and diseases that unfavorably affect the channel-opening equilibrium of the receptor channel.

  2. Effects of gamma-aminobutyric acid on skate retinal horizontal cells: evidence for an electrogenic uptake mechanism.

    PubMed Central

    Malchow, R P; Ripps, H

    1990-01-01

    In the retinae of many vertebrates, there are classes of horizontal cell that probably utilize gamma-aminobutyric acid (GABA) as a neurotransmitter. As with other amino acid transmitter agents, the postsynaptic action of GABA is thought to be terminated by uptake into neurons and glia surrounding the release site. The present study examined whether an uptake system for GABA could be detected in isolated skate horizontal cells by means of electrophysiological methods. Pressure ejection of GABA onto voltage-clamped horizontal cells produced an inward current that showed no sign of desensitization regardless of the GABA concentration. The dose-response relationship followed simple Michaelis-Menten kinetics, with a half-maximal response elicited at approximately 110 microM. Nipecotic acid produced a similar current and reduced the responses to GABA when introduced in the bath solution prior to the GABA pulse. On the other hand, application of 500 microM muscimol or 1 mM baclofen, GABAA and GABAB receptor agonists, respectively, were completely without effect. The GABA-induced current was not blocked by superfusion with 500 microM bicuculline, 500 microM picrotoxin, or 500 microM phaclofen. However, the responses to GABA were abolished when the cells were superfused in Ringer's solution in which choline or lithium had been substituted for sodium, and were reduced when the extracellular chloride concentration was decreased from 266 mM to 16 mM. Current-voltage data showed a maximal response to GABA when the cells were held at or below their resting potential. At more depolarized levels, the inward current became progressively smaller until, near +50 mV, it could no longer be detected; over the range tested (-90 to +50 mV), the response never reversed into an outward current. These findings suggest that the GABA-induced currents in skate horizontal cells are mediated by an electrogenic uptake mechanism. PMID:2247470

  3. Gamma-aminobutyric acid and related molecules in the sea fan Eunicella cavolini (Cnidaria: Octocorallia): a biochemical and immunohistochemical approach.

    PubMed

    Girosi, Laura; Ferrando, Sara; Beltrame, Francesco; Ciarcia, Gaetano; Diaspro, Alberto; Fato, Marco; Magnone, Mirko; Raiteri, Luca; Ramoino, Paola; Tagliafierro, Grazia

    2007-07-01

    The aim of this study has been the biochemical demonstration of the presence of gamma-aminobutyric acid (GABA) in the Mediterranean sea fan Eunicella cavolini by means of high-performance liquid chromatography, and the description of the distribution pattern of GABA and its related molecules, glutamic acid decarboxylase (GAD), vesicular GABA transporter (VGAT) and one of the GABA receptors (GABA(B) R) by immunohistochemical methods. The interrelationships of GABA, GAD and GABA receptor immunoreactivity have been established by using double-immunohistochemical methods and confocal microscopy. The immunodetection of monoclonal and/or polyclonal antibodies has revealed GABA immunoreactivity throughout the polyp tissue, both in neuronal and non-neuronal elements. GAD immunoreactivity has been mostly localized in the neuronal compartment, contacting epithelial and muscular elements. GABA(B) R immunoreactivity appears particularly intense in the nematocytes and in the oocyte envelope; its presence in GAD-immunoreactive neurons in the tentacles suggests an autocrine type of regulation. Western blot analysis has confirmed that a GABA(B) R, with a molecular weight of 142 kDa, similar to that of rat brain, is present in E. cavolini polyp tissue. The identification of the sites of the synthesis, vesicular transport, storage and reception of GABA strongly suggests the presence of an almost complete set of GABA-related molecules for the functioning of the GABAergic system in this simple nervous system. The distribution of these different immunoreactivities has allowed us to hypothesize GABA involvement in nematocyst discharge, in body wall and enteric muscular contraction, in neuronal integration and in male gametocyte differentiation.

  4. Uptake and release of [3H]gamma-aminobutyric acid by embryonic spinal cord neurons in dissociated cell culture

    PubMed Central

    1979-01-01

    We have investigated the uptake and release of [3H]gamma-aminobutyric acid (GABA) by embryonic chick spinal cord cells maintained in culture. Cells dissociated from 4- or 7-d-old embryos were studied between 1 and 3 wk after plating. At 3 degrees C, [3H]GABA was accumulated by a high affinity (Km approximately equal to 4 microM) and a low affinity (Km approximately equal to 100 microM) mechanism. The high affinity transport was markedly inhibited in low Na+ media, by ouabain, at 0 degrees C, and by 2,4-diaminobutyric acid. Autoradiography, after incubation in 0.1 microM [3H]GABA, showed that approximately 50% (range = 30-70%) of the multipolar cells were labeled. These cells were neurons rather than glia; action potentials and/or synaptic potentials were recorded in cells subsequently found to be labeled. Non-neuronal, fibroblast-like cells and co-cultured myotubes were not labeled under the same conditions. The fact that not all of the neurons were labeled is consistent with the suggestion, based on studies of intact adult tissue, that high affinity transport of [3H]GABA may be unique to neurons that use GABA as a neurotransmitter. Our finding that none of fifteen physiologically identified cholinergic neurons, i.e., cells that innervated nearby myotubes, were heavily labeled after incubation in 0.1 microM [3H]GABA is significant in this regard. The newly taken up [3H]GABA was not metabolized in the short run. It was stored in a form that could be released when the neurons were depolarized in a high K+ (100 mM) medium. As expected for a neurotransmitter, the K+-evoked release was reversibly inhibited by reducing the extracellular Ca++/Mg++ ratio. PMID:457763

  5. Effect of paraoxon on muscarinic, dopamine and. gamma. -aminobutyric acid receptors of brain and sensitivity to muscarinic antagonists

    SciTech Connect

    Fernando, J.C.R.; Hoskins, B.; Ho, I.K.

    1986-03-05

    Several acetylcholinesterase (AChE) inhibitors decrease muscarinic cholinergic (mACh) receptors in the brain, alteration of dopamine (DA) and ..gamma..-aminobutyric acid (GABA) receptors after AChE inhibition was also reported. In view of the important interactions among DA, GABA and ACh systems, whether this is a common effect of AChE inhibitors should be established. They report the effect of the AChE inhibitor, paraoxon, on DA, GABA and mACh receptors in the rat. The binding of /sup 3/H-QNB (for mACh), /sup 3/H-spiperone (for DA) and /sup 3/H-muscimol (for GABA) to striatal and hippocampal membranes was analyzed. Also, behavioral sensitivity to atropine was studied. Twenty-four hr after a single dose (0.75 mg/kg, s.c.) of paraoxon, the density of mACh receptors in the striatum was decreased but, at 3 days, no change was seen. In the hippocampus, the mACh receptors were not affected. Repeated treatment with paraoxon (0.3 mg/kg, 48 hourly) for 2 weeks reduced the mACh receptor density in both regions. Neither single nor repeated paraoxon treatment had an effect on DA or GABA receptors. After single or repeated dosing with paraoxon, myoclonus induced by atropine (10 mg/kg, i.p.) was enhanced. The results show rapid downregulation of mACh receptors by paraoxon. DA or GABA, however, appear not to be affected under these treatment regimens.

  6. Determination of theanine and gamma-aminobutyric acid in tea by high performance- liquid chromatography with precolumn derivatization.

    PubMed

    Tu, Yunfei; Yang, Xiufang; Zhang, Shikang; Zhu, Yuejin

    2012-02-01

    A method of precolumn derivatization-high performance liquid chromatography (HPLC) for the determination of theanine and gamma-aminobutyric acid (GABA) in tea was established. o-Phthalaldehyde (OPA) and N-acetyl-L-cysteine (NAC) were chosen as the derivatization reagents. The effects of teapolyphenol (Tp), proline (Pro) and Vitamin C (Vc) on derivatization yields were investigated. The results indicated that Vc not only stabilized the stock solution of OPA, but also enhanced the yield of GABA derivative. However, the yield of theanine derivative was less affected. The HPLC separation system was also optimized. The resolution of the derivatives was improved by adjusting the pH value and phosphate-citric buffer concentration of the mobile phase. The limits of detection (LODs) for GABA and theanine were 3.01 x 10(-5) mmol/L and 7.98 x 10(-5) mmol/L, and the limits of quantification (LOQs) were 9.99 x 10(-5) mmol/L and 2.658 x 10(-4) mmol/L, respectively. The linear ranges of GABA and theanine were 0.01 - 0.4 mmol/L with the correlation coefficient of 0.996 and 0.05 - 0.8 mmol/L with the correlation coefficient of 0.995, respectively. The main recoveries for GABA and theanine in green tea, Oolong tea, and black tea, ranged from 99.29% to 119.60% and from 62.88% to 141.06% respectively. The method with simple procedure and efficient separation was proved to be suitable for the determination of GABA and theanine in tea. PMID:22679834

  7. Concentrative export from the endoplasmic reticulum of the gamma-aminobutyric acid transporter 1 requires binding to SEC24D.

    PubMed

    Farhan, Hesso; Reiterer, Veronika; Korkhov, Vladimir M; Schmid, Johannes A; Freissmuth, Michael; Sitte, Harald H

    2007-03-01

    Re-uptake of gamma-aminobutyric acid (GABA) into presynaptic specializations is mediated by the GABA transporter 1 (GAT1), a member of the SLC6 gene family. Here, we show that a motif in the COOH terminus of GAT1 ((566)RL(567)), which is conserved in SLC6 family members, is a binding site for the COPII coat component Sec24D. We also identified residues in Sec24D ((733)DD(734)) that are required to support the interaction with GAT1 and two additional family members, i.e. the transporters for serotonin and dopamine. We used three strategies to prevent recruitment of Sec24D to GAT1: knock-down of Sec24D by RNA interference, overexpression of Sec24D-VN (replacement of (733)DD(734) by (733)VN(734)), and mutation of (566)RL(567) to (566)AS(567) (GAT1-RL/AS). In each instance, endoplasmic reticulum (ER) export of GAT1 was impaired: in the absence of Sec24D or upon coexpression of dominant negative Sec24D-VN, GAT1 failed to undergo concentrative ER export; GAT1-RL/AS also accumulated in the ER and exerted a dominant negative effect on cell surface targeting of wild type GAT1. Our observations show that concentrative ER-export is contingent on a direct interaction of GAT1 with Sec24D; this also provides a mechanistic explanation for the finding that oligomeric assembly of transporters is required for their ER export: transporter oligomerization supports efficient recruitment of COPII components.

  8. Gamma-aminobutyric acid loaded halloysite nanotubes and in vitro-in vivo evaluation for brain delivery.

    PubMed

    Kırımlıoğlu, Gülsel Yurtdaş; Yazan, Yasemin; Erol, Kevser; Çengelli Ünel, Çiğdem

    2015-11-30

    Gamma-aminobutyric acid (GABA) is a key neurotransmitter where it usually inhibits impulse transmission. GABA release blockage or postsynaptic reaction were determined to provoke epileptic convulsions. The aim of the present study was the development of brain-targeted, nanosized, nontoxic, biocompatible, highly specific formulations. Incorporation of GABA into halloysite nanotubes (HNT) was performed using different methods. Particle size, zeta potential and pH measurements, morphological, thermal, XRD, FTIR analyses and GABA quantification by validated HPLC method were used for the characterization of the systems prepared. Release pattern of GABA from the nanotubes was determined using a dialysis membrane. Following successful incorporation of GABA into HNTs for brain delivery, nanotube formulation coded HNT-GABA H1 was selected for in vivo studies. Smaller particle size with narrow size distribution, possible HNT-GABA interaction indicated by thermal, XRD and FTIR analyses and prolonged release were the parameters considered in this selection. Moreover, HNT-GABA H1 remained stable for 3-month storage period and showed higher cell viability values than GABA. Rats were used in in vivo studies and potential of anticonvulsant effect of GABA was determined in the pentylenetetrazole model of seizure. HNT-GABA H1 was found to increase latency of seizure, decrease ending time of the convulsion, duration of severe convulsion and mortality rate significantly compared to pure GABA. After administration of HNT-GABA H1, GABA concentration in Stratum corsatum measured by enzyme immune assay showed that it was not significantly higher than GABA administered alone. These findings suggest that GABA loaded HNTs reduces the duration of all phases of convulsion indicating efficient delivery of GABA to all brain areas to interfere with epileptic mechanism.

  9. Chronic noise stress-induced alterations of glutamate and gamma-aminobutyric acid and their metabolism in the rat brain.

    PubMed

    Kazi, Amajad Iqbal; Oommen, Anna

    2014-01-01

    Chronic stress induces neurochemical changes that include neurotransmitter imbalance in the brain. Noise is an environmental factor inducing stress. Chronic noise stress affects monoamine neurotransmitter systems in the central nervous system. The effect on other excitatory and inhibitory neurotransmitter systems is not known. The aim was to study the role of chronic noise stress on the glutamatergic and gamma-aminobutyric acid (GABA)ergic systems of the brain. Female Wistar rats (155 ± 5 g) were unintentionally exposed to noise due to construction (75-95 db, 3-4 hours/day, 5 days a week for 7-8 weeks) in the vicinity of the animal care facility. Glutamate/GABA levels and their metabolic enzymes were evaluated in different rat brain regions (cortex, hippocampus, striatum, and cerebellum) and compared with age and gender matched nonexposed rats. Chronic noise stress decreased glutamate levels and glutaminase activity 27% and 33% in the cortex, 15% and 24% in the cerebellum. Glutamate levels increased 10% in the hippocampus, 28% in striatum and glutaminase activity 15% in striatum. Glutamine synthetase activity increased significantly in all brain regions studied, that is, cortex, hippocampus, striatum, and cerebellum (P < 0.05). Noise stress-increased GABA levels and glutamate alpha decarboxylase activity 20% and 45% in the cortex, 13% and 28% in the hippocampus respectively. GABA levels and glutamate alpha decarboxylase activity decreased 15% and 14%, respectively in the striatum. GABA transaminase activity was significantly reduced in the cortex (55%), hippocampus (17%), and cerebellum (33%). Chronic noise stress differentially affected glutamatergic and GABAergic neurotransmitter systems in the rat brain, which may alter glutamate and GABA neurotransmission.

  10. Corticotropin Releasing Factor–Induced Amygdala Gamma-Aminobutyric Acid Release Plays a Key Role in Alcohol Dependence

    PubMed Central

    Roberto, Marisa; Cruz, Maureen T.; Gilpin, Nicholas W.; Sabino, Valentina; Schweitzer, Paul; Bajo, Michal; Cottone, Pietro; Madamba, Samuel G.; Stouffer, David G.; Zorrilla, Eric P.; Koob, George F.; Siggins, George R.; Parsons, Loren H.

    2010-01-01

    Background Corticotropin-releasing factor (CRF) and gamma-aminobutyric acid (GABA)ergic systems in the central amygdala (CeA) are implicated in the high-anxiety, high-drinking profile associated with ethanol dependence. Ethanol augments CeA GABA release in ethanol-naive rats and mice. Methods Using naive and ethanol-dependent rats, we compared electrophysiologic effects and interactions of CRF and ethanol on CeA GABAergic transmission, and we measured GABA dialyzate in CeA after injection of CRF1 antagonists and ethanol. We also compared mRNA expression in CeA for CRF and CRF1 using real-time polymerase chain reaction. We assessed effects of chronic treatment with a CRF1 antagonist on withdrawal-induced increases in alcohol consumption in dependent rats. Results CRF and ethanol augmented CeA GABAergic transmission in naive rats via increased GABA release. Three CRF1 receptor (CRF1) antagonists decreased basal GABAergic responses and abolished ethanol effects. Ethanol-dependent rats exhibited heightened sensitivity to CRF and CRF1 antagonists on CeA GABA release. Intra-CeA CRF1 antagonist administration reversed dependence–related elevations in GABA dialysate and blocked ethanol-induced increases in GABA dialyzate in both dependent and naive rats. Polymerase chain reaction studies indicate increased expression of CRF and CRF1 in CeA of dependent rats. Chronic CRF1 antagonist treatment blocked withdrawal-induced increases in alcohol drinking by dependent rats and tempered moderate increases in alcohol consumption by nondependent rats in intermittent testing. Conclusions These combined findings suggest a key role for specific presynaptic CRF-GABA interactions in CeA in the development and maintenance of ethanol dependence. PMID:20060104

  11. Importance of the gamma-aminobutyric acid(B) receptor C-termini for G-protein coupling.

    PubMed

    Grünewald, Sylvia; Schupp, Bettina J; Ikeda, Stephen R; Kuner, Rohini; Steigerwald, Frank; Kornau, Hans-Christian; Köhr, Georg

    2002-05-01

    Functional gamma-aminobutyric acid(B) (GABA(B)) receptors assemble from two subunits, GABA(B(1)) and GABA(B(2).) This heteromerization, which involves a C-terminal coiled-coil interaction, ensures efficient surface trafficking and agonist-dependent G-protein activation. In the present study, we took a closer look at the implications of the intracellular C termini of GABA(B(1)) and GABA(B(2)) for G-protein coupling. We generated a series of C-terminal mutants of GABA(B(1)) and GABA(B(2)) and tested them for physical interaction, surface trafficking, coupling to adenylyl cyclase, and G-protein-gated inwardly rectifying potassium channels in human embryonic kidney (HEK) 293 cells as well as on endogenous calcium channels in sympathetic neurons of the superior cervical ganglion (SCG). We found that the C-terminal interaction contributes only partly to the heterodimeric assembly of the subunits, indicating the presence of an additional interaction site. The described endoplasmic reticulum retention signal within the C terminus of GABA(B(1)) functioned only in the context of specific amino acids, which constitute part of the GABA(B(1)) coiled-coil sequence. This finding may provide a link between the retention signal and its shielding by the coiled coil of GABA(B(2).) In HEK293 cells, we observed that the two well-known GABA(B) receptor antagonists [S-(R*,R*)]-[3-[[1-(3,4-dichlorophenyl)ethyl]amino]-2-hydroxypropyl](cyclohexylmethyl) phosphinic acid (CGP54626) and (+)-(2S)-5,5-dimethyl-2-morpholineacetic acid (SCH50911) CGP54626 and SCH50911 function as inverse agonists. The C termini of GABA(B(1)) and GABA(B(2)) strongly influenced agonist-independent G-protein coupling, although they were not necessary for agonist-dependent G-protein coupling. The C-terminal GABA(B) receptor mutants described here demonstrate that the active receptor conformation is stabilized by the coiled-coil interaction. Thus, the C-terminal conformation of the GABA(B) receptor may determine its

  12. Binding interaction of a gamma-aminobutyric acid derivative with serum albumin: an insight by fluorescence and molecular modeling analysis.

    PubMed

    Pal, Uttam; Pramanik, Sumit Kumar; Bhattacharya, Baisali; Banerji, Biswadip; C Maiti, Nakul

    2016-01-01

    gamma-Aminobutyric acid (GABA) is a naturally occurring inhibitory neurotransmitter and some of its derivatives showed potential to act as neuroprotective agents. With the aim of developing potential leads for anti-Alzheimer's drugs, in this study we synthesized a novel GABA derivative, methyl 4-(4-((2-(tert-butoxy)-2-oxoethyl)(4-methoxyphenyl)amino)benzamido)butanoate by a unique method of Buchwald-Hartwig cross coupling synthesis; with some modification the yield was significant (97 %) and spectroscopic analysis confirmed that the compound was highly pure (98.8 % by HPLC). The druglikeness properties such as logP, logS, and polar surface area were 3.87, -4.86 and 94.17 Å(2) respectively and it satisfied the Lipinski's rule of five. We examined the binding behavior of the molecule to human serum albumin (HSA) and bovine serum albumin (BSA) which are known as universal drug carrier proteins. The molecule binds to the proteins with low micromolar efficiency and the calculated binding constants were 3.85 and 2.75 micromolar for BSA and HSA, respectively. Temperature dependent study using van't Hoff equation established that the binding was thermodynamically favorable and the changes in the Gibb's free energy, ΔG for the binding process was negative. However, the binding of the molecule to HSA was enthalpy driven and the change of enthalpy (ΔH) was -10.63 kJ/mol, whereas, the binding to BSA was entropy driven and the change in entropy ΔS was 222 J/mol. The molecular docking analysis showed that the binding sites of the molecule lie in the groove between domain I and domain III of BSA, whereas it is within the domain I in case of HSA, which also supported the different thermodynamic nature of binding with HSA and BSA. Molecular dynamics analysis suggested that the binding was stable with time and provided further details of the binding interaction. Molecular dynamics study also highlighted the effect of this ligand binding on the serum albumin structure. PMID

  13. Effects of negative allosteric modulators of gamma-aminobutyric acidA receptors on complex behavioral processes in monkeys.

    PubMed

    Auta, J; Winsauer, P J; Faust, W B; Lambert, P; Moerschbaecher, J M

    1997-01-01

    A multiple schedule of repeated acquisition and performance of conditional discriminations was used to characterize the effects of two negative allosteric modulators of the gamma-aminobutyric acid (GABAA) receptor (ethyl beta-carboline-3-carboxylate [beta-CCE] and N-methyl-beta-carboline-3-carboxamide [FG-7142]), a hallucinogenic beta-carboline derivative (harmine), a benzodiazepine receptor antagonist (flumazenil) and a positive allosteric modulator (alprazolam). In the acquisition component, subjects acquired a different discrimination each session. Acquisition of a discrimination was defined by a decrease in errors as the session progressed. In the performance component, the discrimination was the same each session. Responding in both components was maintained by food presentation under a variable-ratio schedule. Incorrect responses in both components produced a 5-sec timeout. Alprazolam (0.1-18 mg/kg), beta-CCE (0.01-0.32 mg/kg), FG-7142 (0.1-18 mg/kg) and harmine (0.1-1.8 mg/kg) all dose-dependently decreased response rate in both components. However, accuracy of responding-was differentially affected by the drugs. Alprazolam selectively and dose-dependently increased percent errors in acquisition, whereas beta-CCE increased acquisition errors only at the highest doses tested in each subject. In contrast, FG-7142 and harmine had no effects on percent errors at doses that virtually eliminated responding. In all cases, performance accuracy was generally not affected. Flumazenil, at doses that had little or no effect (0.1 and 0.32 mg/kg) or occasionally decreased response rates (1 mg/kg) when administered alone, dose-dependently antagonized the rate-decreasing and error-increasing effects of beta-CCE, FG-7142 and alprazolam. In contrast, flumazenil failed to antagonize the effects of harmine. Thus, the negative allosteric modulators only moderately disrupted acquisition in comparison with the positive allosteric modulator, but the effects of both types of

  14. Influence of cold stress on contents of soluble sugars, vitamin C and free amino acids including gamma-aminobutyric acid (GABA) in spinach (Spinacia oleracea).

    PubMed

    Yoon, Young-Eun; Kuppusamy, Saranya; Cho, Kye Man; Kim, Pil Joo; Kwack, Yong-Bum; Lee, Yong Bok

    2017-01-15

    The contents of soluble sugars (sucrose, fructose, glucose, maltose and raffinose), vitamin C and free amino acids (34 compounds, essential and non-essential) were quantified in open-field and greenhouse-grown spinaches in response to cold stress using liquid chromatography. In general, greenhouse cultivation produced nutritionally high value spinach in a shorter growing period, where the soluble sugars, vitamin C and total amino acids concentrations, including essential were in larger amounts compared to those grown in open-field scenarios. Further, low temperature exposure of spinach during a shorter growth period resulted in the production of spinach with high sucrose, ascorbate, proline, gamma-aminobutyric acid, valine and leucine content, and these constitute the most important energy/nutrient sources. In conclusion, cultivation of spinach in greenhouse at a low temperature (4-7°C) and exposure for a shorter period (7-21days) before harvest is recommended. This strategy will produce a high quality product that people can eat. PMID:27542466

  15. Influence of cold stress on contents of soluble sugars, vitamin C and free amino acids including gamma-aminobutyric acid (GABA) in spinach (Spinacia oleracea).

    PubMed

    Yoon, Young-Eun; Kuppusamy, Saranya; Cho, Kye Man; Kim, Pil Joo; Kwack, Yong-Bum; Lee, Yong Bok

    2017-01-15

    The contents of soluble sugars (sucrose, fructose, glucose, maltose and raffinose), vitamin C and free amino acids (34 compounds, essential and non-essential) were quantified in open-field and greenhouse-grown spinaches in response to cold stress using liquid chromatography. In general, greenhouse cultivation produced nutritionally high value spinach in a shorter growing period, where the soluble sugars, vitamin C and total amino acids concentrations, including essential were in larger amounts compared to those grown in open-field scenarios. Further, low temperature exposure of spinach during a shorter growth period resulted in the production of spinach with high sucrose, ascorbate, proline, gamma-aminobutyric acid, valine and leucine content, and these constitute the most important energy/nutrient sources. In conclusion, cultivation of spinach in greenhouse at a low temperature (4-7°C) and exposure for a shorter period (7-21days) before harvest is recommended. This strategy will produce a high quality product that people can eat.

  16. The central nervous system convulsant pentylenetetrazole stimulates gamma-aminobutyric acid (GABA)-activated current in picrotoxin-resistant GABA(A) receptors in HEK293 cells.

    PubMed

    Dibas, M I; Dillon, G H

    2000-05-19

    We tested the ability of the central nervous system convulsant pentylenetetrazole (PTZ) to inhibit gamma-aminobutyric acid (GABA)-gated current in receptors expressing a mutation that rendered them resistant to picrotoxin. Consistent with previous reports, receptors expressing beta2(T246F), along with alpha3 and gamma2 subunits, resulted in a greatly diminished sensitivity to picrotoxin. Sensitivity to PTZ was completely abolished in the mutant receptor, confirming the hypothesis that PTZ acts at the picrotoxin site. Quite unexpected, however, was our finding that PTZ elicited marked stimulation (up to 400% of control) in the mutated receptors. This stimulatory effect was not mediated via an interaction with the benzodiazepine site, as preincubation with the benzodiazepine antagonist flumazenil did not block the PTZ-induced stimulation. Our results reveal the existence of a novel stimulatory domain of PTZ in GABA(A) receptors.

  17. Imidase catalyzing desymmetric imide hydrolysis forming optically active 3-substituted glutaric acid monoamides for the synthesis of gamma-aminobutyric acid (GABA) analogs.

    PubMed

    Nojiri, Masutoshi; Hibi, Makoto; Shizawa, Hiroaki; Horinouchi, Nobuyuki; Yasohara, Yoshihiko; Takahashi, Satomi; Ogawa, Jun

    2015-12-01

    The recent use of optically active 3-substituted gamma-aminobutyric acid (GABA) analogs in human therapeutics has identified a need for an efficient, stereoselective method of their synthesis. Here, bacterial strains were screened for enzymes capable of stereospecific hydrolysis of 3-substituted glutarimides to generate (R)-3-substituted glutaric acid monoamides. The bacteria Alcaligenes faecalis NBRC13111 and Burkholderia phytofirmans DSM17436 were discovered to hydrolyze 3-(4-chlorophenyl) glutarimide (CGI) to (R)-3-(4-chlorophenyl) glutaric acid monoamide (CGM) with 98.1% enantiomeric excess (e.e.) and 97.5% e.e., respectively. B. phytofirmans DSM17436 could also hydrolyze 3-isobutyl glutarimide (IBI) to produce (R)-3-isobutyl glutaric acid monoamide (IBM) with 94.9% e.e. BpIH, an imidase, was purified from B. phytofirmans DSM17436 and found to generate (R)-CGM from CGI with specific activity of 0.95 U/mg. The amino acid sequence of BpIH had a 75% sequence identity to that of allantoinase from A. faecalis NBRC13111 (AfIH). The purified recombinant BpIH and AfIH catalyzed (R)-selective hydrolysis of CGI and IBI. In addition, a preliminary investigation of the enzymatic properties of BpIH and AfIH revealed that both enzymes were stable in the range of pH 6-10, with an optimal pH of 9.0, stable at temperatures below 40 °C, and were not metalloproteins. These results indicate that the use of this class of hydrolase to generate optically active 3-substituted glutaric acid monoamide could simplify the production of specific chiral GABA analogs for drug therapeutics.

  18. Buffer-free production of gamma-aminobutyric acid using an engineered glutamate decarboxylase from Escherichia coli.

    PubMed

    Kang, Taek Jin; Ho, Ngoc Anh Thu; Pack, Seung Pil

    2013-08-15

    Escherichia coli glutamate decarboxylase (GAD) converts glutamate into γ-aminobutyric acid (GABA) through decarboxylation using proton as a co-substrate. Since GAD is active only at acidic conditions even though pH increases as the reaction proceeds, the conventional practice of using this enzyme involved the use of relatively high concentration of buffers, which might complicate the downstream purification steps. Here we show by simulation and experiments that the free acid substrate, glutamic acid, rather than its monosodium salt can act as a substrate and buffer at the same time. This yielded the buffer- and salt-free synthesis of GABA conveniently in a batch mode. Furthermore, we engineered GAD to hyper active ones by extending or reducing the length of the enzyme by just one residue at its C-terminus. Through the buffer-free reaction with engineered GAD, we could synthesize 1M GABA in 3h, which can be translated into a space-time yield of 34.3g/L/h.

  19. gamma-Aminobutyric acidA (GABAA) receptors modulate [3H]GABA release from isolated neuronal growth cones in the rat.

    PubMed

    Lockerbie, R O; Gordon-Weeks, P R

    1985-04-19

    Potassium-induced release of gamma-[3H]aminobutyric acid [( 3H]GABA) from a growth cone-enriched fraction isolated from neonatal rat forebrain was inhibited by the GABA mimetic muscimol in a dose-dependent manner (IC50 15 nM). The GABA antagonist bicuculline completely reversed the effect of muscimol. Bicuculline alone slightly potentiated the K+-induced release of [3H]GABA. Baclofen, a proposed selective agonist for a bicuculline-insensitive GABAB receptor, was found to cause only a slight reduction in the K+-induced release of [3H]GABA. These results are compatible with the presence of a negative feedback mechanism mediated by GABAA receptors for controlling [3H]GABA release from growth cones of the developing rat forebrain.

  20. Efficient production of gamma-aminobutyric acid using Escherichia coli by co-localization of glutamate synthase, glutamate decarboxylase, and GABA transporter.

    PubMed

    Dung Pham, Van; Somasundaram, Sivachandiran; Lee, Seung Hwan; Park, Si Jae; Hong, Soon Ho

    2016-01-01

    Gamma-aminobutyric acid (GABA) is an important bio-product, which is used in pharmaceutical formulations, nutritional supplements, and biopolymer monomer. The traditional GABA process involves the decarboxylation of glutamate. However, the direct production of GABA from glucose is a more efficient process. To construct the recombinant strains of Escherichia coli, a novel synthetic scaffold was introduced. By carrying out the co-localization of glutamate synthase, glutamate decarboxylase, and GABA transporter, we redirected the TCA cycle flux to GABA pathway. The genetically engineered E. coli strain produced 1.08 g/L of GABA from 10 g/L of initial glucose. Thus, with the introduction of a synthetic scaffold, we increased GABA production by 2.2-fold. The final GABA concentration was increased by 21.8% by inactivating competing pathways.

  1. Production of gamma-aminobutyric acid from glucose by introduction of synthetic scaffolds between isocitrate dehydrogenase, glutamate synthase and glutamate decarboxylase in recombinant Escherichia coli.

    PubMed

    Pham, Van Dung; Lee, Seung Hwan; Park, Si Jae; Hong, Soon Ho

    2015-08-10

    Escherichia coli were engineered for the direct production of gamma-aminobutyric acid from glucose by introduction of synthetic protein scaffold. In this study, three enzymes consisting GABA pathway (isocitrate dehydrogenase, glutamate synthase and glutamate decarboxylase) were connected via synthetic protein scaffold. By introduction of scaffold, 0.92g/L of GABA was produced from 10g/L of glucose while no GABA was produced in wild type E. coli. The optimum pH and temperature for GABA production were 4.5 and 30°C, respectively. When competing metabolic network was inactivated by knockout mutation, maximum GABA concentration of 1.3g/L was obtained from 10g/L glucose. The recombinant E. coli strain which produces GABA directly from glucose was successfully constructed by introduction of protein scaffold.

  2. Redirection of Metabolic Flux into Novel Gamma-Aminobutyric Acid Production Pathway by Introduction of Synthetic Scaffolds Strategy in Escherichia Coli.

    PubMed

    Pham, Van Dung; Somasundaram, Sivachandiran; Lee, Seung Hwan; Park, Si Jae; Hong, Soon Ho

    2016-04-01

    In general, gamma-aminobutyric acid (GABA) pathway involves the decarboxylation of glutamate, which is produced from sugar by Corynebacterium fermentation. GABA can be used for the production of pharmaceuticals and functional foods. Due to the increasing demand of GABA, it is essential to create an effective alternative pathway for the GABA production. In this study, Escherichia coli were engineered to produce GABA from glucose via GABA shunt, which consists of succinate dehydrogenase, succinate-semialdehyde dehydrogenase, and GABA aminotransferase. The three enzymes were physically attached to each other through a synthetic scaffold, and the Krebs cycle flux was redirected to the GABA pathway. By introduction of synthetic scaffold, 0.75 g/l of GABA was produced from 10 g/l of glucose at 30 °C and pH 6.5. The inactivation of competing metabolic pathways provided 15.4 % increase in the final GABA concentration. PMID:26667817

  3. Analogues of gamma-aminobutyric acid (GABA) and trans-4-aminocrotonic acid (TACA) substituted in the 2 position as GABAC receptor antagonists.

    PubMed

    Chebib, M; Vandenberg, R J; Johnston, G A

    1997-12-01

    1. gamma-Aminobutyric acid (GABA) and trans-4-aminocrotonic acid (TACA) have been shown to activate GABAC receptors. In this study, a range of C2, C3, C4 and N-substituted GABA and TACA analogues were examined for activity at GABAC receptors. 2. The effects of these compounds were examined by use of electrophysiological recording from Xenopus oocytes expressing the human rho 1 subunit of GABAC receptors with the two-electrode voltage-clamp method. 3. trans-4-Amino-2-fluorobut-2-enoic acid was found to be a potent agonist (KD = 2.43 microM). In contrast, trans-4-amino-2-methylbut-2-enoic acid was found to be a moderately potent antagonist (IC50 = 31.0 microM and KB = 45.5 microM). These observations highlight the possibility that subtle structural substitutions may change an agonist into an antagonist. 4. 4-Amino-2-methylbutanoic acid (KD = 189 microM), 4-amino-2-methylenebutanoic acid (KD = 182 microM) and 4-amino-2-chlorobutanoic acid (KD = 285 microM) were weak partial agonists. The intrinsic activities of these compounds were 12.1%, 4.4% and 5.2% of the maximal response of GABA, respectively. These compounds more effectively blocked the effects of the agonist, GABA, giving rise to KB values of 53 microM and 101 microM, respectively. 5. The sulphinic acid analogue of GABA, homohypotaurine, was found to be a potent partial agonist (KD = 4.59 microM, intrinsic activity 69%). 6. It was concluded that substitution of a methyl or a halo group in the C2 position of GABA or TACA is tolerated at GABAC receptors. However, there was dramatic loss of activity when these groups were substituted at the C3, C4 and nitrogen positions of GABA and TACA. 7. Molecular modelling studies on a range of active and inactive compounds indicated that the agonist/competitive antagonist binding site of the GABAC receptor may be smaller than that of the GABAA and GABAB receptors. It is suggested that only compounds that can attain relatively flat conformations may bind to the GABAC receptor

  4. Uptake of gamma-aminobutyric acid and L-glutamic acid by synaptosomes from postmortem human cerebral cortex: multiple sites, sodium dependence and effect of tissue preparation.

    PubMed

    Dodd, P R; Watson, W E; Morrison, M M; Johnston, G A; Bird, E D; Cowburn, R F; Hardy, J A

    1989-06-26

    The uptake of gamma-aminobutyric acid (GABA) and L-glutamic acid by synaptosomes prepared from frozen postmortem human brain was shown to be effected via distinct high and low affinity sites. At approximately 17 h postmortem delay, the kinetic parameters for GABA uptake were: high affinity site, Km 7.1 +/- 2.5 microM, Vmax 18.7 +/- 4.8 nmol.min-1 per 100 mg protein; low affinity site, Km 2 +/- 1 mM, Vmax 425 +/- 250 nmol.min-1 per 100 mg protein (means +/- S.E.M., n = 13). Kinetic parameters for L-glutamate uptake were: high affinity site, Km 7.5 +/- 1.0 microM, Vmax 85 +/- 8 nmol.min-1 per 100 mg protein; low affinity site, Km 1.8 +/- 1.2 mM. Vmax 780 +/- 175 nmol.min-1 per 100 mg protein (n = 11). A detailed kinetic analysis of high affinity GABA uptake was performed over a range of sodium ion concentrations. The results were consistent with a coupling ratio of one Na+ ion to one GABA molecule; a similar result was found with rat brain synaptosomes. However, rat and human synaptosomes differed in the degree to which the substrate affinity of the high affinity GABA uptake site varied with decreasing Na+ ion concentration. High affinity GABA uptake was markedly affected by the method used to freeze and divide the tissue, but did not vary greatly in different cortical regions. There was some decline of high affinity GABA uptake activity with postmortem delay, apparently due to a loss of sites rather than a change in site affinity.

  5. Phenotypic consequences of deletion of the {gamma}{sub 3}, {alpha}{sub 5}, or {beta}{sub 3} subunit of the type A {gamma}-aminobutyric acid receptor in mice

    SciTech Connect

    Culia, C.T.; Stubbs, L.J.; Montgomery, C.S.; Russell, L.B.; Rinchik, E.M.

    1994-03-29

    Three genes (Gabrg3, Gabra5, and Gabrb3) encoding the {gamma}{sub 3}, {alpha}{sub 5}, and {beta}{sub 3} subunits of the type A {gamma}-aminobutyric acid receptor, respectively, are known to map near the pink-eyed dilution (p) locus in mouse chromosome 7. This region shares homology with a segment of human chromosome 15 that is implicated in Angelman syndrome, an inherited neurobehavioral disorder. By mapping Gabrg3-Gabra5-Gabrb3-telomere. Like Gabrb3, neither the Gabra5 nor Gabrg3 gene is functionally imprinted in adult mouse brain. Mice deleted for all three subunits die at birth with a cleft palate, although there are rare survivors ({approximately} 5%) that do not have a cleft palate but do exhibit a neurological abnormality characterized by tremor, jerky gait, and runtiness. The authors have previously suggested that deficiency of the {beta}{sub 3} subunit may be responsible for the clefting defect. Most notably, however, in this report they describe mice carrying two overlapping, complementing p deletions that fail to express the {gamma}{sub 3} transcript, as well as mice from another line that express neither the {gamma}{sub 3} nor {alpha}{sub 5} transcripts. Surprisingly, mice from both of these lines are phenotypically normal and do not exhibit any of the neurological symptoms characteristic of the rare survivors that are deleted for all three ({gamma}{sub 3}, {alpha}{sub 5}, and {beta}{sub 3}) subunits. These mice therefore provide a whole-organism type A {gamma}-aminobutyric-acid receptor background that is devoid of any receptor subtypes that normally contain the {gamma}{sub 3} and/or {alpha}{sub 5} subunits. The absence of an overt neurological phenotype in mice lacking the {gamma}{sub 3} and/or {alpha}{sub 5} subunits also suggests that mutations in these genes are unlikely to provide useful animal models for Angelman syndrome in humans.

  6. Compounds exhibiting selective efficacy for different beta subunits of human recombinant gamma-aminobutyric acid A receptors.

    PubMed

    Smith, Alison J; Oxley, Beth; Malpas, Sallie; Pillai, Gopalan V; Simpson, Peter B

    2004-11-01

    Inhibitory GABA(A) receptor modulators are widely used therapeutic agents for a variety of central nervous system disorders. Ltk(-) cells stably expressing human recombinant GABA(A) subunits (alpha1beta1-3gamma2s) were seeded into 96-well plates, loaded with chlorocoumarin-2-dimyristoyl phosphatidylethanolamine and bis(1,3-diethyl-2-thiobarbiturate)trimethineoxonol, and rapid fluorescence resonance energy transfer technique (FRET) measurements were made of GABA-evoked depolarizations in low-Cl(-) buffer using a voltage/ion probe reader. The influence of different betasubunits on the ability of agents to modulate and directly activate the ion channel was examined. GABA evoked concentration-dependent decreases in FRET, increasing fluorescence emission ratio (460/580 nm) at alpha1beta1gamma2, alpha1beta2gamma2, and alpha1beta3gamma2 receptors with similar maximal amplitude (P > 0.05, n = 17) and EC(50) values of 2.4 +/- 0.2, 2.5 +/- 0.2, and 1.3 +/- 0.1 microM, respectively. Piperidine-4-sulfonic acid and 4,5,6,7-tetrahydroisoxazolo[5,4-c]pyridin-3-ol were less potent, with EC(50) values of 8.7 +/- 0.9, 9.2 +/- 0.5, and 11.7 +/- 1.2, and 43.7 +/- 6.4, 24.8 +/- 1.6, and 26.1 +/- 2.4 microM, respectively. Potency and maximal efficacy of propofol, methyl 6,7-dimethoxy-4-ethyl-beta-carboline-3-carboxylate, pentobarbital, and steroids, 5alpha-pregnan-3alpha-ol-20-one and 5beta-pregnan-3alpha-ol-20-one, were unaffected by the beta isoform present in the receptor complex. However, several compounds displayed beta2/3 subunit selectivity, notably loreclezole, R(-)-etomidate, and a group of anti-inflammatory agents including mefenamic acid, flufenamic acid, meclofenamic acid, tolfenamic acid, niflumic acid, and diflunisal. The anti-inflammatories exhibited varying levels of efficacy at beta2/3 subunits, with micromolar potency, while having antagonist or weak inverse agonist profiles at alpha1beta1gamma2. Diflunisal was the most efficacious compound, eliciting greater

  7. Co-localization of Gamma-Aminobutyric Acid and Glutamate in Neurons of the Spider Central Nervous System.

    PubMed

    Fabian-Fine, Ruth; Meisner, Shannon; Torkkeli, Päivi H; Meinertzhagen, Ian A

    2015-12-01

    Spider sensory neurons with cell bodies close to various sensory organs are innervated by putative efferent axons from the central nervous system (CNS). Light and electronmicroscopic imaging of immunolabeled neurons has demonstrated that neurotransmitters present at peripheral synapses include γ-aminobutyric acid (GABA), glutamate and octopamine. Moreover, electrophysiological studies show that these neurotransmitters modulate the sensitivity of peripheral sensory neurons. Here, we undertook immunocytochemical investigations to characterize GABA and glutamate-immunoreactive neurons in three-dimensional reconstructions of the spider CNS. We document that both neurotransmitters are abundant in morphologically distinct neurons throughout the CNS. Labeling for the vesicular transporters, VGAT for GABA and VGLUT for glutamate, showed corresponding patterns, supporting the specificity of antibody binding. Whereas some neurons displayed strong immunolabeling, others were only weakly labeled. Double labeling showed that a subpopulation of weakly labeled neurons present in all ganglia expresses both GABA and glutamate. Double labeled, strongly and weakly labeled GABA and glutamate immunoreactive axons were also observed in the periphery along muscle fibers and peripheral sensory neurons. Electron microscopic investigations showed presynaptic profiles of various diameters with mixed vesicle populations innervating muscle tissue as well as sensory neurons. Our findings provide evidence that: (1) sensory neurons and muscle fibers are innervated by morphologically distinct, centrally located GABA- and glutamate immunoreactive neurons; (2) a subpopulation of these neurons may co-release both neurotransmitters; and (3) sensory neurons and muscles are innervated by all of these neurochemically and morphologically distinct types of neurons. The biochemical diversity of presynaptic innervation may contribute to how spiders filter natural stimuli and coordinate appropriate response

  8. In Silico Prediction of Gamma-Aminobutyric Acid Type-A Receptors Using Novel Machine-Learning-Based SVM and GBDT Approaches

    PubMed Central

    Huang, Yong; Ju, Ying

    2016-01-01

    Gamma-aminobutyric acid type-A receptors (GABAARs) belong to multisubunit membrane spanning ligand-gated ion channels (LGICs) which act as the principal mediators of rapid inhibitory synaptic transmission in the human brain. Therefore, the category prediction of GABAARs just from the protein amino acid sequence would be very helpful for the recognition and research of novel receptors. Based on the proteins' physicochemical properties, amino acids composition and position, a GABAAR classifier was first constructed using a 188-dimensional (188D) algorithm at 90% cd-hit identity and compared with pseudo-amino acid composition (PseAAC) and ProtrWeb web-based algorithms for human GABAAR proteins. Then, four classifiers including gradient boosting decision tree (GBDT), random forest (RF), a library for support vector machine (libSVM), and k-nearest neighbor (k-NN) were compared on the dataset at cd-hit 40% low identity. This work obtained the highest correctly classified rate at 96.8% and the highest specificity at 99.29%. But the values of sensitivity, accuracy, and Matthew's correlation coefficient were a little lower than those of PseAAC and ProtrWeb; GBDT and libSVM can make a little better performance than RF and k-NN at the second dataset. In conclusion, a GABAAR classifier was successfully constructed using only the protein sequence information. PMID:27579307

  9. Cloning of the. gamma. -aminobutyric acid (GABA). rho. sub 1 cDNA: A GABA receptor subunit highly expressed in the retina

    SciTech Connect

    Cutting, G.R.; Lu, Luo; Kasch, L.M.; Montrose-Rafizadeh, C.; Antonarakis, S.E.; Guggino, W.B.; Kazazian, H.H. Jr. ); O'Hara, B.F.; Donovan, D.M.; Shimada, Shoichi ); Uhl, G.R. Johns Hopkins Univ. School of Medicine, Baltimore, MD )

    1991-04-01

    Type A {gamma}-aminobutyric acid (GABA{sub A}) receptors are a family of ligand-gated chloride channels that are the major inhibitory neurotransmitter receptors in the nervous system. Molecular cloning has revealed diversity in the subunits that compose this heterooligomeric receptor, but each previously elucidated subunit displays amino acid similarity in conserved structural elements. The authors have used these highly conserved regions to identify additional members of this family by using the polymerase chain reaction (PCR). One PCR product was used to isolate a full-length cDNA from a human retina cDNA library. The mature protein predicted from this cDNA sequence is 458 amino acids long and displays between 30 and 38% amino acid similarity to the previously identified GABA{sub A} subunits. This gene is expressed primarily in the retina but transcripts are also detected in the brain, lung, and thymus. Injection of Xenopus oocytes with RNA transcribed in vitro produces a GABA-responsive chloride conductance and expression of the cDNA in COS cells yields GABA-displaceable muscimol binding. These features are consistent with our identification of a GABA subunit, GABA {rho}{sub 1}, with prominent retinal expression that increases the diversity and tissue specificity of this ligand-gated ion-channel receptor family.

  10. In Silico Prediction of Gamma-Aminobutyric Acid Type-A Receptors Using Novel Machine-Learning-Based SVM and GBDT Approaches.

    PubMed

    Liao, Zhijun; Huang, Yong; Yue, Xiaodong; Lu, Huijuan; Xuan, Ping; Ju, Ying

    2016-01-01

    Gamma-aminobutyric acid type-A receptors (GABAARs) belong to multisubunit membrane spanning ligand-gated ion channels (LGICs) which act as the principal mediators of rapid inhibitory synaptic transmission in the human brain. Therefore, the category prediction of GABAARs just from the protein amino acid sequence would be very helpful for the recognition and research of novel receptors. Based on the proteins' physicochemical properties, amino acids composition and position, a GABAAR classifier was first constructed using a 188-dimensional (188D) algorithm at 90% cd-hit identity and compared with pseudo-amino acid composition (PseAAC) and ProtrWeb web-based algorithms for human GABAAR proteins. Then, four classifiers including gradient boosting decision tree (GBDT), random forest (RF), a library for support vector machine (libSVM), and k-nearest neighbor (k-NN) were compared on the dataset at cd-hit 40% low identity. This work obtained the highest correctly classified rate at 96.8% and the highest specificity at 99.29%. But the values of sensitivity, accuracy, and Matthew's correlation coefficient were a little lower than those of PseAAC and ProtrWeb; GBDT and libSVM can make a little better performance than RF and k-NN at the second dataset. In conclusion, a GABAAR classifier was successfully constructed using only the protein sequence information. PMID:27579307

  11. Metabolism of gamma-aminobutyric acid during cold acclimation and freezing and its relationship to frost tolerance in barley and wheat.

    PubMed

    Mazzucotelli, Elisabetta; Tartari, Alfredo; Cattivelli, Luigi; Forlani, Giuseppe

    2006-01-01

    Amino acid homeostasis was investigated in frost-resistant barley seedlings under either cold- or freezing-stress conditions. Total free amino acid content varied only slightly, but a substantial conversion of glutamate to gamma-aminobutyric acid (GABA) was found that was proportional to the severity of the stress. Cold acclimation caused a significant increase in amino acid pools, and induced the expression of the GABA-shunt genes. As a consequence, GABA accumulated to a higher extent during the subsequent exposure to lower temperature. A different picture was obtained with a frost-sensitive genotype, in which glutamate decarboxylation occurred during the stress as well, but the activation of the GABA shunt seemed not to take place, and free glutamate was almost depleted. Analogous results were found in frost-resistant and frost-sensitive wheat cultivars. Feeding non-hardened plants with exogenous glutamate resulted in increased GABA accumulation under low temperature. The possibility that glutamate decarboxylation and GABA metabolism would play a role in frost tolerance is discussed. PMID:16997899

  12. Co-Localization of GABA Shunt Enzymes for the Efficient Production of Gamma-Aminobutyric Acid via GABA Shunt Pathway in Escherichia coli.

    PubMed

    Pham, Van Dung; Somasundaram, Sivachandiran; Park, Si Jae; Lee, Seung Hwan; Hong, Soon Ho

    2016-04-28

    Gamma-aminobutyric acid (GABA) is a non-protein amino acid, which is an important inhibitor of neurotransmission in the human brain. GABA is also used as the precursor of biopolymer Nylon-4 production. In this study, the carbon flux from the tricarboxylic acid cycle was directed to the GABA shunt pathway for the production of GABA from glucose. The GABA shunt enzymes succinate-semialdehyde dehydrogenase (GabD) and GABA aminotransferase (GabT) were co-localized along with the GABA transporter (GadC) by using a synthetic scaffold complex. The co-localized enzyme scaffold complex produced 0.71 g/l of GABA from 10 g/l of glucose. Inactivation of competing metabolic pathways in mutant E. coli strains XBM1 and XBM6 increased GABA production 13% to reach 0.80 g/l GABA by the enzymes co-localized and expressed in the mutant strains. The recombinant E. coli system developed in this study demonstrated the possibility of the pathway of the GABA shunt as a novel GABA production pathway.

  13. Use of sourdough fermentation and pseudo-cereals and leguminous flours for the making of a functional bread enriched of gamma-aminobutyric acid (GABA).

    PubMed

    Coda, Rossana; Rizzello, Carlo Giuseppe; Gobbetti, Marco

    2010-02-28

    Lactobacillus plantarum C48 and Lactococcus lactis subsp. lactis PU1, previously selected for the biosynthesis of gamma-aminobutyric acid (GABA), were used for sourdough fermentation of cereal, pseudo-cereal and leguminous flours. Chickpea, amaranth, quinoa and buckwheat were the flours most suitable to be enriched of GABA. The parameters of sourdough fermentation were optimized. Addition of 0.1mM pyridoxal phosphate, dough yield of 160, inoculum of 5 x 10(7)CFU/g of starter bacteria and fermentation for 24h at 30 degrees C were found to be the optimal conditions. A blend of buckwheat, amaranth, chickpea and quinoa flours (ratio 1:1:5.3:1) was selected and fermented with baker's yeast (non-conventional flour bread, NCB) or with Lb. plantarum C48 sourdough (non-conventional flour sourdough bread, NCSB) and compared to baker's yeast started wheat flour bread (WFB). NCSB had the highest concentration of free amino acids and GABA (ca. 4467 and 504 mg/kg, respectively). The concentration of phenolic compounds and antioxidant activity of NCSB bread was the highest, as well as the rate of in vitro starch hydrolysis was the lowest. Texture analysis showed that sourdough fermentation enhances several characteristics of NCSB with respect to NCB, thus approaching the features of WFB. Sensory analysis showed that sourdough fermentation allowed to get good palatability and overall taste appreciation.

  14. A Fluorescence-Coupled Assay for Gamma Aminobutyric Acid (GABA) Reveals Metabolic Stress-Induced Modulation of GABA Content in Neuroendocrine Cancer

    PubMed Central

    Ippolito, Joseph E.; Piwnica-Worms, David

    2014-01-01

    Pathways involved in the synthesis of the neurotransmitter gamma-aminobutyric acid (GABA) have been implicated in the pathogenesis of high grade neuroendocrine (NE) neoplasms as well as neoplasms from a non-NE lineage. Using The Cancer Genome Atlas, overexpression of the GABA synthetic enzyme, glutamate decarboxylase 1 (GAD1), was found to be associated with decreased disease free-survival in prostate adenocarcinoma and decreased overall survival in clear cell renal cell carcinomas. Furthermore, GAD1 was found to be expressed in castrate-resistant prostate cancer cell lines, but not androgen-responsive cell lines. Using a novel fluorescence-coupled enzymatic microplate assay for GABA mediated through reduction of resazurin in a prostate neuroendocrine carcinoma (PNEC) cell line, acid microenvironment-induced stress increased GABA levels while alkaline microenvironment-induced stress decreased GABA through modulation of GAD1 and glutamine synthetase (GLUL) activities. Moreover, glutamine but not glucose deprivation decreased GABA through modulation of GLUL. Consistent with evidence in prokaryotic and eukaryotic organisms that GABA synthesis mediated through GAD1 may play a crucial role in surviving stress, GABA may be an important mediator of stress survival in neoplasms. These findings identify GABA synthesis and metabolism as a potentially important pathway for regulating cancer cell stress response as well as a potential target for therapeutic strategies. PMID:24551133

  15. Efficient gamma-aminobutyric acid bioconversion by employing synthetic complex between glutamate decarboxylase and glutamate/GABA antiporter in engineered Escherichia coli.

    PubMed

    Le Vo, Tam Dinh; Ko, Ji-seun; Park, Si Jae; Lee, Seung Hwan; Hong, Soon Ho

    2013-08-01

    Gamma-aminobutyric acid (GABA) is a precursor of one of the most promising heat-resistant biopolymers, Nylon-4, and can be produced by the decarboxylation of monosodium glutamate (MSG). In this study, a synthetic protein complex was applied to improve the GABA conversion in engineered Escherichia coli. Complexes were constructed by assembling a single protein-protein interaction domain SH3 to the glutamate decarboxylase (GadA and GadB) and attaching a cognate peptide ligand to the glutamate/GABA antiporter (GadC) at the N-terminus, C-terminus, and the 233rd amino acid residue. When GadA and GadC were co-overexpressed via the C-terminus complex, a GABA concentration of 5.65 g/l was obtained from 10 g/l MSG, which corresponds to a GABA yield of 93 %. A significant increase of the GABA productivity was also observed where the GABA productivity increased 2.5-fold in the early culture period due to the introduction of the synthetic protein complex. The GABA pathway efficiency and GABA productivity were enhanced by the introduction of the complex between Gad and glutamate/GABA antiporter.

  16. A fluorescence-coupled assay for gamma aminobutyric acid (GABA) reveals metabolic stress-induced modulation of GABA content in neuroendocrine cancer.

    PubMed

    Ippolito, Joseph E; Piwnica-Worms, David

    2014-01-01

    Pathways involved in the synthesis of the neurotransmitter gamma-aminobutyric acid (GABA) have been implicated in the pathogenesis of high grade neuroendocrine (NE) neoplasms as well as neoplasms from a non-NE lineage. Using The Cancer Genome Atlas, overexpression of the GABA synthetic enzyme, glutamate decarboxylase 1 (GAD1), was found to be associated with decreased disease free-survival in prostate adenocarcinoma and decreased overall survival in clear cell renal cell carcinomas. Furthermore, GAD1 was found to be expressed in castrate-resistant prostate cancer cell lines, but not androgen-responsive cell lines. Using a novel fluorescence-coupled enzymatic microplate assay for GABA mediated through reduction of resazurin in a prostate neuroendocrine carcinoma (PNEC) cell line, acid microenvironment-induced stress increased GABA levels while alkaline microenvironment-induced stress decreased GABA through modulation of GAD1 and glutamine synthetase (GLUL) activities. Moreover, glutamine but not glucose deprivation decreased GABA through modulation of GLUL. Consistent with evidence in prokaryotic and eukaryotic organisms that GABA synthesis mediated through GAD1 may play a crucial role in surviving stress, GABA may be an important mediator of stress survival in neoplasms. These findings identify GABA synthesis and metabolism as a potentially important pathway for regulating cancer cell stress response as well as a potential target for therapeutic strategies.

  17. Vibrational Spectra of γ-Aminobutyric Acid

    NASA Astrophysics Data System (ADS)

    Suresh, D. M.; Sajan, D.; Laladas, K. P.; Joe, I. Hubert; Jayakumar, V. S.

    2008-11-01

    The NIR-FT Raman, FT-IR spectral analysis of γ-Aminobutyric acid (GABA) a simple amino acid is carried out by density functional computations. The vibrational spectra confirm the existence of NH3+ in GABA. Hydroxyl groups H-bonded to the different extents are analysed, supported by computed results.

  18. Effects of electroacupuncture on the levels of retinal gamma-aminobutyric acid and its receptors in a guinea pig model of lens-induced myopia.

    PubMed

    Sha, F; Ye, X; Zhao, W; Xu, C-L; Wang, L; Ding, M-H; Bi, A-L; Wu, J-F; Jiang, W-J; Guo, D-D; Guo, J-G; Bi, H-S

    2015-02-26

    Gamma-aminobutyric acid (GABA) is a major inhibitory neurotransmitter of the retina and affects myopic development. Electroacupuncture (EA) is widely utilized to treat myopia in clinical settings. However, there are few reports on whether EA affects the level of retinal GABA during myopic development. To study this issue, in the present study, we explored the changes of retinal GABA content and the expression of its receptor subtypes, and the effects of EA stimulation on them in a guinea pig model with lens-induced myopia (LIM). Our results showed that the content of GABA and the expression of GABAA and GABAC receptors of retina were up-regulated during the development of myopia, and this up-regulation was inhibited by applying EA to Hegu (LI4) and Taiyang (EX-HN5) acupoints. Moreover, these effects of EA show a positional specificity. While applying EA at a sham acupoint, no apparent change of myopic retinal GABA and its receptor subtypes was observed. Taken together, our findings suggest that LIM is effective to up-regulate the level of retinal GABA, GABAA and GABAC receptors in guinea pigs and the effect may be inhibited by EA stimulation at LI4 and EX-HN5 acupoints.

  19. In vivo effects of ketamine on glutamate-glutamine and gamma-aminobutyric acid in obsessive-compulsive disorder: Proof of concept.

    PubMed

    Rodriguez, Carolyn I; Kegeles, Lawrence S; Levinson, Amanda; Ogden, R Todd; Mao, Xiangling; Milak, Matthew S; Vermes, Donna; Xie, Shan; Hunter, Liane; Flood, Pamela; Moore, Holly; Shungu, Dikoma C; Simpson, Helen B

    2015-08-30

    We previously reported the rapid and robust clinical effects of ketamine versus saline infusions in a proof-of-concept crossover trial in unmedicated adults with obsessive-compulsive disorder (OCD). This study examined the concurrent neurochemical effects of ketamine versus saline infusions using proton magnetic resonance spectroscopy ((1)H MRS) during the clinical proof-of-concept crossover trial. Levels of the inhibitory neurotransmitter gamma-aminobutyric acid (GABA) and the excitatory neurochemicals glutamate+glutamine (Glx) were acquired in the medial prefrontal cortex (MPFC), a region implicated in OCD pathology. Seventeen unmedicated OCD adults received two intravenous infusions at least 1 week apart, one of saline and one of ketamine, while lying supine in a 3.0 T GE MR scanner. The order of each infusion pair was randomized. Levels of GABA and Glx were measured in the MPFC before, during, and after each infusion and normalized to water (W). A mixed effects model found that MPFC GABA/W significantly increased over time in the ketamine compared with the saline infusion. In contrast, there were no significant differences in Glx/W between the ketamine and saline infusions. Together with earlier evidence of low cortical GABA in OCD, our findings suggest that models of OCD pathology should consider the role of GABAergic abnormalities in OCD symptomatology.

  20. Tonic inhibition in mouse hippocampal CA1 pyramidal neurons is mediated by alpha5 subunit-containing gamma-aminobutyric acid type A receptors.

    PubMed

    Caraiscos, Valerie B; Elliott, Erin M; You-Ten, Kong E; Cheng, Victor Y; Belelli, Delia; Newell, J Glen; Jackson, Michael F; Lambert, Jeremy J; Rosahl, Thomas W; Wafford, Keith A; MacDonald, John F; Orser, Beverley A

    2004-03-01

    The principal inhibitory neurotransmitter in the mammalian brain, gamma-aminobutyric acid (GABA), is thought to regulate memory processes by activating transient inhibitory postsynaptic currents. Here we describe a nonsynaptic, tonic form of inhibition in mouse CA1 pyramidal neurons that is generated by a distinct subpopulation of GABA type A receptors (GABA(A)Rs). This tonic inhibitory conductance is predominantly mediated by alpha5 subunit-containing GABA(A)Rs (alpha5GABA(A)Rs) that have different pharmacological and kinetic properties compared to postsynaptic receptors. GABA(A)Rs that mediate the tonic conductance are well suited to detect low, persistent, ambient concentrations of GABA in the extracellular space because they are highly sensitive to GABA and desensitize slowly. Moreover, the tonic current is highly sensitive to enhancement by amnestic drugs. Given the restricted expression of alpha5GABA(A)Rs to the hippocampus and the association between reduced alpha5GABA(A)R function and improved memory performance in behavioral studies, our results suggest that tonic inhibition mediated by alpha5GABA(A)Rs in hippocampal pyramidal neurons plays a key role in cognitive processes.

  1. Distinction between the effects of barbiturates, benzodiazepines and phenytoin on responses to gamma-aminobutyric acid receptor activation and antagonism by bicuculline and picrotoxin.

    PubMed Central

    Simmonds, M. A.

    1981-01-01

    1 Interactions of depressant and anticonvulsant drugs with the neuronal gamma-aminobutyric acid (GABA) receptor + effector system have been examined on afferent fibres to the rat cuneate nucleus in vitro. Three types of interaction have been measured: (a) potentiation of depolarizing responses to the GABA analogue, muscimol: (b) reduction in the potency of bicuculline as an antagonist of muscimol at the GABA receptor: (c) reduction in the potency of picrotoxin as an antagonist of muscimol acting on the effector mechanism. 2 Phenobarbitone reduced the potency of picrotoxin in doses which did not affect the potency of bicuculline and which caused only a small potentiation of muscimol. Pentobarbitone did not show such selectivity, a reduction in potency of picrotoxin always being accompanied by a reduction in potency of bicuculline and a substantial potentiation of muscimol. 3 Flurazepam and lorazepam both reduced the potency of picrotoxin without affecting that of bicuculline and with very little potentiation of muscimol. Phenytoin had no effect on the potency of picrotoxin whilst potentiating muscimol to the same extent as phenobarbitone. 4 The spectrum of drug activity in reducing the potency of picrotoxin correlates well with the reported anticonvulsant effects of these drugs against kindled amygdaloid seizures. Potentiation of muscimol and reduction of bicuculline potency appear more closely related to hypnotic properties. PMID:6265019

  2. Mass transfer characterization of gamma-aminobutyric acid production by Enterococcus faecium CFR 3003: encapsulation improves its survival under simulated gastro-intestinal conditions.

    PubMed

    Divyashri, Gangaraju; Prapulla, Siddalingaiya Gurudatt

    2015-03-01

    Gamma-aminobutyric acid (GABA) production by free and Ca-alginate encapsulated cells of Enterococcus faecium CFR 3003 was investigated. Mass transfer rates characterizing the GABA production process using encapsulated cells were investigated. Experiments were performed to investigate external film and internal pore diffusion mass transfer rates. The Damkohler and Thiele analysis provides a good description of external film and internal pore diffusion resistances, respectively. The experiments revealed that the external film effects could be neglected but the process is affected to the greater extent by internal mass transfer effects and was found to be the principal rate-controlling step. Protective effect of encapsulation on cell survivability was tested under digestive environment, when challenged to salivary α-amylase, simulated gastric fluid and intestinal fluid. Viability of encapsulated cells was significantly higher under simulated gastro-intestinal conditions and could produce higher GABA than those observed with free cells. The results indicate that the Ca-alginate encapsulated probiotics could effectively be delivered to the colonic site for effective inhibitory action.

  3. Identification of clathrin heavy chain as a direct interaction partner for the gamma-aminobutyric acid type A receptor associated protein.

    PubMed

    Mohrlüder, Jeannine; Hoffmann, Yvonne; Stangler, Thomas; Hänel, Karen; Willbold, Dieter

    2007-12-18

    Gamma-aminobutyric acid type A receptors (GABAA receptors) are the major sites of GABA-mediated fast synaptic inhibition in the central nervous system. Variation of the cell surface receptor count is postulated to be of importance in modulating inhibitory synaptic transmission. The GABAA receptor associated protein (GABARAP) is a ubiquitin-like modifier, implicated in GABAA receptor clustering, trafficking, and turnover. GABARAP pull-down experiments with brain lysate identified clathrin heavy chain to be GABARAP-associated. Phage display screening of a randomized peptide library for GABARAP ligands yielded a sequence motif which characterizes the peptide binding specificity of GABARAP. Sequence database searches with this motif revealed clathrin heavy chain as a protein containing the identified sequence motif within its residues 510-522, supporting the result of the pull-down experiments. Calreticulin, which was identified recently as a GABARAP ligand, contains a very similar sequence motif. We demonstrate that calreticulin indeed competes with clathrin heavy chain for GABARAP binding. Finally, employing nuclear magnetic resonance spectroscopy, we mapped the GABARAP residues responsible for binding to clathrin. The hereby mapped GABARAP regions overlap very well with the homologue residues in yeast Atg8 that were recently shown to be important for autophagy. Together with the knowledge that GABARAP and clathrin are known to be involved in GABAA receptor trafficking within the cell, this strongly suggests a clear physiological relevance of the direct interaction of GABARAP with clathrin heavy chain. PMID:18027972

  4. In vivo effects of ketamine on glutamate-glutamine and gamma-aminobutyric acid in obsessive-compulsive disorder: Proof of concept.

    PubMed

    Rodriguez, Carolyn I; Kegeles, Lawrence S; Levinson, Amanda; Ogden, R Todd; Mao, Xiangling; Milak, Matthew S; Vermes, Donna; Xie, Shan; Hunter, Liane; Flood, Pamela; Moore, Holly; Shungu, Dikoma C; Simpson, Helen B

    2015-08-30

    We previously reported the rapid and robust clinical effects of ketamine versus saline infusions in a proof-of-concept crossover trial in unmedicated adults with obsessive-compulsive disorder (OCD). This study examined the concurrent neurochemical effects of ketamine versus saline infusions using proton magnetic resonance spectroscopy ((1)H MRS) during the clinical proof-of-concept crossover trial. Levels of the inhibitory neurotransmitter gamma-aminobutyric acid (GABA) and the excitatory neurochemicals glutamate+glutamine (Glx) were acquired in the medial prefrontal cortex (MPFC), a region implicated in OCD pathology. Seventeen unmedicated OCD adults received two intravenous infusions at least 1 week apart, one of saline and one of ketamine, while lying supine in a 3.0 T GE MR scanner. The order of each infusion pair was randomized. Levels of GABA and Glx were measured in the MPFC before, during, and after each infusion and normalized to water (W). A mixed effects model found that MPFC GABA/W significantly increased over time in the ketamine compared with the saline infusion. In contrast, there were no significant differences in Glx/W between the ketamine and saline infusions. Together with earlier evidence of low cortical GABA in OCD, our findings suggest that models of OCD pathology should consider the role of GABAergic abnormalities in OCD symptomatology. PMID:26104826

  5. Hypergravity exposure decreases gamma-aminobutyric acid immunoreactivity in axon terminals contacting pyramidal cells in the rat somatosensory cortex: a quantitative immunocytochemical image analysis

    NASA Technical Reports Server (NTRS)

    D'Amelio, F.; Wu, L. C.; Fox, R. A.; Daunton, N. G.; Corcoran, M. L.; Polyakov, I.

    1998-01-01

    Quantitative evaluation of gamma-aminobutyric acid immunoreactivity (GABA-IR) in the hindlimb representation of the rat somatosensory cortex after 14 days of exposure to hypergravity (hyper-G) was conducted by using computer-assisted image processing. The area of GABA-IR axosomatic terminals apposed to pyramidal cells of cortical layer V was reduced in rats exposed to hyper-G compared with control rats, which were exposed either to rotation alone or to vivarium conditions. Based on previous immunocytochemical and behavioral studies, we suggest that this reduction is due to changes in sensory feedback information from muscle receptors. Consequently, priorities for muscle recruitment are altered at the cortical level, and a new pattern of muscle activity is thus generated. It is proposed that the reduction observed in GABA-IR of the terminal area around pyramidal neurons is the immunocytochemical expression of changes in the activity of GABAergic cells that participate in reprogramming motor outputs to achieve effective movement control in response to alterations in the afferent information.

  6. Feeding rumen-protected gamma-aminobutyric acid enhances the immune response and antioxidant status of heat-stressed lactating dairy cows.

    PubMed

    Cheng, Jianbo; Zheng, Nan; Sun, Xianzhi; Li, Songli; Wang, Jiaqi; Zhang, Yangdong

    2016-08-01

    This experiment was conducted to investigate the effects of rumen-protected gamma-aminobutyric acid (GABA) on immune function and antioxidant status in heat-stressed dairy cows. Sixty Holstein dairy cows were randomly assigned to 1 of 4 treatments according to a completely randomized block design. The treatments consisted of 0 (control), 40, 80, or 120mg of GABA/kg DM from rumen-protected GABA. The trial lasted 10 weeks. The average temperature-humidity indices at 0700, 1400 and 2200h were 78.4, 80.2 and 78.7, respectively. Rectal temperatures decreased linearly at 0700, 1400, and 2200h with increasing GABA. As the GABA increased, the immunoglobulin (Ig) A and IgG contents and the proportions of CD4(+) and CD8(+) T lymphocytes increased linearly (P<0.05), whereas concentrations of interleukin (IL)-2, IL-4, IL-6 and tumor necrosis factor-α (TNF-α) decreased linearly (P<0.05). The activities of superoxide dismutase (SOD), glutathione-peroxidase (GSH-PX) and total antioxidant capacity (T-AOC) increased linearly (P<0.05), whereas malondialdehyde (MDA) content decreased linearly (P<0.05) with increasing GABA. These results indicate that rumen-protected GABA supplementation to heat-stressed dairy cows can improve their immune function and antioxidant activity. PMID:27503722

  7. A comparison of gamma-aminobutyric acid and the semi-rigid analogues 4-aminotetrolic acid, 4-aminocrotonic acid and imidazole-4-acetic acid on the isolated superior cervical ganglion of the rat.

    PubMed Central

    Bowery, N G; Jones, G P

    1976-01-01

    1 The rat superior cervical ganglion possesses receptors for gamma-aminobutyric acid (GABA). This can be demonstrated in vitro by recording the changes in ganglionic surface potential which occur after the addition of GABA to the bathing solution. 2 The action of three conformationally-restricted analogues of GABA namely 4-aminotetrolic acid (4-ATA), trans 4-aminocrotonic acid (4-ACA) and imidazole-4-acetic acid (IAA) have been examined for activity at this peripheral receptor. 3 All three analogues depolarized the ganglion in a manner similar to GABA. Their actions were transient and were 'occluded' by GABA; also the dose-response curve in each case was parallel to that of GABA. Molar potencies relative to GABA (= 1) were 4-ACA = 1.48, IAA = 0.100, 4-ATA = 0.0028. 4 The action of each analogue could be blocked by the GABA antagonists bicuculline and tetramethylenedisulphotetramine at doses which had relatively little effect on responses to the cholinomimetic carbachol. 5 4-ACA and IAA (1 mM) significantly reduced the ganglionic accumulation of [3H]-GABA (0.2 muM) by 88% and 58% respectively whereas 4-ATA (1 mM), caused no significant reduction in [3H]-GABA accumulation. PMID:1260178

  8. Characterization of the Production of Biogenic Amines and Gamma-Aminobutyric Acid in the Soybean Pastes Fermented by Aspergillus oryzae and Lactobacillus brevis.

    PubMed

    Kim, Nam Yeun; Ji, Geun Eog

    2015-04-01

    The production of gamma-aminobutyric acid (GABA) using GABA-producing lactic acid bacteria (LAB) has been considered to be an attractive strategy. However, some LAB may produce biogenic amines (BA), which may be of concern from the safety viewpoint. The aim of the present study was to characterize the production of GABA and BA in the soybean pastes fermented by Aspergillus oryzae (A. oryzae) FMB S46471 and GABA-producing Lactobacillus brevis (L. brevis) GABA 100. After a ripening period of 90 days, the levels of BA (putrescine, cadaverine, histamine, and tyramine) and GABA in the fermented soybean were assessed by highperformance liquid chromatography. The soybean pastes fermented by A. oryzae and L. brevis showed a range of 7,130-11,592 mg/kg for GABA, 178-305 mg/kg for tyramine, 139-163 mg/kg for putrescine, 7.4-10.8 mg/kg for histamine, and 7.1-7.9 mg/kg for cadaverine, whereas the soybean pastes fermented by A. oryzae only showed a range of 30-1,671 mg/kg for GABA, 0.8-189 mg/kg for tyramine, 1.3-85 mg/kg for putrescine, up to 3.6 mg/kg for histamine, and 0.2-2.4 mg/kg for cadaverine. The results showed that the production of GABA was accompanied by the increase in the production of BA, even though the production levels of histamine and cadaverine were very low. This is the first study to simultaneously characterize the production of BA and GABA in GABA-enriched fermented soybean pastes, and warrants further study to minimize the production of BA while optimizing the production of GABA.

  9. Residues in the extracellular loop 4 are critical for maintaining the conformational equilibrium of the gamma-aminobutyric acid transporter-1.

    PubMed

    MacAulay, Nanna; Meinild, Anne-Kristine; Zeuthen, Thomas; Gether, Ulrik

    2003-08-01

    We mutated residues Met345 and Thr349 in the rat gamma-aminobutyric acid transporter-1 (GAT-1) to histidines (M345H and T349H). These two residues are located four amino acids apart at the extracellular end of transmembrane segment 7 in a region of GAT-1 that we have previously suggested undergoes conformational changes critical for the transport process. The two single mutants and the double mutant (M345H/T349H) were expressed in Xenopus laevis oocytes, and their steady-state and presteady-state kinetics were examined and compared with wild type GAT-1 by using the two-electrode voltage clamp method. Oocytes expressing M345H showed a decrease in apparent GABA affinity, an increase in apparent affinity for Na+, a shift in the charge/voltage (Q/Vm) relationship to more positive membrane potentials, and an increased Li+-induced leak current. Oocytes expressing T349H showed an increase in apparent GABA affinity, a decrease in apparent Na+ affinity, a profound shift in the Q/Vm relationship to more negative potentials, and a decreased Li+-induced leak current. The data are consistent with a shift in the conformational equilibrium of the mutant transporters, with M345H stabilized in an outward-facing conformation and T349H in an inward-facing conformation. These data suggest that the extracellular end of transmembrane domain 7 not only undergoes conformational changes critical for the translocation process but also plays a role in regulating the conformational equilibrium between inward- and outward-facing conformations.

  10. Colocalization of gamma-aminobutyric acid and acetylcholine in neurons in the laterodorsal and pedunculopontine tegmental nuclei in the cat: a light and electron microscopic study.

    PubMed

    Jia, Hong-Ge; Yamuy, Jack; Sampogna, Sharon; Morales, Francisco R; Chase, Michael H

    2003-12-01

    Cholinergic and gamma-aminobutyric acid (GABA) mechanisms in the dorsolateral pontomesencephalic tegmentum have been implicated in the control of active (REM) sleep and wakefulness. To determine the relationships between neurons that contain these neurotransmitters in this region of the brainstem in adult cats, combined light and electron microscopic immunocytochemical procedures were employed. Light microscopic analyses revealed that choline acetyltransferase (ChAT) and GABA immunoreactive neurons were distributed throughout the laterodorsal and pedunculopontine tegmental nuclei (LDT and PPT). Surprisingly, approximately 50% of the ChAT immunoreactive neurons in these nuclei also contained GABA. Using electron microscopic pre-embedding immunocytochemistry, GABA immunoreactivity was observed in somas, dendrites and axon terminals in both the LDT and PPT. Most of the GABA immunoreactive terminals formed symmetrical synapses with non-immunolabeled dendrites. Electron microscopic double-immunolabeling techniques revealed that ChAT and GABA were colocalized in axon terminals in the LDT/PPT. Approximately 30% of the ChAT immunoreactive terminals were also GABA immunoreactive, whereas only 6-8% of the GABA immunoreactive terminals were ChAT immunoreactive. Most of the ChAT/GABA immunoreactive terminals formed symmetrical synapses with non-immunolabeled dendrites; however, ChAT/GABA immunoreactive terminals were also observed that contacted ChAT immunoreactive dendrites. With respect to ChAT immunoreactive postsynaptic profiles, approximately 40% of the somas and 50% of the dendrites received synaptic contact from GABA immunoreactive terminals in both the LDT and PPT. These findings (a) indicate that there are fundamental interactions between cholinergic and GABAergic neurons within the LDT/PPT that play an important role in the control of active sleep and wakefulness and (b) provide an anatomical basis for the intriguing possibility that a mechanism of acetylcholine and

  11. The neuroactive steroid allopregnanolone suppresses hypothalamic gonadotropin-releasing hormone release through a mechanism mediated by the gamma-aminobutyric acidA receptor.

    PubMed

    Calogero, A E; Palumbo, M A; Bosboom, A M; Burrello, N; Ferrara, E; Palumbo, G; Petraglia, F; D'Agata, R

    1998-07-01

    The central nervous system (CNS) is able to synthesize and/or metabolize steroid hormones. These neuroactive steroids are capable of modulating several brain functions and, among these, they seem to regulate the hypothalamic-pituitary-gonadal (HPG) axis. Indeed, recent observations have shown that 5 alpha-pregnane-3 alpha-ol-20-one (allopregnanolone), one of the most abundant naturally occurring neuroactive steroids, suppresses ovulation and sexual behaviour when administered within the CNS. The present study was undertaken to evaluate the effects of allopregnanolone and its inactive stereoisomer, 5 alpha-pregnane-3 beta-ol-20-one, upon the release of gonadotropin-releasing hormone (GnRH) from individually-incubated hemihypothalami. Allopregnanolone suppressed GnRH release in a concentration-dependent manner with maximal activity in the nanomolar range, a range at which this neurosteroid is capable of playing a biological action. The specificity of allopregnanolone suppression of GnRH release was provided by the lack of effect of its known inactive stereoisomer. To evaluate the involvement of gamma-aminobutyric acidA (GABAA) receptor, we examined the effects of two neurosteroids with GABA-antagonistic properties, pregnanolone sulfate (PREG-S) and dehydroepiandrosterone sulfate (DHEAS), and of bicuculline, a selective antagonist of the GABA binding site on the GABAA receptor, on allopregnanolone (10 nM)-suppressed GnRH release. Both PREG-S and bicuculline overcame the inhibitory effects of allopregnanolone on GnRH release, whereas DHEAS did not. To substantiate the involvement of the GABAA receptor further, we tested the effects of muscimol, a selective agonist for this receptor, which suppressed GnRH release. In conclusion, allopregnanolone suppressed hypothalamic GnRH release in vitro and this effect appeared to be mediated by an interaction with the GABAA receptor. We speculate that the inhibitory effect of allopregnanolone on the HPG axis may also be caused by

  12. The role of gamma-aminobutyric acid/glycinergic synaptic transmission in mediating bilirubin-induced hyperexcitation in developing auditory neurons.

    PubMed

    Yin, Xin-Lu; Liang, Min; Shi, Hai-Bo; Wang, Lu-Yang; Li, Chun-Yan; Yin, Shan-Kai

    2016-01-01

    Hyperbilirubinemia is a common clinical phenomenon observed in human newborns. A high level of bilirubin can result in severe jaundice and bilirubin encephalopathy. However, the cellular mechanisms underlying bilirubin excitotoxicity are unclear. Our previous studies showed the action of gamma-aminobutyric acid (GABA)/glycine switches from excitatory to inhibitory during development in the ventral cochlear nucleus (VCN), one of the most sensitive auditory nuclei to bilirubin toxicity. In the present study, we investigated the roles of GABAA/glycine receptors in the induction of bilirubin hyperexcitation in early developing neurons. Using the patch clamp technique, GABAA/glycine receptor-mediated spontaneous inhibitory synaptic currents (sIPSCs) were recorded from bushy and stellate cells in acute brainstem slices from young mice (postnatal day 2-6). Bilirubin significantly increased the frequency of sIPSCs, and this effect was prevented by pretreatments of slices with either fast or slow Ca(2+) chelators BAPTA-AM and EGTA-AM suggesting that bilirubin can increase the release of GABA/glycine via Ca(2+)-dependent mechanisms. Using cell-attached recording configuration, we found that antagonists of GABAA and glycine receptors strongly attenuated spontaneous spiking firings in P2-6 neurons but produced opposite effect in P15-19 neurons. Furthermore, these antagonists reversed bilirubin-evoked hyperexcitability in P2-6 neurons, indicating that excitatory action of GABA/glycinergic transmission specifically contribute to bilirubin-induced hyperexcitability in the early stage of development. Our results suggest that bilirubin-induced enhancement of presynaptic release GABA/Glycine via Ca(2+)-dependent mechanisms may play a critical role in mediating neuronal hyperexcitation associated with jaundice, implicating potential new strategies for predicting, preventing, and treating bilirubin neurotoxicity.

  13. Stoichiometry of expressed alpha(4)beta(2)delta gamma-aminobutyric acid type A receptors depends on the ratio of subunit cDNA transfected.

    PubMed

    Wagoner, Kelly R; Czajkowski, Cynthia

    2010-05-01

    The gamma-aminobutyric acid type A receptor (GABA(A)R) is the target of many depressants, including benzodiazepines, anesthetics, and alcohol. Although the highly prevalent alphabetagamma GABA(A)R subtype mediates the majority of fast synaptic inhibition in the brain, receptors containing delta subunits also play a key role, mediating tonic inhibition and the actions of endogenous neurosteroids and alcohol. However, the fundamental properties of delta-containing GABA(A)Rs, such as subunit stoichiometry, are not well established. To determine subunit stoichiometry of expressed delta-containing GABA(A)Rs, we inserted the alpha-bungarotoxin binding site tag in the alpha(4), beta(2), and delta subunit N termini. An enhanced green fluorescent protein tag was also inserted into the beta(2) subunit to shift its molecular weight, allowing us to separate subunits using SDS-PAGE. Tagged alpha(4)beta(2)delta GABA(A)Rs were expressed in HEK293T cells using various ratios of subunit cDNA, and receptor subunit stoichiometry was determined by quantitating fluorescent alpha-bungarotoxin bound to each subunit on Western blots of surface immunopurified tagged GABA(A)Rs. The results demonstrate that the subunit stoichiometry of alpha(4)beta(2)delta GABA(A)Rs is regulated by the ratio of subunit cDNAs transfected. Increasing the ratio of delta subunit cDNA transfected increased delta subunit incorporation into surface receptors with a concomitant decrease in beta(2) subunit incorporation. Because receptor subunit stoichiometry can directly influence GABA(A)R pharmacological and functional properties, considering how the transfection protocols used affect subunit stoichiometry is essential when studying heterologously expressed alpha(4)beta(2)delta GABA(A)Rs. Successful bungarotoxin binding site tagging of GABA(A)R subunits is a novel tool with which to accurately quantitate subunit stoichiometry and will be useful for monitoring GABA(A)R trafficking in live cells.

  14. Differential expression of gamma-aminobutyric acid type B receptor subunit mRNAs in the developing nervous system and receptor coupling to adenylyl cyclase in embryonic neurons.

    PubMed

    Martin, Stella C; Steiger, Janine L; Gravielle, María Clara; Lyons, Helen R; Russek, Shelley J; Farb, David H

    2004-05-17

    gamma-Aminobutyric acid type B receptors (GABA(B)Rs) mediate both slow inhibitory synaptic activity in the adult nervous system and motility signals for migrating embryonic cortical cells. Previous papers have described the expression of GABA(B)Rs in the adult brain, but the expression and functional significance of these gene products in the embryo are largely unknown. Here we examine GABA(B)R expression from rat embryonic day 10 (E10) to E18 compared with adult and ask whether embryonic cortical neurons contain functional GABA(B)R. GABA(B)R1 transcript levels greatly exceed GABA(B)R2 levels in the developing neural tube at E11, and olfactory bulb and striatum at E17 but equalize in most regions of adult nervous tissue, except for the glomerular and granule cell layers of the main olfactory bulb and the striatum. Consistent with expression differences, the binding affinity of GABA for GABA(B)Rs is significantly lower in adult striatum compared with cerebellum. Multiple lines of evidence from in situ hybridization, RNase protection, and real-time PCR demonstrate that GABA(B)R1a, GABA(B)R1b, GABA(B)R1h (a subunit subtype, lacking a sushi domain, that we have identified in embryonic rat brain), GABA(B)R2, and GABA(B)L transcript levels are not coordinately regulated. Despite the functional requirement for a heterodimer of GABA(B)R subunits, the expression of each subunit mRNA is under independent control during embryonic development, and, by E18, GABA(B)Rs are negatively coupled to adenylyl cyclase in neocortical neurons. The presence of embryonic GABA(B)R transcripts and protein and functional receptor coupling indicates potentially important roles for GABA(B)Rs in modulation of synaptic transmission in the developing embryonic nervous system.

  15. Transporter-associated currents in the gamma-aminobutyric acid transporter GAT-1 are conditionally impaired by mutations of a conserved glycine residue.

    PubMed

    Zhou, Yonggang; Kanner, Baruch I

    2005-05-27

    To determine whether glycine residues play a role in the conformational changes during neurotransmitter transport, we have analyzed site-directed mutants of the gamma-aminobutyric acid (GABA) transporter GAT-1 in a domain containing three consecutive glycines conserved throughout the sodium- and chloride-dependent neurotransmitter transporter family. Only cysteine replacement of glycine 80 resulted in the complete loss of [(3)H]GABA uptake, but oocytes expressing this mutant exhibited the sodium-dependent transient currents thought to reflect a charge-moving conformational change. When sodium was removed and subsequently added back, the transients by G80C did not recover, as opposed to wild type, where recovery was almost complete. Remarkably, the transients by G80C could be restored after exposure of the oocytes to either GABA or a depolarizing pre-pulse. These treatments also resulted in a full recovery of the transients by the wild type. Whereas in wild type lithium leak currents are observed after prior sodium depletion, this was not the case for the glycine 80 mutants unless GABA was added or the oocytes were subjected to a depolarizing pre-pulse. Thus, glycine 80 appears essential for conformational transitions in GAT-1. When this residue is mutated, removal of sodium results in "freezing" the transporter in one conformation from which it can only exit by compensatory changes induced by GABA or depolarization. Our results can be explained by a model invoking two outward-facing states of the empty transporter and a defective transition between these states in the glycine 80 mutants.

  16. Human α1β3γ2L gamma-aminobutyric acid type A receptors: High-level production and purification in a functional state

    PubMed Central

    Dostalova, Zuzana; Zhou, Xiaojuan; Liu, Aiping; Zhang, Xi; Zhang, Yinghui; Desai, Rooma; Forman, Stuart A; Miller, Keith W

    2014-01-01

    Gamma-aminobutyric acid type A receptors (GABAARs) are the most important inhibitory chloride ion channels in the central nervous system and are major targets for a wide variety of drugs. The subunit compositions of GABAARs determine their function and pharmacological profile. GABAARs are heteropentamers of subunits, and (α1)2(β3)2(γ2L)1 is a common subtype. Biochemical and biophysical studies of GABAARs require larger quantities of receptors of defined subunit composition than are currently available. We previously reported high-level production of active human α1β3 GABAAR using tetracycline-inducible stable HEK293 cells. Here we extend the strategy to receptors containing three different subunits. We constructed a stable tetracycline-inducible HEK293-TetR cell line expressing human (N)–FLAG–α1β3γ2L–(C)–(GGS)3GK–1D4 GABAAR. These cells achieved expression levels of 70–90 pmol [3H]muscimol binding sites/15-cm plate at a specific activity of 15–30 pmol/mg of membrane protein. Incorporation of the γ2 subunit was confirmed by the ratio of [3H]flunitrazepam to [3H]muscimol binding sites and sensitivity of GABA-induced currents to benzodiazepines and zinc. The α1β3γ2L GABAARs were solubilized in dodecyl-d-maltoside, purified by anti-FLAG affinity chromatography and reconstituted in CHAPS/asolectin at an overall yield of ∼30%. Typical purifications yielded 1.0–1.5 nmoles of [3H]muscimol binding sites/60 plates. Receptors with similar properties could be purified by 1D4 affinity chromatography with lower overall yield. The composition of the purified, reconstituted receptors was confirmed by ligand binding, Western blot, and proteomics. Allosteric interactions between etomidate and [3H]muscimol binding were maintained in the purified state. PMID:24288268

  17. Modulation of the release of norepinephrine by gamma-aminobutyric acid and morphine in the frontal cerebral cortex of the rat

    SciTech Connect

    Peoples, R.W.

    1989-01-01

    Agents that enhance gamma-aminobutyric acid, or GABA, neurotransmission modulate certain effects of opioids, such as analgesia. Opioid analgesia is mediated in part by norepinephrine in the forebrain. In this study, the interactions between morphine and GABAergic agents on release of ({sup 3}H) norepinephrine from rat frontal cerebral cortical slices were examined. GABA, 5 {times} 10{sup {minus}5}-10{sup {minus}3} M, enhanced potassium stimulated ({sup 3}H) norepinephrine release and reversed the inhibitory effect of morphine in a noncompetitive manner. GABA did not enhance release of ({sup 3}H) norepinephrine stimulated by the calcium ionophore A23187. The effect of GABA was reduced by the GABA{sub A} receptor antagonists bicuculline methiodide or picrotoxin, and by the selective inhibitor of GABA uptake SKF 89976A, but was blocked completely only when bicuculline methiodide and SKF 89976A were used in combination. The GABA{sub A} agonist muscimol, 10{sup {minus}4} M, mimicked the effect of GABA, but the GABA{sub B} agonist ({plus minus})baclofen, 10{sup {minus}4} M, did not affect the release of ({sup 3}H) norepinephrine in the absence or the presence of morphine. Thus GABA appears to produce this effect by stimulating GABA uptake and GABA{sub A}, but not GABA{sub B}, receptors. In contrast to the results that would be predicted for an event involving GABA{sub A} receptors, however, the effect of GABA did not desensitize, and benzodiazepine agonists did not enhance the effect of GABA at any concentration tested between 10{sup {minus}8} and 10{sup {minus}4} M. Thus these receptors may constitute a subclass of GABA{sub A} receptors. These results support a role of GABA uptake and GABA{sub A} receptors in enhancing the release of norepinephrine and modulating its inhibition by opioids in the frontal cortex of the rat.

  18. Propofol induces apoptosis and inhibits the proliferation of rat embryonic neural stem cells via gamma-aminobutyric acid type A receptor.

    PubMed

    Wang, J W; Cheng, W W; Xu, T; Yang, Z Y

    2015-11-23

    We investigated the effect of propofol on the proliferation and viability of rat embryonic neural stem cells (rENSCs) and the potential mechanisms involved. rENSCs were isolated and cultured in vitro and treated with 1, 10, or 50 μM propofol, while the control group was treated with 0.1 μM dimethyl sulfoxide. The effect of propofol on the proliferation and viability of rENSCs was examined by proliferation and apoptosis assays. Real-time polymerase chain reaction was employed to analyze the mRNA expression of checkpoint kinase 1 (Chk1) and p53 in rENSCs exposed to propofol. Immunoprecipitation assay and western blotting analysis were performed to analyze the effect of propofol on Chk1 and p53 activity. The gamma-aminobutyric acid type A (GABAA) receptor antagonist securinine was added to the rENSCs before being treated with propofol to investigate the role of the GABAA receptor in propofol-triggered effects on rENSCs. rENSCs specifically expressing nestin protein were successfully isolated and cultured for experiments. The inhibitory effect of propofol on rENSCs increased dose-dependently. The percentage of apoptotic cells increased to 11.7% and the activity of Chk1 and p53 enhanced after treatment with 50 μM propofol. However, addition of securinine abrogated propofol-induced apoptosis and activation of Chk1. The GABAA receptor mediates propofol-induced apoptosis and proliferation inhibition of rENSCs, possibly by modulating the Chk1/p53 signaling pathway.

  19. Substrate-Na{sup +} complex formation: Coupling mechanism for {gamma}-aminobutyrate symporters

    SciTech Connect

    Pallo, Anna; Simon, Agnes; Bencsura, Akos; Heja, Laszlo; Kardos, Julianna

    2009-07-24

    Crystal structures of transmembrane transport proteins belonging to the important families of neurotransmitter-sodium symporters reveal how they transport neurotransmitters across membranes. Substrate-induced structural conformations of gated neurotransmitter-sodium symporters have been in the focus of research, however, a key question concerning the mechanism of Na{sup +} ion coupling remained unanswered. Homology models of human glial transporter subtypes of the major inhibitory neurotransmitter {gamma}-aminobutyric acid were built. In accordance with selectivity data for subtype 2 vs. 3, docking and molecular dynamics calculations suggest similar orthosteric substrate (inhibitor) conformations and binding crevices but distinguishable allosteric Zn{sup 2+} ion binding motifs. Considering the occluded conformational states of glial human {gamma}-aminobutyric acid transporter subtypes, we found major semi-extended and minor ring-like conformations of zwitterionic {gamma}-aminobutyric acid in complex with Na{sup +} ion. The existence of the minor ring-like conformation of {gamma}-aminobutyric acid in complex with Na{sup +} ion may be attributed to the strengthening of the intramolecular H-bond by the electrostatic effect of Na{sup +} ion. Coupling substrate uptake into cells with the thermodynamically favorable Na{sup +} ion movement through substrate-Na{sup +} ion complex formation may be a mechanistic principle featuring transmembrane neurotransmitter-sodium symporter proteins.

  20. Concordance between isolated cleft palate in mice and alterations within a region including the gene encoding the [beta][sub 3] subunit of the type A [gamma]-aminobutyric acid receptor

    SciTech Connect

    Culiat, C.T.; Stubbs, L.; Nicholls, R.D.; Montgomery, C.S.; Russell, L.B.; Johnson, D.K. ); Rinchik, E.M. Univ. of Florida, Gainesville )

    1993-06-01

    Genetic and molecular analyses of a number of radiation-induced deletion mutations of the pink-eyed dilution (p) locus in mouse chromosome 7 have identified a specific interval on the genetic map associated with a neonatally lethal mutation that results in cleft palate. This interval, closely linked and distal to p, and bracketed by the genes encoding the [alpha][sub 5] and [beta][sub 3] subunits of the type A [gamma]-aminobutyric acid receptor (Gabra5 and Gabrb3, respectively), contains a gene(s) (cp1; cleft palate 1) necessary for normal palate development. The cp1 interval extends from the distal breakpoint of the prenatally lethal p[sup 83FBFo] deletion to the Gabrb3 locus. Among 20 p deletions tested, there was complete concordance between alterations at the Gabrb3 transcription unit and inability to complement the cleft-palate defect. These mapping data, along with previously described in vivo and in vitro teratological effects of [gamma]-aminobutyric acid or its agonists on palate development, suggest the possibility that a particular type A [gamma]-aminobutyric acid receptor that includes the [beta][sub 3] subunit may be necessary for normal palate development. The placement of the cp1 gene within a defined segment of the larger D15S12h (p)-D15S9h-1 interval in the mouse suggests that the highly homologous region of the human genome, 15q11-q13, be evaluated for a role(s) in human fetal facial development. 29 refs., 4 figs., 1 tab.

  1. Use of 3h-. gamma. -aminobutyric acid for transport studies with isolated nerve-terminals from rat brain

    SciTech Connect

    Halvarsson, G.B.; Karlsson, I.; Sellstroem, A.

    1985-07-22

    Isolated synaptosomes were used to study the problem of net accumulation of neurotransmitters. The time-course and the kinetics of exogenous and endogenous GABA transport were studied by liquid-scintillation counting and HPLC-amino acid analysis respectively. Different pools of GABA were suggested by a 6-fold difference in tissue-to-medium-ratio of endogenous vs. exogenous GABA. Net accumulation, exchange and net efflux of GABA was found to be a function of the GABA concentration in the incubation medium. The K/sub m/s for net accumulation and for /sup 3/H-GABA accumulation were 2.68 +/- 1.16 and 6.19 +/- 1.26 ..mu..M respectively, whereas the V/sub max/s were 5.9 +/- 4.9 and 134 +/- 13 pmol/mg w.w min respectively. This means that the transport studies which use exogenous substances (e.g. /sup 3/H-GABA) considerably overestimate the transport by overlooking the magnitude of the counter transport. 22 references, 5 figures, 2 tables.

  2. Structural model for gamma-aminobutyric acid receptor noncompetitive antagonist binding: widely diverse structures fit the same site.

    PubMed

    Chen, Ligong; Durkin, Kathleen A; Casida, John E

    2006-03-28

    Several major insecticides, including alpha-endosulfan, lindane, and fipronil, and the botanical picrotoxinin are noncompetitive antagonists (NCAs) for the GABA receptor. We showed earlier that human beta(3) homopentameric GABA(A) receptor recognizes all of the important GABAergic insecticides and reproduces the high insecticide sensitivity and structure-activity relationships of the native insect receptor. Despite large structural diversity, the NCAs are proposed to fit a single binding site in the chloride channel lumen lined by five transmembrane 2 segments. This hypothesis is examined with the beta(3) homopentamer by mutagenesis, pore structure studies, NCA binding, and molecular modeling. The 15 amino acids in the cytoplasmic half of the pore were mutated to cysteine, serine, or other residue for 22 mutants overall. Localization of A-1'C, A2'C, T6'C, and L9'C (index numbers for the transmembrane 2 region) in the channel lumen was established by disulfide cross-linking. Binding of two NCA radioligands [(3)H]1-(4-ethynylphenyl)-4-n-propyl-2,6,7-trioxabicyclo[2.2.2]octane and [(3)H] 3,3-bis-trifluoromethyl-bicyclo[2,2,1]heptane-2,2-dicarbonitrile was dramatically reduced with 8 of the 15 mutated positions, focusing attention on A2', T6', and L9' as proposed binding sites, consistent with earlier mutagenesis studies. The cytoplasmic half of the beta3 homopentamer pore was modeled as an alpha-helix. The six NCAs listed above plus t-butylbicyclophosphorothionate fit the 2' to 9' pore region forming hydrogen bonds with the T6' hydroxyl and hydrophobic interactions with A2', T6', and L9' alkyl substituents, thereby blocking the channel. Thus, widely diverse NCA structures fit the same GABA receptor beta subunit site with important implications for insecticide cross-resistance and selective toxicity between insects and mammals.

  3. Potentiation by sevoflurane of the gamma-aminobutyric acid-induced chloride current in acutely dissociated CA1 pyramidal neurones from rat hippocampus.

    PubMed Central

    Wu, J.; Harata, N.; Akaike, N.

    1996-01-01

    1. The effects of a new kind of volatile anaesthetic, sevoflurane (Sev), on gamma-aminobutyric acid (GABA)-gated chloride current (Icl) in single neurones dissociated from the rat hippocampal CA1 area were examined using the nystatin perforated patch recording configuration under the voltage-clamp condition. All drugs were applied with a rapid perfusion system, termed the "Y-tube' method. 2. When the concentrations were higher than 3 x 10(-4) M, Sev, itself, induced an inward current (ISev) at a holding potential (VH) of -40 mV. The concentration-response curve of ISev was bell-shaped, with a suppressed peak and plateau currents at high concentrations (above 2 x 10(-3) M). The reversal potential of ISev (ESev) was close to the theoretical Cl- equilibrium potential, indicating that ISev was carried mainly by Cl-. 3. ISev was reversibly blocked by bicuculline (Bic), an antagonist of the GABAA receptor, in a concentration-dependent manner with a half-inhibitory concentration (IC50) of 7.2 x 10(-7) M. But ISev was insensitive to strychnine (Str), an antagonist of the glycine receptor. 4. At low concentrations (between 3 x 10(-4) and 10(-3) M), Sev markedly enhanced the 10(-6) M GABA induced current (IGABA) but reduced the IGABA with accelerating desensitization accompanied by a "hump' current after washout at high concentrations (higher than 2 x 10(-3) M). 5. Sev, 10(-3) M potentiated the current induced by low concentrations of GABA (between 10(-7) and 3 x 10(-6) M) but reduced the current induced by high concentrations (higher than 10(-5) M) of GABA with a clear acceleration of IGABA desensitization. 6. Sev, like pentobarbitone (PB), pregnanolone (PGN) or diazepam (DZP), potentiated the 10(-6) M GABA-induced response without shifting the reversal potential of IGABA. 7. ISev was augmented by PB, PGN, or DZP at concentrations that maximally potentiated IGABA, suggesting that Sev enhanced IGABA at a binding site distinct from that for PB, PGN, or DZP. 8. It is concluded

  4. Specific gamma-aminobutyrate chemotaxis in pseudomonads with different lifestyle.

    PubMed

    Reyes-Darias, Jose Antonio; García, Vanina; Rico-Jiménez, Miriam; Corral-Lugo, Andrés; Lesouhaitier, Olivier; Juárez-Hernández, Dalia; Yang, Yiling; Bi, Shuangyu; Feuilloley, Marc; Muñoz-Rojas, Jesús; Sourjik, Victor; Krell, Tino

    2015-08-01

    The PctC chemoreceptor of Pseudomonas aeruginosa mediates chemotaxis with high specificity to gamma-aminobutyric acid (GABA). This compound is present everywhere in nature and has multiple functions, including being a human neurotransmitter or plant signaling compound. Because P. aeruginosa is ubiquitously distributed in nature and able to infect and colonize different hosts, the physiological relevance of GABA taxis is unclear, but it has been suggested that bacterial attraction to neurotransmitters may enhance virulence. We report the identification of McpG as a specific GABA chemoreceptor in non-pathogenic Pseudomonas putida KT2440. As with PctC, GABA was found to bind McpG tightly. The analysis of chimeras comprising the PctC and McpG ligand-binding domains fused to the Tar signaling domain showed very high GABA sensitivities. We also show that PctC inactivation does not alter virulence in Caenorhabditis elegans. Significant amounts of GABA were detected in tomato root exudates, and deletion of mcpG reduced root colonization that requires chemotaxis through agar. The C. elegans data and the detection of a GABA receptor in non-pathogenic species indicate that GABA taxis may not be related to virulence in animal systems but may be of importance in the context of colonization and infection of plant roots by soil-dwelling pseudomonads.

  5. Interactive effects of glutamine and gamma-aminobutyric acid on growth performance and skeletal muscle amino acid metabolism of 22-42-day-old broilers exposed to hot environment

    NASA Astrophysics Data System (ADS)

    Hu, Hong; Bai, Xi; Shah, Assar Ali; Dai, Sifa; Wang, Like; Hua, Jinling; Che, Chuanyan; He, Shaojun; Wen, Aiyou; Jiang, Jinpeng

    2016-06-01

    The present experiment was conducted to investigate the interactive effects between dietary glutamine (Gln, 0 and 5 g/kg) and gamma-aminobutyric acid (GABA, 0 and 100 mg/kg) on growth performance and amino acid (AA) metabolism of broilers under hot environment. A total of 360 22-day-old Arbor Acres male chickens were randomly assigned to five treatment groups under thermoneutral chamber (PC, 23 °C) and cyclic heat stress (HS, 30-34 °C cycling) conditions. Compared with the PC group, cyclic HS decreased ( P < 0.05) daily weight gain (DWG), daily feed consumption (DFC), the concentrations of Gln, glutamate (Glu), and GABA, and the activities of glutaminase and glutamic acid decarboxylase (GAD) in breast muscle at 28, 35, and 42 days, while it increased ( P < 0.05) the activities of glutamine synthetase (GS) and gamma-aminobutyric acid transaminase (GABA-T) at 28, 35, and 42 days. Dietary Gln and GABA improved ( P < 0.05) DWG and DFC of broilers under cyclic HS during 28-42 days. In breast muscle, the Gln supplementation increased ( P < 0.05) the concentrations of Gln (28, 35, and 42 days), Glu (28, 35, and 42 days), and GABA (42 days) and the activities of glutaminase (28, 35, and 42 days) and GAD (28, 35, and 42 days) but decreased ( P < 0.05) GS activities at 28, 35, and 42 days and GABA-T activities at 28 days. The addition of GABA increased ( P < 0.05) the concentrations of Gln and Glu and activities of glutaminase and GAD, while it decreased ( P < 0.05) GABA-T activities at 28, 35, and 42 days. Significant interactions ( P < 0.05) between Gln and GABA were found on breast skeletal muscle Gln concentrations, glutaminase activities, GS activities at 28 and 35 days, and DWG, GABA concentrations, and GABA-T activities at 28, 35, and 42 days in broilers under cyclic HS. In conclusion, the present results indicated that the interactions of exogenous Gln and GABA could offer a potential nutritional strategy to prevent HS-related depression in skeletal muscle Gln and

  6. A novel, rapid, inhibitory effect of insulin on alpha1beta2gamma2s gamma-aminobutyric acid type A receptors.

    PubMed

    Williams, Daniel B

    2008-09-26

    In the CNS, GABA and insulin seem to contribute to similar processes, including neuronal survival; learning and reward; and energy balance and food intake. It is likely then that insulin and GABA may interact, perhaps at the GABA(A) receptor. One such interaction has already been described [Q. Wan, Z.G. Xiong, H.Y. Man, C.A. Ackerley, J. Braunton, W.Y. Lu, L.E. Becker, J.F. MacDonald, Y.T. Wang, Recruitment of functional GABA(A) receptors to postsynaptic domains by insulin, Nature 388 (1997) 686-690]; in it a micromolar concentration of insulin causes the insertion of GABA(A) receptors into the cell membrane, increasing GABA current. I have discovered another effect of insulin on GABA(A) currents. Using a receptor isoform, alpha(1)beta(2)gamma(2s) that is the likely main neuronal GABA(A) isoform expressed recombinantly in Xenopus oocytes, insulin inhibits GABA-induced current when applied simultaneously with low concentrations of GABA. Insulin will significantly inhibit currents induced by EC(30-50) concentrations of GABA by about 38%. Insulin is potent in this effect; IC(50) of insulin was found to be about 4.3 x 10(-10) M. The insulin effect on the GABA dose responses looked like that of an antagonist similar to bicuculline or beta-carbolines. However, an effect of phosphorylation on the GABA(A) receptor from the insulin receptor signal transduction pathway cannot yet be dismissed.

  7. Dual mechanisms regulating glutamate decarboxylases and accumulation of gamma-aminobutyric acid in tea (Camellia sinensis) leaves exposed to multiple stresses.

    PubMed

    Mei, Xin; Chen, Yiyong; Zhang, Lingyun; Fu, Xiumin; Wei, Qing; Grierson, Don; Zhou, Ying; Huang, Yahui; Dong, Fang; Yang, Ziyin

    2016-01-01

    γ-Aminobutyric acid (GABA) is one of the major inhibitory neurotransmitters in the central nervous system. It has multiple positive effects on mammalian physiology and is an important bioactive component of tea (Camellia sinensis). GABA generally occurs at a very low level in plants but GABA content increases substantially after exposure to a range of stresses, especially oxygen-deficiency. During processing of tea leaves, a combination of anoxic stress and mechanical damage are essential for the high accumulation of GABA. This is believed to be initiated by a change in glutamate decarboxylase activity, but the underlying mechanisms are unclear. In the present study we characterized factors regulating the expression and activity of three tea glutamate decarboxylase genes (CsGAD1, 2, and 3), and their encoded enzymes. The results suggests that, unlike the model plant Arabidopsis thaliana, there are dual mechanisms regulating the accumulation of GABA in tea leaves exposed to multiple stresses, including activation of CsGAD1 enzymatic activity by calmodulin upon the onset of the stress and accumulation of high levels of CsGAD2 mRNA induced by a combination of anoxic stress and mechanical damage. PMID:27021285

  8. Dual mechanisms regulating glutamate decarboxylases and accumulation of gamma-aminobutyric acid in tea (Camellia sinensis) leaves exposed to multiple stresses

    PubMed Central

    Mei, Xin; Chen, Yiyong; Zhang, Lingyun; Fu, Xiumin; Wei, Qing; Grierson, Don; Zhou, Ying; Huang, Yahui; Dong, Fang; Yang, Ziyin

    2016-01-01

    γ-Aminobutyric acid (GABA) is one of the major inhibitory neurotransmitters in the central nervous system. It has multiple positive effects on mammalian physiology and is an important bioactive component of tea (Camellia sinensis). GABA generally occurs at a very low level in plants but GABA content increases substantially after exposure to a range of stresses, especially oxygen-deficiency. During processing of tea leaves, a combination of anoxic stress and mechanical damage are essential for the high accumulation of GABA. This is believed to be initiated by a change in glutamate decarboxylase activity, but the underlying mechanisms are unclear. In the present study we characterized factors regulating the expression and activity of three tea glutamate decarboxylase genes (CsGAD1, 2, and 3), and their encoded enzymes. The results suggests that, unlike the model plant Arabidopsis thaliana, there are dual mechanisms regulating the accumulation of GABA in tea leaves exposed to multiple stresses, including activation of CsGAD1 enzymatic activity by calmodulin upon the onset of the stress and accumulation of high levels of CsGAD2 mRNA induced by a combination of anoxic stress and mechanical damage. PMID:27021285

  9. A high-throughput functional assay for characterization of gamma-aminobutyric acid(A) channel modulators using cryopreserved transiently transfected cells.

    PubMed

    Liu, Jay; Chen, Tongming; Norris, Tyrell; Knappenberger, Kathy; Huston, Julie; Wood, Michael; Bostwick, Robert

    2008-12-01

    The ionotropic -aminobutyric acid (GABA)A receptors are an important family of drug targets for a variety of neurological and psychiatric disorders. Selective modulation of certain subtypes of the receptor could lead to novel or improved therapies. However, the discovery of subtype-selective compounds has been hampered by the lack of a high-throughput functional assay and the difficulty in establishing stable cell lines expressing GABAA receptor in a proper subunit composition. To meet drug discovery need we developed a fluorescent imaging plate reader(FLIPR)-based membrane potential assay with sufficient robustness and reproducibility for use in a high-throughput format. Two major subtypes of GABAA receptor were used: GABAA1 and GABAA2, which are composed of (alpha1)2(beta2)2gama2 and (alpha1)2(beta3)2gama2, respectively. We expressed the receptors by transiently co-transfecting cells with the three subunit DNAs in separate constructs, and by controlling the ratio of the DNA amount for each subunit transfected we forced the cells to express GABAA receptors in a pharmacologically relevant form. A large batch of transfected human embryonic kidney 293 cells were cryopreserved and used to screen and evaluate GABAA modulators.In these cells, agonist activation of GABAA receptor resulted in Cl- efflux and membrane depolarization, which was detected by FLIPR as an increase in fluorescence signal. Based on our characterization of several known GABAA modulators and a test set of compounds known to bind to the GABAA benzodiazepine site, we have demonstrated the validity and utility of this assay for discovery of novel pharmacological agents acting at GABAA receptors.

  10. In vivo simultaneous monitoring of gamma-aminobutyric acid, glutamate, and L-aspartate using brain microdialysis and capillary electrophoresis with laser-induced fluorescence detection: Analytical developments and in vitro/in vivo validations.

    PubMed

    Sauvinet, Valérie; Parrot, Sandrine; Benturquia, Nadia; Bravo-Moratón, Eva; Renaud, Bernard; Denoroy, Luc

    2003-09-01

    gamma-Aminobutyric acid (GABA), glutamate (Glu), and L-aspartate (L-Asp) are three major amino acid neurotransmitters in the central nervous system. In this work, a method for the separation of these three neurotransmitters in brain microdialysis samples using a commercially available capillary electrophoresis (CE) system has been developed. Molecules were tagged on their primary amine function with the fluorogene agent naphthalene-2,3-dicarboxaldehyde (NDA), and, after separation by micellar electrokinetic chromatography, were detected by laser-induced fluorescence using a 442 nm helium-cadmium laser. The separation conditions for the analysis of derivatized neurotransmitters in standard solutions and microdialysates have been optimized, and this method has been validated on both pharmacological and analytical basis. The separation of GABA, Glu, and L-Asp takes less than 10 min by using a 75 mmol/L borate buffer, pH 9.2, containing 70 mmol/L SDS and 10 mmol/L hydroxypropyl-beta-cyclodextrin and + 25 kV voltage. The detection limits were 3, 15 nmol/L and, 5 nmol/L for GABA, Glu, and L-Asp, respectively. Moreover, submicroliter samples can be analyzed. This method allows a simple, rapid and accurate measurement of the three amino acid neurotransmitters for the in vivo brain monitoring using microdialysis sampling.

  11. [Effect of vitamin B3-active compounds on the content of free and combined gamma-aminobutyric acid and glutamic acid in the brain of mice].

    PubMed

    Rozanov, V A; Reĭtarova, T E

    1983-01-01

    The bound and free GABA and glutamic acid content in the brain of F1 (CBA X C57B1/6) hybrid mice was investigated by the Eliott method. A tendency to a decrease of GABA and glutamate content in the brain with their practically constant bound/free ratio was observed 24 h after calcium-D-pantothenate injections (150 mumole/kg, 9 injections for 3 days). Calcium-D-homopantothenate injected in the same way caused a significant decrease in the GABA content, and a sharp drop of the bound/free GABA ratio. The effect is not associated with the influence of calcium ions in the composition of the injected compounds. PMID:6140785

  12. The human [gamma]-aminobutyric acid receptor subunit [beta]3 and [alpha]5 gene cluster in chromosome 15q11-q13 is rich in highly polymorphic (CA)[sub n] repeats

    SciTech Connect

    Glatt, K.; Lalande, M. ); Sinnett, D. )

    1994-01-01

    The [gamma]-aminobutyric acid (GABA[sub A]) receptor [beta]33 (GABRB3) and [alpha]5 (GABRA5) subunit genes have been localized to the Angelman and Prader-Willi syndrome region of chromosome 15q11-q13. GABRB3, which encompasses 250 kb, is located 100 kb proximal of GABRA5, with the two genes arranged in head-to-head transcriptional orientation. In screening 135 kb of cloned DNA within a 260-kb interval extending from within GABRB3 to the 5[prime] end of GABRA5, 10 new (CA), repeats have been identified. Five of these have been analyzed in detail and found to be highly polymorphic, with the polymorphism information content (PIC) ranging from 0.7 to 0.85 and with heterozygosities of 67 to 94%. In the clones from GABRB3/GABRA5 region, therefore, the frequency of (CA)[sub n] with PICs [ge] 0.7 is 1 per 27 kb. Previous estimates of the density of (CA)[sub n] with PICs [ge] 0.7 in the human genome have been approximately 10-fold lower. The GABRB3/GABRA5 region appears, therefore, to be enriched for highly informative (CA)[sub n]. This set of closely spaced, short tandem repeat polymorphisms will be useful in the molecular analyses of Prader-Willi and Angelman syndromes and in high-resolution studies of genetic recombination within this region. 21 refs., 2 figs., 1 tab.

  13. Antagonistic properties of a natural product - Bicuculline with the gamma-aminobutyric acid receptor: Studied through electrostatic potential mapping, electronic and vibrational spectra using ab initio and density functional theory

    NASA Astrophysics Data System (ADS)

    Srivastava, Anubha; Tandon, Poonam; Jain, Sudha; Asthana, B. P.

    2011-12-01

    (+)-Bicuculline (hereinafter referred to as bicuculline), a phthalide isoquinoline alkaloid is of current interest as an antagonist of gamma-aminobutyric acid (GABA). Its inhibitor properties have been studied through molecular electrostatic potential (MEP) mapping of this molecule and GABA receptor. The hot site on the potential surface of bicuculline, which is also isosteric with GABA receptor, has been used to interpret the inhibitor property. A systematic quantum chemical study of the possible conformations, their relative stabilities, FT-Raman, FT-IR and UV-vis spectroscopic analysis of bicuculline has been reported. The optimized geometries, wavenumber and intensity of the vibrational bands of all the conformers of bicuculline have been calculated using ab initio Hartree-Fock (HF) and density functional theory (DFT) employing B3LYP functional and 6-311G(d,p) basis set. Mulliken atomic charges, HOMO-LUMO gap Δ E, ionization potential, dipole moments and total energy have also been obtained for the optimized geometries of both the molecules. TD-DFT method is used to calculate the electronic absorption parameters in gas phase as well as in solvent environment using integral equation formalism-polarizable continuum model (IEF-PCM) employing 6-31G basis set and the results thus obtained are compared with the UV absorption spectra. The combination of experimental and calculated results provides an insight into the structural and vibrational spectroscopic properties of bicuculline.

  14. Perinatal exposure to germinated brown rice and its gamma amino-butyric acid-rich extract prevents high fat diet-induced insulin resistance in first generation rat offspring

    PubMed Central

    Adamu, Hadiza Altine; Imam, Mustapha Umar; Ooi, Der-Jiun; Esa, Norhaizan Mohd; Rosli, Rozita; Ismail, Maznah

    2016-01-01

    Background Evidence suggests perinatal environments influence the risk of developing insulin resistance. Objective The present study was aimed at determining the effects of intrauterine exposure to germinated brown rice (GBR) and GBR-derived gamma (γ) aminobutyric acid (GABA) extract on epigenetically mediated high fat diet–induced insulin resistance. Design Pregnant Sprague Dawley rats were fed high-fat diet (HFD), HFD+GBR, or HFD+GABA throughout pregnancy until 4 weeks postdelivery. The pups were weighed weekly and maintained on normal pellet until 8 weeks postdelivery. After sacrifice, biochemical markers of obesity and insulin resistance including oral glucose tolerance test, adiponectin, leptin, and retinol binding protein-4 (RBP4) were measured. Hepatic gene expression changes and the global methylation and histone acetylation levels were also evaluated. Results Detailed analyses revealed that mothers given GBR and GABA extract, and their offspring had increased adiponectin levels and reduced insulin, homeostasis model assessment of insulin resistance, leptin, oxidative stress, and RBP4 levels, while their hepatic mRNA levels of GLUT2 and IPF1 were increased. Furthermore, GBR and GABA extract lowered global DNA methylation levels and modulated H3 and H4 acetylation levels. Conclusions These results showed that intrauterine exposure to GBR-influenced metabolic outcomes in offspring of rats with underlying epigenetic changes and transcriptional implications that led to improved glucose homeostasis. PMID:26842399

  15. Cloning, Expression Analysis, and Molecular Modeling of the Gamma-Aminobutyric Acid Receptor Alpha2 Subunit Gene from the Common Cutworm, Spodoptera litura

    PubMed Central

    Zuo, Hongliang; Gao, Lu; Hu, Zhen; Liu, Haiyuan; Zhong, Guohua

    2013-01-01

    Intensive research on the molecule structures of the gamma-nminobutyric acid (GABA) receptor in agricultural pests has great significance to the mechanism investigation, resistance prevention, and molecular design of novel pesticides. The GABA receptor a2 (SlGABARα2) subunit gene in Spodoptera litura (Fabricius) (Lepidoptera: Noctuidae) was cloned using the technologies of reverse transcription PCR and rapid amplification of cDNA ends. The gemonic DNA sequence of SlGABARα2 has 5164 bp with 8 exons and 7 introns that were in accordance with the GT-AG splicing formula. The complete mRNA sequence of SlGABARα2 was 1965 bp, with an open reading frame of 1500 bp encoding a protein of 499 amino acids. The GABA receptor is highly conserved among insects. The conserved regions include several N-glycosylation, Oglycosylation, and phosphorylation sites, as well as 4 transmembrane domains. The identities that SlGABARα2 shared with the GABA receptor a2 subunit of Spodoptera exigua, Heliothis virescens, Chilo suppressalis, Plutella xylostella, Bombyx mori ranged from 99.2% to 87.2% at the amino acid level. The comparative 3-dimensional model of SlGABARα2 showed that its tertiary structure was composed of 4 major α-helixes located at the 4 putative transmembrane domains on one side, with some β-sheets and 1 small α-helix on the other side. SlGABARα2 may be attached to the membrane by 4 α-helixes that bind ions in other conserved domains to transport them through the membrane. The results of quantitative real time PCR demonstrated that SlGABARα2 was expressed in all developmental stages of S. litura. The relative expression level of SlGABARα2 was the lowest in eggs and increased with larval growth, while it declined slightly in pupae and reached the peak in adults. The expressions of SlGABARα2 in larvae varied among different tissues; it was extremely high in the brain but was low in the midgut, epicuticle, Malpighian tube, and fat body. PMID:23909412

  16. Cloning, expression analysis, and molecular modeling of the gamma-aminobutyric acid receptor alpha2 subunit gene from the common cutworm, Spodoptera litura.

    PubMed

    Zuo, Hongliang; Gao, Lu; Hu, Zhen; Liu, Haiyuan; Zhong, Guohua

    2013-01-01

    Intensive research on the molecule structures of the gamma-nminobutyric acid (GABA) receptor in agricultural pests has great significance to the mechanism investigation, resistance prevention, and molecular design of novel pesticides. The GABA receptor a2 (SlGABARα2) subunit gene in Spodoptera litura (Fabricius) (Lepidoptera: Noctuidae) was cloned using the technologies of reverse transcription PCR and rapid amplification of cDNA ends. The gemonic DNA sequence of SlGABARα2 has 5164 bp with 8 exons and 7 introns that were in accordance with the GT-AG splicing formula. The complete mRNA sequence of SlGABARα2 was 1965 bp, with an open reading frame of 1500 bp encoding a protein of 499 amino acids. The GABA receptor is highly conserved among insects. The conserved regions include several N-glycosylation, Oglycosylation, and phosphorylation sites, as well as 4 transmembrane domains. The identities that SlGABARα2 shared with the GABA receptor a2 subunit of Spodoptera exigua, Heliothis virescens, Chilo suppressalis, Plutella xylostella, Bombyx mori ranged from 99.2% to 87.2% at the amino acid level. The comparative 3-dimensional model of SlGABARα2 showed that its tertiary structure was composed of 4 major α-helixes located at the 4 putative transmembrane domains on one side, with some β-sheets and 1 small α-helix on the other side. SlGABARα2 may be attached to the membrane by 4 α-helixes that bind ions in other conserved domains to transport them through the membrane. The results of quantitative real time PCR demonstrated that SlGABARα2 was expressed in all developmental stages of S. litura. The relative expression level of SlGABARα2 was the lowest in eggs and increased with larval growth, while it declined slightly in pupae and reached the peak in adults. The expressions of SlGABARα2 in larvae varied among different tissues; it was extremely high in the brain but was low in the midgut, epicuticle, Malpighian tube, and fat body. PMID:23909412

  17. Invasive ability of human renal cell carcinoma cell line Caki-2 is accelerated by gamma-aminobutyric acid, via sustained activation of ERK1/2 inducible matrix metalloproteinases.

    PubMed

    Inamoto, Teruo; Azuma, Haruhito; Sakamoto, Takeshi; Kiyama, Satoshi; Ubai, Takanobu; Kotake, Yatsugu; Watanabe, Masahito; Katsuoka, Yoji

    2007-10-01

    Gamma-aminobutyric acid (GABA) was first discovered as an inhibitory neurotransmitter in the central nervous system (CNS) and has been reported to have a variety of functions, including regulation of cell division, cell differentiation and maturation, and to be involved in the development of certain cancers outside the CNS. In the present study, using the human renal cell carcinoma cell line Caki-2, we demonstrated that GABA stimulation significantly increased the expression of MMP-2 and -9 and subsequently increased the invasive activity of the cancer cells. Because MAPK signaling is one of the key regulators of MMP expression, we further evaluated MAPK signaling after stimulation with GABA. It was found that GABA stimulation promoted the phosphorylation of MAPKs, including ERK1/2, JNK, and p38. ERK1/2 phosphorylation was sustained for up to 12 h, while phosphorylation of JNK and p38 returned to the endogenous level by 30 min. It was noteworthy that the ras/raf/MEK/ERK pathway inhibitor PD98059 attenuated GABA-induced MMP-9 expression and that both PD98059 and MMP inhibitors attenuated the GABA-induced invasive activity of Caki-2 cells. Moreover, data obtained by depletion of the MEK/ERK pathway using interfering RNA transfection of Caki-2 cells clearly corroborated the above results, as both MMP-9 expression and GABA-induced invasive ability were decreased significantly. We also demonstrated that the GABA-induced increase in invasive ability via ERK1/2 up-regulation was mediated mainly through the GABA-B receptor. These results indicate that GABA stimulation promotes cancer cell invasion and that the effect is partly due to ERK1/2-dependent up-regulation of MMPs.

  18. Functional modulation of cerebral gamma-aminobutyric acidA receptor/benzodiazepine receptor/chloride ion channel complex with ethyl beta-carboline-3-carboxylate: Presence of independent binding site for ethyl beta-carboline-3-carboxylate

    SciTech Connect

    Taguchi, J.; Kuriyama, K. )

    1990-05-01

    Effect of ethyl beta-carboline-3-carboxylate (beta-CCE) on the function of gamma-aminobutyric acid (GABA)A receptor/benzodiazepine receptor/chloride ion channel complex was studied. Beta-CCE noncompetitively and competitively inhibited (3H)flunitrazepam binding to benzodiazepine receptor, but not (3H)muscimol binding to GABAA receptor as well as t-(3H)butylbicycloorthobenzoate (( 3H) TBOB) binding to chloride ion channel, in particulate fraction of the mouse brain. Ro15-1788 also inhibited competitively (3H) flunitrazepam binding. On the other hand, the binding of beta-(3H)CCE was inhibited noncompetitively and competitively by clonazepam and competitively by Ro15-1788. In agreement with these results, benzodiazepines-stimulated (3H)muscimol binding was antagonized by beta-CCE and Ro15-1788. Gel column chromatography for the solubilized fraction from cerebral particulate fraction by 0.2% sodium deoxycholate (DOC-Na) in the presence of 1 M KCl indicated that beta-(3H)CCE binding site was eluted in the same fraction (molecular weight, 250,000) as the binding sites for (3H)flunitrazepam, (3H)muscimol and (3H)TBOB. GABA-stimulated 36Cl- influx into membrane vesicles prepared from the bovine cerebral cortex was stimulated and attenuated by flunitrazepam and beta-CCE, respectively. These effects of flunitrazepam and beta-CCE on the GABA-stimulated 36Cl- influx were antagonized by Ro15-1788. The present results suggest that the binding site for beta-CCE, which resides on GABAA receptor/benzodiazepine receptor/chloride ion channel complex, may be different from that for benzodiazepine. Possible roles of beta-CCE binding site in the allosteric inhibitions on benzodiazepine binding site as well as on the functional coupling between chloride ion channel and GABAA receptor are also suggested.

  19. Immunohistochemical evidence for colocalization of gamma-aminobutyric acid and serotonin in neurons of the ventral medulla oblongata projecting to the spinal cord.

    PubMed

    Millhorn, D E; Hökfelt, T; Seroogy, K; Oertel, W; Verhofstad, A A; Wu, J Y

    1987-04-28

    Fluorescence immunohistochemistry was used to analyze the medulla oblongata of colchicine-treated rats that had been incubated with guinea pig antibodies to serotonin (5-HT) and either rabbit or sheep antibodies to glutamic acid decarboxylase (GAD). Numerous cells in the rostral ventrolateral medulla in the region of nucleus raphe magnus were immunostained for either 5-HT or GAD. A substantial number of neurons showed positive immunoreactivity for both substances, and were most frequently observed in the lateral aspect of nucleus raphe magnus. In addition, a number of the 5-HT/GAD-containing neurons were retrogradely labelled with Fast blue dye that had been injected into the thoracic spinal cord. This work provides evidence for colocalization of the classical neurotransmitters 5-HT and GABA in single cells of the ventral medulla oblongata, some of which project to the spinal cord. PMID:3555707

  20. Effects of NaCl Replacement with Gamma-Aminobutyric acid (GABA) on the Quality Characteristics and Sensorial Properties of Model Meat Products

    PubMed Central

    Chun, Ji-Yeon; Cho, Hyung-Yong; Min, Sang-Gi

    2014-01-01

    This study investigated the effects of γ-aminobutylic acid (GABA) on the quality and sensorial properties of both the GABA/NaCl complex and model meat products. GABA/NaCl complex was prepared by spray-drying, and the surface dimensions, morphology, rheology, and saltiness were characterized. For model meat products, pork patties were prepared by replacing NaCl with GABA. For characteristics of the complex, increasing GABA concentration increased the surface dimensions of the complex. However, GABA did not affect the rheological properties of solutions containing the complex. The addition of 2% GABA exhibited significantly higher saltiness than the control (no GABA treatment). In the case of pork patties, sensory testing indicated that the addition of GABA decreased the saltiness intensity. Both the intensity of juiciness and tenderness of patties containing GABA also scored lower than the control, based on the NaCl reduction. These results were consistent with the quality characteristics (cooking loss and texture profile analysis). Nevertheless, overall acceptability of the pork patties showed that up to 1.5%, patties containing GABA did not significantly differ from the control. Consequently, the results indicated that GABA has a potential application in meat products, but also manifested a deterioration of quality by the NaCl reduction, which warrants further exploration. PMID:26761294

  1. The effects of lowered extracellular sodium on gamma-aminobutyric acid (GABA)-induced currents of Muller (glial) cells of the skate retina.

    PubMed

    Qian, H; Malchow, R P; Ripps, H

    1993-04-01

    1. The effects of external sodium on GABA-induced chloride currents were examined with whole-cell voltage-clamp recordings obtained from enzymatically dissociated solitary Muller cells in culture. Our goal was to determine whether a sodium-dependent GABA uptake mechanism influences the GABAa-mediated responses of skate Muller cells. 2. At low concentrations of GABA (0.01 to 0.5 microM), removal of sodium from the external solution resulted in a marked increase in the ligand-gated currents mediated by activation of GABAa receptors. The enhancement by lowered sodium was greatest at hyperpolarizing potentials and decreased progressively as the cell was depolarized. 3. The reversal potential for the GABA-induced response was not significantly altered by the removal of sodium, suggesting that sodium ions did not directly contribute to the GABAa-mediated current. 4. Lowering external sodium had no effect on the currents induced by the GABAa-agonist muscimol, consistent with its much lower affinity for the GABA transport carrier. 5. Application of the GABA uptake blocker nipecotic acid also abolished the effects of lowered sodium. 6. These findings suggest that the effects of lowered external sodium resulted from a decrease in the uptake of GABA into the Muller cells, thus raising the effective concentration of GABA acting upon the GABAa receptors. PMID:8394215

  2. DNA Methylation at the Neonatal State and at the Time of Diagnosis: Preliminary Support for an Association with the Estrogen Receptor 1, Gamma-Aminobutyric Acid B Receptor 1, and Myelin Oligodendrocyte Glycoprotein in Female Adolescent Patients with OCD.

    PubMed

    Nissen, Judith Becker; Hansen, Christine Søholm; Starnawska, Anna; Mattheisen, Manuel; Børglum, Anders Dupont; Buttenschøn, Henriette Nørmølle; Hollegaard, Mads

    2016-01-01

    Obsessive-compulsive disorder (OCD) is a neuropsychiatric disorder. Non-genetic factors and their interaction with genes have attracted increasing attention. Epigenetics is regarded an important interface between environmental signals and activation/repression of genomic responses. Epigenetic mechanisms have not previously been examined in OCD in children and adolescents. The aim of the present study was to examine the DNA methylation profile of selected genes in blood spots from neonates later diagnosed with OCD and in the same children/adolescents at the time of diagnosis compared with age- and sex-matched controls. Furthermore, we wanted to characterize the association of the differential methylation profiles with the severity of OCD and treatment outcome. Dried and new blood spot samples were obtained from 21 female children/adolescents with verified OCD and 12 female controls. The differential methylation was analyzed using a linear model and the correlation with the severity of OCD and treatment outcome was analyzed using the Pearson correlation. We evaluated selected Illumina Infinium HumanMethylation450 BeadChip probes within and up to 100,000 bp up- and downstream of 14 genes previously associated with OCD (SLC1A1, SLC25A12, GABBR1, GAD1, DLGAP1, MOG, BDNF, OLIG2, NTRK2 and 3, ESR1, SL6A4, TPH2, and COMT). The study found no significantly differential methylation. However, preliminary support for a difference was found for the gamma-aminobutyric acid (GABA) B receptor 1 (cg10234998, cg17099072) in blood samples at birth and for the estrogen receptor 1 (ESR1) (cg10939667), the myelin oligodendrocyte glycoprotein (MOG) (cg16650906), and the brain-derived neurotrophic factor (BDNF) (cg14080521) in blood samples at the time of diagnosis. Preliminary support for an association was observed between the methylation profiles of GABBR1 and MOG and baseline severity, treatment effect, and responder status; and between the methylation profile of ESR1 and baseline

  3. DNA Methylation at the Neonatal State and at the Time of Diagnosis: Preliminary Support for an Association with the Estrogen Receptor 1, Gamma-Aminobutyric Acid B Receptor 1, and Myelin Oligodendrocyte Glycoprotein in Female Adolescent Patients with OCD

    PubMed Central

    Nissen, Judith Becker; Hansen, Christine Søholm; Starnawska, Anna; Mattheisen, Manuel; Børglum, Anders Dupont; Buttenschøn, Henriette Nørmølle; Hollegaard, Mads

    2016-01-01

    Obsessive–compulsive disorder (OCD) is a neuropsychiatric disorder. Non-genetic factors and their interaction with genes have attracted increasing attention. Epigenetics is regarded an important interface between environmental signals and activation/repression of genomic responses. Epigenetic mechanisms have not previously been examined in OCD in children and adolescents. The aim of the present study was to examine the DNA methylation profile of selected genes in blood spots from neonates later diagnosed with OCD and in the same children/adolescents at the time of diagnosis compared with age- and sex-matched controls. Furthermore, we wanted to characterize the association of the differential methylation profiles with the severity of OCD and treatment outcome. Dried and new blood spot samples were obtained from 21 female children/adolescents with verified OCD and 12 female controls. The differential methylation was analyzed using a linear model and the correlation with the severity of OCD and treatment outcome was analyzed using the Pearson correlation. We evaluated selected Illumina Infinium HumanMethylation450 BeadChip probes within and up to 100,000 bp up- and downstream of 14 genes previously associated with OCD (SLC1A1, SLC25A12, GABBR1, GAD1, DLGAP1, MOG, BDNF, OLIG2, NTRK2 and 3, ESR1, SL6A4, TPH2, and COMT). The study found no significantly differential methylation. However, preliminary support for a difference was found for the gamma-aminobutyric acid (GABA) B receptor 1 (cg10234998, cg17099072) in blood samples at birth and for the estrogen receptor 1 (ESR1) (cg10939667), the myelin oligodendrocyte glycoprotein (MOG) (cg16650906), and the brain-derived neurotrophic factor (BDNF) (cg14080521) in blood samples at the time of diagnosis. Preliminary support for an association was observed between the methylation profiles of GABBR1 and MOG and baseline severity, treatment effect, and responder status; and between the methylation profile of ESR1 and baseline

  4. Another mechanism for creating diversity in gamma-aminobutyrate type A receptors: RNA splicing directs expression of two forms of gamma 2 phosphorylation site.

    PubMed Central

    Whiting, P; McKernan, R M; Iversen, L L

    1990-01-01

    Diversity of gamma-aminobutyrate type A (GABAA) receptors has recently been proposed to be achieved by assembly of receptor subtypes from a multitude of subunits (alpha 1-6, beta 1-3, gamma 1-2, and delta) encoded by different genes. Here we report a further mechanism for creating GABAA receptor diversity: alternative RNA splicing. Two forms of bovine gamma 2 subunit cDNA were isolated (gamma 2S and gamma 2L) that differed by the presence or absence of a 24-base-pair (8-amino acid) insertion in the cytoplasmic domain between the third and fourth putative membrane-spanning regions. Polymerase chain reaction from RNA demonstrated that the two forms of gamma 2 subunit are expressed in bovine, human, and rat brain. Sequencing of genomic DNA clones encoding the gamma 2 subunit demonstrated that the 24-base-pair insert is organized as a separate exon. Analysis of the sequence of the 8-amino acid insert revealed that it contains a protein kinase C consensus phosphorylation site. Expression of the large cytoplasmic loop domains of gamma 2S and gamma 2L in Escherichia coli, followed by phosphorylation of the recombinant proteins by protein kinase C, demonstrated that gamma 2L, but not gamma 2S, could be phosphorylated. Thus the two forms of gamma 2 subunit differ by the presence or absence of a protein kinase C phosphorylation site. This mechanism for creating GABAA receptor diversity may allow differential regulation of the function of receptor subtypes. Images PMID:1702226

  5. Comparative proteomic analysis of differentially expressed proteins in β-aminobutyric acid enhanced Arabidopsis thaliana tolerance to simulated acid rain.

    PubMed

    Liu, Tingwu; Jiang, Xinwu; Shi, Wuliang; Chen, Juan; Pei, Zhenming; Zheng, Hailei

    2011-05-01

    Acid rain is a worldwide environmental issue that has seriously destroyed forest ecosystems. As a highly effective and broad-spectrum plant resistance-inducing agent, β-aminobutyric acid could elevate the tolerance of Arabidopsis when subjected to simulated acid rain. Using comparative proteomic strategies, we analyzed 203 significantly varied proteins of which 175 proteins were identified responding to β-aminobutyric acid in the absence and presence of simulated acid rain. They could be divided into ten groups according to their biological functions. Among them, the majority was cell rescue, development and defense-related proteins, followed by transcription, protein synthesis, folding, modification and destination-associated proteins. Our conclusion is β-aminobutyric acid can lead to a large-scale primary metabolism change and simultaneously activate antioxidant system and salicylic acid, jasmonic acid, abscisic acid signaling pathways. In addition, β-aminobutyric acid can reinforce physical barriers to defend simulated acid rain stress.

  6. Neuronal and glial localization of GAT-1, a high-affinity gamma-aminobutyric acid plasma membrane transporter, in human cerebral cortex: with a note on its distribution in monkey cortex.

    PubMed

    Conti, F; Melone, M; De Biasi, S; Minelli, A; Brecha, N C; Ducati, A

    1998-06-22

    High-affinity gamma-aminobutyric (GABA) plasma membrane transporters (GATs) influence the action of GABA, the main inhibitory neurotransmitter in the human cerebral cortex. In this study, the cellular expression of GAT-1, the main cortical GABA transporter, was investigated in the human cerebral cortex by using immunocytochemistry with affinity-purified polyclonal antibodies directed to the C-terminus of rat GAT-1. In temporal and prefrontal association cortex (Brodmann's areas 21 and 46) and in cingulofrontal transition cortex (area 32), specific GAT-1 immunoreactivity (ir) was localized to numerous puncta and fibers in all cortical layers. GAT-1+ puncta were distributed homogeneously in all cortical layers, although they were slightly more numerous in layers II-IV, and appeared to have a preferential relationship to the somata and proximal dendrites of unlabeled pyramidal cells, even though, in many cases, they were also observed around nonpyramidal cells. Electron microscopic observations showed that GAT-1+ puncta were axon terminals that formed exclusively symmetric synapses. In addition, some distal astrocytic processes also contained immunoreaction product. Analysis of the patterns of GAT-1 labeling in temporal and prefrontal association areas (21 and 46), in cingulofrontal transition areas (32), and in somatic sensory and motor areas (1 and 4) of the monkey cortex revealed that its distribution varies according to the type of cortex examined and indicated that the distribution of GAT-1 is similar in anatomically corresponding areas of different species. The present study demonstrates that, in the human homotypical cortex, GAT-1 is expressed by both inhibitory axon terminals and astrocytic processes. This localization of GAT-1 is compatible with a major role for this transporter in GABA uptake at GABAergic synapses and suggests that GAT-1 may contribute to determining GABA levels in the extracellular space.

  7. Relating MEG measured motor cortical oscillations to resting γ-aminobutyric acid (GABA) concentration.

    PubMed

    Gaetz, W; Edgar, J C; Wang, D J; Roberts, T P L

    2011-03-15

    The human motor cortex exhibits characteristic beta (15-30 Hz) and gamma oscillations (60-90 Hz), typically observed in the context of transient finger movement tasks. The functional significance of these oscillations, such as post-movement beta rebound (PMBR) and movement-related gamma synchrony (MRGS) remains unclear. Considerable animal and human non-invasive studies, however, suggest that the networks supporting these motor cortex oscillations depend critically on the inhibitory neurotransmitter γ-Aminobutyric acid (GABA). Despite such speculation, a direct relation between MEG measured motor cortex oscillatory power and frequency with resting GABA concentrations has not been demonstrated. In the present study, motor cortical responses were measured from 9 healthy adults while they performed a cued button-press task using their right index finger. In each participant, PMBR and MRGS measures were obtained from time-frequency plots obtained from primary motor (MI) sources, localized using beamformer differential source localization. For each participant, complimentary magnetic resonance spectroscopy (MRS) GABA measures aligned to the motor hand knob of the left central sulcus were also obtained. GABA concentration was estimated as the ratio of the motor cortex GABA integral to a cortical reference NAA resonance at 2 ppm. A significant linear relation was observed between MI GABA concentration and MRGS frequency (R(2)=0.46, p<0.05), with no association observed between GABA concentration and MRGS power. Conversely, a significant linear relation was observed between MI GABA concentration and PMBR power (R(2)=0.34, p<0.05), with no relation observed for GABA concentration and PMBR frequency. Finally, a significant negative linear relation between the participant's age and MI gamma frequency was observed, such that older participants had a lower gamma frequency (R(2)=0.40, p<0.05). Present findings support a role for GABA in the generation and modulation of

  8. Enhancement of α5-Containing Gamma-Aminobutyric Acid TypeAReceptors by the Nonimmobilizer 1,2-Dichlorohexafluorocyclobutane (F6) is Abolished by the β3(N265M) Mutation

    PubMed Central

    Burkat, Paul M.; Lor, Chong; Perouansky, Misha; Pearce, Robert A.

    2014-01-01

    Background Modulation of γ-aminobutyric acid type A receptors (GABAARs) by general anesthetics may contribute to their ability to produce amnesia. Receptors containing α5 subunits, which mediate tonic and slow synaptic inhibition, are co-localized with β3 and γ2 subunits in dendritic layers of the hippocampus and are sensitive to low (amnestic) concentrations of anesthetics. Since α5 and β3 subunits influence performance in hippocampus-dependent learning tasks in the presence and absence of general anesthetics, and the experimental inhaled drug 1,2-dichlorohexafluorocyclobutane (F6) impairs hippocampus-dependent learning, we hypothesized that F6 would modulate receptors that incorporate α5 and β3 subunits. We hypothesized further that the β3(N265M) mutation, which controls receptor modulation by general anesthetics, would similarly influence modulation by F6. Methods Using whole-cell electrophysiological recording techniques, we tested the effects of F6 at concentrations ranging from 4 μM to 16 μM on receptors expressed in human embryonic kidney293 cells. We measured drug modulation of wild type α5β3 and α5β3γ2L GABAARs, and receptors harboring the β3(N265M) mutation. We also tested the effects of F6 on α1β2γ2L receptors, which were reported previously to be insensitive to this drug when expressed in Xenopus oocytes. Results F6 enhanced the responses of wild type α5β3γ2L but not α1β2γ2L receptors to low concentrations of GABA in a concentration-dependent manner. Receptors that incorporated the mutant β3(N265M) subunit were insensitive to F6. When applied together with a high concentration of GABA, F6 blocked currents through α5β3 but not α5β3γ2L receptors. F6 did not alter deactivation of α5β3γ2L receptors after brief, high concentration pulses of GABA. Conclusions The nonimmobilizer F6 modulates GABAARs in a manner that depends on subunit composition and on mode of receptor activation by GABA, supporting a possible role for α5

  9. PROTEOMIC ANALYSIS OF B-AMINOBUTYRIC ACID-PRIMED DROUGHT RESISTANCE IN CRABAPPLE SEEDLINGS

    Technology Transfer Automated Retrieval System (TEKTRAN)

    In a variety of annual crops and some model plant species, the non-protein, amino acid, DL-B-aminobutyric acid (BABA), has been shown to enhance disease resistance and increase salt and drought tolerance, through sensitization, and not direct induction of defense genes. This process is referred to a...

  10. Standard enthalpies of formation of γ-aminobutyric acid and the products of its dissociation in aqueous solution

    NASA Astrophysics Data System (ADS)

    Lytkin, A. I.; Chernikov, V. V.; Krutova, O. N.; Skvortsov, I. A.; Korchagina, A. S.

    2016-09-01

    Heat effects of the dissolution of crystalline γ-aminobutyric acid in water and potassium hydroxide solutions are determined by direct colorimetry at 298.15 K. Standard enthalpies of formation of γ-aminobutyric acid and the products of its dissociation in aqueous solution are calculated.

  11. Proteomic analysis of drought resistance in crabapple seedlings primed by the xenobiotic Beta-aminobutyric acid

    Technology Transfer Automated Retrieval System (TEKTRAN)

    In a variety of annual crops and model plants, the xenobiotic DL-Beta-aminobutyric acid (BABA) has been shown to enhance disease resistance and increase salt, drought and thermotolerance. BABA does not activate stress genes directly, but sensitizes plants to respond more quickly and strongly to biot...

  12. Ultrasonic Studies of 4-Aminobutyric Acid in Aqueous Metformin Hydrochloride Solutions at Different Temperatures

    NASA Astrophysics Data System (ADS)

    Rajagopal, K.; Jayabalakrishnan, S. S.

    2010-12-01

    Ultrasonic speeds and density data of 4-aminobutyric acid in 0.05 M, 0.10 M, and 0.15 M aqueous metformin hydrochloride (MFHCl) solutions are measured at 308.15 K, 313.15 K, and 318.15 K. The isentropic compressibility ( k S ), the change in isentropic compressibility (Δ k S ), the relative change in isentropic compressibility ({Δ k_S/k_S^0}), the apparent molal compressibility ({k_φ}), the limiting apparent molal compressibility ({k_φ^0 }), the transfer limiting apparent molal compressibility ({Δ k_φ^0}), the hydration number ( n H), and the pair and triplet interaction parameters ( k AH, k AHH) are estimated. The above parameters are used to interpret the solute-solute and solute-solvent interactions of 4-aminobutyric acid in aqueous MFHCl solutions.

  13. Beta-aminobutyric acid priming of plant defense: the role of ABA and other hormones.

    PubMed

    Baccelli, Ivan; Mauch-Mani, Brigitte

    2016-08-01

    Plants are exposed to recurring biotic and abiotic stresses that can, in extreme situations, lead to substantial yield losses. With the changing environment, the stress pressure is likely to increase and sustainable measures to alleviate the effect on our crops are sought. Priming plants for better stress resistance is one of the sustainable possibilities to reach this goal. Here, we report on the effects of beta-aminobutyric acid, a priming agent with an exceptionally wide range of action and describe its way of preparing plants to defend themselves against various attacks, among others through the modulation of their hormonal defense signaling, and highlight the special role of abscisic acid in this process.

  14. Statistical Mechanics Model for the Interaction between the Neurotransmitter γ-Aminobutyric acid and GABAA Receptors

    NASA Astrophysics Data System (ADS)

    Zafar, Sufi; Saxena, Nina C.; Conrad, Kevin A.; Hussain, Arif

    2004-07-01

    Interactions between the neurotransmitter γ-aminobutyric acid (GABA) and GABAA receptor ion channels play an important role in the central nervous system. A statistical mechanics model is proposed for the interaction between GABA and GABAA receptors. The model provides good fits to the electrophysiology data as well as an estimation of receptor activation energies, and predicts the temperature dependence consistent with measurements. In addition, the model provides insights into single channel conductance measurements. This model is also applicable to other ligand-gated ion channels with similar pentameric structures.

  15. Alpha-amino acid behaves differently from beta- or gamma-amino acids as treated by trimetaphosphate.

    PubMed

    Gao, X; Liu, Y; Xu, P X; Cai, Y M; Zhao, Y F

    2008-01-01

    The condensation reactions of sodium trimetaphosphate with single amino acids, namely glycine, L-alanine, beta-alanine and gamma-aminobutyric acid or pairs of these amino acids were reinvestigated by electrospray ion-trap mass spectrometry and high performance liquid chromatography. It was found when mixtures were treated by sodium trimetaphosphate only in the presence of alpha-amino acid dipeptides were formed. Without addition of alpha-amino acids, the beta-amino acid or gamma-aminobutyric acid could not form peptide either by themselves or with their mixtures under the same conditions. From the data it is concluded that phosphate might select alpha-amino acids to produce the peptides being important precursors for the origin of life. PMID:17973074

  16. Design and mechanism of tetrahydrothiophene-based γ-aminobutyric acid aminotransferase inactivators.

    PubMed

    Le, Hoang V; Hawker, Dustin D; Wu, Rui; Doud, Emma; Widom, Julia; Sanishvili, Ruslan; Liu, Dali; Kelleher, Neil L; Silverman, Richard B

    2015-04-01

    Low levels of γ-aminobutyric acid (GABA), one of two major neurotransmitters that regulate brain neuronal activity, are associated with many neurological disorders, such as epilepsy, Parkinson's disease, Alzheimer's disease, Huntington's disease, and cocaine addiction. One of the main methods to raise the GABA level in human brain is to use small molecules that cross the blood-brain barrier and inhibit the activity of γ-aminobutyric acid aminotransferase (GABA-AT), the enzyme that degrades GABA. We have designed a series of conformationally restricted tetrahydrothiophene-based GABA analogues with a properly positioned leaving group that could facilitate a ring-opening mechanism, leading to inactivation of GABA-AT. One compound in the series is 8 times more efficient an inactivator of GABA-AT than vigabatrin, the only FDA-approved inactivator of GABA-AT. Our mechanistic studies show that the compound inactivates GABA-AT by a new mechanism. The metabolite resulting from inactivation does not covalently bind to amino acid residues of GABA-AT but stays in the active site via H-bonding interactions with Arg-192, a π-π interaction with Phe-189, and a weak nonbonded S···O═C interaction with Glu-270, thereby inactivating the enzyme.

  17. Standard enthalpies of formation of α-aminobutyric acid and products of its dissociation in an aqueous solution

    NASA Astrophysics Data System (ADS)

    Lytkin, A. I.; Chernikov, V. V.; Krutova, O. N.

    2016-08-01

    Heats of solution of crystalline α-aminobutyric acid in water and in aqueous solutions of potassium hydroxide at 298.15 K are measured by means of direct calorimetry. Standard enthalpies of formation of the amino acid and products of its dissociation in an aqueous solution are calculated.

  18. Effect of the treatment by slightly acidic electrolyzed water on the accumulation of γ-aminobutyric acid in germinated brown millet.

    PubMed

    Li, Xingfeng; Hao, Jianxiong; Liu, Xianggui; Liu, Haijie; Ning, Yawei; Cheng, Ruhong; Tan, Bin; Jia, Yingmin

    2015-11-01

    The accumulation of γ-aminobutyric acid and the microbial decontamination are concerned increasingly in the production of sprouts. In this work, the effect of the treatment by slightly acidic electrolyzed water on the accumulation of γ-aminobutyric acid in the germinated brown millet was evaluated by high performance liquid chromatography during germination. The results showed that slightly acidic electrolyzed water with appropriate available chlorine (15 or 30 mg/L) could promote the accumulation of γ-aminobutyric acid by up to 21% (P < 0.05). However, the treatment with slightly acidic electrolyzed water could not enhance the sprouts growth of the germinated brown millet. The catalase and peroxidase activities of the germinated brown millet during germination were in agreement with the sprouts growth. Our results suggested that the accumulation of γ-aminobutyric acid was independent of the length of sprouts in germinated grains. Moreover, the treatment with slightly acidic electrolyzed water significantly reduced the microbial counts in the germinated millet (P < 0.05) and the treatment with high available chlorine concentration (15 and 30 mg/L) showed stronger anti-infection potential in the germinated brown millet than that of lower available chlorine concentration (5 mg/L). In conclusion, the treatment with slightly acidic electrolyzed water is an available approach to improve the accumulation of γ-aminobutyric acid and anti-infection potential in the germinated brown millet, and it can avoid too long millet sprouts.

  19. Simultaneous Spectral Editing for γ-Aminobutyric Acid and Taurine Using Double Quantum Coherence Transfer

    NASA Astrophysics Data System (ADS)

    Lei, Hao; Peeling, James

    2000-03-01

    Conventional double quantum (DQ) editing techniques recover resonances of one metabolite at a time and are thus inefficient for monitoring metabolic changes involving several metabolites. A DQ coherence transfer double editing sequence using a dual-band DQ coherence read pulse is described here. The sequence permits simultaneous spectral editing for two metabolites with similar J coupling constants in a single scan. Simultaneous editing for taurine and γ-aminobutyric acid (GABA) is demonstrated using solution phantoms and rat brain tissue. Selectivity of the double editing sequence for the target metabolites is as good as that achieved using conventional DQ editing which selects each metabolite individually. With experimental parameters of the double editing sequence chosen to optimize GABA editing, the sensitivity for GABA detection is the same as that with GABA editing only, while the sensitivity for taurine detection is decreased slightly compared to that with taurine editing only.

  20. Theoretical study of γ-aminobutyric acid conformers: Intramolecular interactions and ionization energies

    NASA Astrophysics Data System (ADS)

    Wang, Ke-Dong; Wang, Mei-Ting; Meng, Ju

    2014-10-01

    Allowing for all combinations of internal single-bond rotamers, 1,296 unique trial structures of γ-Aminobutyric acid (GABA) are obtained. All of these structures are optimized at the M06-2X level of theory and a total of 68 local minimal conformers are found. The nine low-lying conformers are used for further studies. According to the calculated relative Gibbs free energies at M06-2X level of theory, we find that the dispersion is important for the relative energy of GABA. The intramolecular hydrogen bonds and hyperconjugative interaction and their effects on the conformational stability are studied. The results show that both of them have great influence on the conformers. The vertical ionization energies (VIE) are calculated and match the experimental data well. The results show that the neutral GABA in the gas phase is a multi-conformer system and at least four conformations exist.

  1. Acidification and γ-aminobutyric acid independently alter kairomone-induced behaviour

    PubMed Central

    Cohen, Jonathan H.

    2016-01-01

    Exposure to high pCO2 or low pH alters sensation and behaviour in many marine animals. We show that crab larvae lose their ability to detect and/or process predator kairomones after exposure to low pH over a time scale relevant to diel pH cycles in coastal environments. Previous work suggests that acidification affects sensation and behaviour through altered neural function, specifically the action of γ-aminobutyric acid (GABA), because a GABA antagonist, gabazine, restores the original behaviour. Here, however, gabazine resulted in a loss of kairomone detection/processing, regardless of pH. Our results also suggest that GABAergic signalling is necessary for kairomone identification in these larvae. Hence, the mechanism for the observed pH effect varies from the original GABA hypothesis. Furthermore, we suggest that this pH effect is adaptive under diel-cycling pH. PMID:27703697

  2. Γ-aminobutyric acid receptors affect the progression and migration of tumor cells.

    PubMed

    Zhang, Xiaoxue; Du, Zuoyi; Liu, Jun; He, Jianxing

    2014-12-01

    Γ-aminobutyric acid (GABA) is a multifunctional molecule found in the nervous system and non-neuronal tissues. GABA receptors combine with GABA molecules and transmit signal stimuli into cells. In addition to traditional neurotransmission and regulation of secretion, GABA and GABA receptors are involved in cell differentiation and proliferation throughout peripheral organs, as well as in tumorigenesis. The exact mechanism of the GABAergic system in regulating tumor development is unclear, but many studies have revealed that GABA receptors exert critical regulative effects on tumor cell proliferation and migration. In this review, the molecular structure, distribution and biological function of GABA receptors associated with tumorigenesis are described. Recent advances in the elucidation of mechanisms underlying GABAergic signaling control over tumor growth are also discussed.

  3. Optimization of γ-aminobutyric acid production by Lactobacillus plantarum Taj-Apis362 from honeybees.

    PubMed

    Tajabadi, Naser; Ebrahimpour, Afshin; Baradaran, Ali; Rahim, Raha Abdul; Mahyudin, Nor Ainy; Manap, Mohd Yazid Abdul; Bakar, Fatimah Abu; Saari, Nazamid

    2015-04-15

    Dominant strains of lactic acid bacteria (LAB) isolated from honey bees were evaluated for their γ-aminobutyric acid (GABA)-producing ability. Out of 24 strains, strain Taj-Apis362 showed the highest GABA-producing ability (1.76 mM) in MRS broth containing 50 mM initial glutamic acid cultured for 60 h. Effects of fermentation parameters, including initial glutamic acid level, culture temperature, initial pH and incubation time on GABA production were investigated via a single parameter optimization strategy. The optimal fermentation condition for GABA production was modeled using response surface methodology (RSM). The results showed that the culture temperature was the most significant factor for GABA production. The optimum conditions for maximum GABA production by Lactobacillus plantarum Taj-Apis362 were an initial glutamic acid concentration of 497.97 mM, culture temperature of 36 °C, initial pH of 5.31 and incubation time of 60 h, which produced 7.15 mM of GABA. The value is comparable with the predicted value of 7.21 mM.

  4. Optimization of γ-aminobutyric acid production by Lactobacillus plantarum Taj-Apis362 from honeybees.

    PubMed

    Tajabadi, Naser; Ebrahimpour, Afshin; Baradaran, Ali; Rahim, Raha Abdul; Mahyudin, Nor Ainy; Manap, Mohd Yazid Abdul; Bakar, Fatimah Abu; Saari, Nazamid

    2015-01-01

    Dominant strains of lactic acid bacteria (LAB) isolated from honey bees were evaluated for their γ-aminobutyric acid (GABA)-producing ability. Out of 24 strains, strain Taj-Apis362 showed the highest GABA-producing ability (1.76 mM) in MRS broth containing 50 mM initial glutamic acid cultured for 60 h. Effects of fermentation parameters, including initial glutamic acid level, culture temperature, initial pH and incubation time on GABA production were investigated via a single parameter optimization strategy. The optimal fermentation condition for GABA production was modeled using response surface methodology (RSM). The results showed that the culture temperature was the most significant factor for GABA production. The optimum conditions for maximum GABA production by Lactobacillus plantarum Taj-Apis362 were an initial glutamic acid concentration of 497.97 mM, culture temperature of 36 °C, initial pH of 5.31 and incubation time of 60 h, which produced 7.15 mM of GABA. The value is comparable with the predicted value of 7.21 mM. PMID:25884548

  5. Production of gaba (γ – Aminobutyric acid) by microorganisms: a review

    PubMed Central

    Dhakal, Radhika; Bajpai, Vivek K.; Baek, Kwang-Hyun

    2012-01-01

    GABA (γ-aminobutyric acid) is a four carbon non-protein amino acid that is widely distributed in plants, animals and microorganisms. As a metabolic product of plants and microorganisms produced by the decarboxylation of glutamic acid, GABA functions as an inhibitory neurotransmitter in the brain that directly affects the personality and the stress management. A wide range of traditional foods produced by microbial fermentation contain GABA, in which GABA is safe and eco-friendly, and also has the possibility of providing new health-benefited products enriched with GABA. Synthesis of GABA is catalyzed by glutamate decarboxylase, therefore, the optimal fermentation condition is mainly based on the biochemical properties of the enzyme. Major GABA producing microorganisms are lactic acid bacteria (LAB), which make food spoilage pathogens unable to grow and act as probiotics in the gastrointestinal tract. The major factors affecting the production of GABA by microbial fermentation are temperature, pH, fermentation time and different media additives, therefore, these factors are summarized to provide the most up-dated information for effective GABA synthesis. There has been a huge accumulation of knowledge on GABA application for human health accompanying with a demand on natural GABA supply. Only the GABA production by microorganisms can fulfill the demand with GABA-enriched health beneficial foods. PMID:24031948

  6. Effects of γ-Aminobutyric acid transporter 1 inhibition by tiagabine on brain glutamate and γ-Aminobutyric acid metabolism in the anesthetized rat In vivo.

    PubMed

    Patel, Anant B; de Graaf, Robin A; Rothman, Douglas L; Behar, Kevin L

    2015-07-01

    γ-Aminobutyric acid (GABA) clearance from the extracellular space after release from neurons involves reuptake into terminals and astrocytes through GABA transporters (GATs). The relative flows through these two pathways for GABA released from neurons remains unclear. This study determines the effect of tiagabine, a selective inhibitor of neuronal GAT-1, on the rates of glutamate (Glu) and GABA metabolism and GABA resynthesis via the GABA-glutamine (Gln) cycle. Halothane-anesthetized rats were administered tiagabine (30 mg/kg, i.p.) and 45 min later received an intravenous infusion of either [1,6-(13)C2]glucose (in vivo) or [2-(13)C]acetate (ex vivo). Nontreated rats served as controls. Metabolites and (13)C enrichments were measured with (1)H-[(13)C]-nuclear magnetic resonance spectroscopy and referenced to their corresponding endpoint values measured in extracts from in situ frozen brain. Metabolic flux estimates of GABAergic and glutamatergic neurons were determined by fitting a metabolic model to the (13)C turnover data measured in vivo during [1,6-(13)C2]glucose infusion. Tiagabine-treated rats were indistinguishable (P > 0.05) from controls in tissue amino acid levels and in (13)C enrichments from [2-(13)C]acetate. Tiagabine reduced average rates of glucose oxidation and neurotransmitter cycling in both glutamatergic neurons (↓18%, CMR(glc(ox)Glu): control, 0.27 ± 0.05 vs. tiagabine, 0.22 ± 0.04 µmol/g/min; ↓11%, V(cyc(Glu-Gln)): control 0.23 ± 0.05 vs. tiagabine 0.21 ± 0.04 µmol/g/min and GABAergic neurons (↓18-25%, CMR(glc(ox)GABA): control 0.09 ± 0.02 vs. tiagabine 0.07 ± 0.03 µmol/g/min; V(cyc(GABA-Gln)): control 0.08 ± 0.02 vs. tiagabine 0.07 ± 0.03 µmol/g/min), but the changes in glutamatergic and GABAergic fluxes were not significant (P > 0.10). The results suggest that any reduction in GABA metabolism by tiagabine might be an indirect response to reduced glutamatergic drive rather than direct compensatory effects.

  7. γ-Aminobutyric acid (GABA) homeostasis regulates pollen germination and polarized growth in Picea wilsonii.

    PubMed

    Ling, Yu; Chen, Tong; Jing, Yanping; Fan, Lusheng; Wan, Yinglang; Lin, Jinxing

    2013-11-01

    γ-Aminobutyric acid (GABA) is a four-carbon non-protein amino acid found in a wide range of organisms. Recently, GABA accumulation has been shown to play a role in the stress response and cell growth in angiosperms. However, the effect of GABA deficiency on pollen tube development remains unclear. Here, we demonstrated that specific concentrations of exogenous GABA stimulated pollen tube growth in Picea wilsonii, while an overdose suppressed pollen tube elongation. The germination percentage of pollen grains and morphological variations in pollen tubes responded in a dose-dependent manner to treatment with 3-mercaptopropionic acid (3-MP), a glutamate decarboxylase inhibitor, while the inhibitory effects could be recovered in calcium-containing medium supplemented with GABA. Using immunofluorescence labeling, we found that the actin cables were disorganized in 3-MP treated cells, followed by the transition of endo/exocytosis activating sites from the apex to the whole tube shank. In addition, variations in the deposition of cell wall components were detected upon labeling with JIM5, JIM7, and aniline blue. Our results demonstrated that calcium-dependent GABA signaling regulates pollen germination and polarized tube growth in P. wilsonii by affecting actin filament patterns, vesicle trafficking, and the configuration and distribution of cell wall components.

  8. Use of Lactococcus lactis to enrich sourdough bread with γ-aminobutyric acid.

    PubMed

    Bhanwar, Seema; Bamnia, Meenakshi; Ghosh, Moushumi; Ganguli, Abhijit

    2013-02-01

    Fried sourdough bread (bhatura) with an elevated amount of γ-aminobutyric acid (GABA) was produced using lactic acid bacteria (LAB). The LAB starter was screened and isolated from pickled yam showing highest GABA content and was identified as Lactococcus lactis subsp. lactis. The maximum GABA production in de Man Rogosa Sharpe (MRS) media supplemented with monosodium glutamate (MSG) was 110 mg/100 ml at pH 5, and 1-3% NaCl did not change the production of GABA significantly (p>0.05). When MSG was replaced with Vigna mungo in sourdough, the amount of GABA for bhatura was 226.22 mg/100 g representing about 10-fold increase. A sensory evaluation resulted as the overall general acceptability of bhatura to be 4.91 ± 0.03 on a five-point hedonic scale. Thus, the results indicated the potential of L. lactis as a LAB starter for the production of GABA-enriched bhatura. Although other physiological effects can be expected in the product, animal and clinical studies are mandatory prior to application of this food.

  9. Subchronic toxicity evaluation of γ-aminobutyric acid (GABA) in rats.

    PubMed

    Takeshima, Kazuhito; Yamatsu, Atsushi; Yamashita, Yusuke; Watabe, Kazuya; Horie, Noriko; Masuda, Kazuyuki; Kim, Mujo

    2014-06-01

    γ-Aminobutyric acid (GABA) is an amino acid compound contained in vegetables such as tomatoes and also widely distributed in mammals. GABA acts as an inhibitory neurotransmitter and promotes parasympathetic activity to provide several beneficial effects, for instance, relaxation, anti-stress, and insomnia. GABA, produced via a fermentation process, has been available as a functional food ingredient. As part of a program to assess its safety, GABA was administered by oral gavage at doses of 500, 1250, and 2500mg/kg body weight to groups of 10 male and 10 female Sprague-Dawley rats for 13weeks. Treatment was not associated with the test substance-related mortality and appeared to be well tolerated. There were no toxicologically and statistically significant changes in urinalysis, hematology, clinical chemistry parameters, and in necropsy findings. A few statistically significant changes in food consumption and body weights were noted in the male groups while any significant changes were not noted in female groups. There was no effect of treatment on organ weights or on the results of the histopathological examinations. The results of toxicity evaluation support the safety use of GABA and the potential use as a functional food ingredient.

  10. Connecting Proline and γ-Aminobutyric Acid in Stressed Plants through Non-Enzymatic Reactions

    PubMed Central

    Signorelli, Santiago; Dans, Pablo D.; Coitiño, E. Laura; Borsani, Omar; Monza, Jorge

    2015-01-01

    The accumulation of proline (Pro) in plants exposed to biotic/abiotic stress is a well-documented and conserved response in most vegetal species. Stress conditions induce the overproduction of reactive oxygen species which can lead to cellular damage. In vitro assays have shown that enzyme inactivation by hydroxyl radicals (·OH) can be avoided in presence of Pro, suggesting that this amino acid could act as an ·OH scavenger. We applied Density Functional Theory coupled with a polarizable continuum model to elucidate how Pro reacts with ·OH. In this work we suggest that Pro reacts favourably with ·OH by H–abstraction on the amine group. This reaction produces the spontaneous decarboxylation of Pro leading to the formation of pyrrolidin-1-yl. In turn, pyrrolidin-1-yl can easily be converted to Δ1-pyrroline, the substrate of the enzyme Δ1-pyrroline dehydrogenase, which produces γ-aminobutyric acid (GABA). GABA and Pro are frequently accumulated in stressed plants and several protective roles have been assigned to these molecules. Thereby we present an alternative non-enzymatic way to synthetize GABA under oxidative stress. Finally this work sheds light on a new beneficial role of Pro accumulation in the maintenance of photosynthetic activity. PMID:25775459

  11. Evaluation of improved γ-aminobutyric acid production in yogurt using Lactobacillus plantarum NDC75017.

    PubMed

    Shan, Y; Man, C X; Han, X; Li, L; Guo, Y; Deng, Y; Li, T; Zhang, L W; Jiang, Y J

    2015-04-01

    Most γ-aminobutyric acid (GABA)-producing microorganisms are lactic acid bacteria (LAB), but the yield of GABA is limited in most of these GABA-producing strains. In this study, the production of GABA was carried out by using Lactobacillus plantarum NDC75017, a strain screened from traditional fermented dairy products in China. Concentrations of substrate (l-monosodium glutamate, L-MSG) and coenzyme (pyridoxal-5-phosphate, PLP) of glutamate decarboxylase (GAD) and culture temperature were investigated to evaluate their effects on GABA yield of Lb. plantarum NDC75017. The results indicated that GABA production was related to GAD activity and biomass of Lb. plantarum NDC75017. Response surface methodology was used to optimize conditions of GABA production. The optimal factors for GABA production were L-MSG at 80 mM, PLP at 18 μM, and a culture temperature of 36 °C. Under these conditions, production of GABA was maximized at 314.56 mg/100 g. Addition of Lb. plantarum NDC75017 to a commercial starter culture led to higher GABA production in fermented yogurt. Flavor and texture of the prepared yogurt and the control yogurt did not differ significantly. Thus, Lb. plantarum NDC75017 has good potential for manufacture of GABA-enriched fermented milk products.

  12. γ-Aminobutyric acid (GABA) concentration inversely correlates with basal perfusion in human occipital lobe.

    PubMed

    Donahue, Manus J; Rane, Swati; Hussey, Erin; Mason, Emily; Pradhan, Subechhya; Waddell, Kevin W; Ally, Brandon A

    2014-03-01

    Commonly used neuroimaging approaches in humans exploit hemodynamic or metabolic indicators of brain function. However, fundamental gaps remain in our ability to relate such hemo-metabolic reactivity to neurotransmission, with recent reports providing paradoxical information regarding the relationship among basal perfusion, functional imaging contrast, and neurotransmission in awake humans. Here, sequential magnetic resonance spectroscopy (MRS) measurements of the primary inhibitory neurotransmitter, γ-aminobutyric acid (GABA+macromolecules normalized by the complex N-acetyl aspartate-N-acetyl aspartyl glutamic acid: [GABA(+)]/[NAA-NAAG]), and magnetic resonance imaging (MRI) measurements of perfusion, fractional gray-matter volume, and arterial arrival time (AAT) are recorded in human visual cortex from a controlled cohort of young adult male volunteers with neurocognitive battery-confirmed comparable cognitive capacity (3 T; n=16; age=23±3 years). Regression analyses reveal an inverse correlation between [GABA(+)]/[NAA-NAAG] and perfusion (R=-0.46; P=0.037), yet no relationship between AAT and [GABA(+)]/[NAA-NAAG] (R=-0.12; P=0.33). Perfusion measurements that do not control for AAT variations reveal reduced correlations between [GABA(+)]/[NAA-NAAG] and perfusion (R=-0.13; P=0.32). These findings largely reconcile contradictory reports between perfusion and inhibitory tone, and underscore the physiologic origins of the growing literature relating functional imaging signals, hemodynamics, and neurotransmission.

  13. Enzymatic production of γ-aminobutyric acid in soybeans using high hydrostatic pressure and precursor feeding.

    PubMed

    Ueno, Shigeaki; Katayama, Takumi; Watanabe, Takae; Nakajima, Kanako; Hayashi, Mayumi; Shigematsu, Toru; Fujii, Tomoyuki

    2013-01-01

    The effects were investigated of the glutamic acid (Glu) substrate concentration on the generation and kinetics of γ-aminobutyric acid (GABA) in soybeans treated under high hydrostatic pressure (HHP; 200 MPa for 10 min at 25 °C). The conversion of Glu to GABA decreased with increasing initial Glu concentration in the soybeans. The crude glutamate decarboxylase (GAD) obtained from the HHP-treated soybeans showed substrate inhibition. The GABA production rate in the HHP-treated soybeans fitted the following substrate inhibition kinetic equation: v0=(VmaxS0)/(Km+S0+(S0)2/Ki). The Km value for the HHP-treated soybeans was significantly higher than that of the untreated soybeans. The Km values in this study show the affinity between Glu and GAD, and indicate that the HHP-treated soybeans had lower affinity between Glu and GAD than the untreated soybeans. GAD extracted from the HHP-treated soybeans showed a similar value to that in the HHP-treated soybeans. The intact biochemical system was so damaged in the HHP-treated soybeans that it showed substrate inhibition kinetics similar to that of the extracted GAD. The combination of HHP and precursor feeding proved to be a novel tool that can be used to increase the concentration of a target component.

  14. Connecting proline and γ-aminobutyric acid in stressed plants through non-enzymatic reactions.

    PubMed

    Signorelli, Santiago; Dans, Pablo D; Coitiño, E Laura; Borsani, Omar; Monza, Jorge

    2015-01-01

    The accumulation of proline (Pro) in plants exposed to biotic/abiotic stress is a well-documented and conserved response in most vegetal species. Stress conditions induce the overproduction of reactive oxygen species which can lead to cellular damage. In vitro assays have shown that enzyme inactivation by hydroxyl radicals (·OH) can be avoided in presence of Pro, suggesting that this amino acid could act as an ·OH scavenger. We applied Density Functional Theory coupled with a polarizable continuum model to elucidate how Pro reacts with ·OH. In this work we suggest that Pro reacts favourably with ·OH by H-abstraction on the amine group. This reaction produces the spontaneous decarboxylation of Pro leading to the formation of pyrrolidin-1-yl. In turn, pyrrolidin-1-yl can easily be converted to Δ1-pyrroline, the substrate of the enzyme Δ1-pyrroline dehydrogenase, which produces γ-aminobutyric acid (GABA). GABA and Pro are frequently accumulated in stressed plants and several protective roles have been assigned to these molecules. Thereby we present an alternative non-enzymatic way to synthetize GABA under oxidative stress. Finally this work sheds light on a new beneficial role of Pro accumulation in the maintenance of photosynthetic activity.

  15. Functionalities of conjugated compounds of γ-aminobutyric acid with salicylaldehyde or cinnamaldehyde.

    PubMed

    Liu, Tai-Ti; Tseng, Yi-Wei; Yang, Tsung-Shi

    2016-01-01

    Aldehydes or ketones can react with amino compounds to form Schiff base adducts, which have been widely studied and shown to exhibit antimicrobial, antioxidant or antiviral activity. Salicylaldehyde (SA) and cinnamaldehyde (CA) are components of plant essential oils. γ-Aminobutyric acid (GA) is an important substance in the mammalian central nervous system and responsible for many bioactivities. This study aimed to synthesize functional Schiff base adducts using GA and SA or CA; to study the antimicrobial activity, antioxidant activity and tyrosinase-inhibition activity of these adducts (SA-GA and CA-GA) and their metal complexes--SA-GA-Cu (or Zn) and CA-GA-Cu (or Zn); and to find their applications in food systems. SA-GA and CA-GA both exhibited good antibacterial effects, and so did their Cu complexes. As for antioxidant activity, SA-GA and CA-GA were superior to their metal complexes in most tests. Regarding inhibition of enzymatic browning of mushrooms, both SA-GA-Cu and CA-GA-Cu could inhibit tyrosinase activity effectively.

  16. Contents of Neo-flavored Tea (GABA Kintaro) Containing γ-Aminobutyric Acid

    NASA Astrophysics Data System (ADS)

    Shiraki, Yoshiya

    The contents of γ-aminobutyric acid (GABA), catechins, theaflavins, caffeine and pheophorbide-a in neo-flavored tea (GABA Kintaro tea) were analyzed. 1)The amounts of GABA were increased over 1.5mg/g by means of infrared ray irradiation with agitation treatment. 2)There was a tendency for the amount of catechins to be decreased by this treatment, whereas the amount of theaflavins tended to increase with the same treatment. The composition of these contents in this GABA Kintaro tea was almost the same as that of black tea. 3)There was a tendency for the amount of caffeine to be decreased by this treatment. 4)There was a tendency for the amount of pheophorbide-a to be increased by this treatment. 5)The result of this study showed that the amounts of GABA and theaflavins in this GABA Kintaro tea were higher than ordinary green tea but contained few catechins.It became clear that the amount of pheophorbide-a in this GABA Kintaro tea was less than the standard value established in processed chlorella.

  17. Application of MS Transport Assays to the Four Human γ-Aminobutyric Acid Transporters.

    PubMed

    Schmitt, Sebastian; Höfner, Georg; Wanner, Klaus T

    2015-09-01

    γ-Aminobutyric acid (GABA) transporters (GATs) are promising drug targets for various diseases associated with imbalances in GABAergic neurotransmission. For the development of new drugs or pharmacological tools addressing GATs, screening techniques to identify new inhibitors and to characterize their potency at each GAT subtype are indispensable. By now, the technique by far dominating is based on radiolabeled GABA. We recently described "MS Transport Assays" for hGAT-1 by employing ((2) H6 )GABA as the substrate. In the present study, we applied this approach to all four human GAT subtypes and determined the KM values for GAT-mediated transport of ((2) H6 )GABA at each subtype. Furthermore, a comprehensive set of GAT inhibitors reflecting the whole range of potency and subtype selectivity known so far was evaluated for their potency. The comparison of pIC50 values obtained in conventional [(3) H]GABA uptake assays with those obtained in MS Transport Assays indicated the reliability of the latter. The MS Transport Assays enable a throughput similar to that of conventional radiometric transport assays performed in a 96-well format but avoid the use of radiolabeled substrates.

  18. The inhibitory role of γ-aminobutyric acid (GABA) on immunomodulation of Pacific oyster Crassostrea gigas.

    PubMed

    Li, Meijia; Qiu, Limei; Wang, Lingling; Wang, Weilin; Xin, Lusheng; Li, Yiqun; Liu, Zhaoqun; Song, Linsheng

    2016-05-01

    γ-aminobutyric acid (GABA) is an inhibitory neurotransmitter to suppress the immune-mediated pro-inflammatory reactions, and it has been used in the treatment of many inflammation-related diseases in vertebrates, while its immunomodulatory role in invertebrates has never been reported. In the present study, GABA was found to exist in the hemolymph of Pacific oyster Crassostrea gigas, and its concentration decreased slightly from 8.00 ± 0.37 μmol L(-1) at normal condition to 7.73 ± 0.15 μmol L(-1) at 6 h after LPS stimulation, and then increased to 9.34 ± 0.15 μmol L(-1), 8.86 ± 0.68 μmol L(-1) at 12 h and 48 h, respectively. After LPS stimulation, the mRNA expressions of pro-inflammatory cytokines (CgIL-17 and CgTNF) and immune effectors (CgSOD and CgBPI), and the protein expression of NOS increased significantly, and these increased trends were remarkably inhibited by GABA stimulation. At the same time, the phagocytosis rate and apoptosis rate of immunocytes also increased obviously after LPS stimulation, whereas the increase was repressed with the addition of GABA. The results collectively demonstrated that GABA was an indispensable inhibitory agent for both humoral and cellular immune response, which mainly functioned at the late phase of immune response to avoid the excess immune reactions and maintain the immune homeostasis.

  19. Utilization of barley or wheat bran to bioconvert glutamate to γ-aminobutyric acid (GABA).

    PubMed

    Jin, Wen-Jie; Kim, Min-Ju; Kim, Keun-Sung

    2013-09-01

    This study deals with the utilization of agro-industrial wastes created by barley and wheat bran in the production of a value-added product, γ-aminobutyric acid (GABA). The simple and eco-friendly reaction requires no pretreatment or microbial fermentation steps but uses barley or wheat bran as an enzyme source, glutamate as a substrate, and pyridoxal 5'-phosphate (PLP) as a cofactor. The optimal reaction conditions were determined on the basis of the temperatures and times used for the decarboxylation reactions and the initial concentrations of barley or wheat bran, glutamate, and PLP. The optimal reactions produced 9.2 mM of GABA from 10 mM glutamate, yielding a 92% GABA conversion rate, when barley bran was used and 6.0 mM of GABA from 10 mM glutamate, yielding a 60% GABA conversion rate, when wheat bran was used. The results imply that barley bran is more efficient than wheat bran in the production of GABA.

  20. Vesicular Inhibitory Amino Acid Transporter Is a Cl−/γ-Aminobutyrate Co-transporter*

    PubMed Central

    Juge, Narinobu; Muroyama, Akiko; Hiasa, Miki; Omote, Hiroshi; Moriyama, Yoshinori

    2009-01-01

    The vesicular inhibitory amino acid transporter (VIAAT) is a synaptic vesicle protein responsible for the vesicular storage of γ-aminobutyrate (GABA) and glycine which plays an essential role in GABAergic and glycinergic neurotransmission. The transport mechanism of VIAAT remains largely unknown. Here, we show that proteoliposomes containing purified VIAAT actively took up GABA upon formation of membrane potential (Δψ) (positive inside) but not ΔpH. VIAAT-mediated GABA uptake had an absolute requirement for Cl− and actually accompanied Cl− movement. Kinetic analysis indicated that one GABA molecule and two Cl− equivalents were transported during one transport cycle. VIAAT in which Glu213 was specifically mutated to alanine completely lost the ability to take up both GABA and Cl−. Essentially the same results were obtained with glycine, another substrate of VIAAT. These results demonstrated that VIAAT is a vesicular Cl− transporter that co-transports Cl− with GABA or glycine in a Δψ dependent manner. It is concluded that Cl− plays an essential role in vesicular storage of GABA and glycine. PMID:19843525

  1. Functionalities of conjugated compounds of γ-aminobutyric acid with salicylaldehyde or cinnamaldehyde.

    PubMed

    Liu, Tai-Ti; Tseng, Yi-Wei; Yang, Tsung-Shi

    2016-01-01

    Aldehydes or ketones can react with amino compounds to form Schiff base adducts, which have been widely studied and shown to exhibit antimicrobial, antioxidant or antiviral activity. Salicylaldehyde (SA) and cinnamaldehyde (CA) are components of plant essential oils. γ-Aminobutyric acid (GA) is an important substance in the mammalian central nervous system and responsible for many bioactivities. This study aimed to synthesize functional Schiff base adducts using GA and SA or CA; to study the antimicrobial activity, antioxidant activity and tyrosinase-inhibition activity of these adducts (SA-GA and CA-GA) and their metal complexes--SA-GA-Cu (or Zn) and CA-GA-Cu (or Zn); and to find their applications in food systems. SA-GA and CA-GA both exhibited good antibacterial effects, and so did their Cu complexes. As for antioxidant activity, SA-GA and CA-GA were superior to their metal complexes in most tests. Regarding inhibition of enzymatic browning of mushrooms, both SA-GA-Cu and CA-GA-Cu could inhibit tyrosinase activity effectively. PMID:26213082

  2. Induced resistance against the Asian citrus psyllid, Diaphorina citri, by β-aminobutyric acid in citrus.

    PubMed

    Tiwari, Siddharth; Meyer, Wendy L; Stelinski, Lukasz L

    2013-10-01

    β-Aminobutyric acid (BABA) is known to induce resistance to microbial pathogens, nematodes and insects in several host plant/pest systems. The present study was undertaken to determine whether a similar effect of BABA occurred against the Asian citrus psyllid, Diaphorina citri Kuwayama, in citrus. A 25 mM drench application of BABA significantly reduced the number of eggs/plant as compared with a water control, whereas 200 and 100 mM applications of BABA reduced the numbers of nymphs/plant and adults/plants, respectively. A 5 mM foliar application of BABA significantly reduced the number of adults but not eggs or nymphs when compared with a water control treatment. In addition, leaf-dip bioassays using various concentrations (25–500 mM) of BABA indicated no direct toxic effect on 2nd and 5th instar nymphs or adult D. citri. BABA-treated plants were characterized by significantly lower levels of iron, magnesium, phosphorus, sodium, sulfur and zinc as compared with control plants. The expression level of the PR-2 gene (β-1,3-glucanase) in BABA-treated plants that were also damaged by D. citri adult feeding was significantly higher than in plants exposed to BABA, D. citri feeding alone or control plants. Our results indicate the potential for using BABA as a systemic acquired resistance management tool for D. citri.

  3. The Central Amygdala and Alcohol: Role of γ-Aminobutyric Acid, Glutamate, and Neuropeptides

    PubMed Central

    Roberto, Marisa; Gilpin, Nicholas W.; Siggins, George R.

    2012-01-01

    Alcohol dependence is a chronically relapsing disorder characterized by compulsive drug seeking and drug taking, loss of control in limiting intake, and the emergence of a withdrawal syndrome in the absence of the drug. Accumulating evidence suggests an important role for synaptic transmission in the central amygdala (CeA) in mediating alcohol-related behaviors and neuroadaptative mechanisms associated with alcohol dependence. Acute alcohol facilitates γ-aminobutyric acid-ergic (GABAergic) transmission in CeA via both pre- and postsynaptic mechanisms, and chronic alcohol increases baseline GABAergic transmission. Acute alcohol inhibits glutamatergic transmission via effects at N-methyl-d-aspartate (NMDA) and AMPA receptors in CeA, whereas chronic alcohol up-regulates N-methyl-d-aspartate receptor (NMDAR)-mediated transmission. Pro- (e.g., corticotropin-releasing factor [CRF]) and anti-stress (e.g., NPY, nociceptin) neuropeptides affect alcohol- and anxiety-related behaviors, and also alter the alcohol-induced effects on CeA neurotransmission. Alcohol dependence produces plasticity in these neuropeptide systems, reflecting a recruitment of those systems during the transition to alcohol dependence. PMID:23085848

  4. Independent Effects of γ-Aminobutyric Acid Transaminase (GABAT) on Metabolic and Sleep Homeostasis.

    PubMed

    Maguire, Sarah E; Rhoades, Seth; Chen, Wen-Feng; Sengupta, Arjun; Yue, Zhifeng; Lim, Jason C; Mitchell, Claire H; Weljie, Aalim M; Sehgal, Amita

    2015-08-14

    Breakdown of the major sleep-promoting neurotransmitter, γ-aminobutyric acid (GABA), in the GABA shunt generates catabolites that may enter the tricarboxylic acid cycle, but it is unknown whether catabolic by-products of the GABA shunt actually support metabolic homeostasis. In Drosophila, the loss of the specific enzyme that degrades GABA, GABA transaminase (GABAT), increases sleep, and we show here that it also affects metabolism such that flies lacking GABAT fail to survive on carbohydrate media. Expression of GABAT in neurons or glia rescues this phenotype, indicating a general metabolic function for this enzyme in the brain. As GABA degradation produces two catabolic products, glutamate and succinic semialdehyde, we sought to determine which was responsible for the metabolic phenotype. Through genetic and pharmacological experiments, we determined that glutamate, rather than succinic semialdehyde, accounts for the metabolic phenotype of gabat mutants. This is supported by biochemical measurements of catabolites in wild-type and mutant animals. Using in vitro labeling assays, we found that inhibition of GABAT affects energetic pathways. Interestingly, we also observed that gaba mutants display a general disruption in bioenergetics as measured by altered levels of tricarboxylic acid cycle intermediates, NAD(+)/NADH, and ATP levels. Finally, we report that the effects of GABAT on sleep do not depend upon glutamate, indicating that GABAT regulates metabolic and sleep homeostasis through independent mechanisms. These data indicate a role of the GABA shunt in the development of metabolic risk and suggest that neurological disorders caused by altered glutamate or GABA may be associated with metabolic disruption.

  5. Vesicular γ-Aminobutyric Acid Transporter Expression in Amacrine and Horizontal Cells

    PubMed Central

    Cueva, Juan G.; Haverkamp, Silke; Reimer, Richard J.; Edwards, Robert; Wässle, Heinz; Brecha, Nicholas C.

    2010-01-01

    The vesicular γ-aminobutyric acid (GABA) transporter (VGAT), which transports the inhibitory amino acid transmitters GABA and glycine, is localized to synaptic vesicles in axon terminals. The localization of VGAT immunoreactivity to mouse and rat retina was evaluated with light and electron microscopy by using well-characterized VGAT antibodies. Specific VGAT immunoreactivity was localized to numerous varicose processes in all laminae of the inner plexiform layer (IPL) and to the outer plexiform layer (OPL). Amacrine cell somata characterized by weak VGAT immunoreactivity in the cytoplasm were located in the ganglion cell layer and proximal inner nuclear layer (INL) adjacent to the IPL. In rat retina, VGAT-immunoreactive cell bodies also contained GABA, glycine, or parvalbumin (PV) immunoreactivity, suggesting vesicular uptake of GABA or glycine by these cells. A few varicose VGAT-immunoreactive processes entered the OPL from the IPL. VGAT immunoreactivity in the OPL was predominantly localized to horizontal cell processes. VGAT and calcium binding protein-28K immunoreactivities (CaBP; a marker for horizontal cells) were colocalized in processes and terminals distributed to the OPL. Furthermore, VGAT immunoreactivity overlapped or was immediately adjacent to postsynaptic density-95 (PSD-95) immunoreactivity, which is prominent in photoreceptor terminals. Preem-bedding immunoelectron microscopy of mouse and rat retinae showed that VGAT immunoreactivity was localized to horizontal cell processes and their terminals. Immunoreactivity was distributed throughout the cytoplasm of the horizontal cell processes. Taken together, these findings demonstrate VGAT immunoreactivity in both amacrine and horizontal cell processes, suggesting these cells contain vesicles that accumulate GABA and glycine, possibly for vesicular release. PMID:11920703

  6. Comparative proteomic analysis of β-aminobutyric acid-mediated alleviation of salt stress in barley.

    PubMed

    Mostek, Agnieszka; Börner, Andreas; Weidner, Stanisław

    2016-02-01

    The non-protein amino acid β-aminobutyric acid (BABA) is known to induce plant resistance to a broad spectrum of biotic and abiotic stresses. This is the first study describing the effect of BABA seed priming on physiological and proteomic changes under salt stress conditions in barley (Hordeum vulgare). The aim of our study was to investigate the changes of fresh weight, dry weight and relative water content (RWC) as well as root proteome changes of two barley lines contrasting in salt tolerance (DH14, DH 187) in response to salt stress after seed priming in water or in 800 μM BABA. Seed priming with BABA significantly increased (p ≤ 0.05) RWC in both barley lines, which indicates considerably lower water loss in BABA-primed plants than in the non-primed control plants. Dry and fresh matter increased significantly in line DH 187, whereas no changes were detected in line DH14. BABA-primed plants of both lines showed different proteomic patterns than the non-primed control plants. The root proteins exhibiting significant abundance changes (1.75-fold change, p ≤ 0.05) were separated by two-dimensional polyacrylamide gel electrophoresis (2D- PAGE). Thirty-one spots, representing 24 proteins, were successfully identified by MALDI TOF/TOF mass spectrometry. The most prominent differences include the up-regulation of antioxidant enzymes (catalase, peroxidase and superoxide dismutase), PR proteins (chitinase, endo-1,3-β-glucosidase), and chaperones (cyclophilin, HSC 70). Our results indicate that BABA induces defence and detoxification processes which may enable faster and more effective responses to salt stress, increasing the chances of survival under adverse environmental conditions.

  7. Plasmalemmal and Vesicular γ-Aminobutyric Acid Transporter Expression in the Developing Mouse Retina

    PubMed Central

    GUO, CHENYING; STELLA, SALVATORE L.; HIRANO, ARLENE A.; BRECHA, NICHOLAS C.

    2009-01-01

    Plasmalemmal and vesicular γ-aminobutyric acid (GABA) transporters influence neurotransmission by regulating high-affinity GABA uptake and GABA release into the synaptic cleft and extracellular space. Postnatal expression of the plasmalemmal GABA transporter-1 (GAT-1), GAT-3, and the vesicular GABA/glycine transporter (VGAT) were evaluated in the developing mouse retina by using immunohistochemistry with affinity-purified antibodies. Weak transporter immunoreactivity was observed in the inner retina at postnatal day 0 (P0). GAT-1 immunostaining at P0 and at older ages was in amacrine and displaced amacrine cells in the inner nuclear layer (INL) and ganglion cell layer (GCL), respectively, and in their processes in the inner plexiform layer (IPL). At P10, weak GAT-1 immunostaining was in Müller cell processes. GAT-3 immunostaining at P0 and older ages was in amacrine cells and their processes, as well as in Müller cells and their processes that extended radially across the retina. At P10, Müller cell somata were observed in the middle of the INL. VGAT immunostaining was present at P0 and older ages in amacrine cells in the INL as well as processes in the IPL. At P5, weak VGAT immunostaining was also observed in horizontal cell somata and processes. By P15, the GAT and VGAT immunostaining patterns appear similar to the adult immunostaining patterns; they reached adult levels by about P20. These findings demonstrate that GABA uptake and release are initially established in the inner retina during the first postnatal week and that these systems subsequently mature in the outer retina during the second postnatal week. PMID:18975268

  8. Efficient Modulation of γ-Aminobutyric Acid Type A Receptors by Piperine Derivatives

    PubMed Central

    2014-01-01

    Piperine activates TRPV1 (transient receptor potential vanilloid type 1 receptor) receptors and modulates γ-aminobutyric acid type A receptors (GABAAR). We have synthesized a library of 76 piperine analogues and analyzed their effects on GABAAR by means of a two-microelectrode voltage-clamp technique. GABAAR were expressed in Xenopus laevis oocytes. Structure–activity relationships (SARs) were established to identify structural elements essential for efficiency and potency. Efficiency of piperine derivatives was significantly increased by exchanging the piperidine moiety with either N,N-dipropyl, N,N-diisopropyl, N,N-dibutyl, p-methylpiperidine, or N,N-bis(trifluoroethyl) groups. Potency was enhanced by replacing the piperidine moiety by N,N-dibutyl, N,N-diisobutyl, or N,N-bistrifluoroethyl groups. Linker modifications did not substantially enhance the effect on GABAAR. Compound 23 [(2E,4E)-5-(1,3-benzodioxol-5-yl)-N,N-dipropyl-2,4-pentadienamide] induced the strongest modulation of GABAA (maximal GABA-induced chloride current modulation (IGABA-max = 1673% ± 146%, EC50 = 51.7 ± 9.5 μM), while 25 [(2E,4E)-5-(1,3-benzodioxol-5-yl)-N,N-dibutyl-2,4-pentadienamide] displayed the highest potency (EC50 = 13.8 ± 1.8 μM, IGABA-max = 760% ± 47%). Compound 23 induced significantly stronger anxiolysis in mice than piperine and thus may serve as a starting point for developing novel GABAAR modulators. PMID:24905252

  9. Conformational preferences of γ-aminobutyric acid in the gas phase and in water

    NASA Astrophysics Data System (ADS)

    Song, Il Keun; Kang, Young Kee

    2012-09-01

    The conformational study of γ-aminobutyric acid (GABA) has been carried out at the M06-2X/cc-pVTZ level of theory in the gas phase and the SMD M06-2X/cc-pVTZ level of theory in water. In the gas phase, the folded conformation gG1 with gauche- and gauche+ conformations for the Cβsbnd Cα and Cγsbnd Cβ bonds, respectively, is found to be lowest in energy and enthalpy, which can be ascribed to the favored hyperconjugative n → π* interaction between the lone electron pair of the amine nitrogen atom and the Cdbnd O bond of the carboxylic group and the favored antiparallel dipole-dipole interaction between the Nsbnd H bond and the Cdbnd O bond. In addition, the intramolecular hydrogen bonds between the carboxylic group and the amine Nsbnd H group have contributed to stabilize some low-energy conformers. However, the most preferred conformation is found to be tG1 and more stable by 0.4 kcal/mol in ΔG than the conformer gG1, in which the favored entropic term due to the conformational flexibility and the other favored n → σ*, σ → σ*, and π → σ* interactions seem to play a role. The conformational preferences of the neutral GABA calculated by ΔG's are reasonably consistent with the populations deduced from FT microwave spectroscopy in supersonic jets combined with laser ablation. In water, the two folded conformers Gg and gG of the zwitterionic GABA are dominantly populated, each of which has the population of 47%, and the hydrogen bond between the ammonium Nsbnd H group and the lone electron pair of the Csbnd O- group seems to be crucial in stabilizing these conformers. Our calculated result that the folded conformers preferentially exist in water is consistent with the 1H NMR experiments in D2O.

  10. Simultaneous detection of resolved glutamate, glutamine, and γ-aminobutyric acid at 4 T

    NASA Astrophysics Data System (ADS)

    Hu, Jiani; Yang, Shaolin; Xuan, Yang; Jiang, Quan; Yang, Yihong; Haacke, E. Mark

    2007-04-01

    A new approach is introduced to simultaneously detect resolved glutamate (Glu), glutamine (Gln), and γ-aminobutyric acid (GABA) using a standard STEAM localization pulse sequence with the optimized sequence timing parameters. This approach exploits the dependence of the STEAM spectra of the strongly coupled spin systems of Glu, Gln, and GABA on the echo time TE and the mixing time TM at 4 T to find an optimized sequence parameter set, i.e., {TE, TM}, where the outer-wings of the Glu C4 multiplet resonances around 2.35 ppm, the Gln C4 multiplet resonances around 2.45 ppm, and the GABA C2 multiplet resonance around 2.28 ppm are significantly suppressed and the three resonances become virtual singlets simultaneously and thus resolved. Spectral simulation and optimization were conducted to find the optimized sequence parameters, and phantom and in vivo experiments (on normal human brains, one patient with traumatic brain injury, and one patient with brain tumor) were carried out for verification. The results have demonstrated that the Gln, Glu, and GABA signals at 2.2-2.5 ppm can be well resolved using a standard STEAM sequence with the optimized sequence timing parameters around {82 ms, 48 ms} at 4 T, while the other main metabolites, such as N-acetyl aspartate (NAA), choline (tCho), and creatine (tCr), are still preserved in the same spectrum. The technique can be easily implemented and should prove to be a useful tool for the basic and clinical studies associated with metabolism of Glu, Gln, and/or GABA.

  11. Metabolome analysis of milk fermented by γ-aminobutyric acid-producing Lactococcus lactis.

    PubMed

    Hagi, Tatsuro; Kobayashi, Miho; Nomura, Masaru

    2016-02-01

    γ-Aminobutyric acid (GABA) is one of the most important functional components in fermented foods because of its physiological functions, such as neurotransmission and antihypertensive activities. However, little is known about components other than GABA in GABA-rich fermented foods. A metabolomic approach offers an opportunity to discover bioactive and flavor components in fermented food. To find specific components in milk fermented with GABA-producing Lactococcus lactis 01-7, we compared the components found in GABA-rich fermented milk with those found in control milk fermented without GABA production using capillary electrophoresis time-of-flight mass spectrometry. A principal component analysis score plot showed a clear differentiation between the control milk fermented with L. lactis 01-1, which does not produce GABA, and GABA-rich milk fermented with a combination of L. lactis strains 01-1 and 01-7. As expected, the amount of GABA in GABA-rich fermented milk was much higher (1,216-fold) than that of the control milk. Interestingly, the amount of Orn was also much higher (27-fold) than that of the control milk. Peptide analysis showed that levels of 6 putative angiotensin-I-converting enzyme (ACE)-inhibitory peptides were also higher in the GABA-rich fermented milk. Furthermore, ACE-inhibitory activity of GABA-rich fermented milk tended to be higher than that of the control milk. These results indicate that the GABA-producing strain 01-7 provides fermented milk with other functional components in addition to GABA.

  12. Promoter variants determine γ-aminobutyric acid homeostasis-related gene transcription in human epileptic hippocampi.

    PubMed

    Pernhorst, Katharina; Raabe, Anna; Niehusmann, Pitt; van Loo, Karen M J; Grote, Alexander; Hoffmann, Per; Cichon, Sven; Sander, Thomas; Schoch, Susanne; Becker, Albert J

    2011-12-01

    The functional consequences of single nucleotide polymorphisms associated with episodic brain disorders such as epilepsy and depression are unclear. Allelic associations with generalized epilepsies have been reported for single nucleotide polymorphisms rs1883415 (ALDH5A1; succinic semialdehyde dehydrogenase) and rs4906902 (GABRB3; GABAA β3), both of which are present in the 5' regulatory region of genes involved in γ-aminobutyric acid (GABA) homeostasis. To address their allelic association with episodic brain disorders and allele-specific impact on the transcriptional regulation of these genes in human brain tissue, DNA and messenger RNA (mRNA) isolated from hippocampi were obtained at epilepsy surgery of 146 pharmacoresistant mesial temporal lobe epilepsy (mTLE) patients and from 651 healthy controls. We found that the C allele of rs1883415 is accumulated to a greater extentin mTLE versus controls. By real-time quantitative reverse transcription-polymerase chain reaction analyses, individuals homozygous for the C allele showed higher ALDH5A1 mRNA expression. The rs4906902 G allele of the GABRB3 gene was overrepresented in mTLE patients with depression; individuals homozygous for the G allele showed reduced GABRB3 mRNA expression. Bioinformatic analyses suggest that rs1883415 and rs4906902 alter the DNA binding affinity of the transcription factors Egr-3 in ALDH5A1 and MEF-2 in GABRB3 promoters, respectively. Using in vitro luciferase transfection assays, we observed that, in both cases, the transcription factors regulate gene expression depending on the allelic variant in the same direction as in the human hippocampi. Our data suggest that distinct promoter variants may sensitize individuals for differential, potentially stimulus-induced alterations of GABA homeostasis-relevant gene expression. This might contribute to the episodic onset of symptoms and point to new targets for pharmacotherapies.

  13. β-aminobutyric acid mediated drought stress alleviation in maize (Zea mays L.).

    PubMed

    Shaw, Arun K; Bhardwaj, Pardeep K; Ghosh, Supriya; Roy, Sankhajit; Saha, Suman; Sherpa, Ang R; Saha, Samir K; Hossain, Zahed

    2016-02-01

    The present study highlights the role of β-aminobutyric acid (BABA) in alleviating drought stress effects in maize (Zea mays L.). Chemical priming was imposed by pretreating 1-week-old plants with 600 μM BABA prior to applying drought stress. Specific activities of key antioxidant enzymes and metabolites (ascorbate and glutathione) levels of ascorbate-glutathione cycle were studied to unravel the priming-induced modulation of plant defense system. Furthermore, changes in endogenous ABA and JA concentrations as well as mRNA expressions of key genes involved in their respective biosynthesis pathways were monitored in BABA-primed (BABA+) and non-primed (BABA-) leaves of drought-challenged plants to better understand the mechanistic insights into the BABA-induced hormonal regulation of plant response to water-deficit stress. Accelerated stomatal closure, high relative water content, and less membrane damage were observed in BABA-primed leaves under water-deficit condition. Elevated APX and SOD activity in non-primed leaves found to be insufficient to scavenge all H2O2 and O2 (·-) resulting in oxidative burst as evident after histochemical staining with NBT and DAB. A higher proline accumulation in non-primed leaves also does not give much protection against drought stress. Increased GR activity supported with the enhanced mRNA and protein expressions might help the BABA-primed plants to maintain a high GSH pool essential for sustaining balanced redox status to counter drought-induced oxidative stress damages. Hormonal analysis suggests that in maize, BABA-potentiated drought tolerance is primarily mediated through JA-dependent pathway by the activation of antioxidant defense systems while ABA biosynthesis pathway also plays an important role in fine-tuning of drought stress response.

  14. Role of a gamma-aminobutryic acid (GABA) receptor mutation in the evolution and spread of Diabrotica virgifera virgifera resistance to cyclodiene insecticides

    Technology Transfer Automated Retrieval System (TEKTRAN)

    An alanine to serine amino acid substitution within the Rdl subunit of the gamma-aminobutyric acid (GABA) receptor confers resistance to cyclodiene insecticides in many species. The corn rootworm, Diabrotica virgifera virgifera, is a damaging pest of cultivated corn that was partially controlled by ...

  15. Development of "Pinceaux" formations and dendritic translocation of climbing fibers during the acquisition of the balance between glutamatergic and gamma-aminobutyric acidergic inputs in developing Purkinje cells.

    PubMed

    Sotelo, Constantino

    2008-01-10

    The acquisition of the dynamic balance between excitation and inhibition in developing Purkinje cells, necessary for their proper function, is analyzed. Newborn (P0) mouse cerebellum contains glutamatergic (VGLUT2-IR) and gamma-aminobutyric acid (GABA)-ergic (VIAAT-IR) axons. The former prevail and belong to climbing fibers, whereas the latter neither colabel with calbindin-expressing fibers nor belong to axons of the cortical GABAergic interneurons. During the first postnatal week, VIAAT-IR axons in the Purkinje cell neighborhood remains very low, and the first synapses with basket fibers are formed at P7, when climbing fibers have already established dense pericellular nets. The descending basket fibers reach the Purkinje cell axon initial segment by P9, immediately establishing axoaxonic synapses. The pinceaux appear as primitive vortex-like arrangements by P12, and by P20 interbasket fiber septate-like junctions, typical of fully mature pinceaux, are still missing. The climbing fiber's somatodendritic translocation occurs later than expected, after the regression of the multiple innervation, and follows the ascending collaterals of the basket axons, which are apparently the optimal substrate for the proper subcellular targeting of the climbing fibers. These results emphasize that chemical transmission in the axon initial segment precedes the electrical inhibition generated by field effects. In addition, GABAergic Purkinje cells, as opposed to glutamatergic projection neurons in other cortical structures, do not begin to receive their excitation to inhibition balance until the end of the first postnatal week, despite the early presence of potentially functional GABAergic axons that possess the required vesicular transport system.

  16. Biodiversity and γ-aminobutyric acid production by lactic acid bacteria isolated from traditional alpine raw cow's milk cheeses.

    PubMed

    Franciosi, Elena; Carafa, Ilaria; Nardin, Tiziana; Schiavon, Silvia; Poznanski, Elisa; Cavazza, Agostino; Larcher, Roberto; Tuohy, Kieran M

    2015-01-01

    "Nostrano-cheeses" are traditional alpine cheeses made from raw cow's milk in Trentino-Alto Adige, Italy. This study identified lactic acid bacteria (LAB) developing during maturation of "Nostrano-cheeses" and evaluated their potential to produce γ-aminobutyric acid (GABA), an immunologically active compound and neurotransmitter. Cheese samples were collected on six cheese-making days, in three dairy factories located in different areas of Trentino and at different stages of cheese ripening (24 h, 15 days, and 1, 2, 3, 6, and 8 months). A total of 1,059 LAB isolates were screened using Random Amplified Polymorphic DNA-PCR (RAPD-PCR) and differentiated into 583 clusters. LAB strains from dominant clusters (n = 97) were genetically identified to species level by partial 16S rRNA gene sequencing. LAB species most frequently isolated were Lactobacillus paracasei, Streptococcus thermophilus, and Leuconostoc mesenteroides. The 97 dominant clusters were also characterized for their ability in producing GABA by high-performance liquid chromatography (HPLC). About 71% of the dominant bacteria clusters evolving during cheeses ripening were able to produce GABA. Most GABA producers were Lactobacillus paracasei but other GABA producing species included Lactococcus lactis, Lactobacillus plantarum, Lactobacillus rhamnosus, Pediococcus pentosaceus, and Streptococcus thermophilus. No Enterococcus faecalis or Sc. macedonicus isolates produced GABA. The isolate producing the highest amount of GABA (80.0±2.7 mg/kg) was a Sc. thermophilus.

  17. Lactic Acid Bacterial Starter Culture with Antioxidant and γ-Aminobutyric Acid Biosynthetic Activities Isolated from Flatfish-Sikhae Fermentation.

    PubMed

    Won, Yeong Geol; Yu, Hyun-Hee; Chang, Young-Hyo; Hwang, Han-Joon

    2015-12-01

    The aim of this study is to select a lactic acid bacterial strain as a starter culture for flatfish-Sikhae fermentation and to evaluate its suitability for application in a food system. Four strains of lactic acid bacteria isolated from commercial flatfish-Sikhae were identified and selected as starter culture candidates through investigation of growth rates, salt tolerance, food safety, and functional properties such as antioxidative and antimicrobial activities. The fermentation properties of the starter candidates were also examined in food systems prepared with these strains (candidate batch) in comparison with a spontaneous fermentation process without starter culture (control batch) at 15°C. The results showed that the candidate YG331 batch had better fermentation properties such as viable cell count, pH, and acidity than the other experimental batches, including the control batch. The results are expressed according to selection criteria based on a preliminary sensory evaluation and physiochemical investigation. Also, only a small amount of histamine was detected with the candidate YG331 batch. The radical scavenging activity of the candidate batches was better compared with the control batch, and especially candidate YG331 batch showed the best radical scavenging activity. Also, we isolated another starter candidate (identified as Lactobacillus brevis PM03) with γ-aminobutyric acid (GABA)-producing activity from commercial flatfish-Sikhae products. The sensory scores of the candidate YG331 batch were better than those of the other experimental batches in terms of flavor, color, and overall acceptance. In this study, we established selection criteria for the lactic acid bacterial starter for the flatfish-Sikhae production and finally selected candidate YG331 as the most suitable starter.

  18. Focused pseudostatic hydrazone libraries screened by mass spectrometry binding assay: optimizing affinities toward γ-aminobutyric acid transporter 1.

    PubMed

    Sindelar, Miriam; Lutz, Toni A; Petrera, Marilena; Wanner, Klaus T

    2013-02-14

    Mass spectrometric (MS) binding assays, a powerful tool to determine affinities of single drug candidates toward chosen targets, were recently demonstrated to be suitable for the screening of compound libraries generated with reactions of dynamic combinatorial chemistry when rendering libraries pseudostatic. Screening of small hydrazone libraries targeting γ-aminobutyric acid transporter 1 (GAT1), the most abundant γ-aminobutyric acid (GABA) transporter in the central nervous system, revealed two nipecotic acid derived binders with submicromolar affinities. Starting from the biphenyl carrying hit as lead structure, the objective of the present study was to discover novel high affinity GAT1 binders by screening of biphenyl focused pseudostatic hydrazone libraries formed from hydrazine 10 and 36 biphenylcarbaldehydes 11c-al. Hydrazone 12z that carried a 2',4'-dichlorobiphenyl residue was found to be the most potent binder with low nanomolar affinity (pK(i) = 8.094 ± 0.098). When stable carba analogues of representative hydrazones were synthesized and evaluated, the best binder 13z was again displaying the 2',4'-dichlorobiphenyl moiety (pK(i) = 6.930 ± 0.021).

  19. Metabolic pathways regulated by γ-aminobutyric acid (GABA) contributing to heat tolerance in creeping bentgrass (Agrostis stolonifera)

    PubMed Central

    Li, Zhou; Yu, Jingjin; Peng, Yan; Huang, Bingru

    2016-01-01

    γ-Aminobutyric acid is a non-protein amino acid involved in various metabolic processes. The objectives of this study were to examine whether increased GABA could improve heat tolerance in cool-season creeping bentgrass through physiological analysis, and to determine major metabolic pathways regulated by GABA through metabolic profiling. Plants were pretreated with 0.5 mM GABA or water before exposed to non-stressed condition (21/19 °C) or heat stress (35/30 °C) in controlled growth chambers for 35 d. The growth and physiological analysis demonstrated that exogenous GABA application significantly improved heat tolerance of creeping bentgrass. Metabolic profiling found that exogenous application of GABA led to increases in accumulations of amino acids (glutamic acid, aspartic acid, alanine, threonine, serine, and valine), organic acids (aconitic acid, malic acid, succinic acid, oxalic acid, and threonic acid), sugars (sucrose, fructose, glucose, galactose, and maltose), and sugar alcohols (mannitol and myo-inositol). These findings suggest that GABA-induced heat tolerance in creeping bentgrass could involve the enhancement of photosynthesis and ascorbate-glutathione cycle, the maintenance of osmotic adjustment, and the increase in GABA shunt. The increased GABA shunt could be the supply of intermediates to feed the tricarboxylic acid cycle of respiration metabolism during a long-term heat stress, thereby maintaining metabolic homeostasis. PMID:27455877

  20. Metabolic pathways regulated by γ-aminobutyric acid (GABA) contributing to heat tolerance in creeping bentgrass (Agrostis stolonifera).

    PubMed

    Li, Zhou; Yu, Jingjin; Peng, Yan; Huang, Bingru

    2016-07-26

    γ-Aminobutyric acid is a non-protein amino acid involved in various metabolic processes. The objectives of this study were to examine whether increased GABA could improve heat tolerance in cool-season creeping bentgrass through physiological analysis, and to determine major metabolic pathways regulated by GABA through metabolic profiling. Plants were pretreated with 0.5 mM GABA or water before exposed to non-stressed condition (21/19 °C) or heat stress (35/30 °C) in controlled growth chambers for 35 d. The growth and physiological analysis demonstrated that exogenous GABA application significantly improved heat tolerance of creeping bentgrass. Metabolic profiling found that exogenous application of GABA led to increases in accumulations of amino acids (glutamic acid, aspartic acid, alanine, threonine, serine, and valine), organic acids (aconitic acid, malic acid, succinic acid, oxalic acid, and threonic acid), sugars (sucrose, fructose, glucose, galactose, and maltose), and sugar alcohols (mannitol and myo-inositol). These findings suggest that GABA-induced heat tolerance in creeping bentgrass could involve the enhancement of photosynthesis and ascorbate-glutathione cycle, the maintenance of osmotic adjustment, and the increase in GABA shunt. The increased GABA shunt could be the supply of intermediates to feed the tricarboxylic acid cycle of respiration metabolism during a long-term heat stress, thereby maintaining metabolic homeostasis.

  1. Metabolic pathways regulated by γ-aminobutyric acid (GABA) contributing to heat tolerance in creeping bentgrass (Agrostis stolonifera).

    PubMed

    Li, Zhou; Yu, Jingjin; Peng, Yan; Huang, Bingru

    2016-01-01

    γ-Aminobutyric acid is a non-protein amino acid involved in various metabolic processes. The objectives of this study were to examine whether increased GABA could improve heat tolerance in cool-season creeping bentgrass through physiological analysis, and to determine major metabolic pathways regulated by GABA through metabolic profiling. Plants were pretreated with 0.5 mM GABA or water before exposed to non-stressed condition (21/19 °C) or heat stress (35/30 °C) in controlled growth chambers for 35 d. The growth and physiological analysis demonstrated that exogenous GABA application significantly improved heat tolerance of creeping bentgrass. Metabolic profiling found that exogenous application of GABA led to increases in accumulations of amino acids (glutamic acid, aspartic acid, alanine, threonine, serine, and valine), organic acids (aconitic acid, malic acid, succinic acid, oxalic acid, and threonic acid), sugars (sucrose, fructose, glucose, galactose, and maltose), and sugar alcohols (mannitol and myo-inositol). These findings suggest that GABA-induced heat tolerance in creeping bentgrass could involve the enhancement of photosynthesis and ascorbate-glutathione cycle, the maintenance of osmotic adjustment, and the increase in GABA shunt. The increased GABA shunt could be the supply of intermediates to feed the tricarboxylic acid cycle of respiration metabolism during a long-term heat stress, thereby maintaining metabolic homeostasis. PMID:27455877

  2. A comparative density functional theory study of electronic structure and optical properties of γ-aminobutyric acid and its cocrystals with oxalic and benzoic acid

    NASA Astrophysics Data System (ADS)

    da Silva Filho, J. G.; Freire, V. N.; Caetano, E. W. S.; Ladeira, L. O.; Fulco, U. L.; Albuquerque, E. L.

    2013-11-01

    In this letter, we study the electronic structure and optical properties of the active medicinal component γ-aminobutyric acid (GABA) and its cocrystals with oxalic (OXA) and benzoic (BZA) acid by means of the density functional theory formalism. It is shown that the cocrystallization strongly weakens the zwitterionic character of the GABA molecule leading to striking differences among the electronic band structures and optical absorption spectra of the GABA crystal and GABA:OXA, GABA:BZA cocrystals, originating from distinct sets of hydrogen bonds. Calculated band widths and Δ-sol band gap estimates indicate that both GABA and GABA:OXA, GABA:BZA cocrystals are indirect gap insulators.

  3. Exogenous γ-Aminobutyric Acid Improves the Structure and Function of Photosystem II in Muskmelon Seedlings Exposed to Salinity-Alkalinity Stress

    PubMed Central

    Xu, Weinan; Zhen, Ai; Zhang, Liang; Hu, Xiaohui

    2016-01-01

    Gamma-aminobutyric acid (GABA) is important in plant responses to environmental stresses. We wished to clarify the role of GABA in maintenance of photosynthesis in muskmelon seedlings (Cucumis melo L., cv. Yipintianxia) during saline-alkaline stress. To this end, we assessed the effect of GABA on the structure and function of the photosynthetic apparatus in muskmelon seedlings grown under saline-alkaline stress. These stresses in combination reduced net photosynthetic rate, gas-exchange, and inhibited photosystem II (PSII) electron transport as measured by the JIP-test. They also reduced the activity of chloroplast ATPases and disrupted the internal lamellar system of the thylakoids. Exogenous GABA alleviated the stress-induced reduction of net photosynthesis, the activity of chloroplast ATPases, and overcame some of the damaging effects of stress on the chloroplast structure. Based on interpretation of the JIP-test, we conclude that exogenous GABA alleviated stress-related damage on the acceptor side of PSII. It also restored energy distribution, the reaction center status, and enhanced the ability of PSII to repair reaction centers in stressed seedlings. GABA may play a crucial role in protecting the chloroplast structure and function of PSII against the deleterious effects of salinity-alkalinity stress. PMID:27764179

  4. γ-Aminobutyric acid induces resistance against Penicillium expansum by priming of defence responses in pear fruit.

    PubMed

    Yu, Chen; Zeng, Lizhen; Sheng, Kuang; Chen, Fangxia; Zhou, Tao; Zheng, Xiaodong; Yu, Ting

    2014-09-15

    The results from this study showed that treatment with γ-aminobutyric acid (GABA), at 100-1000 μg/ml, induced strong resistance against blue mould rot caused by Penicillium expansum in pear fruit. Moreover, the activities of five defence-related enzymes (including chitinase, β-1,3-glucanase, phenylalnine ammonialyase, peroxidase and polyphenol oxidase) and the expression of these corresponding genes were markedly and/or promptly enhanced in the treatment with GABA and inoculation with P. expansum compared with those that were treated with GABA or inoculated with pathogen alone. In addition, the treatment of pear with GABA had little adverse effect on the edible quality of the fruit. To the best of our knowledge, this is the first report that GABA can effectively reduce fungal disease of harvested fruit. Its mechanisms may be closely correlated with the induction of fruit resistance by priming activation and expression of defence-related enzymes and genes upon challenge with pathogen.

  5. Quantification of γ-Aminobutyric Acid in Cerebrospinal Fluid Using Liquid Chromatography-Electrospray Tandem Mass Spectrometry.

    PubMed

    Arning, Erland; Bottiglieri, Teodoro

    2016-01-01

    We describe a simple stable isotope dilution method for accurate and precise measurement of γ-aminobutyric acid (GABA), a major inhibitory neurotransmitter in human cerebrospinal fluid (CSF) as a clinical diagnostic test. Determination of GABA in CSF (50 μL) was performed utilizing high performance liquid chromatography coupled with electrospray positive ionization tandem mass spectrometry (HPLC-ESI-MS/MS). Analysis of free and total GABA requires two individual sample preparations and mass spectrometry analyses. Free GABA in CSF is determined by a 1:2 dilution with internal standard (GABA-D2) and injected directly onto the HPLC-ESI-MS/MS system. Determination of total GABA in CSF requires additional sample preparation in order to hydrolyze all the bound GABA in the sample to the free form. This requires hydrolyzing the sample by boiling in acidic conditions (hydrochloric acid) for 4 h. The sample is then further diluted 1:10 with a 90 % acetonitrile/0.1 % formic acid solution and injected into the HPLC-ESI-MS/MS system. Each assay is quantified using a five-point standard curve and is linear from 6 nM to 1000 nM and 0.63 μM to 80 μM for free and total GABA, respectively.

  6. Quantification of γ-Aminobutyric Acid in Cerebrospinal Fluid Using Liquid Chromatography-Electrospray Tandem Mass Spectrometry.

    PubMed

    Arning, Erland; Bottiglieri, Teodoro

    2016-01-01

    We describe a simple stable isotope dilution method for accurate and precise measurement of γ-aminobutyric acid (GABA), a major inhibitory neurotransmitter in human cerebrospinal fluid (CSF) as a clinical diagnostic test. Determination of GABA in CSF (50 μL) was performed utilizing high performance liquid chromatography coupled with electrospray positive ionization tandem mass spectrometry (HPLC-ESI-MS/MS). Analysis of free and total GABA requires two individual sample preparations and mass spectrometry analyses. Free GABA in CSF is determined by a 1:2 dilution with internal standard (GABA-D2) and injected directly onto the HPLC-ESI-MS/MS system. Determination of total GABA in CSF requires additional sample preparation in order to hydrolyze all the bound GABA in the sample to the free form. This requires hydrolyzing the sample by boiling in acidic conditions (hydrochloric acid) for 4 h. The sample is then further diluted 1:10 with a 90 % acetonitrile/0.1 % formic acid solution and injected into the HPLC-ESI-MS/MS system. Each assay is quantified using a five-point standard curve and is linear from 6 nM to 1000 nM and 0.63 μM to 80 μM for free and total GABA, respectively. PMID:26602123

  7. Bicyclic γ-amino acids as inhibitors of γ-aminobutyrate aminotransferase.

    PubMed

    Pinto, Andrea; Tamborini, Lucia; Pennacchietti, Eugenia; Coluccia, Antonio; Silvestri, Romano; Cullia, Gregorio; De Micheli, Carlo; Conti, Paola; De Biase, Daniela

    2016-01-01

    The γ-aminobutyrate (GABA)-degradative enzyme GABA aminotransferase (GABA-AT) is regarded as an attractive target to control GABA levels in the central nervous system: this has important implications in the treatment of several neurological disorders and drug dependencies. We have investigated the ability of newly synthesized compounds to act as GABA-AT inhibitors. These compounds have a unique bicyclic structure: the carbocyclic ring bears the GABA skeleton, while the fused 3-Br-isoxazoline ring contains an electrophilic warhead susceptible of nucleophilic attack by an active site residue of the target enzyme. Out of the four compounds tested, only the one named (+)-3 was found to significantly inhibit mammalian GABA-AT in vitro. Docking studies, performed on the available structures of GABA-AT, support the experimental findings: out of the four tested compounds, only (+)-3 suitably orients the electrophilic 3-Br-isoxazoline warhead towards the active site nucleophilic residue Lys329, thereby explaining the irreversible inhibition of GABA-AT observed experimentally. PMID:25807299

  8. Exogenous γ-aminobutyric acid treatment affects citrate and amino acid accumulation to improve fruit quality and storage performance of postharvest citrus fruit.

    PubMed

    Sheng, Ling; Shen, Dandan; Luo, Yi; Sun, Xiaohua; Wang, Jinqiu; Luo, Tao; Zeng, Yunliu; Xu, Juan; Deng, Xiuxin; Cheng, Yunjiang

    2017-02-01

    The loss of organic acids during postharvest storage is one of the major factors that reduces the fruit quality and economic value of citrus. Citrate is the most important organic acid in citrus fruits. Molecular evidence has proved that γ-aminobutyric acid (GABA) shunt plays a key role in citrate metabolism. Here, we investigated the effects of exogenous GABA treatment on citrate metabolism and storage quality of postharvest citrus fruit. The content of citrate was significantly increased, which was primarily attributed to the inhibition of the expression of glutamate decarboxylase (GAD). Amino acids, including glutamate, alanine, serine, aspartate and proline, were also increased. Moreover, GABA treatment decreased the fruit rot rate. The activities of antioxidant enzymes and the content of energy source ATP were affected by the treatment. Our results indicate that GABA treatment is a very effective approach for postharvest quality maintenance and improvement of storage performance in citrus production. PMID:27596402

  9. Oligomerization of L-gamma-carboxyglutamic acid

    NASA Technical Reports Server (NTRS)

    Hill, A. R. Jr; Orgel, L. E.; Bada, J. L. (Principal Investigator)

    1999-01-01

    Unlike glutamic acid, L-gamma-carboxyglutamic acid does not oligomerize efficiently when treated with carbonyldiimidazole in aqueous solution. However, divalent ions such as Mg2+ catalyze the reaction, and lead to the formation of oligomers in good yield. In the presence of hydroxylapatite, L-gamma-carboxyglutamic acid oligomerizes efficiently in a reaction that proceeds in the absence of divalent ions but is further catalyzed when they are present. After 'feeding' 50 times with activated amino acid in the presence of the Mg2+ ion, oligomers longer than the 20-mer could be detected. The effect of hydroxylapatite on peptide elongation is very sensitive to the nature of the activated amino acid and the acceptor peptide. Glutamic acid oligomerizes more efficiently than L-gamma-carboxyglutamic acid on hydroxylapatite and adds more efficiently to decaglutamic acid in solution. One might, therefore, expect that glutamic acid would add more efficiently than L-gamma-carboxyglutamic acid to decaglutamic acid on hydroxylapatite. The contrary is true--the addition of L-gamma-carboxyglutamic acid is substantially more efficient. This suggests that oligomerization on the surface of hydroxylapatite depends on the detailed match between the structure of the surface of the mineral and the structure of the oligomer.

  10. Thermodynamics of DL-α-aminobutyric acid induced solvation mechanism in aqueous KCl solutions at 288.15-308.15 K

    NASA Astrophysics Data System (ADS)

    Mondal, S.; Ghosh, S.; Hossain, A.; Mahali, K.; Roy, S.; Dolui, B. K.

    2016-09-01

    The solubilities of DL-α-aminobutyric acid in KCl solutions of different concentrations are measured at 288.15-308.15 K. Gibbs energies and entropies have been determined for transfer of α-aminobutyric acid form water to aqueous KCl solution at 298.15 K. The cavity, dipole-dipole and other interactions affecting the solubility, as well as stability of the amino acid in solution are also evaluated. Gibbs energy and entropy of transfer due to interactions are computed to create the model of the complex solute-solvent and solventsolvent interactions. Molar volume, densities, dipole moment of solvent and diameter of co-solvent in aqueous potassium chloride are also evaluated.

  11. Analysis of cerebrospinal fluid γ-aminobutyric acid by capillary electrophoresis with laser-induced fluorescence detection.

    PubMed

    Casado, Mercedes; Molero, Marta; Sierra, Cristina; García-Cazorla, Angels; Ormazabal, Aida; Artuch, Rafael

    2014-04-01

    The measurement of γ-aminobutyric acid (GABA) is suitable for investigating various neurological disorders. In this study, a sensitive and selective method for free GABA quantification in cerebrospinal fluid (CSF) has been standardised. This method is based on CE with LIF detection using 4-fluoro-7-nitrobenzo-2-oxa-1,3-diazole (NBD-F) as a derivatisating agent. The reaction conditions (NBD-F concentration, pH, temperature and reaction time) and the electrophoretic parameters (run buffer composition and pH and separation voltage) were optimised to obtain the maximum derivatisation efficiency and electrophoretic resolution. The best resolution was obtained using 200 mM sodium borate, 10 mM SDS, 8.5 mM β-CD, pH 10 and 20 kV voltage. The method was linear in the concentration range of 2.5-1000 nM with good inter- and intra-assay precision values. The effects of CSF handling on free GABA concentrations were also evaluated. Our results show that the time delay between CSF collection and freezing strongly increases the CSF GABA values. Age-related reference values were established in 55 paediatric controls. The influence of antiepileptic therapy on free CSF GABA was studied in 38 neuropaediatric patients. Significantly, higher GABA values were obtained in patients taking valproic acid or vigabatrin therapy, which are antiepileptic drugs that modulate GABA metabolism.

  12. Novel fermented chickpea milk with enhanced level of γ-aminobutyric acid and neuroprotective effect on PC12 cells

    PubMed Central

    Li, Wen; Wei, Mingming; Wu, Junjun; Rui, Xin

    2016-01-01

    In this study, novel fermented chickpea milk with high γ -aminobutyric acid (GABA) content and potential neuroprotective activity was developed. Fermentation starter that can produce GABA was selected from 377 strains of lactic acid bacteria isolated from traditional Chinese fermented foods. Among the screened strains, strain M-6 showed the highest GABA-producing capacity in De Man–Rogosa and Sharp (MRS) broth and chickpea milk. M-6 was identified as Lactobacillus plantarum based on Gram staining, API carbohydrate fermentation pattern testing, and 16s rDNA sequencing. The complete gene encoding glutamate decarboxylase was cloned to confirm the presence of the gene in L. plantarum M-6. The fermentation condition was optimized by response surface methodology. Results demonstrated that L. plantarum M-6 produced the highest GABA content of 537.23 mg/L. The optimal condition included an inoculum concentration of 7%, presence of 0.2% (m/v) monosodium glutamate and 55 µ M pyridoxal-5-phosphate, incubation temperature of 39 °C and fermentation time of 48 h . GABA-enriched chickpea milk exerted protective effects on PC12 cells against MnCl2 -induced injury. GABA-enriched chickpea milk improved cell viability and markedly attenuated the release of lactate dehydrogenase compared with the impaired cells. PMID:27602272

  13. Evaluation of commercial soy sauce koji strains of Aspergillus oryzae for γ-aminobutyric acid (GABA) production.

    PubMed

    Ab Kadir, Safuan; Wan-Mohtar, Wan Abd Al Qadr Imad; Mohammad, Rosfarizan; Abdul Halim Lim, Sarina; Sabo Mohammed, Abdulkarim; Saari, Nazamid

    2016-10-01

    In this study, four selected commercial strains of Aspergillus oryzae were collected from soy sauce koji. These A. oryzae strains designated as NSK, NSZ, NSJ and NST shared similar morphological characteristics with the reference strain (A. oryzae FRR 1675) which confirmed them as A. oryzae species. They were further evaluated for their ability to produce γ-aminobutyric acid (GABA) by cultivating the spore suspension in a broth medium containing 0.4 % (w/v) of glutamic acid as a substrate for GABA production. The results showed that these strains were capable of producing GABA; however, the concentrations differed significantly (P < 0.05) among themselves. Based on the A. oryzae strains, highest GABA concentration was obtained from NSK (194 mg/L) followed by NSZ (63 mg/L), NSJ (51.53 mg/L) and NST (31.66 mg/L). Therefore, A. oryzae NSK was characterized and the sequence was found to be similar to A. oryzae and A. flavus with 99 % similarity. The evolutionary distance (K nuc) between sequences of identical fungal species was calculated and a phylogenetic tree prepared from the K nuc data showed that the isolate belonged to the A. oryzae species. This finding may allow the development of GABA-rich ingredients using A. oryzae NSK as a starter culture for soy sauce production.

  14. Evaluation of commercial soy sauce koji strains of Aspergillus oryzae for γ-aminobutyric acid (GABA) production.

    PubMed

    Ab Kadir, Safuan; Wan-Mohtar, Wan Abd Al Qadr Imad; Mohammad, Rosfarizan; Abdul Halim Lim, Sarina; Sabo Mohammed, Abdulkarim; Saari, Nazamid

    2016-10-01

    In this study, four selected commercial strains of Aspergillus oryzae were collected from soy sauce koji. These A. oryzae strains designated as NSK, NSZ, NSJ and NST shared similar morphological characteristics with the reference strain (A. oryzae FRR 1675) which confirmed them as A. oryzae species. They were further evaluated for their ability to produce γ-aminobutyric acid (GABA) by cultivating the spore suspension in a broth medium containing 0.4 % (w/v) of glutamic acid as a substrate for GABA production. The results showed that these strains were capable of producing GABA; however, the concentrations differed significantly (P < 0.05) among themselves. Based on the A. oryzae strains, highest GABA concentration was obtained from NSK (194 mg/L) followed by NSZ (63 mg/L), NSJ (51.53 mg/L) and NST (31.66 mg/L). Therefore, A. oryzae NSK was characterized and the sequence was found to be similar to A. oryzae and A. flavus with 99 % similarity. The evolutionary distance (K nuc) between sequences of identical fungal species was calculated and a phylogenetic tree prepared from the K nuc data showed that the isolate belonged to the A. oryzae species. This finding may allow the development of GABA-rich ingredients using A. oryzae NSK as a starter culture for soy sauce production. PMID:27541157

  15. Novel fermented chickpea milk with enhanced level of γ-aminobutyric acid and neuroprotective effect on PC12 cells.

    PubMed

    Li, Wen; Wei, Mingming; Wu, Junjun; Rui, Xin; Dong, Mingsheng

    2016-01-01

    In this study, novel fermented chickpea milk with high γ -aminobutyric acid (GABA) content and potential neuroprotective activity was developed. Fermentation starter that can produce GABA was selected from 377 strains of lactic acid bacteria isolated from traditional Chinese fermented foods. Among the screened strains, strain M-6 showed the highest GABA-producing capacity in De Man-Rogosa and Sharp (MRS) broth and chickpea milk. M-6 was identified as Lactobacillus plantarum based on Gram staining, API carbohydrate fermentation pattern testing, and 16s rDNA sequencing. The complete gene encoding glutamate decarboxylase was cloned to confirm the presence of the gene in L. plantarum M-6. The fermentation condition was optimized by response surface methodology. Results demonstrated that L. plantarum M-6 produced the highest GABA content of 537.23 mg/L. The optimal condition included an inoculum concentration of 7%, presence of 0.2% (m/v) monosodium glutamate and 55 µ M pyridoxal-5-phosphate, incubation temperature of 39 °C and fermentation time of 48 h . GABA-enriched chickpea milk exerted protective effects on PC12 cells against MnCl2 -induced injury. GABA-enriched chickpea milk improved cell viability and markedly attenuated the release of lactate dehydrogenase compared with the impaired cells. PMID:27602272

  16. Analysis of cerebrospinal fluid γ-aminobutyric acid by capillary electrophoresis with laser-induced fluorescence detection.

    PubMed

    Casado, Mercedes; Molero, Marta; Sierra, Cristina; García-Cazorla, Angels; Ormazabal, Aida; Artuch, Rafael

    2014-04-01

    The measurement of γ-aminobutyric acid (GABA) is suitable for investigating various neurological disorders. In this study, a sensitive and selective method for free GABA quantification in cerebrospinal fluid (CSF) has been standardised. This method is based on CE with LIF detection using 4-fluoro-7-nitrobenzo-2-oxa-1,3-diazole (NBD-F) as a derivatisating agent. The reaction conditions (NBD-F concentration, pH, temperature and reaction time) and the electrophoretic parameters (run buffer composition and pH and separation voltage) were optimised to obtain the maximum derivatisation efficiency and electrophoretic resolution. The best resolution was obtained using 200 mM sodium borate, 10 mM SDS, 8.5 mM β-CD, pH 10 and 20 kV voltage. The method was linear in the concentration range of 2.5-1000 nM with good inter- and intra-assay precision values. The effects of CSF handling on free GABA concentrations were also evaluated. Our results show that the time delay between CSF collection and freezing strongly increases the CSF GABA values. Age-related reference values were established in 55 paediatric controls. The influence of antiepileptic therapy on free CSF GABA was studied in 38 neuropaediatric patients. Significantly, higher GABA values were obtained in patients taking valproic acid or vigabatrin therapy, which are antiepileptic drugs that modulate GABA metabolism. PMID:24338894

  17. Novel fermented chickpea milk with enhanced level of γ-aminobutyric acid and neuroprotective effect on PC12 cells

    PubMed Central

    Li, Wen; Wei, Mingming; Wu, Junjun; Rui, Xin

    2016-01-01

    In this study, novel fermented chickpea milk with high γ -aminobutyric acid (GABA) content and potential neuroprotective activity was developed. Fermentation starter that can produce GABA was selected from 377 strains of lactic acid bacteria isolated from traditional Chinese fermented foods. Among the screened strains, strain M-6 showed the highest GABA-producing capacity in De Man–Rogosa and Sharp (MRS) broth and chickpea milk. M-6 was identified as Lactobacillus plantarum based on Gram staining, API carbohydrate fermentation pattern testing, and 16s rDNA sequencing. The complete gene encoding glutamate decarboxylase was cloned to confirm the presence of the gene in L. plantarum M-6. The fermentation condition was optimized by response surface methodology. Results demonstrated that L. plantarum M-6 produced the highest GABA content of 537.23 mg/L. The optimal condition included an inoculum concentration of 7%, presence of 0.2% (m/v) monosodium glutamate and 55 µ M pyridoxal-5-phosphate, incubation temperature of 39 °C and fermentation time of 48 h . GABA-enriched chickpea milk exerted protective effects on PC12 cells against MnCl2 -induced injury. GABA-enriched chickpea milk improved cell viability and markedly attenuated the release of lactate dehydrogenase compared with the impaired cells.

  18. Identification and Characterization of γ-Aminobutyric Acid Uptake System GabPCg (NCgl0464) in Corynebacterium glutamicum

    PubMed Central

    Zhao, Zhi; Ma, Wen-hua; Zhou, Ning-Yi

    2012-01-01

    Corynebacterium glutamicum is widely used for industrial production of various amino acids and vitamins, and there is growing interest in engineering this bacterium for more commercial bioproducts such as γ-aminobutyric acid (GABA). In this study, a C. glutamicum GABA-specific transporter (GabPCg) encoded by ncgl0464 was identified and characterized. GabPCg plays a major role in GABA uptake and is essential to C. glutamicum growing on GABA. GABA uptake by GabPCg was weakly competed by l-Asn and l-Gln and stimulated by sodium ion (Na+). The Km and Vmax values were determined to be 41.1 ± 4.5 μM and 36.8 ± 2.6 nmol min−1 (mg dry weight [DW])−1, respectively, at pH 6.5 and 34.2 ± 1.1 μM and 67.3 ± 1.0 nmol min−1 (mg DW)−1, respectively, at pH 7.5. GabPCg has 29% amino acid sequence identity to a previously and functionally identified aromatic amino acid transporter (TyrP) of Escherichia coli but low identities to the currently known GABA transporters (17% and 15% to E. coli GabP and Bacillus subtilis GabP, respectively). The mutant RES167 Δncgl0464/pGXKZ9 with the GabPCg deletion showed 12.5% higher productivity of GABA than RES167/pGXKZ9. It is concluded that GabPCg represents a new type of GABA transporter and is potentially important for engineering GABA-producing C. glutamicum strains. PMID:22307305

  19. Genetic manipulation of the γ-aminobutyric acid (GABA) shunt in rice: overexpression of truncated glutamate decarboxylase (GAD2) and knockdown of γ-aminobutyric acid transaminase (GABA-T) lead to sustained and high levels of GABA accumulation in rice kernels.

    PubMed

    Shimajiri, Yasuka; Oonishi, Takayuki; Ozaki, Kae; Kainou, Kumiko; Akama, Kazuhito

    2013-06-01

    Gamma-aminobutyric acid (GABA) is a non-protein amino acid commonly present in all organisms. Because cellular levels of GABA in plants are mainly regulated by synthesis (glutamate decarboxylase, GAD) and catabolism (GABA-transaminase, GABA-T), we attempted seed-specific manipulation of the GABA shunt to achieve stable GABA accumulation in rice. A truncated GAD2 sequence, one of five GAD genes, controlled by the glutelin (GluB-1) or rice embryo globulin promoters (REG) and GABA-T-based trigger sequences in RNA interference (RNAi) cassettes controlled by one of these promoters as well, was introduced into rice (cv. Koshihikari) to establish stable transgenic lines under herbicide selection using pyriminobac. T₁ and T₂ generations of rice lines displayed high GABA concentrations (2-100 mg/100 g grain). In analyses of two selected lines from the T₃ generation, there was a strong correlation between GABA level and the expression of truncated GAD2, whereas the inhibitory effect of GABA-T expression was relatively weak. In these two lines both with two T-DNA copies, their starch, amylose, and protein levels were slightly lower than non-transformed cv. Koshihikari. Free amino acid analysis of mature kernels of these lines demonstrated elevated levels of GABA (75-350 mg/100 g polished rice) and also high levels of several amino acids, such as Ala, Ser, and Val. Because these lines of seeds could sustain their GABA content after harvest (up to 6 months), the strategy in this study could lead to the accumulation GABA and for these to be sustained in the edible parts.

  20. Extraction, purification and anti-fatigue activity of γ-aminobutyric acid from mulberry (Morus alba L.) leaves

    NASA Astrophysics Data System (ADS)

    Chen, Hengwen; He, Xuanhui; Liu, Yan; Li, Jun; He, Qingyong; Zhang, Cuiying; Wei, Benjun; Zhang, Ye; Wang, Jie

    2016-01-01

    Mulberry (Morus alba L.) is a tree species of Moraceae widely distributed in Southern China. In the present study, the white crystal of γ-aminobutyric acid (GABA) was purified from mulberry leaves, and its bioactivity was also investigated. The main results were as follows: first, the crude GABA was extracted from mulberry leaves by using biochemical methods. Then, the crude was purified by chromatography over an S-8 macroporous resin, Sephadex G-10, and 732 cation exchange resin to yield a white crystal. Lavage administration and exposure of GABA to male NIH mice showed no adverse effects on their growth and development. In an endurance capacity test, the average loaded-swimming time of medium dose was 111.60% longer than the control (P < 0.01). Further investigations showed that relative to that of model control, the respective blood lactate (BL) concentrations of low- and medium-dose were 28.52% and 28.81% lower (P < 0.05), whereas the levels of blood urea nitrogen (BUN) were 36.83% and 40.54% lower (P < 0.05), and that of liver glycogen (LG) levels were 12.81% and 17.22% lower (P < 0.05). The results indicated that GABA has an advantage over taurine of anti-fatigue effect. These findings were indicative of the anti-fatigue activity of GABA.

  1. Extraction, purification and anti-fatigue activity of γ-aminobutyric acid from mulberry (Morus alba L.) leaves.

    PubMed

    Chen, Hengwen; He, Xuanhui; Liu, Yan; Li, Jun; He, Qingyong; Zhang, Cuiying; Wei, Benjun; Zhang, Ye; Wang, Jie

    2016-01-01

    Mulberry (Morus alba L.) is a tree species of Moraceae widely distributed in Southern China. In the present study, the white crystal of γ-aminobutyric acid (GABA) was purified from mulberry leaves, and its bioactivity was also investigated. The main results were as follows: first, the crude GABA was extracted from mulberry leaves by using biochemical methods. Then, the crude was purified by chromatography over an S-8 macroporous resin, Sephadex G-10, and 732 cation exchange resin to yield a white crystal. Lavage administration and exposure of GABA to male NIH mice showed no adverse effects on their growth and development. In an endurance capacity test, the average loaded-swimming time of medium dose was 111.60% longer than the control (P < 0.01). Further investigations showed that relative to that of model control, the respective blood lactate (BL) concentrations of low- and medium-dose were 28.52% and 28.81% lower (P < 0.05), whereas the levels of blood urea nitrogen (BUN) were 36.83% and 40.54% lower (P < 0.05), and that of liver glycogen (LG) levels were 12.81% and 17.22% lower (P < 0.05). The results indicated that GABA has an advantage over taurine of anti-fatigue effect. These findings were indicative of the anti-fatigue activity of GABA. PMID:26743028

  2. Extraction, purification and anti-fatigue activity of γ-aminobutyric acid from mulberry (Morus alba L.) leaves

    PubMed Central

    Chen, Hengwen; He, Xuanhui; Liu, Yan; Li, Jun; He, Qingyong; Zhang, Cuiying; Wei, Benjun; Zhang, Ye; Wang, Jie

    2016-01-01

    Mulberry (Morus alba L.) is a tree species of Moraceae widely distributed in Southern China. In the present study, the white crystal of γ-aminobutyric acid (GABA) was purified from mulberry leaves, and its bioactivity was also investigated. The main results were as follows: first, the crude GABA was extracted from mulberry leaves by using biochemical methods. Then, the crude was purified by chromatography over an S-8 macroporous resin, Sephadex G-10, and 732 cation exchange resin to yield a white crystal. Lavage administration and exposure of GABA to male NIH mice showed no adverse effects on their growth and development. In an endurance capacity test, the average loaded-swimming time of medium dose was 111.60% longer than the control (P < 0.01). Further investigations showed that relative to that of model control, the respective blood lactate (BL) concentrations of low- and medium-dose were 28.52% and 28.81% lower (P < 0.05), whereas the levels of blood urea nitrogen (BUN) were 36.83% and 40.54% lower (P < 0.05), and that of liver glycogen (LG) levels were 12.81% and 17.22% lower (P < 0.05). The results indicated that GABA has an advantage over taurine of anti-fatigue effect. These findings were indicative of the anti-fatigue activity of GABA. PMID:26743028

  3. Pyridoxine Supplementation Improves the Activity of Recombinant Glutamate Decarboxylase and the Enzymatic Production of Gama-Aminobutyric Acid

    PubMed Central

    Huang, Yan; Su, Lingqia; Wu, Jing

    2016-01-01

    Glutamate decarboxylase (GAD) catalyzes the irreversible decarboxylation of L-glutamate to the valuable food supplement γ-aminobutyric acid (GABA). In this study, GAD from Escherichia coli K12, a pyridoxal phosphate (PLP)-dependent enzyme, was overexpressed in E. coli. The GAD produced in media supplemented with 0.05 mM soluble vitamin B6 analog pyridoxine hydrochloride (GAD-V) activity was 154.8 U mL-1, 1.8-fold higher than that of GAD obtained without supplementation (GAD-C). Purified GAD-V exhibited increased activity (193.4 U mg-1, 1.5-fold higher than that of GAD-C), superior thermostability (2.8-fold greater than that of GAD-C), and higher kcat/Km (1.6-fold higher than that of GAD-C). Under optimal conditions in reactions mixtures lacking added PLP, crude GAD-V converted 500 g L-1 monosodium glutamate (MSG) to GABA with a yield of 100%, and 750 g L-1 MSG with a yield of 88.7%. These results establish the utility of pyridoxine supplementation and lay the foundation for large-scale enzymatic production of GABA. PMID:27438707

  4. Factors influencing diamine oxidase activity and γ-aminobutyric acid content of fava bean (Vicia faba L.) during germination.

    PubMed

    Yang, Runqiang; Chen, Hui; Gu, Zhenxin

    2011-11-01

    Factors (germination time, spectra, temperature, pH, and chemical inhibitors) influencing diamine oxidase (DAO, EC 1.4.3.6) activity and γ-aminobutyric acid (GABA) content of fava bean (Vicia faba L.) during germination were investigated in this study. DAO activity significantly increased in germinating seeds but varied with different organs. The enzyme activity was higher in shoot than that in cotyledon, hypocotyl, and radicle. When seeds were germinated in the dark, DAO activity was 2.35-, 2.00-, 2.36-, 4.40-, and 1.67-fold of that under white, red, blue, green, and yellow spectra, respectively. The optimum germination temperature and pH value for increasing DAO activity were 30 °C and 3.0, respectively. The DAO activity was inhibited significantly by aminoguanidine and sodium ethylenediamine tetracetate, while it was activated by CuCl(2) and CaCl(2). Germinating at an appropriate temperature and pH, 30% of GABA formation was supplied by DAO. Calcium was related to the regulation of DAO activity and GABA accumulation. PMID:21942768

  5. Factors influencing diamine oxidase activity and γ-aminobutyric acid content of fava bean (Vicia faba L.) during germination.

    PubMed

    Yang, Runqiang; Chen, Hui; Gu, Zhenxin

    2011-11-01

    Factors (germination time, spectra, temperature, pH, and chemical inhibitors) influencing diamine oxidase (DAO, EC 1.4.3.6) activity and γ-aminobutyric acid (GABA) content of fava bean (Vicia faba L.) during germination were investigated in this study. DAO activity significantly increased in germinating seeds but varied with different organs. The enzyme activity was higher in shoot than that in cotyledon, hypocotyl, and radicle. When seeds were germinated in the dark, DAO activity was 2.35-, 2.00-, 2.36-, 4.40-, and 1.67-fold of that under white, red, blue, green, and yellow spectra, respectively. The optimum germination temperature and pH value for increasing DAO activity were 30 °C and 3.0, respectively. The DAO activity was inhibited significantly by aminoguanidine and sodium ethylenediamine tetracetate, while it was activated by CuCl(2) and CaCl(2). Germinating at an appropriate temperature and pH, 30% of GABA formation was supplied by DAO. Calcium was related to the regulation of DAO activity and GABA accumulation.

  6. Pyridoxine Supplementation Improves the Activity of Recombinant Glutamate Decarboxylase and the Enzymatic Production of Gama-Aminobutyric Acid.

    PubMed

    Huang, Yan; Su, Lingqia; Wu, Jing

    2016-01-01

    Glutamate decarboxylase (GAD) catalyzes the irreversible decarboxylation of L-glutamate to the valuable food supplement γ-aminobutyric acid (GABA). In this study, GAD from Escherichia coli K12, a pyridoxal phosphate (PLP)-dependent enzyme, was overexpressed in E. coli. The GAD produced in media supplemented with 0.05 mM soluble vitamin B6 analog pyridoxine hydrochloride (GAD-V) activity was 154.8 U mL-1, 1.8-fold higher than that of GAD obtained without supplementation (GAD-C). Purified GAD-V exhibited increased activity (193.4 U mg-1, 1.5-fold higher than that of GAD-C), superior thermostability (2.8-fold greater than that of GAD-C), and higher kcat/Km (1.6-fold higher than that of GAD-C). Under optimal conditions in reactions mixtures lacking added PLP, crude GAD-V converted 500 g L-1 monosodium glutamate (MSG) to GABA with a yield of 100%, and 750 g L-1 MSG with a yield of 88.7%. These results establish the utility of pyridoxine supplementation and lay the foundation for large-scale enzymatic production of GABA. PMID:27438707

  7. Determination and comparison of γ-aminobutyric acid (GABA) content in pu-erh and other types of Chinese tea.

    PubMed

    Zhao, Ming; Ma, Yan; Wei, Zhen-zhen; Yuan, Wen-xia; Li, Ya-li; Zhang, Chun-hua; Xue, Xiao-ting; Zhou, Hong-jie

    2011-04-27

    Two previous studies have reported that pu-erh tea contains a high level of γ-aminobutyric acid (GABA), which is the major inhibitory neurotransmitter in the central nervous system and has several physiological functions. However, two other researchers have demonstrated that the GABA content of several pu-erh teas was low. Due to the high value and health benefits of GABA, analysis of mass-produced pu-erh tea is necessary to determine whether it is actually enriched with GABA. A high-performance liquid chromatography (HPLC) method was developed for the determination of GABA in tea, the results of which were verified by amino acid analysis using an Amino Acid Analyzer (AAA). A total of 114 samples of various types of Chinese tea, including 62 pu-erh teas, 13 green teas, 8 oolong teas, 8 black teas, 3 white teas, 4 GABA teas, and 16 process samples from two industrial fermentations of pu-erh tea (including the raw material and the first to seventh turnings), were analyzed using HPLC. Statistical analysis demonstrated that the GABA content in pu-erh tea was significantly lower than that in other types of tea (p < 0.05) and that the GABA content decreased during industrial fermentation of pu-erh tea (p < 0.05). This mass analysis and comparison suggested GABA was not a major bioactive constituent and resolved the disagreement GABA content in pu-erh tea. In addition, the GABA content in white tea was found to be significantly higher than that in the other types of tea (p < 0.05), leading to the possibility of producing GABA-enriched white tea.

  8. Propofol differentially inhibits the release of glutamate, γ-aminobutyric acid and glycine in the spinal dorsal horn of rats

    PubMed Central

    Yang, Jing; Wang, Wei; Yong, Zheng; Mi, Weidong; Zhang, Hong

    2015-01-01

    Objective(s): Propofol (2, 6-diisopropylphenol) is an intravenous anesthetic that is commonly used for the general anesthesia. It is well known that the spinal cord is one of the working targets of general anesthesia including propofol. However, there is a lack of investigation of the effects of propofol on spinal dorsal horn which is important for the sensory transmission of nociceptive signals. The objective of this study was to investigate the effects of increasing dosage of propofol on the release of glutamate (Glu), γ-aminobutyric acid (GABA) and glycine (Gly) in the spinal dorsal horn. Materials and Methods: The efflux of Glu, GABA or Gly in the spinal dorsal horn of rats was detected using transverse spinal microdialysis under an awake condition and various depths of propofol anesthesia. The infusion rates of propofol were, in order, 400 µg/(kg·min), 600 µg/(kg·min) and 800 µg/(kg·min), with a 20 min infusion period being maintained at each infusion rate. Results: Propofol decreased the glutamate efflux within spinal dorsal horn in a dose-dependent manner, and the maximum decrease was 56.8 ± 6.0% at high-dose propofol infusion producing immobility. The inhibitory GABA and Gly efflux was also decreased about 15–20% at low-dose propofol infusion only producing sedation, but did not continue to drop with higher doses of propofol. Conclusion: Propofol decreased both excitatory and inhibitory amino acids efflux in spinal dorsal horn, and the preferential suppression of the excitatory amino acid might be associated with the analgesic effect of propofol. PMID:26557972

  9. 2-Guanidine-4-methylquinazoline acts as a novel competitive antagonist of A type γ-aminobutyric acid receptors.

    PubMed

    Xiao, Xian; Zhu, Michael X; Xu, Tian-Le

    2013-12-01

    The pentameric A type γ-aminobutyric acid receptors (GABAARs) are the major inhibitory neurotransmitter receptors in the nervous system and have long been considered as important pharmaceutical targets for the treatment of multiple neurological or psychological disorders. Here, we show that 2-guanidine-4-methylquinazoline (GMQ), a recently identified acid-sensing ion channel (ASIC) modulator, strongly and preferentially inhibits GABAAR among the major neurotransmitter-gated ion channels in cultured rat hippocampal neurons. GMQ inhibited GABA (1 μM)-induced currents in a competitive manner, with an IC50 (0.39±0.05 μM) comparable to that of bicuculline. Schild analysis revealed a slope of 1.04±0.06 for GMQ on α1β2 GABAARs expressed in HEK293T cells. Single-channel analysis showed that GMQ decreased open probability of GABAARs without affecting conductance. Moreover, GMQ inhibited GABAergic neurotransmission in hippocampal neurons, while having no significant effect on the basal field excitatory postsynaptic potentials (fEPSPs) and the intrinsic excitability of neurons. Using site-directed mutagenesis, we further demonstrated that mutations at Glu155 of β2 subunit and Phe64 of α1 subunit, both located inside the GABA binding pocket, profoundly decreased the sensitivity of the receptor to both GABA and GMQ. Interestingly, these mutations did not significantly affect the inhibition by amiloride, a diuretic structurally similar to GMQ and a known GABAAR inhibitor. We conclude that GMQ represents a novel chemical structure that acts, possibly, by competing with GABA binding to GABAARs. It is anticipated that GMQ and its analogs will facilitate the development of new chemical probes for GABAARs.

  10. Effects of rumen-protected γ-aminobutyric acid on performance and nutrient digestibility in heat-stressed dairy cows.

    PubMed

    Cheng, J B; Bu, D P; Wang, J Q; Sun, X Z; Pan, L; Zhou, L Y; Liu, W

    2014-09-01

    This experiment was conducted to investigate the effects of rumen-protected γ-aminobutyric acid (GABA) on performance and nutrient digestibility in heat-stressed dairy cows. Sixty Holstein dairy cows (141±15 d in milk, 35.9±4.3kg of milk/d, and parity 2.0±1.1) were randomly assigned to 1 of 4 treatments according to a completely randomized block design. Treatments consisted of 0 (control), 40, 80, or 120mg of true GABA/kg of dry matter (DM). The trial lasted 10wk. The average temperature-humidity indices at 0700, 1400, and 2200h were 78.4, 80.2, and 78.7, respectively. Rectal temperatures decreased linearly at 0700, 1400, and 2200h with increasing GABA concentration. Supplementation of GABA had no effect on respiration rates at any time point. Dry matter intake, energy-corrected milk, 4% fat-corrected milk, and milk fat yield tended to increase linearly with increasing GABA concentration. Supplementation of GABA affected, in a quadratic manner, milk protein and lactose concentrations, and milk protein yield, and the peak values were reached at a dose of 40mg of GABA/kg. Milk urea nitrogen concentration responded quadratically. Total solids content increased linearly with increasing GABA concentration. Supplementation of GABA had no effect on milk yield, lactose production, total solids, milk fat concentration, somatic cell score, or feed efficiency. Apparent total-tract digestibilities of DM, organic matter, crude protein, neutral detergent fiber, and acid detergent fiber were similar among treatments. These results indicate that rumen-protected GABA supplementation to dairy cows can alleviate heat stress by reducing rectal temperature, increase DM intake and milk production, and improve milk composition. The appropriate supplemental GABA level for heat-stressed dairy cows is 40mg/kg of DM.

  11. Phosphatase inhibitors remove the run-down of γ-aminobutyric acid type A receptors in the human epileptic brain

    PubMed Central

    Palma, E.; Ragozzino, D. A.; Di Angelantonio, S.; Spinelli, G.; Trettel, F.; Martinez-Torres, A.; Torchia, G.; Arcella, A.; Di Gennaro, G.; Quarato, P. P.; Esposito, V.; Cantore, G.; Miledi, R.; Eusebi, F.

    2004-01-01

    The properties of γ-aminobutyric acid (GABA) type A receptors (GABAA receptors) microtransplanted from the human epileptic brain to the plasma membrane of Xenopus oocytes were compared with those recorded directly from neurons, or glial cells, in human brains slices. Cell membranes isolated from brain specimens, surgically obtained from six patients afflicted with drug-resistant temporal lobe epilepsy (TLE) were injected into frog oocytes. Within a few hours, these oocytes acquired GABAA receptors that generated GABA currents with an unusual run-down, which was inhibited by orthovanadate and okadaic acid. In contrast, receptors derived from membranes of a nonepileptic hippocampal uncus, membranes from mouse brain, or recombinant rat α1β2γ2-GABA receptors exhibited a much less pronounced GABA-current run-down. Moreover, the GABAA receptors of pyramidal neurons in temporal neocortex slices from the same six epileptic patients exhibited a stronger run-down than the receptors of rat pyramidal neurons. Interestingly, the GABAA receptors of neighboring glial cells remained substantially stable after repetitive activation. Therefore, the excessive GABA-current run-down observed in the membrane-injected oocytes recapitulates essentially what occurs in neurons, rather than in glial cells. Quantitative RT-PCR analyses from the same TLE neocortex specimens revealed that GABAA-receptor β1, β2, β3, and γ2 subunit mRNAs were significantly overexpressed (8- to 33-fold) compared with control autopsy tissues. Our results suggest that an abnormal GABA-receptor subunit transcription in the TLE brain leads to the expression of run-down-enhanced GABAA receptors. Blockage of phosphatases stabilizes the TLE GABAA receptors and strengthens GABAergic inhibition. It may be that this process can be targeted to develop new treatments for intractable epilepsy. PMID:15218107

  12. 2-Guanidine-4-methylquinazoline acts as a novel competitive antagonist of A type γ-aminobutyric acid receptors.

    PubMed

    Xiao, Xian; Zhu, Michael X; Xu, Tian-Le

    2013-12-01

    The pentameric A type γ-aminobutyric acid receptors (GABAARs) are the major inhibitory neurotransmitter receptors in the nervous system and have long been considered as important pharmaceutical targets for the treatment of multiple neurological or psychological disorders. Here, we show that 2-guanidine-4-methylquinazoline (GMQ), a recently identified acid-sensing ion channel (ASIC) modulator, strongly and preferentially inhibits GABAAR among the major neurotransmitter-gated ion channels in cultured rat hippocampal neurons. GMQ inhibited GABA (1 μM)-induced currents in a competitive manner, with an IC50 (0.39±0.05 μM) comparable to that of bicuculline. Schild analysis revealed a slope of 1.04±0.06 for GMQ on α1β2 GABAARs expressed in HEK293T cells. Single-channel analysis showed that GMQ decreased open probability of GABAARs without affecting conductance. Moreover, GMQ inhibited GABAergic neurotransmission in hippocampal neurons, while having no significant effect on the basal field excitatory postsynaptic potentials (fEPSPs) and the intrinsic excitability of neurons. Using site-directed mutagenesis, we further demonstrated that mutations at Glu155 of β2 subunit and Phe64 of α1 subunit, both located inside the GABA binding pocket, profoundly decreased the sensitivity of the receptor to both GABA and GMQ. Interestingly, these mutations did not significantly affect the inhibition by amiloride, a diuretic structurally similar to GMQ and a known GABAAR inhibitor. We conclude that GMQ represents a novel chemical structure that acts, possibly, by competing with GABA binding to GABAARs. It is anticipated that GMQ and its analogs will facilitate the development of new chemical probes for GABAARs. PMID:23916476

  13. Exogenous γ-aminobutyric Acid (GABA) Application Improved Early Growth, Net Photosynthesis, and Associated Physio-Biochemical Events in Maize

    PubMed Central

    Li, Wu; Liu, Jianhua; Ashraf, Umair; Li, Gaoke; Li, Yuliang; Lu, Wenjia; Gao, Lei; Han, Fuguang; Hu, Jianguang

    2016-01-01

    γ-aminobutyric acid (GABA) is an endogenous signaling molecule and involved in growth regulations and plant development, however, a little information is available on the consequences of exogenous GABA application on growth, development, and associated physio-biochemical processes in maize. The present study examined the GABA-induced regulations in early growth, net photosynthetic rate, gas exchange, osmoregulation, and enzymatic activities in three maize cultivars, i.e., Yuecainuo 6, Zhengtian 68, and Yuecainuo 2. Two levels of GABA, i.e., 0 mg L-1 and 50 mg L-1, in solution form, with total application volume of 100 ml per pot containing 15 maize seedlings were exogenously applied. Results revealed that exogenous GABA application improved seedling growth in terms of seedling length and biomass accumulation in all maize cultivars at both 3 and 7 days after treatment (DAT). It also promoted net photosynthesis and variably affected gas exchange attributes, i.e., stomatal conductance (Gs), intercellular CO2 concentration (Ci), and transpiration rate (Tr), as well as leaves SPAD value. Furthermore, lipid peroxidation [in terms of malondialdehyde (MDA)] under GABA treated maize seedlings were also remained variable; however, osmolyte accumulation (protein and proline) and activities of anti-oxidants enzymes, i.e., super-oxide dismutase and peroxidase were also affected differently at both 3 and 7 DAT in all maize cultivars. Furthermore, enzymes involved in nitrogen metabolism, e.g., nitrate reductase and glutamine synthetase were improved. These results suggest the involvement of GABA in various physio-metablical mechanisms which might lead to improvement in morphological growth of maize. In future, research is still needed at molecular and genetic levels to unravel the involvement of GABA-mediated regulations in growth and its associated physio-biochemical mechanisms. PMID:27446149

  14. Expression of the γ-Aminobutyric Acid (GABA) Plasma Membrane Transporter-1 in Monkey and Human Retina

    PubMed Central

    Casini, Giovanni; Rickman, Dennis W.; Brecha, Nicholas C.

    2010-01-01

    Purpose To determine the expression pattern of the predominant γ-aminobutyric acid (GABA) plasma membrane transporter GAT-1 in Old World monkey (Macaca mulatta) and human retina. Methods GAT-1 was localized in retinal sections by using immunohistochemical techniques with fluorescence and confocal microscopy. Double-labeling studies were performed with the GAT-1 antibody using antibodies to GABA, vasoactive intestinal polypeptide (VIP), tyrosine hydroxylase (TH), and the bipolar cell marker Mab115A10. Results The pattern of GAT-1 immunostaining was similar in human and monkey retinas. Numerous small immunoreactive somata were in the inner nuclear layer (INL) and were present rarely in the inner plexiform layer (IPL) of all retinal regions. Medium GAT-1 somata were in the ganglion cell layer in the parafoveal and peripheral retinal regions. GAT-1 fibers were densely distributed throughout the IPL. Varicose processes, originating from both the IPL and somata in the INL, arborized in the outer plexiform layer (OPL), forming a sparse network in all retinal regions, except the fovea. Sparsely occurring GAT-1 processes were in the nerve fiber layer in parafoveal regions and near the optic nerve head but not in the optic nerve. In the INL, 99% of the GAT-1 somata contained GABA, and 66% of the GABA immunoreactive somata expressed GAT-1. GAT-1 immunoreactivity was in all VIP-containing cells, but it was absent in TH-immunoreactive amacrine cells and in Mab115A10 immunoreactive bipolar cells. Conclusions GAT-1 in primate retinas is expressed by amacrine and displaced amacrine cells. The predominant expression of GAT-1 in the inner retina is consistent with the idea that GABA transporters influence neurotransmission and thus participate in visual information processing in the retina. PMID:16565409

  15. Development of early-born γ-Aminobutyric acid hub neurons in mouse hippocampus from embryogenesis to adulthood.

    PubMed

    Villette, Vincent; Guigue, Philippe; Picardo, Michel Aimé; Sousa, Vitor Hugo; Leprince, Erwan; Lachamp, Philippe; Malvache, Arnaud; Tressard, Thomas; Cossart, Rosa; Baude, Agnès

    2016-08-15

    Early-born γ-aminobutyric acid (GABA) neurons (EBGNs) are major components of the hippocampal circuit because at early postnatal stages they form a subpopulation of "hub cells" transiently supporting CA3 network synchronization (Picardo et al. [2011] Neuron 71:695-709). It is therefore essential to determine when these cells acquire the remarkable morphofunctional attributes supporting their network function and whether they develop into a specific subtype of interneuron into adulthood. Inducible genetic fate mapping conveniently allows for the labeling of EBGNs throughout their life. EBGNs were first analyzed during the perinatal week. We observed that EBGNs acquired mature characteristics at the time when the first synapse-driven synchronous activities appeared in the form of giant depolarizing potentials. The fate of EBGNs was next analyzed in the adult hippocampus by using anatomical characterization. Adult EBGNs included a significant proportion of cells projecting selectively to the septum; in turn, EBGNs were targeted by septal and entorhinal inputs. In addition, most EBGNs were strongly targeted by cholinergic and monoaminergic terminals, suggesting significant subcortical innervation. Finally, we found that some EBGNs located in the septum or the entorhinal cortex also displayed a long-range projection that we traced to the hippocampus. Therefore, this study shows that the maturation of the morphophysiological properties of EBGNs mirrors the evolution of early network dynamics, suggesting that both phenomena may be causally linked. We propose that a subpopulation of EBGNs forms into adulthood a scaffold of GABAergic projection neurons linking the hippocampus to distant structures. J. Comp. Neurol. 524:2440-2461, 2016. © 2016 Wiley Periodicals, Inc.

  16. Transient Receptor Potential Vanilloid 4 Inhibits γ-Aminobutyric Acid-Activated Current in Hippocampal Pyramidal Neurons

    PubMed Central

    Hong, Zhiwen; Tian, Yujing; Qi, Mengwen; Li, Yingchun; Du, Yimei; Chen, Lei; Liu, Wentao; Chen, Ling

    2016-01-01

    The balance between excitatory and inhibitory neurotransmitter systems is crucial for the modulation of neuronal excitability in the central nervous system (CNS). The activation of transient receptor potential vanilloid 4 (TRPV4) is reported to enhance the response of hippocampal glutamate receptors, but whether the inhibitory neurotransmitter system can be regulated by TRPV4 remains unknown. γ-aminobutyric acid (GABA) is the major inhibitory neurotransmitter in the CNS. Here, we show that application of transient receptor potential vanilloid 4 (TRPV4) synthetic (GSK1016790A or 4α-PDD) or endogenous agonist (5,6-EET) inhibited GABA-activated current (IGABA) in hippocampal CA1 pyramidal neurons, which was blocked by specific antagonists of TRPV4 and of GABAA receptors. GSK1016790A increased the phosphorylated AMP-activated protein kinase (p-AMPK) and decreased the phosphorylated protein kinase B (p-Akt) protein levels, which was attenuated by removing extracellular calcium or by a calcium/calmodulin-dependent protein kinase kinase-β antagonist. GSK1016790A-induced decrease of p-Akt protein level was sensitive to an AMPK antagonist. GSK1016790A-inhibited IGABA was blocked by an AMPK antagonist or a phosphatidyl inositol 3 kinase (PI3K) agonist. GSK1016790A-induced inhibition of IGABA was also significantly attenuated by a protein kinase C (PKC) antagonist but was unaffected by protein kinase A or calcium/calmodulin-dependent protein kinase II antagonist. We conclude that activation of TRPV4 inhibits GABAA receptor, which may be mediated by activation of AMPK and subsequent down-regulation of PI3K/Akt signaling and activation of PKC signaling. Inhibition of GABAA receptors may account for the neuronal hyperexcitability caused by TRPV4 activation. PMID:27616980

  17. Transient Receptor Potential Vanilloid 4 Inhibits γ-Aminobutyric Acid-Activated Current in Hippocampal Pyramidal Neurons

    PubMed Central

    Hong, Zhiwen; Tian, Yujing; Qi, Mengwen; Li, Yingchun; Du, Yimei; Chen, Lei; Liu, Wentao; Chen, Ling

    2016-01-01

    The balance between excitatory and inhibitory neurotransmitter systems is crucial for the modulation of neuronal excitability in the central nervous system (CNS). The activation of transient receptor potential vanilloid 4 (TRPV4) is reported to enhance the response of hippocampal glutamate receptors, but whether the inhibitory neurotransmitter system can be regulated by TRPV4 remains unknown. γ-aminobutyric acid (GABA) is the major inhibitory neurotransmitter in the CNS. Here, we show that application of transient receptor potential vanilloid 4 (TRPV4) synthetic (GSK1016790A or 4α-PDD) or endogenous agonist (5,6-EET) inhibited GABA-activated current (IGABA) in hippocampal CA1 pyramidal neurons, which was blocked by specific antagonists of TRPV4 and of GABAA receptors. GSK1016790A increased the phosphorylated AMP-activated protein kinase (p-AMPK) and decreased the phosphorylated protein kinase B (p-Akt) protein levels, which was attenuated by removing extracellular calcium or by a calcium/calmodulin-dependent protein kinase kinase-β antagonist. GSK1016790A-induced decrease of p-Akt protein level was sensitive to an AMPK antagonist. GSK1016790A-inhibited IGABA was blocked by an AMPK antagonist or a phosphatidyl inositol 3 kinase (PI3K) agonist. GSK1016790A-induced inhibition of IGABA was also significantly attenuated by a protein kinase C (PKC) antagonist but was unaffected by protein kinase A or calcium/calmodulin-dependent protein kinase II antagonist. We conclude that activation of TRPV4 inhibits GABAA receptor, which may be mediated by activation of AMPK and subsequent down-regulation of PI3K/Akt signaling and activation of PKC signaling. Inhibition of GABAA receptors may account for the neuronal hyperexcitability caused by TRPV4 activation.

  18. Exogenous γ-aminobutyric Acid (GABA) Application Improved Early Growth, Net Photosynthesis, and Associated Physio-Biochemical Events in Maize.

    PubMed

    Li, Wu; Liu, Jianhua; Ashraf, Umair; Li, Gaoke; Li, Yuliang; Lu, Wenjia; Gao, Lei; Han, Fuguang; Hu, Jianguang

    2016-01-01

    γ-aminobutyric acid (GABA) is an endogenous signaling molecule and involved in growth regulations and plant development, however, a little information is available on the consequences of exogenous GABA application on growth, development, and associated physio-biochemical processes in maize. The present study examined the GABA-induced regulations in early growth, net photosynthetic rate, gas exchange, osmoregulation, and enzymatic activities in three maize cultivars, i.e., Yuecainuo 6, Zhengtian 68, and Yuecainuo 2. Two levels of GABA, i.e., 0 mg L(-1) and 50 mg L(-1), in solution form, with total application volume of 100 ml per pot containing 15 maize seedlings were exogenously applied. Results revealed that exogenous GABA application improved seedling growth in terms of seedling length and biomass accumulation in all maize cultivars at both 3 and 7 days after treatment (DAT). It also promoted net photosynthesis and variably affected gas exchange attributes, i.e., stomatal conductance (Gs), intercellular CO2 concentration (Ci), and transpiration rate (Tr), as well as leaves SPAD value. Furthermore, lipid peroxidation [in terms of malondialdehyde (MDA)] under GABA treated maize seedlings were also remained variable; however, osmolyte accumulation (protein and proline) and activities of anti-oxidants enzymes, i.e., super-oxide dismutase and peroxidase were also affected differently at both 3 and 7 DAT in all maize cultivars. Furthermore, enzymes involved in nitrogen metabolism, e.g., nitrate reductase and glutamine synthetase were improved. These results suggest the involvement of GABA in various physio-metablical mechanisms which might lead to improvement in morphological growth of maize. In future, research is still needed at molecular and genetic levels to unravel the involvement of GABA-mediated regulations in growth and its associated physio-biochemical mechanisms.

  19. Linking γ-aminobutyric acid A receptor to epidermal growth factor receptor pathways activation in human prostate cancer.

    PubMed

    Wu, Weijuan; Yang, Qing; Fung, Kar-Ming; Humphreys, Mitchell R; Brame, Lacy S; Cao, Amy; Fang, Yu-Ting; Shih, Pin-Tsen; Kropp, Bradley P; Lin, Hsueh-Kung

    2014-03-01

    Neuroendocrine (NE) differentiation has been attributed to the progression of castration-resistant prostate cancer (CRPC). Growth factor pathways including the epidermal growth factor receptor (EGFR) signaling have been implicated in the development of NE features and progression to a castration-resistant phenotype. However, upstream molecules that regulate the growth factor pathway remain largely unknown. Using androgen-insensitive bone metastasis PC-3 cells and androgen-sensitive lymph node metastasis LNCaP cells derived from human prostate cancer (PCa) patients, we demonstrated that γ-aminobutyric acid A receptor (GABA(A)R) ligand (GABA) and agonist (isoguvacine) stimulate cell proliferation, enhance EGF family members expression, and activate EGFR and a downstream signaling molecule, Src, in both PC-3 and LNCaP cells. Inclusion of a GABA(A)R antagonist, picrotoxin, or an EGFR tyrosine kinase inhibitor, Gefitinib (ZD1839 or Iressa), blocked isoguvacine and GABA-stimulated cell growth, trans-phospohorylation of EGFR, and tyrosyl phosphorylation of Src in both PCa cell lines. Spatial distributions of GABAAR α₁ and phosphorylated Src (Tyr416) were studied in human prostate tissues by immunohistochemistry. In contrast to extremely low or absence of GABA(A)R α₁-positive immunoreactivity in normal prostate epithelium, elevated GABA(A)R α₁ immunoreactivity was detected in prostate carcinomatous glands. Similarly, immunoreactivity of phospho-Src (Tyr416) was specifically localized and limited to the nucleoli of all invasive prostate carcinoma cells, but negative in normal tissues. Strong GABAAR α₁ immunoreactivity was spatially adjacent to the neoplastic glands where strong phospho-Src (Tyr416)-positive immunoreactivity was demonstrated, but not in adjacent to normal glands. These results suggest that the GABA signaling is linked to the EGFR pathway and may work through autocrine or paracine mechanism to promote CRPC progression.

  20. Effects of γ-aminobutyric acid on feed intake, growth performance and expression of related genes in growing lambs.

    PubMed

    Wang, D M; Chacher, B; Liu, H Y; Wang, J K; Lin, J; Liu, J X

    2015-03-01

    This study was conducted to investigate the effects of rumen-protected γ-aminobutyric acid (GABA) on feed intake, growth performance and expression of related genes in growing lambs. A total of 24 lambs weaned at age of 50 days were divided into four block of six based on their BW, six lambs within a block were allocated to three pairs, which were then assigned randomly to three treatments with addition of rumen-protected GABA at levels of 0, 70 or 140 mg/day for 6 weeks. Dry matter intake was recorded weekly in three consecutive days, and BW was recorded every two weeks. At the end of the trial, four lambs from each group were slaughtered, and duodenum and ileum mucosa were obtained for measurement of mRNA abundance of GABA receptor and cholecystokinin receptor. Dry matter intake was higher (P<0.01) in the lambs fed 140 mg/day GABA than that in the control or 70 mg GABA-fed lambs. Average daily gain and nutrients digestibility were not different (P>0.05) among treatments. Lambs fed 140 mg/day GABA had higher mRNA abundance of GABA-B receptor (P<0.01) and lower mRNA abundance of cholecystokinin-2 receptor (P<0.01) in duodenum mucosa. Serum CCK content was lower (P<0.01) in lambs fed 140 mg/day GABA than that in control. It is indicated that GABA may enhance feed intake by regulating GABA- and cholecystokinin-related genes.

  1. Differential regulation of the postsynaptic clustering of γ-aminobutyric acid type A (GABAA) receptors by collybistin isoforms.

    PubMed

    Chiou, Tzu-Ting; Bonhomme, Bevan; Jin, Hongbing; Miralles, Celia P; Xiao, Haiyan; Fu, Zhanyan; Harvey, Robert J; Harvey, Kirsten; Vicini, Stefano; De Blas, Angel L

    2011-06-24

    Collybistin promotes submembrane clustering of gephyrin and is essential for the postsynaptic localization of gephyrin and γ-aminobutyric acid type A (GABA(A)) receptors at GABAergic synapses in hippocampus and amygdala. Four collybistin isoforms are expressed in brain neurons; CB2 and CB3 differ in the C terminus and occur with and without the Src homology 3 (SH3) domain. We have found that in transfected hippocampal neurons, all collybistin isoforms (CB2(SH3+), CB2(SH3-), CB3(SH3+), and CB3(SH3-)) target to and concentrate at GABAergic postsynapses. Moreover, in non-transfected neurons, collybistin concentrates at GABAergic synapses. Hippocampal neurons co-transfected with CB2(SH3-) and gephyrin developed very large postsynaptic gephyrin and GABA(A) receptor clusters (superclusters). This effect was accompanied by a significant increase in the amplitude of miniature inhibitory postsynaptic currents. Co-transfection with CB2(SH3+) and gephyrin induced the formation of many (supernumerary) non-synaptic clusters. Transfection with gephyrin alone did not affect cluster number or size, but gephyrin potentiated the clustering effect of CB2(SH3-) or CB2(SH3+). Co-transfection with CB2(SH3-) or CB2(SH3+) and gephyrin did not affect the density of presynaptic GABAergic terminals contacting the transfected cells, indicating that collybistin is not synaptogenic. Nevertheless, the synaptic superclusters induced by CB2(SH3-) and gephyrin were accompanied by enlarged presynaptic GABAergic terminals. The enhanced clustering of gephyrin and GABA(A) receptors induced by collybistin isoforms was not accompanied by enhanced clustering of neuroligin 2. Moreover, during the development of GABAergic synapses, the clustering of gephyrin and GABA(A) receptors preceded the clustering of neuroligin 2. We propose a model in which the SH3- isoforms play a major role in the postsynaptic accumulation of GABA(A) receptors and in GABAergic synaptic strength.

  2. α-Thujone (the active component of absinthe): γ-Aminobutyric acid type A receptor modulation and metabolic detoxification

    PubMed Central

    Höld, Karin M.; Sirisoma, Nilantha S.; Ikeda, Tomoko; Narahashi, Toshio; Casida, John E.

    2000-01-01

    α-Thujone is the toxic agent in absinthe, a liqueur popular in the 19th and early 20th centuries that has adverse health effects. It is also the active ingredient of wormwood oil and some other herbal medicines and is reported to have antinociceptive, insecticidal, and anthelmintic activity. This study elucidates the mechanism of α-thujone neurotoxicity and identifies its major metabolites and their role in the poisoning process. Four observations establish that α-thujone is a modulator of the γ-aminobutyric acid (GABA) type A receptor. First, the poisoning signs (and their alleviation by diazepam and phenobarbital) in mice are similar to those of the classical antagonist picrotoxinin. Second, a strain of Drosophila specifically resistant to chloride channel blockers is also tolerant to α-thujone. Third, α-thujone is a competitive inhibitor of [3H]ethynylbicycloorthobenzoate binding to mouse brain membranes. Most definitively, GABA-induced peak currents in rat dorsal root ganglion neurons are suppressed by α-thujone with complete reversal after washout. α-Thujone is quickly metabolized in vitro by mouse liver microsomes with NADPH (cytochrome P450) forming 7-hydroxy-α-thujone as the major product plus five minor ones (4-hydroxy-α-thujone, 4-hydroxy-β-thujone, two other hydroxythujones, and 7,8-dehydro-α-thujone), several of which also are detected in the brain of mice treated i.p. with α-thujone. The major 7-hydroxy metabolite attains much higher brain levels than α-thujone but is less toxic to mice and Drosophila and less potent in the binding assay. The other metabolites assayed are also detoxification products. Thus, α-thujone in absinthe and herbal medicines is a rapid-acting and readily detoxified modulator of the GABA-gated chloride channel. PMID:10725394

  3. Transient Receptor Potential Vanilloid 4 Inhibits γ-Aminobutyric Acid-Activated Current in Hippocampal Pyramidal Neurons.

    PubMed

    Hong, Zhiwen; Tian, Yujing; Qi, Mengwen; Li, Yingchun; Du, Yimei; Chen, Lei; Liu, Wentao; Chen, Ling

    2016-01-01

    The balance between excitatory and inhibitory neurotransmitter systems is crucial for the modulation of neuronal excitability in the central nervous system (CNS). The activation of transient receptor potential vanilloid 4 (TRPV4) is reported to enhance the response of hippocampal glutamate receptors, but whether the inhibitory neurotransmitter system can be regulated by TRPV4 remains unknown. γ-aminobutyric acid (GABA) is the major inhibitory neurotransmitter in the CNS. Here, we show that application of transient receptor potential vanilloid 4 (TRPV4) synthetic (GSK1016790A or 4α-PDD) or endogenous agonist (5,6-EET) inhibited GABA-activated current (I GABA) in hippocampal CA1 pyramidal neurons, which was blocked by specific antagonists of TRPV4 and of GABAA receptors. GSK1016790A increased the phosphorylated AMP-activated protein kinase (p-AMPK) and decreased the phosphorylated protein kinase B (p-Akt) protein levels, which was attenuated by removing extracellular calcium or by a calcium/calmodulin-dependent protein kinase kinase-β antagonist. GSK1016790A-induced decrease of p-Akt protein level was sensitive to an AMPK antagonist. GSK1016790A-inhibited I GABA was blocked by an AMPK antagonist or a phosphatidyl inositol 3 kinase (PI3K) agonist. GSK1016790A-induced inhibition of I GABA was also significantly attenuated by a protein kinase C (PKC) antagonist but was unaffected by protein kinase A or calcium/calmodulin-dependent protein kinase II antagonist. We conclude that activation of TRPV4 inhibits GABAA receptor, which may be mediated by activation of AMPK and subsequent down-regulation of PI3K/Akt signaling and activation of PKC signaling. Inhibition of GABAA receptors may account for the neuronal hyperexcitability caused by TRPV4 activation. PMID:27616980

  4. Exogenous γ-aminobutyric Acid (GABA) Application Improved Early Growth, Net Photosynthesis, and Associated Physio-Biochemical Events in Maize.

    PubMed

    Li, Wu; Liu, Jianhua; Ashraf, Umair; Li, Gaoke; Li, Yuliang; Lu, Wenjia; Gao, Lei; Han, Fuguang; Hu, Jianguang

    2016-01-01

    γ-aminobutyric acid (GABA) is an endogenous signaling molecule and involved in growth regulations and plant development, however, a little information is available on the consequences of exogenous GABA application on growth, development, and associated physio-biochemical processes in maize. The present study examined the GABA-induced regulations in early growth, net photosynthetic rate, gas exchange, osmoregulation, and enzymatic activities in three maize cultivars, i.e., Yuecainuo 6, Zhengtian 68, and Yuecainuo 2. Two levels of GABA, i.e., 0 mg L(-1) and 50 mg L(-1), in solution form, with total application volume of 100 ml per pot containing 15 maize seedlings were exogenously applied. Results revealed that exogenous GABA application improved seedling growth in terms of seedling length and biomass accumulation in all maize cultivars at both 3 and 7 days after treatment (DAT). It also promoted net photosynthesis and variably affected gas exchange attributes, i.e., stomatal conductance (Gs), intercellular CO2 concentration (Ci), and transpiration rate (Tr), as well as leaves SPAD value. Furthermore, lipid peroxidation [in terms of malondialdehyde (MDA)] under GABA treated maize seedlings were also remained variable; however, osmolyte accumulation (protein and proline) and activities of anti-oxidants enzymes, i.e., super-oxide dismutase and peroxidase were also affected differently at both 3 and 7 DAT in all maize cultivars. Furthermore, enzymes involved in nitrogen metabolism, e.g., nitrate reductase and glutamine synthetase were improved. These results suggest the involvement of GABA in various physio-metablical mechanisms which might lead to improvement in morphological growth of maize. In future, research is still needed at molecular and genetic levels to unravel the involvement of GABA-mediated regulations in growth and its associated physio-biochemical mechanisms. PMID:27446149

  5. Development of early‐born γ‐Aminobutyric acid hub neurons in mouse hippocampus from embryogenesis to adulthood

    PubMed Central

    Villette, Vincent; Guigue, Philippe; Picardo, Michel Aimé; Sousa, Vitor Hugo; Leprince, Erwan; Lachamp, Philippe; Malvache, Arnaud; Tressard, Thomas; Cossart, Rosa

    2016-01-01

    ABSTRACT Early‐born γ‐aminobutyric acid (GABA) neurons (EBGNs) are major components of the hippocampal circuit because at early postnatal stages they form a subpopulation of “hub cells” transiently supporting CA3 network synchronization (Picardo et al. [2011] Neuron 71:695–709). It is therefore essential to determine when these cells acquire the remarkable morphofunctional attributes supporting their network function and whether they develop into a specific subtype of interneuron into adulthood. Inducible genetic fate mapping conveniently allows for the labeling of EBGNs throughout their life. EBGNs were first analyzed during the perinatal week. We observed that EBGNs acquired mature characteristics at the time when the first synapse‐driven synchronous activities appeared in the form of giant depolarizing potentials. The fate of EBGNs was next analyzed in the adult hippocampus by using anatomical characterization. Adult EBGNs included a significant proportion of cells projecting selectively to the septum; in turn, EBGNs were targeted by septal and entorhinal inputs. In addition, most EBGNs were strongly targeted by cholinergic and monoaminergic terminals, suggesting significant subcortical innervation. Finally, we found that some EBGNs located in the septum or the entorhinal cortex also displayed a long‐range projection that we traced to the hippocampus. Therefore, this study shows that the maturation of the morphophysiological properties of EBGNs mirrors the evolution of early network dynamics, suggesting that both phenomena may be causally linked. We propose that a subpopulation of EBGNs forms into adulthood a scaffold of GABAergic projection neurons linking the hippocampus to distant structures. J. Comp. Neurol. 524:2440–2461, 2016. © 2016 Wiley Periodicals, Inc. PMID:26779909

  6. Electrophysiological characterization of ivermectin triple actions on Musca chloride channels gated by l-glutamic acid and γ-aminobutyric acid.

    PubMed

    Fuse, Toshinori; Kita, Tomo; Nakata, Yunosuke; Ozoe, Fumiyo; Ozoe, Yoshihisa

    2016-10-01

    Ivermectin (IVM) is a macrocyclic lactone that exerts antifilarial, antiparasitic, and insecticidal effects on nematodes and insects by acting on l-glutamic acid-gated chloride channels (GluCls). IVM also acts as an allosteric modulator of various other ion channels. Although the IVM binding site in the Caenorhabditis elegans GluCl was identified by X-ray crystallographic analysis, the mechanism of action of IVM in insects is not well defined. We therefore examined the action of IVM on the housefly (Musca domestica) GluCl and γ-aminobutyric acid (GABA)-gated ion channel (GABACl). For both channels, IVM induced currents by itself, potentiated currents induced by low concentrations of agonists, and inhibited currents induced by high concentrations of agonists. Despite exerting common actions on both types of channels, GluCls were more susceptible to IVM actions than GABACls, indicating that GluCls are the primary target of IVM. Substitution of an amino acid residue in the third transmembrane segment (G312M in GluCls, and G333A and G333M in GABACls) resulted in significantly reduced levels or loss of activation, potentiation, and antagonism of the channels, indicating that these three actions result from the interaction of IVM with amino acid residues in the transmembrane intersubunit crevice.

  7. Electrophysiological characterization of ivermectin triple actions on Musca chloride channels gated by l-glutamic acid and γ-aminobutyric acid.

    PubMed

    Fuse, Toshinori; Kita, Tomo; Nakata, Yunosuke; Ozoe, Fumiyo; Ozoe, Yoshihisa

    2016-10-01

    Ivermectin (IVM) is a macrocyclic lactone that exerts antifilarial, antiparasitic, and insecticidal effects on nematodes and insects by acting on l-glutamic acid-gated chloride channels (GluCls). IVM also acts as an allosteric modulator of various other ion channels. Although the IVM binding site in the Caenorhabditis elegans GluCl was identified by X-ray crystallographic analysis, the mechanism of action of IVM in insects is not well defined. We therefore examined the action of IVM on the housefly (Musca domestica) GluCl and γ-aminobutyric acid (GABA)-gated ion channel (GABACl). For both channels, IVM induced currents by itself, potentiated currents induced by low concentrations of agonists, and inhibited currents induced by high concentrations of agonists. Despite exerting common actions on both types of channels, GluCls were more susceptible to IVM actions than GABACls, indicating that GluCls are the primary target of IVM. Substitution of an amino acid residue in the third transmembrane segment (G312M in GluCls, and G333A and G333M in GABACls) resulted in significantly reduced levels or loss of activation, potentiation, and antagonism of the channels, indicating that these three actions result from the interaction of IVM with amino acid residues in the transmembrane intersubunit crevice. PMID:27543424

  8. Widely expressed transcripts for chemokine receptor CXCR1 in identified glutamatergic, gamma-aminobutyric acidergic, and cholinergic neurons and astrocytes of the rat brain: a single-cell reverse transcription-multiplex polymerase chain reaction study.

    PubMed

    Danik, M; Puma, C; Quirion, R; Williams, S

    2003-10-15

    Increasing evidence suggests that the chemokine interleukin (IL)-8/CXCL8 plays important roles in CNS development, neuronal survival, modulation of excitability, and neuroimmune response. Recently, we have shown that CXCL8 can acutely modulate ion channel activity in septal neurons expressing receptors CXCR1 and/or CXCR2. This was a surprising finding, insofar as CXCR1 expression had not been described for the mammalian brain. Here we investigated whether CXCR1 transcripts are present in other brain regions, whether they are expressed at the single-cell level in molecularly identified neurons and astrocytes, and how they are regulated during early postnatal development. In addition, possible cellular colocalization of CXCR1 and CXCR2 transcripts was examined. Semiquantitative reverse transcription-polymerase chain reaction (RT-PCR) revealed that CXCR1 mRNAs were expressed in the septum, striatum, hippocampus, cerebellum, and cortex (temporoparietal and entorhinal) at different levels and appeared to be regulated independently from CXCR2 during development. By using RT multiplex PCR on acutely dissociated cells from these brain regions, we show that CXCR1 transcripts were expressed in 83% of 84 sampled neurons displaying cholinergic (choline acetyltransferase mRNAs), gamma-aminobutyric acidergic (glutamic acid decarboxylases 65 and 67 mRNAs), or glutamatergic (vesicular glutamate transporters 1 and 2 mRNAs) phenotypes. CXCR1 and CXCR2 transcripts were colocalized in 45% of neurons sampled and also were present in some glial fibrillary acidic protein mRNA-expressing astrocytes. This is the first study to demonstrate the widespread expression of CXCR1 transcripts in the brain and suggests that CXCR1 may have hitherto unsuspected roles in neuromodulation and inflammation. PMID:14515358

  9. DL-β-Aminobutyric Acid-Induced Resistance in Soybean against Aphis glycines Matsumura (Hemiptera: Aphididae)

    PubMed Central

    Zhong, Yunpeng; Wang, Biao; Yan, Junhui; Cheng, Linjing; Yao, Luming; Xiao, Liang; Wu, Tianlong

    2014-01-01

    Priming can improve plant innate capability to deal with the stresses caused by both biotic and abiotic factors. In this study, the effect of DL-β-amino-n-butyric acid (BABA) against Aphis glycines Matsumura, the soybean aphid (SA) was evaluated. We found that 25 mM BABA as a root drench had minimal adverse impact on plant growth and also efficiently protected soybean from SA infestation. In both choice and non-choice tests, SA number was significantly decreased to a low level in soybean seedlings drenched with 25 mM BABA compared to the control counterparts. BABA treatment resulted in a significant increase in the activities of several defense enzymes, such as phenylalanine ammonia-lyase (PAL), peroxidase (POX), polyphenol oxidase (PPO), chitinase (CHI), and β-1, 3-glucanase (GLU) in soybean seedlings attacked by aphid. Meanwhile, the induction of 15 defense-related genes by aphid, such as AOS, CHS, MMP2, NPR1-1, NPR1-2, and PR genes, were significantly augmented in BABA-treated soybean seedlings. Our study suggest that BABA application is a promising way to enhance soybean resistance against SA. PMID:24454805

  10. Molecular recognition of nitrated fatty acids by PPAR[gamma

    SciTech Connect

    Li, Yong; Zhang, Jifeng; Schopfer, Francisco J.; Martynowski, Dariusz; Garcia-Barrio, Minerva T.; Kovach, Amanda; Suino-Powell, Kelly; Baker, Paul R.S.; Freeman, Bruce A.; Chen, Y. Eugene; Xu, H. Eric

    2010-03-08

    Peroxisome proliferator activated receptor-{gamma} (PPAR{gamma}) regulates metabolic homeostasis and adipocyte differentiation, and it is activated by oxidized and nitrated fatty acids. Here we report the crystal structure of the PPAR{gamma} ligand binding domain bound to nitrated linoleic acid, a potent endogenous ligand of PPAR{gamma}. Structural and functional studies of receptor-ligand interactions reveal the molecular basis of PPAR{gamma} discrimination of various naturally occurring fatty acid derivatives.

  11. Dual effects of slightly acidic electrolyzed water (SAEW) treatment on the accumulation of γ-aminobutyric acid (GABA) and rutin in germinated buckwheat.

    PubMed

    Hao, Jianxiong; Wu, Tongjiao; Li, Huiying; Wang, Wei; Liu, Haijie

    2016-06-15

    In the present study, the dual effects of slightly acidic electrolyzed water (SAEW) treatment on γ-aminobutyric acid (GABA) and rutin accumulation of germinated buckwheat were evaluated during germination. The results showed that SAEW treatment (pH 5.83, ACC of 20.3 mg/L) could promote the accumulation of GABA and rutin in germinated buckwheat. The GABA and rutin contents of SAEW-germinated buckwheat reached 143.20 and 739.9 mg/100 g respectively, which is significantly higher than those of control (P<0.05). Moreover, SAEW treatment could increase the activity of glutamic acid decarboxylase (GAD) and phenylalanine ammonialyase (PAL) and thus result in the GABA and rutin accumulation of germinated buckwheat. The results suggested that SAEW treatment could promote the rutin accumulation of germinated buckwheat by influencing phenylpropanoid secondary metabolic pathway instead of the inhibition of rutin degrading enzyme (RDE) activity. In addition, SAEW treatment had no adverse impact on the sprouts growth and could reduce the microbial populations of germinated buckwheat during germination.

  12. Dual effects of slightly acidic electrolyzed water (SAEW) treatment on the accumulation of γ-aminobutyric acid (GABA) and rutin in germinated buckwheat.

    PubMed

    Hao, Jianxiong; Wu, Tongjiao; Li, Huiying; Wang, Wei; Liu, Haijie

    2016-06-15

    In the present study, the dual effects of slightly acidic electrolyzed water (SAEW) treatment on γ-aminobutyric acid (GABA) and rutin accumulation of germinated buckwheat were evaluated during germination. The results showed that SAEW treatment (pH 5.83, ACC of 20.3 mg/L) could promote the accumulation of GABA and rutin in germinated buckwheat. The GABA and rutin contents of SAEW-germinated buckwheat reached 143.20 and 739.9 mg/100 g respectively, which is significantly higher than those of control (P<0.05). Moreover, SAEW treatment could increase the activity of glutamic acid decarboxylase (GAD) and phenylalanine ammonialyase (PAL) and thus result in the GABA and rutin accumulation of germinated buckwheat. The results suggested that SAEW treatment could promote the rutin accumulation of germinated buckwheat by influencing phenylpropanoid secondary metabolic pathway instead of the inhibition of rutin degrading enzyme (RDE) activity. In addition, SAEW treatment had no adverse impact on the sprouts growth and could reduce the microbial populations of germinated buckwheat during germination. PMID:26868552

  13. Gamma-aminobutyric acid-modulated benzodiazepine binding sites in bacteria

    SciTech Connect

    Lummis, S.C.R.; Johnston, G.A.R. ); Nicoletti, G. ); Holan, G. )

    1991-01-01

    Benzodiazepine binding sites, which were once considered to exist only in higher vertebrates, are here demonstrated in the bacteria E. coli. The bacterial ({sup 3}H)diazepam binding sites are modulated by GABA; the modulation is dose dependent and is reduced at high concentrations. The most potent competitors of E.Coli ({sup 3}H)diazepam binding are those that are active in displacing ({sup 3}H)benzodiazepines from vertebrate peripheral benzodiazepine binding sites. These vertebrate sites are not modulated by GABA, in contrast to vertebrate neuronal benzodiazepine binding sites. The E.coli benzodiazepine binding sites therefore differ from both classes of vertebrate benzodiazepine binding sites; however the ligand spectrum and GABA-modulatory properties of the E.coli sites are similar to those found in insects. This intermediate type of receptor in lower species suggests a precursor for at least one class of vertebrate benzodiazepine binding sites may have existed.

  14. Reproducibility and effect of tissue composition on cerebellar γ-aminobutyric acid (GABA) MRS in an elderly population.

    PubMed

    Long, Zaiyang; Dyke, Jonathan P; Ma, Ruoyun; Huang, Chaorui C; Louis, Elan D; Dydak, Ulrike

    2015-10-01

    MRS provides a valuable tool for the non-invasive detection of brain γ-aminobutyric acid (GABA) in vivo. GABAergic dysfunction has been observed in the aging cerebellum. The study of cerebellar GABA changes is of considerable interest in understanding certain age-related motor disorders. However, little is known about the reproducibility of GABA MRS in an aged population. Therefore, this study aimed to explore the feasibility and reproducibility of GABA MRS in the aged cerebellum at 3.0 T and to examine the effect of differing tissue composition on GABA measurements. MRI and (1)H MRS examinations were performed on 10 healthy elderly volunteers (mean age, 75.2 ± 6.5 years) using a 3.0-T Siemens Tim Trio scanner. Among them, five subjects were scanned twice to assess the short-term reproducibility. The MEGA-PRESS (Mescher-Garwood point-resolved spectroscopy) J-editing sequence was used for GABA detection in two volumes of interest (VOIs) in the left and right cerebellar dentate. MRS data processing and quantification were performed with LCModel 6.3-0L using two separate basis sets, generated from density matrix simulations using published values for chemical shifts and J couplings. Raw metabolite levels from LCModel outputs were corrected for cerebrospinal fluid contamination and relaxation. GABA-edited spectra yielded robust and stable GABA measurements with averaged intra-individual coefficients of variation for corrected GABA+ between 4.0 ± 2.8% and 13.4 ± 6.3%, and inter-individual coefficients of variation between 12.6% and 24.2%. In addition, there was a significant correlation between GABA+ obtained with the two LCModel basis sets. Overall, our results demonstrated the feasibility and reproducibility of cerebellar GABA-edited MRS at 3.0 T in an elderly population. This information might be helpful for studies using this technique to study GABA changes in normal or diseased aging brain, e.g. for power calculations and the interpretation of longitudinal

  15. Peripartum neuroactive steroid and γ-aminobutyric acid profiles in women at-risk for postpartum depression.

    PubMed

    Deligiannidis, Kristina M; Kroll-Desrosiers, Aimee R; Mo, Shunyan; Nguyen, Hien P; Svenson, Abby; Jaitly, Nina; Hall, Janet E; Barton, Bruce A; Rothschild, Anthony J; Shaffer, Scott A

    2016-08-01

    Neuroactive steroids (NAS) are allosteric modulators of the γ-aminobutyric acid (GABA) system. NAS and GABA are implicated in depression. The peripartum period involves physiologic changes in NAS which may be associated with peripartum depression and anxiety. We measured peripartum plasma NAS and GABA in healthy comparison subjects (HCS) and those at-risk for postpartum depression (AR-PPD) due to current mild depressive or anxiety symptoms or a history of depression. We evaluated 56 peripartum medication-free subjects. We measured symptoms with the Hamilton Depression Rating Scale (HAM-D17), Hamilton Anxiety Rating Scale (HAM-A) and Spielberger State-Trait Anxiety Inventory-State (STAI-S). Plasma NAS and GABA were quantified by liquid chromatography-mass spectrometry. We examined the associations between longitudinal changes in NAS, GABA and depressive and anxiety symptoms using generalized estimating equation methods. Peripartum GABA concentration was 1.9±0.7ng/mL (p=0.004) lower and progesterone and pregnanolone were 15.8±7.5 (p=0.04) and 1.5±0.7ng/mL (p=0.03) higher in AR-PPD versus HCS, respectively. HAM-D17 was negatively associated with GABA (β=-0.14±0.05, p=0.01) and positively associated with pregnanolone (β=0.16±0.06, p=0.01). STAI-S was positively associated with pregnanolone (β=0.11±0.04, p=0.004), allopregnanolone (β=0.13±0.05, p=0.006) and pregnenolone (β=0.02±0.01, p=0.04). HAM-A was negatively associated with GABA (β=-0.12±0.04, p=0.004) and positively associated with pregnanolone (β=0.11±0.05, p=0.05). Altered peripartum NAS and GABA profiles in AR-PPD women suggest that their interaction may play an important role in the pathophysiology of peripartum depression and anxiety. PMID:27209438

  16. Reproducibility and effect of tissue composition on cerebellar γ-aminobutyric acid (GABA) MRS in an elderly population.

    PubMed

    Long, Zaiyang; Dyke, Jonathan P; Ma, Ruoyun; Huang, Chaorui C; Louis, Elan D; Dydak, Ulrike

    2015-10-01

    MRS provides a valuable tool for the non-invasive detection of brain γ-aminobutyric acid (GABA) in vivo. GABAergic dysfunction has been observed in the aging cerebellum. The study of cerebellar GABA changes is of considerable interest in understanding certain age-related motor disorders. However, little is known about the reproducibility of GABA MRS in an aged population. Therefore, this study aimed to explore the feasibility and reproducibility of GABA MRS in the aged cerebellum at 3.0 T and to examine the effect of differing tissue composition on GABA measurements. MRI and (1)H MRS examinations were performed on 10 healthy elderly volunteers (mean age, 75.2 ± 6.5 years) using a 3.0-T Siemens Tim Trio scanner. Among them, five subjects were scanned twice to assess the short-term reproducibility. The MEGA-PRESS (Mescher-Garwood point-resolved spectroscopy) J-editing sequence was used for GABA detection in two volumes of interest (VOIs) in the left and right cerebellar dentate. MRS data processing and quantification were performed with LCModel 6.3-0L using two separate basis sets, generated from density matrix simulations using published values for chemical shifts and J couplings. Raw metabolite levels from LCModel outputs were corrected for cerebrospinal fluid contamination and relaxation. GABA-edited spectra yielded robust and stable GABA measurements with averaged intra-individual coefficients of variation for corrected GABA+ between 4.0 ± 2.8% and 13.4 ± 6.3%, and inter-individual coefficients of variation between 12.6% and 24.2%. In addition, there was a significant correlation between GABA+ obtained with the two LCModel basis sets. Overall, our results demonstrated the feasibility and reproducibility of cerebellar GABA-edited MRS at 3.0 T in an elderly population. This information might be helpful for studies using this technique to study GABA changes in normal or diseased aging brain, e.g. for power calculations and the interpretation of longitudinal

  17. Gas release-based prescreening combined with reversed-phase HPLC quantitation for efficient selection of high-γ-aminobutyric acid (GABA)-producing lactic acid bacteria.

    PubMed

    Wu, Qinglong; Shah, Nagendra P

    2015-02-01

    High γ-aminobutyric acid (GABA)-producing lactobacilli are promising for the manufacture of GABA-rich foods and to synthesize GRAS (generally recognized as safe)-grade GABA. However, common chromatography-based screening is time-consuming and inefficient. In the present study, Korean kimchi was used as a model of lactic acid-based fermented foods, and a gas release-based prescreening of potential GABA producers was developed. The ability to produce GABA by potential GABA producers in de Man, Rogosa, and Sharpe medium supplemented with or without monosodium glutamate was further determined by HPLC. Based on the results, 9 isolates were regarded as high GABA producers, and were further genetically identified as Lactobacillus brevis based on the sequences of 16S rRNA gene. Gas release-based prescreening combined with reversed-phase HPLC confirmation was an efficient and cost-effective method to identify high-GABA-producing LAB, which could be good candidates for probiotics. The GABA that is naturally produced by these high-GABA-producing LAB could be used as a food additive.

  18. Enzymatic preparation of. cap alpha. - and. beta. -deuterated or tritiated amino acids with l-methionine. gamma. -lyase

    SciTech Connect

    Esaki, N.; Sawada, S.; Tanaka, H.; Soda, K.

    1982-01-15

    L-Methionine ..gamma..-lyase catalyzes the exchange of ..cap alpha..- and ..beta..-hydrogens of L-methionine and S-methyl-L-cysteine with deuterium or tritium of solvents. The rate of ..cap alpha..-hydrogen exchange with deuterium was about 40 times faster than that of the elimination reactions. The deuterium and tritium were exchanged also with the ..cap alpha..- and ..beta..-hydrogens of the straight-chain amino acids which do not undergo the elimination: L-alanine, L-..cap alpha..-aminobutyrate, L-norvaline, and L-norleucine. No exchange occurs for the D-isomers, acidic L-amino acids, basic L-amino acids, and branched-chain L-amino acids, although ..cap alpha..-hydrogen of glycine, L-trypotophan, and L-phenylalanine is exchanged slowly. These enzymatic hydrogen-exchange reactions facilitate specific labeling of the L-amino acids with deuterium and tritium.

  19. Mechanism of Inactivation of γ-Aminobutyric Acid Aminotransferase by (1S ,3S)-3-Amino-4-difluoromethylene-1-cyclopentanoic Acid (CPP-115)

    DOE PAGESBeta

    Lee, Hyunbeom; Doud, Emma H.; Wu, Rui; Sanishvili, Ruslan; Juncosa, Jose I.; Liu, Dali; Kelleher, Neil L.; Silverman, Richard B.

    2015-01-23

    γ-Aminobutyric acid aminotransferase (GABA-AT) is a pyridoxal 5'-phosphate (PLP)-dependent enzyme that degrades GABA, the principal inhibitory neurotransmitter in mammalian cells. When the concentration of GABA falls below a threshold level, convulsions can occur. Inhibition of GABA-AT raises GABA levels in the brain, which can terminate seizures as well as have potential therapeutic applications in treating other neurological disorders, including drug addiction. Among the analogues that we previously developed, (1S,3S)-3-amino-4-difluoromethylene-1-cyclopentanoic acid (CPP-115) showed 187 times greater potency than that of vigabatrin, a known inactivator of GABA-AT and approved drug (Sabril) for the treatment of infantile spasms and refractory adult epilepsy. Recently,more » CPP-115 was shown to have no adverse effects in a Phase I clinical trial. Here we report a novel inactivation mechanism for CPP-115, a mechanism-based inactivator that undergoes GABA-AT-catalyzed hydrolysis of the difluoromethylene group to a carboxylic acid with concomitant loss of two fluoride ions and coenzyme conversion to pyridoxamine 5'-phosphate (PMP). The partition ratio for CPP-115 with GABA-AT is about 2000, releasing cyclopentanone-2,4-dicarboxylate (22) and two other precursors of this compound (20 and 21). Time-dependent inactivation occurs by a conformational change induced by the formation of the aldimine of 4-aminocyclopentane-1,3-dicarboxylic acid and PMP (20), which disrupts an electrostatic interaction between Glu270 and Arg445 to form an electrostatic interaction between Arg445 and the newly formed carboxylate produced by hydrolysis of the difluoromethylene group in CPP-115, resulting in a noncovalent, tightly bound complex. Ultimately, this represents a novel mechanism for inactivation of GABA-AT and a new approach for the design of mechanism-based inactivators in general.« less

  20. Mechanism of Inactivation of γ-Aminobutyric Acid Aminotransferase by (1S,3S)-3-Amino-4-difluoromethylene-1-cyclopentanoic Acid (CPP-115)

    PubMed Central

    2016-01-01

    γ-Aminobutyric acid aminotransferase (GABA-AT) is a pyridoxal 5′-phosphate (PLP)-dependent enzyme that degrades GABA, the principal inhibitory neurotransmitter in mammalian cells. When the concentration of GABA falls below a threshold level, convulsions can occur. Inhibition of GABA-AT raises GABA levels in the brain, which can terminate seizures as well as have potential therapeutic applications in treating other neurological disorders, including drug addiction. Among the analogues that we previously developed, (1S,3S)-3-amino-4-difluoromethylene-1-cyclopentanoic acid (CPP-115) showed 187 times greater potency than that of vigabatrin, a known inactivator of GABA-AT and approved drug (Sabril) for the treatment of infantile spasms and refractory adult epilepsy. Recently, CPP-115 was shown to have no adverse effects in a Phase I clinical trial. Here we report a novel inactivation mechanism for CPP-115, a mechanism-based inactivator that undergoes GABA-AT-catalyzed hydrolysis of the difluoromethylene group to a carboxylic acid with concomitant loss of two fluoride ions and coenzyme conversion to pyridoxamine 5′-phosphate (PMP). The partition ratio for CPP-115 with GABA-AT is about 2000, releasing cyclopentanone-2,4-dicarboxylate (22) and two other precursors of this compound (20 and 21). Time-dependent inactivation occurs by a conformational change induced by the formation of the aldimine of 4-aminocyclopentane-1,3-dicarboxylic acid and PMP (20), which disrupts an electrostatic interaction between Glu270 and Arg445 to form an electrostatic interaction between Arg445 and the newly formed carboxylate produced by hydrolysis of the difluoromethylene group in CPP-115, resulting in a noncovalent, tightly bound complex. This represents a novel mechanism for inactivation of GABA-AT and a new approach for the design of mechanism-based inactivators in general. PMID:25616005

  1. Straightforward and effective synthesis of γ-aminobutyric acid transporter subtype 2-selective acyl-substituted azaspiro[4.5]decanes.

    PubMed

    Ma, Xiaofeng; Lubin, Hodney; Ioja, Enikő; Kékesi, Orsolya; Simon, Ágnes; Apáti, Ágota; Orbán, Tamás I; Héja, László; Kardos, Julianna; Markó, István E

    2016-01-15

    Supply of major metabolites such as γ-aminobutyric acid (GABA), β-alanine and taurine is an essential instrument that shapes signalling, proper cell functioning and survival in the brain and peripheral organs. This background motivates the synthesis of novel classes of compounds regulating their selective transport through various fluid-organ barriers via the low-affinity γ-aminobutyric acid (GABA) transporter subtype 2 (GAT2). Natural and synthetic spirocyclic compounds or therapeutics with a range of structures and biological activity are increasingly recognised in this regard. Based on pre-validated GABA transport activity, straightforward and efficient synthesis method was developed to provide an azaspiro[4.5]decane scaffold, holding a variety of charge, substituent and 3D constrain of spirocyclic amine. Investigation of the azaspiro[4.5]decane scaffold in cell lines expressing the four GABA transporter subtypes led to the discovery of a subclass of a GAT2-selective compounds with acyl-substituted azaspiro[4.5]decane core.

  2. Straightforward and effective synthesis of γ-aminobutyric acid transporter subtype 2-selective acyl-substituted azaspiro[4.5]decanes.

    PubMed

    Ma, Xiaofeng; Lubin, Hodney; Ioja, Enikő; Kékesi, Orsolya; Simon, Ágnes; Apáti, Ágota; Orbán, Tamás I; Héja, László; Kardos, Julianna; Markó, István E

    2016-01-15

    Supply of major metabolites such as γ-aminobutyric acid (GABA), β-alanine and taurine is an essential instrument that shapes signalling, proper cell functioning and survival in the brain and peripheral organs. This background motivates the synthesis of novel classes of compounds regulating their selective transport through various fluid-organ barriers via the low-affinity γ-aminobutyric acid (GABA) transporter subtype 2 (GAT2). Natural and synthetic spirocyclic compounds or therapeutics with a range of structures and biological activity are increasingly recognised in this regard. Based on pre-validated GABA transport activity, straightforward and efficient synthesis method was developed to provide an azaspiro[4.5]decane scaffold, holding a variety of charge, substituent and 3D constrain of spirocyclic amine. Investigation of the azaspiro[4.5]decane scaffold in cell lines expressing the four GABA transporter subtypes led to the discovery of a subclass of a GAT2-selective compounds with acyl-substituted azaspiro[4.5]decane core. PMID:26706177

  3. Proteomic analysis of B-aminobutyric acid priming and aba-induction of drought resistance in crabapple (Malus pumila): effect on general metabolism, the phenylpropanoid pathway and cell wall enzymes

    Technology Transfer Automated Retrieval System (TEKTRAN)

    In a variety of annual crops and model plants, the xenobiotic compound, DL-beta-aminobutyric acid (BABA), has been shown to enhance disease resistance and increase salt, drought, and thermotolerance. BABA does not activate stress genes directly but rather sensitizes plants to respond more quickly a...

  4. Effect of β-aminobutyric acid on cell wall modification and senescence in sweet cherry during storage at 20°C.

    PubMed

    Wang, Lei; Jin, Peng; Wang, Jing; Jiang, Lulu; Shan, Timin; Zheng, Yonghua

    2015-05-15

    The effects of postharvest β-aminobutyric acid (BABA) treatment on fruit firmness, pectin degrading enzymes, cell wall constituents and microstructural alterations of pericarp in sweet cherry fruit were investigated. BABA significantly delayed the decline of fruit firmness and inhibited the increase of membrane permeability and the accumulation of malondialdehyde in cherries. The BABA-treated fruit exhibited significantly higher contents of water-soluble pectin, CDTA-soluble pectin, Na2CO3-soluble pectin, total pectin, cellulose and hemicellulose than the control during storage. Activities of pectin degrading enzymes including polygalacturonase and pectinmethylesterase were markedly reduced by BABA treatment. Observations by scanning electron microscopy showed BABA maintained smooth cuticle and integrated structure of subepidermal cell in sweet cherry. These results suggest that the delay in fruit senescence by BABA may be due to depressed membrane permeability and malondialdehyde content, reduced activities of polygalacturonase and pectinmethylesterase, enhanced cell wall polysaccharides content, and integrated subepidermal cell structure in sweet cherry.

  5. Exogenous γ-aminobutyric acid (GABA) affects pollen tube growth via modulating putative Ca2+-permeable membrane channels and is coupled to negative regulation on glutamate decarboxylase.

    PubMed

    Yu, Guang-Hui; Zou, Jie; Feng, Jing; Peng, Xiong-Bo; Wu, Ju-You; Wu, Ying-Liang; Palanivelu, Ravishankar; Sun, Meng-Xiang

    2014-07-01

    γ-Aminobutyric acid (GABA) is implicated in pollen tube growth, but the molecular and cellular mechanisms that it mediates are largely unknown. Here, it is shown that exogenous GABA modulates putative Ca(2+)-permeable channels on the plasma membranes of tobacco pollen grains and pollen tubes. Whole-cell voltage-clamp experiments and non-invasive micromeasurement technology (NMT) revealed that the influx of Ca(2+) increases in pollen tubes in response to exogenous GABA. It is also demonstrated that glutamate decarboxylase (GAD), the rate-limiting enzyme of GABA biosynthesis, is involved in feedback controls of Ca(2+)-permeable channels to fluctuate intracellular GABA levels and thus modulate pollen tube growth. The findings suggest that GAD activity linked with Ca(2+)-permeable channels relays an extracellular GABA signal and integrates multiple signal pathways to modulate tobacco pollen tube growth. Thus, the data explain how GABA mediates the communication between the style and the growing pollen tubes.

  6. GABA shunt and polyamine degradation pathway on γ-aminobutyric acid accumulation in germinating fava bean (Vicia faba L.) under hypoxia.

    PubMed

    Yang, Runqiang; Guo, Qianghui; Gu, Zhenxin

    2013-01-01

    GABA shunt and polyamine degradation pathway on γ-aminobutyric acid (GABA) accumulation in germinating fava bean under hypoxia was investigated. GABA content, GAD and DAO activity were significantly increased under hypoxia treatment. Glu and polyamine contents enhanced largely and thus supplied as sufficient substrates for GABA formation. In contrast, GABA content decreased, mainly in the embryo, after removing the hypoxia stress. DAO activity, Glu and polyamines contents decreased, while an increment of GAD activity was observed. This indicated that GAD activity can be not only regulated by hypoxia, but by the rapid growth of embryo after the recovery from hypoxia stress. When treated with AG, DAO activity was almost inhibited completely, and the GABA content decreased by 32.96% and 32.07% after treated for 3 and 5 days, respectively. Hence, it can be inferred that about 30% of GABA formed in germinating fava bean under hypoxia was supplied by polyamine degradation pathway. PMID:23017406

  7. GABA shunt and polyamine degradation pathway on γ-aminobutyric acid accumulation in germinating fava bean (Vicia faba L.) under hypoxia.

    PubMed

    Yang, Runqiang; Guo, Qianghui; Gu, Zhenxin

    2013-01-01

    GABA shunt and polyamine degradation pathway on γ-aminobutyric acid (GABA) accumulation in germinating fava bean under hypoxia was investigated. GABA content, GAD and DAO activity were significantly increased under hypoxia treatment. Glu and polyamine contents enhanced largely and thus supplied as sufficient substrates for GABA formation. In contrast, GABA content decreased, mainly in the embryo, after removing the hypoxia stress. DAO activity, Glu and polyamines contents decreased, while an increment of GAD activity was observed. This indicated that GAD activity can be not only regulated by hypoxia, but by the rapid growth of embryo after the recovery from hypoxia stress. When treated with AG, DAO activity was almost inhibited completely, and the GABA content decreased by 32.96% and 32.07% after treated for 3 and 5 days, respectively. Hence, it can be inferred that about 30% of GABA formed in germinating fava bean under hypoxia was supplied by polyamine degradation pathway.

  8. Effect of γ-aminobutyric acid on digestive enzymes, absorption function, and immune function of intestinal mucosa in heat-stressed chicken.

    PubMed

    Chen, Z; Xie, J; Wang, B; Tang, J

    2014-10-01

    To explore the effect of dietary γ-aminobutyric acid (GABA) on digestive enzyme activity, absorption function and immune function of intestinal mucosa in heat-stressed Wenchang chicken were studied. One-day-old male Wenchang chickens were randomly divided into a control group (CK), heat stress group (HS), and GABA+HS group. The chickens from the GABA+HS group were administered with 0.2 mL of GABA solution daily. Chickens from HS and GABA+HS groups were subjected to heat stress treatment at 40 ± 0.5°C for 2 h during 1300 to 1500 h every day. Blood was drawn and 0.5 cm-long duodenum, jejunum, and ileum were collected from the chickens on d 3, 5, 7, 9, 12, and 15. Results showed that the activity of Ca²⁺-Mg²⁺-adenosine triphosphatase (ATPase), Na⁺-K⁺-ATPase, maltase, sucrase, and alkaline phosphatase, the contents of secretory IgA, glutathione, and d-xylose, and the number of lymphocytes in HS group were significantly lower than those in the CK group. Among them, some were rescued after the treatment of GABA as the time extension. For maltase, d-xylose, alkaline phosphatase, and Na⁺-K⁺-ATPase, it required 5 to 7 d for achieving the significant effect. For sucrase, 12 d for the alleviation effect was required. In the case of other parameters, no alleviation was observed during the whole period of the study. We have concluded that HS can inhibit the activity of digestive enzymes and reduce absorption and immune functions of intestinal mucosa. γ-Aminobutyric acid can effectively alleviate these inhibitory effects.

  9. GABAB receptor-mediated activation of astrocytes by gamma-hydroxybutyric acid

    PubMed Central

    Gould, Timothy; Chen, Lixin; Emri, Zsuzsa; Pirttimaki, Tiina; Errington, Adam C.; Crunelli, Vincenzo; Parri, H. Rheinallt

    2014-01-01

    The gamma-aminobutyric acid (GABA) metabolite gamma-hydroxybutyric acid (GHB) shows a variety of behavioural effects when administered to animals and humans, including reward/addiction properties and absence seizures. At the cellular level, these actions of GHB are mediated by activation of neuronal GABAB receptors (GABABRs) where it acts as a weak agonist. Because astrocytes respond to endogenous and exogenously applied GABA by activation of both GABAA and GABABRs, here we investigated the action of GHB on astrocytes on the ventral tegmental area (VTA) and the ventrobasal (VB) thalamic nucleus, two brain areas involved in the reward and proepileptic action of GHB, respectively, and compared it with that of the potent GABABR agonist baclofen. We found that GHB and baclofen elicited dose-dependent (ED50: 1.6 mM and 1.3 µM, respectively) transient increases in intracellular Ca2+ in VTA and VB astrocytes of young mice and rats, which were accounted for by activation of their GABABRs and mediated by Ca2+ release from intracellular store release. In contrast, prolonged GHB and baclofen exposure caused a reduction in spontaneous astrocyte activity and glutamate release from VTA astrocytes. These findings have key (patho)physiological implications for our understanding of the addictive and proepileptic actions of GHB. PMID:25225100

  10. Enhancement of γ-aminobutyric acid (GABA) in Nham (Thai fermented pork sausage) using starter cultures of Lactobacillus namurensis NH2 and Pediococcus pentosaceus HN8.

    PubMed

    Ratanaburee, Anussara; Kantachote, Duangporn; Charernjiratrakul, Wilawan; Sukhoom, Ampaitip

    2013-10-15

    The aim was to produce Nham that was enriched with γ-aminobutyric acid (GABA); therefore two GABA producing lactic acid bacteria (Pediococcus pentosaceus HN8 and Lactobacillus namurensis NH2) were used as starter cultures. By using the central composite design (CCD) we showed that addition of 0.5% monosodium glutamate (MSG) together with an inoculum size of roughly 6logCFU/g of each of the two strains produced a maximal amounts of GABA (4051 mg/kg) in the 'GABA Nham' product. This was higher than any current popular commercial Nham product by roughly 8 times. 'GABA Nham' with the additions of both starters and MSG (TSM) supported maximum populations of lactic acid bacteria (LAB) with a minimum of yeasts and no staphylococci or molds when compared to the controls that had no addition of any starters or MSG (TNN), or only the addition of MSG (TNM), or with only the starter (TSN). Based on proximate analysis among the Nham sets, 'GABA Nham' was low in fat, carbohydrate and energy although its texture and color were slightly different from the control (TNN). However, sensory evaluations of 'GABA Nham' were more acceptable than the controls and commercial Nham products for all tested parameters. Hence, a unique novel 'GABA Nham' fermented pork sausage was successfully developed.

  11. Gamma linolenic acid: an antiinflammatory omega-6 fatty acid.

    PubMed

    Kapoor, Rakesh; Huang, Yung-Sheng

    2006-12-01

    Inflammation plays an important role in health and disease. Most of the chronic diseases of modern society, including cancer, diabetes, heart disease, arthritis, Alzheimer's disease, etc. have inflammatory component. At the same time, the link between diet and disease is also being recognized. Amongst dietary constituents, fat has gained most recognition in affecting health. Saturated and trans fatty acids have been implicated in obesity, heart disease, diabetes and cancer while polyunsaturated fatty acids (PUFAs) generally have a positive effect on health. The PUFAs of omega-3 and omega-6 series play a significant role in health and disease by generating potent modulatory molecules for inflammatory responses, including eicosanoids (prostaglandins, and leukotrienes), and cytokines (interleukins) and affecting the gene expression of various bioactive molecules. Gamma linolenic acid (GLA, all cis 6, 9, 12-Octadecatrienoic acid, C18:3, n-6), is produced in the body from linoleic acid (all cis 6,9-octadecadienoic acid), an essential fatty acid of omega-6 series by the enzyme delta-6-desaturase. Preformed GLA is present in trace amounts in green leafy vegetables and in nuts. The most significant source of GLA for infants is breast milk. GLA is further metabolized to dihomogamma linlenic acid (DGLA) which undergoes oxidative metabolism by cyclooxygenases and lipoxygenases to produce anti-inflammatory eicosanoids (prostaglandins of series 1 and leukotrienes of series 3). GLA and its metabolites also affect expression of various genes where by regulating the levels of gene products including matrix proteins. These gene products play a significant role in immune functions and also in cell death (apoptosis). The present review will emphasize the role of GLA in modulating inflammatory response, and hence its potential applications as an anti-inflammatory nutrient or adjuvant.

  12. Dairy Streptococcus thermophilus improves cell viability of Lactobacillus brevis NPS-QW-145 and its γ-aminobutyric acid biosynthesis ability in milk

    PubMed Central

    Wu, Qinglong; Law, Yee-Song; Shah, Nagendra P.

    2015-01-01

    Most high γ-aminobutyric acid (GABA) producers are Lactobacillus brevis of plant origin, which may be not able to ferment milk well due to its poor proteolytic nature as evidenced by the absence of genes encoding extracellular proteinases in its genome. In the present study, two glutamic acid decarboxylase (GAD) genes, gadA and gadB, were found in high GABA-producing L. brevis NPS-QW-145. Co-culturing of this organism with conventional dairy starters was carried out to manufacture GABA-rich fermented milk. It was observed that all the selected strains of Streptococcus thermophilus, but not Lactobacillus delbrueckii subsp. bulgaricus, improved the viability of L. brevis NPS-QW-145 in milk. Only certain strains of S. thermophilus improved the gadA mRNA level in L. brevis NPS-QW-145, thus enhanced GABA biosynthesis by the latter. These results suggest that certain S. thermophilus strains are highly recommended to co-culture with high GABA producer for manufacturing GABA-rich fermented milk. PMID:26245488

  13. Comparison of γ-Aminobutyric Acid and Biogenic Amine Content of Different types of Ewe’s Milk Cheese Produced in Sardinia, Italy

    PubMed Central

    Porcu, Arianna; Ru, Antonio; Salaris, Margherita; Franco, Mario A.; De Santis, Enrico P.L.

    2015-01-01

    The bioactive compounds γ-aminobutyric acid (GABA) and biogenic amines (BA), together with protein-free amino acids, were measured by high-performance liquid chromatography in ewe’s milk cheeses produced in Sardinia with different technological traits. The study included three types of cheese: Pecorino Sardo PDO, Pecorino and Casu Marzu. Farmhouse Casu Marzu and Pecorino showed GABA content (maximum levels: 1001.3 and 378.1 mg 100 g–1 respectively) that had never been found so high in cheese before, suggesting that these types of cheese present ideal conditions to produce GABA. These two types of cheese also showed high levels of BA (their total maximum levels were 1035.7 and 288.0 mg 100 g–1 respectively). Pearson correlation analysis detected significant correlation between GABA and the main BA present in the cheeses (tyramine, cadaverine and putrescine), suggesting that the factors affecting the production of GABA are the same as those influencing BA formation. PMID:27800396

  14. Oral intake of γ-aminobutyric acid affects mood and activities of central nervous system during stressed condition induced by mental tasks.

    PubMed

    Yoto, A; Murao, S; Motoki, M; Yokoyama, Y; Horie, N; Takeshima, K; Masuda, K; Kim, M; Yokogoshi, H

    2012-09-01

    γ-Aminobutyric acid (GABA) is a kind of amino acid contained in green tea leaves and other foods. Several reports have shown that GABA might affect brain protein synthesis, improve many brain functions such as memory and study capability, lower the blood pressure of spontaneously hypertensive rats, and may also have a relaxation effect in humans. However, the evidence for its mood-improving function is still not sufficient. In this study, we investigated how the oral intake of GABA influences human adults psychologically and physiologically under a condition of mental stress. Sixty-three adults (28 males, 35 females) participated in a randomized, single blind, placebo-controlled, crossover-designed study over two experiment days. Capsules containing 100 mg of GABA or dextrin as a placebo were used as test samples. The results showed that EEG activities including alpha band and beta band brain waves decreased depending on the mental stress task loads, and the condition of 30 min after GABA intake diminished this decrease compared with the placebo condition. That is to say, GABA might have alleviated the stress induced by the mental tasks. This effect also corresponded with the results of the POMS scores. PMID:22203366

  15. Dairy Streptococcus thermophilus improves cell viability of Lactobacillus brevis NPS-QW-145 and its γ-aminobutyric acid biosynthesis ability in milk.

    PubMed

    Wu, Qinglong; Law, Yee-Song; Shah, Nagendra P

    2015-08-06

    Most high γ-aminobutyric acid (GABA) producers are Lactobacillus brevis of plant origin, which may be not able to ferment milk well due to its poor proteolytic nature as evidenced by the absence of genes encoding extracellular proteinases in its genome. In the present study, two glutamic acid decarboxylase (GAD) genes, gadA and gadB, were found in high GABA-producing L. brevis NPS-QW-145. Co-culturing of this organism with conventional dairy starters was carried out to manufacture GABA-rich fermented milk. It was observed that all the selected strains of Streptococcus thermophilus, but not Lactobacillus delbrueckii subsp. bulgaricus, improved the viability of L. brevis NPS-QW-145 in milk. Only certain strains of S. thermophilus improved the gadA mRNA level in L. brevis NPS-QW-145, thus enhanced GABA biosynthesis by the latter. These results suggest that certain S. thermophilus strains are highly recommended to co-culture with high GABA producer for manufacturing GABA-rich fermented milk.

  16. [Response of reactive oxygen metabolism in melon chloroplasts to short-term salinity-alkalinity stress regulated by exogenous γ-aminobutyric acid].

    PubMed

    Xiang, Li-xia; Hu, Li-pan; Hu, Xiao-hui; Pan, Xiong-bo; Ren, Wen-qi

    2015-12-01

    The regulatory effect of exogenous γ-aminobutyric acid (GABA) on metabolism of reactive oxygen species (ROS) in melon chloroplasts under short-term salinity-alkalinity stress were investigated in melon variety 'Jinhui No. 1', which was cultured with deep flow hydroponics. The result showed that under salinity-alkalinity stress, the photosynthetic pigment content, MDA content, superoxide anion (O₂·) production rate and hydrogen peroxide (H₂O₂) content in chloroplast increased significantly, the contents of antioxidants ascorbic acid (AsA) and glutathione (GSH) increased, and the activities of H⁺-ATPase and H⁺-PPiase were inhibited obviously. With exogenous GABA application, the accumulations of O₂·, MDA and H₂O₂ induced by salinity-alkalinity stress were inhibited. Exogenous GABA alleviated the increase of photosynthetic pigment content, improved the activity of SOD, enzymes of AsA-GSH cycle, total AsA and total GSH while decreased the AsA/DHA ratio and GSH/GSSH ratio. Foliar GABA could enhance the H⁺-ATPase and H⁺-PPiase activities. Our results suggested that the exogenous GABA could accelerate the ROS metabolism in chloroplast, promote the recycle of AsA-GSH, and maintain the permeability of cell membrane to improve the ability of melon chloroplast against salinity-alkalinity stress.

  17. Oral intake of γ-aminobutyric acid affects mood and activities of central nervous system during stressed condition induced by mental tasks.

    PubMed

    Yoto, A; Murao, S; Motoki, M; Yokoyama, Y; Horie, N; Takeshima, K; Masuda, K; Kim, M; Yokogoshi, H

    2012-09-01

    γ-Aminobutyric acid (GABA) is a kind of amino acid contained in green tea leaves and other foods. Several reports have shown that GABA might affect brain protein synthesis, improve many brain functions such as memory and study capability, lower the blood pressure of spontaneously hypertensive rats, and may also have a relaxation effect in humans. However, the evidence for its mood-improving function is still not sufficient. In this study, we investigated how the oral intake of GABA influences human adults psychologically and physiologically under a condition of mental stress. Sixty-three adults (28 males, 35 females) participated in a randomized, single blind, placebo-controlled, crossover-designed study over two experiment days. Capsules containing 100 mg of GABA or dextrin as a placebo were used as test samples. The results showed that EEG activities including alpha band and beta band brain waves decreased depending on the mental stress task loads, and the condition of 30 min after GABA intake diminished this decrease compared with the placebo condition. That is to say, GABA might have alleviated the stress induced by the mental tasks. This effect also corresponded with the results of the POMS scores.

  18. [Response of reactive oxygen metabolism in melon chloroplasts to short-term salinity-alkalinity stress regulated by exogenous γ-aminobutyric acid].

    PubMed

    Xiang, Li-xia; Hu, Li-pan; Hu, Xiao-hui; Pan, Xiong-bo; Ren, Wen-qi

    2015-12-01

    The regulatory effect of exogenous γ-aminobutyric acid (GABA) on metabolism of reactive oxygen species (ROS) in melon chloroplasts under short-term salinity-alkalinity stress were investigated in melon variety 'Jinhui No. 1', which was cultured with deep flow hydroponics. The result showed that under salinity-alkalinity stress, the photosynthetic pigment content, MDA content, superoxide anion (O₂·) production rate and hydrogen peroxide (H₂O₂) content in chloroplast increased significantly, the contents of antioxidants ascorbic acid (AsA) and glutathione (GSH) increased, and the activities of H⁺-ATPase and H⁺-PPiase were inhibited obviously. With exogenous GABA application, the accumulations of O₂·, MDA and H₂O₂ induced by salinity-alkalinity stress were inhibited. Exogenous GABA alleviated the increase of photosynthetic pigment content, improved the activity of SOD, enzymes of AsA-GSH cycle, total AsA and total GSH while decreased the AsA/DHA ratio and GSH/GSSH ratio. Foliar GABA could enhance the H⁺-ATPase and H⁺-PPiase activities. Our results suggested that the exogenous GABA could accelerate the ROS metabolism in chloroplast, promote the recycle of AsA-GSH, and maintain the permeability of cell membrane to improve the ability of melon chloroplast against salinity-alkalinity stress. PMID:27112014

  19. Mutations in y-aminobutyric acid (GABA) transaminase genes in plants or Pseudomonas syringae reduce bacterial virulence

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Pseudomonas syringae pv. tomato DC3000 is a bacterial pathogen of Arabidopsis and tomato that grows in the apoplast. The non-protein amino acid '-amino butyric acid (GABA) is produced by Arabidopsis and tomato and is the most abundant amino acid in the apoplastic fluid of tomato. The DC3000 genome h...

  20. Metabolism and transport of gamma-carboxyglutamic acid.

    PubMed

    Shah, D V; Tews, J K; Harper, A E; Suttie, J W

    1978-03-01

    gamma-Carboxyglutamic acid residues have beeh shown to be present in prothrombin, the other vitamin K-dependent clotting factors, and more recently in bone and kidney proteins. This amino acid is formed by a posttranslational vitamin K-dependent carboxylation of glutamyl residues in polypeptide precursors of these protens. It has now been demonstrated that this amino acid, either in the free or peptide-bound form, is not metabolically degraded by the rat, but is quantitatively excreted in the urine. In nephrectomized rats, the tissue concentration of intravenously administered gamma-carboxyglutamic acid is increased, but there is still no evidence of any oxidative metabolism of this amino acid. These amino acid is transported by kidney slices against a concentration gradient, but does not accumulate in liver, intestinal or brain tissues. Preliminary data suggest that gamma-carboxyglutamic acid may be concentrated by a carrier system different from that utilized by other amino acids. PMID:629998

  1. Impact of Precooling and Controlled-Atmosphere Storage on γ-Aminobutyric Acid (GABA) Accumulation in Longan (Dimocarpus longan Lour.) Fruit.

    PubMed

    Zhou, Molin; Ndeurumio, Kessy H; Zhao, Lei; Hu, Zhuoyan

    2016-08-24

    Longan (Dimocarpus longan Lour.) fruit cultivars 'Chuliang' and 'Shixia' were analyzed for γ-aminobutyric acid (GABA) accumulation after precooling and in controlled-atmosphere storage. Fruit were exposed to 5% O2 plus 3%, 5%, or 10% CO2 at 4 °C, and GABA and associated enzymes, aril firmness, and pericarp color were measured. Aril softening and pericarp browning were delayed by 5% CO2 + 5% O2. GABA concentrations and glutamate decarboxylase (GAD; EC 4.1.1.15) activities declined during storage at the higher-CO2 treatments. However, GABA aminotransferase (GABA-T; EC 2.6.1.19) activities in elevated CO2-treated fruit fluctuated during storage. GABA concentrations increased after precooling treatments. GAD activity and GABA-T activity were different between cultivars after precooling. GABA concentrations in fruit increased after 3 days of 10% CO2 + 5% O2 treatment and then declined as storage time increased. GABA accumulation was associated with stimulation of GAD activity rather than inhibition of GABA-T activity.

  2. Evaluation of γ- aminobutyric acid, phytate and antioxidant activity of tempeh-like fermented oats (Avena sativa L.) prepared with different filamentous fungi.

    PubMed

    Cai, Shengbao; Gao, Fengyi; Zhang, Xudong; Wang, Ou; Wu, Wei; Zhu, Songjie; Zhang, Di; Zhou, Feng; Ji, Baoping

    2014-10-01

    Tempeh is a popular traditional fermented food in Asia. Many tempeh-like foods are made from cereal grains. However, the information of γ-aminobutyric acid (GABA) accumulation in those tempeh-like cereal grains during fermentation is lacking. Meanwhile, little information is available on the anti-nutrient contents and antioxidant activity of tempeh-like fermented oats. The aim of the present work was to study the changes of GABA, phytate, natural antioxidants and antioxidant activity of tempeh-like fermented oats. As fermentation time progressed, the GABA, total phenolics content (TPC) and flavonoids increased rapidly. The Aspergillus oryzae-fermented oats had the highest GABA, whereas Rhizopus oryzae-fermented oats had the highest TPC. Phytate, an anti-nutrient component, was dramatically reduced in the fermented oats, especially those by A. oryzae (reduced by about 63 %). The antioxidant activities of fermented oats were also significantly enhanced after 72 h fermentation (p < 0.05). This study demonstrated that oats fermented by generally recognized as safe (GRAS) fungi can be recommended as tempeh-like functional foods with higher GABA, more natural antioxidants and lower phytate compared with native oats.

  3. Determination of therapeutic γ-aminobutyric acid analogs in forensic whole blood by hydrophilic interaction liquid chromatography-electrospray tandem mass spectrometry.

    PubMed

    Sørensen, Lambert K; Hasselstrøm, Jørgen B

    2014-05-01

    Vigabatrin, pregabalin, gabapentin and baclofen are γ-aminobutyric acid analogs that are used in the treatment of epileptic seizures (vigabatrin, pregabalin and gabapentin) and spasticity (baclofen). The intake of these drugs may induce adverse reactions and impair the ability of an individual to drive a vehicle. There have also been reports of cases of intoxication and fatalities from overdoses. For rapid and accurate quantification of these drugs in forensic cases, an ultraperformance liquid chromatography tandem mass spectrometry method using pneumatically assisted electrospray ionization has been developed. The technique has been validated on both ante- and postmortem human whole blood. The protein in the blood samples was removed by the addition of a mixture of methanol and acetonitrile, and the extract was ultrafiltered and diluted with acetonitrile. The separation was performed by hydrophilic interaction liquid chromatography. Calibration of the system was achieved through use of matrix-matched calibrants combined with isotope dilution. The lower limits of quantification were 0.02-0.04 mg/L, and the relative intra-laboratory reproducibility standard deviations were <4 and 8% at concentrations of 10 and 1 mg/L, respectively. The mean true recoveries were >89%. The trueness expressed as the relative bias of the test results was within ±7% at concentrations of 1-40 mg/L for vigabatrin, pregabalin and gabapentin and of 0.1-4 mg/L for baclofen.

  4. Accumulation of γ‐aminobutyric acid by E nterococcus avium 9184 in scallop solution in a two‐stage fermentation strategy

    PubMed Central

    Yang, Haoyue; Hu, Linfeng; Liu, Song

    2015-01-01

    Summary In this study, a new bacterial strain having a high ability to produce γ‐aminobutyric acid (GABA) was isolated from naturally fermented scallop solution and was identified as E nterococcus avium. To the best of our knowledge, this is the first study to prove that E . avium possesses glutamate decarboxylase activity. The strain was then mutagenized with UV radiation and was designated as E . avium 9184. Scallop solution was used as the culture medium to produce GABA. A two‐stage fermentation strategy was applied to accumulate GABA. In the first stage, cell growth was regulated. Optimum conditions for cell growth were pH, 6.5; temperature, 37°C; and glucose concentration, 10 g·L−1. This produced a maximum dry cell mass of 2.10 g·L−1. In the second stage, GABA formation was regulated. GABA concentration reached 3.71 g·L−1 at 96 h pH 6.0, 37°C and initial l‐monosodium glutamate concentration of 10 g·L−1. Thus, compared with traditional one‐stage fermentation, the two‐stage fermentation significantly increased GABA accumulation. These results provide preliminary data to produce GABA using E . avium and also provide a new approach to process and utilize shellfish. PMID:26200650

  5. Further evidence for involvement of the dorsal hippocampus serotonergic and γ-aminobutyric acid (GABA)ergic pathways in the expression of contextual fear conditioning in rats.

    PubMed

    Almada, Rafael C; Albrechet-Souza, Lucas; Brandão, Marcus L

    2013-12-01

    Intra-dorsal hippocampus (DH) injections of 8-hydroxy-2-(di-n-propylamino) tetralin (8-OH-DPAT), a serotonin-1A (5-hydroxytryptamine (5-HT)-1A) receptor agonist, were previously shown to inhibit the expression of contextual fear when administered six hours after conditioning. However, further understanding of the consolidation and expression of aversive memories requires investigations of these and other mechanisms at distinct time points and the regions of the brain to which they are transferred. Thus, the purpose of the present study was to investigate the role of DH serotonergic and γ-aminobutyric acid (GABA)ergic mechanisms in the expression of contextual fear 24 h after conditioning, reflected by fear-potentiated startle (FPS) and freezing behavior. The recruitment of the amygdala and medial prefrontal cortex (mPFC) in these processes was also evaluated by measuring Fos protein immunoreactivity. Although intra-DH injections of 8-OH-DPAT did not produce behavioral changes, muscimol reduced both FPS and the freezing response. Fos protein immunoreactivity revealed that contextual fear promoted wide activation of the mPFC, which was significantly reduced after intra-DH infusions of muscimol. The present findings, together with previous data, indicate that in contrast to 5-HT, which appears to play a role during the early phases of contextual aversive memory consolidation, longer-lasting GABA-mediated mechanisms are recruited during the expression of contextual fear memories.

  6. Virus-induced gene silencing reveals control of reactive oxygen species accumulation and salt tolerance in tomato by γ-aminobutyric acid metabolic pathway.

    PubMed

    Bao, Hexigeduleng; Chen, Xianyang; Lv, Sulian; Jiang, Ping; Feng, Juanjuan; Fan, Pengxiang; Nie, Lingling; Li, Yinxin

    2015-03-01

    γ-Aminobutyric acid (GABA) accumulates in many plant species in response to environmental stress. However, the physiological function of GABA or its metabolic pathway (GABA shunt) in plants remains largely unclear. Here, the genes, including glutamate decarboxylases (SlGADs), GABA transaminases (SlGABA-Ts) and succinic semialdehyde dehydrogenase (SlSSADH), controlling three steps of the metabolic pathway of GABA, were studied through virus-induced gene silencing approach in tomato. Silencing of SlGADs (GABA biosynthetic genes) and SlGABA-Ts (GABA catabolic genes) led to increased accumulation of reactive oxygen species (ROS) as well as salt sensitivity under 200 mm NaCl treatment. Targeted quantitative analysis of metabolites revealed that GABA decreased and increased in the SlGADs- and SlGABA-Ts-silenced plants, respectively, whereas succinate (the final product of GABA metabolism) decreased in both silenced plants. Contrarily, SlSSADH-silenced plants, also defective in GABA degradation process, showed dwarf phenotype, curled leaves and enhanced accumulation of ROS in normal conditions, suggesting the involvement of a bypath for succinic semialdehyde catabolism to γ-hydroxybutyrate as reported previously in Arabidopsis, were less sensitive to salt stress. These results suggest that GABA shunt is involved in salt tolerance of tomato, probably by affecting the homeostasis of metabolites such as succinate and γ-hydroxybutyrate and subsequent ROS accumulation under salt stress.

  7. Enhancing Contents of γ-Aminobutyric Acid (GABA) and Other Micronutrients in Dehulled Rice during Germination under Normoxic and Hypoxic Conditions.

    PubMed

    Ding, Junzhou; Yang, Tewu; Feng, Hao; Dong, Mengyi; Slavin, Margaret; Xiong, Shanbai; Zhao, Siming

    2016-02-10

    Biofortification of staple grains with high contents of essential micronutrients is an important strategy to overcome micronutrient malnutrition. However, few attempts have targeted at γ-aminobutyric acid (GABA), a functional nutrient for aging populations. In this study, two rice cultivars, Heinuo and Xianhui 207, were used to investigate changes in GABA and other nutritional compounds of dehulled rice after germination under normoxic and hypoxic conditions. Forty-one metabolites were identified in both cultivars treated by normoxic germination, whereas the germinated dehulled rice of Heinuo and Xianhui 207 under hypoxic treatment had 43 and 41 metabolites identified, respectively. GABA increased in dehulled rice after germination, especially under hypoxia. Meanwhile, a number of other health-beneficial and/or flavor-related compounds such as lysine and d-mannose increased after the hypoxic treatment. The accumulation of GABA exhibited genotype-specific modes in both normoxic and hypoxic treatments. With regard to GABA production, Xianhui 207 was more responsive to the germination process than Heinuo, whereas Heinuo was more responsive to hypoxia than Xianhui 207. This study provides a promising approach to biofortify dehulled rice with increased GABA and other nutrients through metabolomic-based regulation.

  8. Posterior cingulate γ-aminobutyric acid and glutamate/glutamine are reduced in amnestic mild cognitive impairment and are unrelated to amyloid deposition and apolipoprotein E genotype.

    PubMed

    Riese, Florian; Gietl, Anton; Zölch, Niklaus; Henning, Anke; O'Gorman, Ruth; Kälin, Andrea M; Leh, Sandra E; Buck, Alfred; Warnock, Geoffrey; Edden, Richard A E; Luechinger, Roger; Hock, Christoph; Kollias, Spyros; Michels, Lars

    2015-01-01

    The biomarker potential of the inhibitory neurotransmitter γ-aminobutyric acid (GABA) for the in vivo characterization of preclinical stages in Alzheimer's disease has not yet been explored. We measured GABA, glutamate + glutamine (Glx), and N-acetyl-aspartate (NAA) levels by single-voxel MEGA-PRESS magnetic resonance spectroscopy in the posterior cingulate cortex of 21 elderly subjects and 15 patients with amnestic mild cognitive impairment. Participants underwent Pittsburgh Compound B positron emission tomography, apolipoprotein E (APOE) genotyping, and neuropsychological examination. GABA, Glx, and NAA levels were significantly lower in patients. NAA was lower in Pittsburgh Compound B-positive subjects and APOE ε4 allele carriers. GABA, Glx, and NAA levels were positively correlated to CERAD word learning scores. Reductions in GABA, Glx, and NAA levels may serve as metabolic biomarkers for cognitive impairment in amnestic mild cognitive impairment. Because GABA and Glx do not seem to reflect amyloid β deposition or APOE genotype, they are less likely biomarker candidates for preclinical Alzheimer's disease.

  9. Quantification of γ-aminobutyric acid in the heads of houseflies (Musca domestica) and diamondback moths (Plutella xylostella (L.)), using capillary electrophoresis with laser-induced fluorescence detection.

    PubMed

    Shi, Xueyan; Liang, Pei; Song, Dunlun; Yang, Wenling; Gao, Xiwu

    2012-02-01

    A novel method was developed for quantifying the levels of γ-aminobutyric acid (GABA) in the heads of houseflies (Musca domestica) and diamondback moths (Plutella xylostella (L.)), using capillary electrophoresis with laser-induced fluorescence detection (CE-LIF). The GABA in sample was derivatized with 4-chloro-7-nitro-2,1,3-benzoxadiazole (NBD-Cl) prior to CE-LIF analysis. In total, 32 mmol/L borate buffer, at pH 9.2 and containing 5.3 mmol/L β-cyclodextrin (β-CD) and 10.4 mmol/L sodium dodecyl sulfate (SDS), was determined to be the optimum CE background electrolyte (BGE) for GABA analysis. The detection limit of GABA was 0.016 μmol/L. The relative standard deviations (RSDs) of the migration time and peak area of GABA were 1.78 and 4.93%, respectively. The average recoveries of 0.97, 3.88, and 5.83 μmol/L of GABA, each added to the head sample of housefly, ranged from 88.9 to 110.5%. This method is simple and applicable to GABA assays of the heads of insects. With this newly developed CE-LIF method, the amounts of GABA in the heads of houseflies (M. domestica) and diamondback moths (P. xylostella (L.)) were measured. The results are relevant to the understandings of some insecticides and insecticide-resistance mechanisms in pests.

  10. Impact of Precooling and Controlled-Atmosphere Storage on γ-Aminobutyric Acid (GABA) Accumulation in Longan (Dimocarpus longan Lour.) Fruit.

    PubMed

    Zhou, Molin; Ndeurumio, Kessy H; Zhao, Lei; Hu, Zhuoyan

    2016-08-24

    Longan (Dimocarpus longan Lour.) fruit cultivars 'Chuliang' and 'Shixia' were analyzed for γ-aminobutyric acid (GABA) accumulation after precooling and in controlled-atmosphere storage. Fruit were exposed to 5% O2 plus 3%, 5%, or 10% CO2 at 4 °C, and GABA and associated enzymes, aril firmness, and pericarp color were measured. Aril softening and pericarp browning were delayed by 5% CO2 + 5% O2. GABA concentrations and glutamate decarboxylase (GAD; EC 4.1.1.15) activities declined during storage at the higher-CO2 treatments. However, GABA aminotransferase (GABA-T; EC 2.6.1.19) activities in elevated CO2-treated fruit fluctuated during storage. GABA concentrations increased after precooling treatments. GAD activity and GABA-T activity were different between cultivars after precooling. GABA concentrations in fruit increased after 3 days of 10% CO2 + 5% O2 treatment and then declined as storage time increased. GABA accumulation was associated with stimulation of GAD activity rather than inhibition of GABA-T activity. PMID:27412947

  11. Analysis of persistent changes to γ-aminobutyric acid receptor gene expression in Plutella xylostella subjected to sublethal amounts of spinosad.

    PubMed

    Yin, X-H; Wu, Q-J; Zhang, Y-J; Long, Y-H; Wu, X-M; Li, R-Y; Wang, M; Tian, X-L; Jiao, X-G

    2016-01-01

    A multi-generational approach was used to investigate the persistent effects of a sub-lethal dose of spinosad in Plutella xylostella. The susceptibility of various sub-populations of P. xylostella to spinosad and the effects of the insecticide on the gene expression of γ-aminobutyric acid receptor (GABAR) were determined. The results of a leaf dip bioassay showed that the sensitivity of P. xylostella to spinosad decreased across generations. The sub-strains had been previously selected based on a determined LC25 of spinosad. Considering that GABA-gated chloride channels are the primary targets of spinosad, the cDNA of P. xylostella was used to clone GABARα by using reverse transcription-polymerase chain reaction (RT-PCR). The mature peptide cDNA was 1477-bp long and contained a 1449-bp open reading frame encoding a protein of 483 amino acids. The resulting amino acid sequence was used to generate a neighbor-joining dendrogram, and homology search was conducted using NCBI BLAST. The protein had high similarity with the known GABAR sequence from P. xylostella. Subsequent semi-quantitative RT-PCR and real-time PCR analyses indicated that the GABAR transcript levels in the spinosad-resistant strain (RR, 145.82-fold) and in Sub1 strain (selected with LC25 spinosad for one generation) were the highest, followed by those in the spinosad-susceptible strain, the Sub10 strain (selected for ten generations), and the Sub5 strain (selected for five generations). This multi-generational study found significant correlations between spinosad susceptibility and GABAR gene expression, providing insights into the long-term effects of sub-lethal insecticide exposure and its potential to lead to the development of insecticide-resistant insect populations. PMID:27525859

  12. Library screening by means of mass spectrometry (MS) binding assays-exemplarily demonstrated for a pseudostatic library addressing γ-aminobutyric acid (GABA) transporter 1 (GAT1).

    PubMed

    Sindelar, Miriam; Wanner, Klaus T

    2012-09-01

    In the present study, the application of mass spectrometry (MS) binding assays as a tool for library screening is reported. For library generation, dynamic combinatorial chemistry (DCC) was used. These libraries can be screened by means of MS binding assays when appropriate measures are taken to render the libraries pseudostatic. That way, the efficiency of MS binding assays to determine ligand binding in compound screening with the ease of library generation by DCC is combined. The feasibility of this approach is shown for γ-aminobutyric acid (GABA) transporter 1 (GAT1) as a target, representing the most important subtype of the GABA transporters. For the screening, hydrazone libraries were employed that were generated in the presence of the target by reacting various sets of aldehydes with a hydrazine derivative that is delineated from piperidine-3-carboxylic acid (nipecotic acid), a common fragment of known GAT1 inhibitors. To ensure that the library generated is pseudostatic, a large excess of the nipecotic acid derivative is employed. As the library is generated in a buffer system suitable for binding and the target is already present, the mixtures can be directly analyzed by MS binding assays-the process of library generation and screening thus becoming simple to perform. The binding affinities of the hits identified by deconvolution were confirmed in conventional competitive MS binding assays performed with single compounds obtained by separate synthesis. In this way, two nipecotic acid derivatives exhibiting a biaryl moiety, 1-{2-[2'-(1,1'-biphenyl-2-ylmethylidene)hydrazine]ethyl}piperidine-3-carboxylic acid and 1-(2-{2'-[1-(2-thiophenylphenyl)methylidene]hydrazine}ethyl)piperidine-3-carboxylic acid, were found to be potent GAT1 ligands exhibiting pK(i) values of 6.186 ± 0.028 and 6.229 ± 0.039, respectively. This method enables screening of libraries, whether generated by conventional chemistry or DCC, and is applicable to all kinds of targets including

  13. Library screening by means of mass spectrometry (MS) binding assays-exemplarily demonstrated for a pseudostatic library addressing γ-aminobutyric acid (GABA) transporter 1 (GAT1).

    PubMed

    Sindelar, Miriam; Wanner, Klaus T

    2012-09-01

    In the present study, the application of mass spectrometry (MS) binding assays as a tool for library screening is reported. For library generation, dynamic combinatorial chemistry (DCC) was used. These libraries can be screened by means of MS binding assays when appropriate measures are taken to render the libraries pseudostatic. That way, the efficiency of MS binding assays to determine ligand binding in compound screening with the ease of library generation by DCC is combined. The feasibility of this approach is shown for γ-aminobutyric acid (GABA) transporter 1 (GAT1) as a target, representing the most important subtype of the GABA transporters. For the screening, hydrazone libraries were employed that were generated in the presence of the target by reacting various sets of aldehydes with a hydrazine derivative that is delineated from piperidine-3-carboxylic acid (nipecotic acid), a common fragment of known GAT1 inhibitors. To ensure that the library generated is pseudostatic, a large excess of the nipecotic acid derivative is employed. As the library is generated in a buffer system suitable for binding and the target is already present, the mixtures can be directly analyzed by MS binding assays-the process of library generation and screening thus becoming simple to perform. The binding affinities of the hits identified by deconvolution were confirmed in conventional competitive MS binding assays performed with single compounds obtained by separate synthesis. In this way, two nipecotic acid derivatives exhibiting a biaryl moiety, 1-{2-[2'-(1,1'-biphenyl-2-ylmethylidene)hydrazine]ethyl}piperidine-3-carboxylic acid and 1-(2-{2'-[1-(2-thiophenylphenyl)methylidene]hydrazine}ethyl)piperidine-3-carboxylic acid, were found to be potent GAT1 ligands exhibiting pK(i) values of 6.186 ± 0.028 and 6.229 ± 0.039, respectively. This method enables screening of libraries, whether generated by conventional chemistry or DCC, and is applicable to all kinds of targets including

  14. Altered γ-aminobutyric acid neurotransmission in major depressive disorder: a critical review of the supporting evidence and the influence of serotonergic antidepressants

    PubMed Central

    Pehrson, Alan L; Sanchez, Connie

    2015-01-01

    Evidence suggesting that central nervous system γ-aminobutyric acid (GABA) concentrations are reduced in patients with major depressive disorder (MDD) has been present since at least 1980, and this idea has recently gained support from more recent magnetic resonance spectroscopy data. These observations have led to the assumption that MDD’s underlying etiology is tied to an overall reduction in GABA-mediated inhibitory neurotransmission. In this paper, we review the mechanisms that govern GABA and glutamate concentrations in the brain, and provide a comprehensive and critical evaluation of the clinical data supporting reduced GABA neurotransmission in MDD. This review includes an evaluation of magnetic resonance spectroscopy data, as well as data on the expression and function of the GABA-synthesizing enzyme glutamic acid decarboxylase, GABA neuron-specific cell markers, such as parvalbumin, calretinin and calbindin, and the GABAA and GABAB receptors in clinical MDD populations. We explore a potential role for glial pathology in MDD-related reductions in GABA concentrations, and evidence of a connection between neurosteroids, GABA neurotransmission, and hormone-related mood disorders. Additionally, we investigate the effects of GABAergic pharmacological agents on mood, and demonstrate that these compounds have complex effects that do not universally support the idea that reduced GABA neurotransmission is at the root of MDD. Finally, we discuss the connections between serotonergic and GABAergic neurotransmission, and show that two serotonin-focused antidepressants – the selective serotonin-reuptake inhibitor fluoxetine and the multimodal antidepressant vortioxetine – modulate GABA neurotransmission in opposing ways, despite both being effective MDD treatments. Altogether, this review demonstrates that there are large gaps in our understanding of the relationship between GABA physiology and MDD, which must be remedied with more data from well

  15. Enhancement of γ-aminobutyric acid production in recombinant Corynebacterium glutamicum by co-expressing two glutamate decarboxylase genes from Lactobacillus brevis.

    PubMed

    Shi, Feng; Jiang, Junjun; Li, Yongfu; Li, Youxin; Xie, Yilong

    2013-11-01

    γ-Aminobutyric acid (GABA), a non-protein amino acid, is a bioactive component in the food, feed and pharmaceutical fields. To establish an effective single-step production system for GABA, a recombinant Corynebacterium glutamicum strain co-expressing two glutamate decarboxylase (GAD) genes (gadB1 and gadB2) derived from Lactobacillus brevis Lb85 was constructed. Compared with the GABA production of the gadB1 or gadB2 single-expressing strains, GABA production by the gadB1-gadB2 co-expressing strain increased more than twofold. By optimising urea supplementation, the total production of L-glutamate and GABA increased from 22.57 ± 1.24 to 30.18 ± 1.33 g L⁻¹, and GABA production increased from 4.02 ± 0.95 to 18.66 ± 2.11 g L⁻¹ after 84-h cultivation. Under optimal urea supplementation, L-glutamate continued to be consumed, GABA continued to accumulate after 36 h of fermentation, and the pH level fluctuated. GABA production increased to a maximum level of 27.13 ± 0.54 g L⁻¹ after 120-h flask cultivation and 26.32 g L⁻¹ after 60-h fed-batch fermentation. The conversion ratio of L-glutamate to GABA reached 0.60-0.74 mol mol⁻¹. By co-expressing gadB1 and gadB2 and optimising the urea addition method, C. glutamicum was genetically improved for de novo biosynthesis of GABA from its own accumulated L-glutamate.

  16. Altered γ-aminobutyric acid neurotransmission in major depressive disorder: a critical review of the supporting evidence and the influence of serotonergic antidepressants.

    PubMed

    Pehrson, Alan L; Sanchez, Connie

    2015-01-01

    Evidence suggesting that central nervous system γ-aminobutyric acid (GABA) concentrations are reduced in patients with major depressive disorder (MDD) has been present since at least 1980, and this idea has recently gained support from more recent magnetic resonance spectroscopy data. These observations have led to the assumption that MDD's underlying etiology is tied to an overall reduction in GABA-mediated inhibitory neurotransmission. In this paper, we review the mechanisms that govern GABA and glutamate concentrations in the brain, and provide a comprehensive and critical evaluation of the clinical data supporting reduced GABA neurotransmission in MDD. This review includes an evaluation of magnetic resonance spectroscopy data, as well as data on the expression and function of the GABA-synthesizing enzyme glutamic acid decarboxylase, GABA neuron-specific cell markers, such as parvalbumin, calretinin and calbindin, and the GABAA and GABAB receptors in clinical MDD populations. We explore a potential role for glial pathology in MDD-related reductions in GABA concentrations, and evidence of a connection between neurosteroids, GABA neurotransmission, and hormone-related mood disorders. Additionally, we investigate the effects of GABAergic pharmacological agents on mood, and demonstrate that these compounds have complex effects that do not universally support the idea that reduced GABA neurotransmission is at the root of MDD. Finally, we discuss the connections between serotonergic and GABAergic neurotransmission, and show that two serotonin-focused antidepressants - the selective serotonin-reuptake inhibitor fluoxetine and the multimodal antidepressant vortioxetine - modulate GABA neurotransmission in opposing ways, despite both being effective MDD treatments. Altogether, this review demonstrates that there are large gaps in our understanding of the relationship between GABA physiology and MDD, which must be remedied with more data from well-controlled empirical

  17. Aspartate-containing neurons of the brainstem and rostral spinal cord of the sea lamprey Petromyzon marinus: distribution and comparison with γ-aminobutyric acid.

    PubMed

    Villar-Cerviño, Verona; Fernández-López, Blanca; Rodicio, María Celina; Anadón, Ramón

    2014-04-15

    The amino acid L-aspartate (ASP) is one of the most abundant excitatory neurotransmitters in the mammalian brain, but its distribution in other vertebrates has not yet been well characterized. We investigated the distribution of ASP in the brainstem and rostral spinal cord of the adult sea lamprey by using ASP immunohistochemistry. Our results indicate that ASP is accumulated in specific neurons, but not in glia (tanycytes). ASP-immunoreactive neuronal populations were rather similar as the glutamatergic populations reported in the adult sea lamprey (Villar-Cerviño et al. [2013] J Comp Neurol 521:522-557), although some important differences were noted. Characteristically, the largest reticular neurons of the lamprey brainstem (Müller cells) showed ASP immunoreactivity in perikarya and processes, in contrast to the absence or faint glutamate immunoreactivity reported in these perikarya. We also compared the distribution of ASP and γ-aminobutyric acid (GABA) in brainstem neurons by using double immunofluorescence methods. In regions such as the midbrain tectum, dorsal isthmus, and motor nuclei, ASP and GABA immunoreactivity was mostly located in different neurons, whereas in other nuclei (torus semicircularis, octavolateralis area, parvocellular reticular formation), many of the ASP-immunonegative neurons displayed colocalization with GABA. These results, together with those of our previous studies of colocalization of glutamate and GABA, suggest that some lamprey neurons may co-release both excitatory and inhibitory neurotransmitters. Further investigation is needed to elucidate the pathways of uptake and release of ASP by ASP-immunoreactive neurons. Our results indicate that ASP is a neurotransmitter in the central nervous system representative of agnathans, the earliest vertebrate group.

  18. A third human retinoic acid receptor, hRAR-. gamma

    SciTech Connect

    Krust, A.; Kastner, Ph.; Petkovich, M.; Zelent, A.; Chambon, P. )

    1989-07-01

    Retinoic acid receptors (RARs) are retinoic acid (RA)-inducible enhancer factors belonging to the superfamily of steroid/thyroid nuclear receptors. The authors have previously characterized two human RAR (hRAR-{alpha} and hRAR-{beta}) cDNAs and have recently cloned their murine cognates (mRAR-{alpha} and mRAR-{beta}) together with a third RAR (mRAR-{gamma}) whose RNA was detected predominantly in skin, a well-known target for RA. mRAR-{gamma} cDNA was used here to clone its human counterpart (hRAR-{gamma}) from a T47D breast cancer cell cDNA library. Using a transient transfection assay in HeLa cells and a reporter gene harboring a synthetic RA responsive element, they demonstrate that hRAR-{gamma} cDNA indeed encodes a RA-inducible transcriptional trans-activator. Interestingly, comparisons of the amino acid sequences of all six human and mouse RARs indicate that the interspecies conservation of a given member of the RAR subfamily (either {alpha}, {beta}, or {gamma}) is much higher than the conservation of all three receptors within a given species. These observations indicate that RAR-{alpha}, -{beta}, and -{gamma} may perform specific functions. They show also that hRAR-{gamma} RNA is the predominant RAR RNA species in human skin, which suggests that hRAR-{gamma} mediates some of the retinoid effects in this tissue.

  19. Multiple Propofol-binding Sites in a γ-Aminobutyric Acid Type A Receptor (GABAAR) Identified Using a Photoreactive Propofol Analog*♦

    PubMed Central

    Jayakar, Selwyn S.; Zhou, Xiaojuan; Chiara, David C.; Dostalova, Zuzana; Savechenkov, Pavel Y.; Bruzik, Karol S.; Dailey, William P.; Miller, Keith W.; Eckenhoff, Roderic G.; Cohen, Jonathan B.

    2014-01-01

    Propofol acts as a positive allosteric modulator of γ-aminobutyric acid type A receptors (GABAARs), an interaction necessary for its anesthetic potency in vivo as a general anesthetic. Identifying the location of propofol-binding sites is necessary to understand its mechanism of GABAAR modulation. [3H]2-(3-Methyl-3H-diaziren-3-yl)ethyl 1-(phenylethyl)-1H-imidazole-5-carboxylate (azietomidate) and R-[3H]5-allyl-1-methyl-5-(m-trifluoromethyl-diazirynylphenyl)barbituric acid (mTFD-MPAB), photoreactive analogs of 2-ethyl 1-(phenylethyl)-1H-imidazole-5-carboxylate (etomidate) and mephobarbital, respectively, have identified two homologous but pharmacologically distinct classes of intersubunit-binding sites for general anesthetics in the GABAAR transmembrane domain. Here, we use a photoreactive analog of propofol (2-isopropyl-5-[3-(trifluoromethyl)-3H-diazirin-3-yl]phenol ([3H]AziPm)) to identify propofol-binding sites in heterologously expressed human α1β3 GABAARs. Propofol, AziPm, etomidate, and R-mTFD-MPAB each inhibited [3H]AziPm photoincorporation into GABAAR subunits maximally by ∼50%. When the amino acids photolabeled by [3H]AziPm were identified by protein microsequencing, we found propofol-inhibitable photolabeling of amino acids in the β3-α1 subunit interface (β3Met-286 in β3M3 and α1Met-236 in α1M1), previously photolabeled by [3H]azietomidate, and α1Ile-239, located one helical turn below α1Met-236. There was also propofol-inhibitable [3H]AziPm photolabeling of β3Met-227 in βM1, the amino acid in the α1-β3 subunit interface photolabeled by R-[3H]mTFD-MPAB. The propofol-inhibitable [3H]AziPm photolabeling in the GABAAR β3 subunit in conjunction with the concentration dependence of inhibition of that photolabeling by etomidate or R-mTFD-MPAB also establish that each anesthetic binds to the homologous site at the β3-β3 subunit interface. These results establish that AziPm as well as propofol bind to the homologous intersubunit sites in the

  20. The novel isoxazoline ectoparasiticide fluralaner: selective inhibition of arthropod γ-aminobutyric acid- and L-glutamate-gated chloride channels and insecticidal/acaricidal activity.

    PubMed

    Gassel, Michael; Wolf, Christian; Noack, Sandra; Williams, Heike; Ilg, Thomas

    2014-02-01

    Isoxazolines are a novel class of parasiticides that are potent inhibitors of γ-aminobutyric acid (GABA)-gated chloride channels (GABACls) and L-glutamate-gated chloride channels (GluCls). In this study, the effects of the isoxazoline drug fluralaner on insect and acarid GABACl (RDL) and GluCl and its parasiticidal potency were investigated. We report the identification and cDNA cloning of Rhipicephalus (R.) microplus RDL and GluCl genes, and their functional expression in Xenopus laevis oocytes. The generation of six clonal HEK293 cell lines expressing Rhipicephalus microplus RDL and GluCl, Ctenocephalides felis RDL-A285 and RDL-S285, as well as Drosophila melanogaster RDLCl-A302 and RDL-S302, combined with the development of a membrane potential fluorescence dye assay allowed the comparison of ion channel inhibition by fluralaner with that of established insecticides addressing RDL and GluCl as targets. In these assays fluralaner was several orders of magnitude more potent than picrotoxinin and dieldrin, and performed 5-236 fold better than fipronil on the arthropod RDLs, while a rat GABACl remained unaffected. Comparative studies showed that R. microplus RDL is 52-fold more sensitive than R. microplus GluCl to fluralaner inhibition, confirming that the GABA-gated chloride channel is the primary target of this new parasiticide. In agreement with the superior RDL on-target activity, fluralaner outperformed dieldrin and fipronil in insecticidal screens on cat fleas (Ctenocephalides felis), yellow fever mosquito larvae (Aedes aegypti) and sheep blowfly larvae (Lucilia cuprina), as well as in acaricidal screens on cattle tick (R. microplus) adult females, brown dog tick (Rhipicephalus sanguineus) adult females and Ornithodoros moubata nymphs. These findings highlight the potential of fluralaner as a novel ectoparasiticide. PMID:24365472

  1. The Protective Effect of γ-aminobutyric Acid on Kidney Injury Induced by Renal Ischemia-reperfusion in Ovariectomized Estradiol-treated Rats

    PubMed Central

    Talebi, Nahid; Nematbakhsh, Mehdi; Monajemi, Ramesh; Mazaheri, Safoora; Talebi, Ardeshir; Vafapour, Marzieh

    2016-01-01

    Background: Renal ischemia-reperfusion injury (IRI) is one of the most important causes of kidney injury, which is possibly gender-related. This study was designed to investigate the role of γ-aminobutyric acid (GABA) against IRI in ovariectomized estradiol-treated rats. Methods: Thirty-five ovariectomized Wistar rats were used in six experimental groups. The first three groups did not subject to estradiol treatment and assigned as sham-operated, control, and GABA-treated groups. GABA (50 μmol/kg) and saline were injected in the treated and control groups 30 min before the surgery, respectively. The second three groups received the same treatments but received estradiol valerate (500 μg/kg, intramuscularly) 3 days prior to the surgery. The IRI was induced in the control and treated groups by clamping the renal artery for 45 min and then 24 h of reperfusion. All animals were sacrificed for the measurements. Results: The serum levels of creatinine and blood urea nitrogen, kidney weight, and kidney tissue damage score significantly increased in the IRI rats (P < 0.05). GABA significantly decreased the aforementioned parameters (P < 0.05). The uterus weight increased significantly in rats that received estradiol (P < 0.05). Serum and kidney levels of nitrite (nitric oxide metabolite) did not alter significantly. Serum level of malondialdehyde increased significantly in the ovariectomized rats exposed to IRI (P < 0.05). Conclusions: It seems that GABA improved IRI in ovariectomized rats. Estradiol was also nephroprotective against IRI. However, co-administration of estradiol and GABA could not protect the kidney against IRI. PMID:26941907

  2. Spinal Nerve Ligation Decreases γ-Aminobutyric AcidB Receptors on Specific Populations of Immunohistochemically Identified Neurons in L5 Dorsal Root Ganglion of the Rat

    PubMed Central

    Engle, Mitchell P.; Merrill, Michelle A.; De Prado, Blanca Marquez; Hammond, Donna L.

    2014-01-01

    This study examined the distribution of γ-aminobutyric acid (GABA)B receptors on immunohistochemically identified neurons, and levels of GABAB(1) and GABAB(2) mRNA, in the L4 and L5 dorsal root ganglia (DRG) of the rat in the absence of injury and 2 weeks after L5 spinal nerve ligation. In uninjured DRG, GABAB(1) immunoreactivity colocalized exclusively with the neuronal marker (NeuN) and did not colocalize with the satellite cell marker S-100. The GABAB(1) subunit colocalized to >97% of DRG neurons immunoreactive (IR) for neurofilament 200 (N52) or calcitonin gene-related peptide (CGRP), or labeled by isolectin B4 (IB4). Immunoreactivity for GABAB(2) was not detectable. L5 spinal nerve ligation did not alter the number of GABAB(1)-IR neurons or its colocalization pattern in the L4 DRG. However, ligation reduced the number of GABAB(1)-IR neurons in the L5 DRG by ≈38% compared with sham-operated and naïve rats. Specifically, ligation decreased the number of CGRP-IR neurons in the L5 DRG by 75%, but did not decrease the percent colocalization of GABAB(1) in those that remained. In the few IB4-positive neurons that remained in the L5 DRG, colocalization of GABAB(1)-IR decreased to 75%. Ligation also decreased levels of GABAB(1) and GABAB(2) mRNA in the L5, but not the L4 DRG compared with sham-operated or naïve rats. These findings indicate that the GABAB receptor is positioned to presynaptically modulate afferent transmission by myelinated, unmyelinated, and peptidergic afferents in the dorsal horn. Loss of GABAB receptors on primary afferent neurons may contribute to the development of mechanical allodynia after L5 spinal nerve ligation. PMID:22120979

  3. Tomato Glutamate Decarboxylase Genes SlGAD2 and SlGAD3 Play Key Roles in Regulating γ-Aminobutyric Acid Levels in Tomato (Solanum lycopersicum).

    PubMed

    Takayama, Mariko; Koike, Satoshi; Kusano, Miyako; Matsukura, Chiaki; Saito, Kazuki; Ariizumi, Tohru; Ezura, Hiroshi

    2015-08-01

    Tomato (Solanum lycopersicum) can accumulate relatively high levels of γ-aminobutyric acid (GABA) during fruit development. However, the molecular mechanism underlying GABA accumulation and its physiological function in tomato fruits remain elusive. We previously identified three tomato genes (SlGAD1, SlGAD2 and SlGAD3) encoding glutamate decarboxylase (GAD), likely the key enzyme for GABA biosynthesis in tomato fruits. In this study, we generated transgenic tomato plants in which each SlGAD was suppressed and those in which all three SlGADs were simultaneously suppressed. A significant decrease in GABA levels, i.e. 50-81% compared with wild-type (WT) levels, was observed in mature green (MG) fruits of the SlGAD2-suppressed lines, while a more drastic reduction (up to <10% of WT levels) was observed in the SlGAD3- and triple SlGAD-suppressed lines. These findings suggest that both SlGAD2 and SlGAD3 expression are crucial for GABA biosynthesis in tomato fruits. The importance of SlGAD3 expression was also confirmed by generating transgenic tomato plants that over-expressed SlGAD3. The MG and red fruits of the over-expressing transgenic lines contained higher levels of GABA (2.7- to 5.2-fold) than those of the WT. We also determined that strong down-regulation of the SlGADs had little effect on overall plant growth, fruit development or primary fruit metabolism under normal growth conditions.

  4. Unedited in vivo detection and quantification of γ-aminobutyric acid in the occipital cortex using short-TE MRS at 3 T.

    PubMed

    Near, Jamie; Andersson, Jesper; Maron, Eduard; Mekle, Ralf; Gruetter, Rolf; Cowen, Philip; Jezzard, Peter

    2013-11-01

    Short-TE MRS has been proposed recently as a method for the in vivo detection and quantification of γ-aminobutyric acid (GABA) in the human brain at 3 T. In this study, we investigated the accuracy and reproducibility of short-TE MRS measurements of GABA at 3 T using both simulations and experiments. LCModel analysis was performed on a large number of simulated spectra with known metabolite input concentrations. Simulated spectra were generated using a range of spectral linewidths and signal-to-noise ratios to investigate the effect of varying experimental conditions, and analyses were performed using two different baseline models to investigate the effect of an inaccurate baseline model on GABA quantification. The results of these analyses indicated that, under experimental conditions corresponding to those typically observed in the occipital cortex, GABA concentration estimates are reproducible (mean reproducibility error, <20%), even when an incorrect baseline model is used. However, simulations indicate that the accuracy of GABA concentration estimates depends strongly on the experimental conditions (linewidth and signal-to-noise ratio). In addition to simulations, in vivo GABA measurements were performed using both spectral editing and short-TE MRS in the occipital cortex of 14 healthy volunteers. Short-TE MRS measurements of GABA exhibited a significant positive correlation with edited GABA measurements (R = 0.58, p < 0.05), suggesting that short-TE measurements of GABA correspond well with measurements made using spectral editing techniques. Finally, within-session reproducibility was assessed in the same 14 subjects using four consecutive short-TE GABA measurements in the occipital cortex. Across all subjects, the average coefficient of variation of these four GABA measurements was 8.7 ± 4.9%. This study demonstrates that, under some experimental conditions, short-TE MRS can be employed for the reproducible detection of GABA at 3 T, but that the technique

  5. Application of γ-aminobutyric acid demonstrates a protective role of polyamine and GABA metabolism in muskmelon seedlings under Ca(NO3)2 stress.

    PubMed

    Hu, Xiaohui; Xu, Zhiran; Xu, Weinan; Li, Jianming; Zhao, Ning; Zhou, Yue

    2015-07-01

    The effects of exogenous γ-aminobutyric acid (GABA) application on growth, polyamine and endogenous GABA metabolism in muskmelon leaves and roots were measured. Plants were treated under control or 80 mM Ca(NO3)2 stress conditions with or without foliar spraying 50 mM GABA. Ca(NO3)2 stress significantly suppressed seedling growth and GABA transaminase activity, and enhanced glutamate decarboxylase (GAD) activity and endogenous GABA levels. Polyamine (PA) biosynthesis and degradation capacity increased in parallel with increasing GAD activity. Exogenous GABA application effectively alleviated the growth inhibition caused by Ca(NO3)2 stress, and significantly enhanced the activities of arginine decarboxylase (ADC), ornithine decarboxylase (ODC), S-adenosylmethionine decarboxylase (SAMDC), polyamine oxidase (PAO), and diamine oxidase (DAO). Exogenous GABA also significantly reduced the accumulation of free putrescine (Put) and increased the levels of free spermidine (Spd) and spermine (Spm) in leaves, which improved the capacity for polyamine biosynthesis. Application of exogenous GABA under Ca(NO3)2 stress enables the plants to maintain a higher ratio of free Spd and free Spm with respect to free Put. Our data suggest that exogenous GABA has an important role in improving muskmelon seedling tolerance to Ca(NO3)2 stress by improving biosynthesis of PAs and GABA, and by preventing PA degradation. There is a potential positive feedback mechanism that results from higher endogenous GABA content and the combined effects of Ca(NO3)2 stress and exogenous GABA, which coordinately alleviate Ca(NO3)2 stress injury by enhancing PA biosynthesis and converting free Put to an insoluble bound PA form, and reduce PA degradation in muskmelon seedlings.

  6. Attenuation of γ-aminobutyric acid (GABA) transaminase activity contributes to GABA increase in the cerebral cortex of mice exposed to β-cypermethrin.

    PubMed

    Han, Y; Cao, D; Li, X; Zhang, R; Yu, F; Ren, Y; An, L

    2014-03-01

    The current study investigated the γ-aminobutyric acid (GABA) levels and GABA metabolic enzymes (GABA transaminase (GABA(T)) and glutamate decarboxylase (GAD)) activities at 2 and 4 h after treatment, using a high-performance liquid chromatography with ultraviolet detectors and colorimetric assay, in the cerebral cortex of mice treated with 20, 40 or 80 mg/kg β-cypermethrin by a single oral gavage, with corn oil as vehicle control. In addition, GABA protein (4 h after treatment), GABA(T) protein (2 h after treatment) and GABA receptors messenger RNA (mRNA) expression were detected by immunohistochemistry, Western blot and real-time quantitative reverse transcriptase polymerase chain reaction, respectively. β-Cypermethrin (80 mg/kg) significantly increased GABA levels in the cerebral cortex of mice, at both 2 and 4 h after treatment, compared with the control. Also, GABA immunohistochemistry results suggested that the number of positive granules was increased in the cerebral cortex of mice 4 h after exposure to 80 mg/kg β-cypermethrin when compared with the control. Furthermore, the results also showed that GABA(T) activity detected was significantly decreased in the cerebral cortex of mice 2 h after β-cypermethrin administration (40 or 80 mg/kg). No significant changes were found in GAD activity, or the expression of GABA(T) protein and GABAB receptors mRNA, in the cerebral cortex of mice, except that 80 mg/kg β-cypermethrin caused a significant decrease, compared with the vehicle control, in GABAA receptors mRNA expression 4 h after administration. These results suggested that attenuated GABA(T) activity induced by β-cypermethrin contributed to increased GABA levels in the mouse brain. The downregulated GABAA receptors mRNA expression is most likely a downstream event.

  7. Suppression of γ-aminobutyric acid (GABA) transaminases induces prominent GABA accumulation, dwarfism and infertility in the tomato (Solanum lycopersicum L.).

    PubMed

    Koike, Satoshi; Matsukura, Chiaki; Takayama, Mariko; Asamizu, Erika; Ezura, Hiroshi

    2013-05-01

    Tomatoes accumulate γ-aminobutyric acid (GABA) at high levels in the immature fruits. GABA is rapidly converted to succinate during fruit ripening through the activities of GABA transaminase (GABA-T) and succinate semialdehyde dehydrogenase (SSADH). Although three genes encoding GABA-T and both pyruvate- and α-ketoglutarate-dependent GABA-T activities have been detected in tomato fruits, the mechanism underlying the GABA-T-mediated conversion of GABA has not been fully understood. In this work, we conducted loss-of-function analyses utilizing RNA interference (RNAi) transgenic plants with suppressed pyruvate- and glyoxylate-dependent GABA-T gene expression to clarify which GABA-T isoforms are essential for its function. The RNAi plants with suppressed SlGABA-T gene expression, particularly SlGABA-T1, showed severe dwarfism and infertility. SlGABA-T1 expression was inversely associated with GABA levels in the fruit at the red ripe stage. The GABA contents in 35S::SlGABA-T1(RNAi) lines were 1.3-2.0 times and 6.8-9.2 times higher in mature green and red ripe fruits, respectively, than the contents in wild-type fruits. In addition, SlGABA-T1 expression was strongly suppressed in the GABA-accumulating lines. These results indicate that pyruvate- and glyoxylate-dependent GABA-T is the essential isoform for GABA metabolism in tomato plants and that GABA-T1 primarily contributes to GABA reduction in the ripening fruits.

  8. Relationship of γ-aminobutyric acid and glutamate+glutamine concentrations in the perigenual anterior cingulate cortex with performance of Cambridge Gambling Task.

    PubMed

    Fujihara, Kazuyuki; Narita, Kosuke; Suzuki, Yusuke; Takei, Yuichi; Suda, Masashi; Tagawa, Minami; Ujita, Koichi; Sakai, Yuki; Narumoto, Jin; Near, Jamie; Fukuda, Masato

    2015-04-01

    The anterior cingulate cortex (ACC), consisting of the perigenual ACC (pgACC) and mid-ACC (i.e., affective and cognitive areas, respectively), plays a significant role in the performance of gambling tasks, which are used to measure decision-making behavior under conditions of risk. Although recent neuroimaging studies have suggested that the γ-aminobutyric acid (GABA) concentration in the pgACC is associated with decision-making behavior, knowledge regarding the relationship of GABA concentrations in subdivisions of the ACC with gambling task performance is still limited. The aim of our magnetic resonance spectroscopy study is to investigate in 20 healthy males the relationship of concentrations of GABA and glutamate+glutamine (Glx) in the pgACC, mid-ACC, and occipital cortex (OC) with multiple indexes of decision-making behavior under conditions of risk, using the Cambridge Gambling Task (CGT). The GABA/creatine (Cr) ratio in the pgACC negatively correlated with delay aversion score, which corresponds to the impulsivity index. The Glx/Cr ratio in the pgACC negatively correlated with risk adjustment score, which is reported to reflect the ability to change the amount of the bet depending on the probability of winning or losing. The scores of CGT did not significantly correlate with the GABA/Cr or Glx/Cr ratio in the mid-ACC or OC. Results of this study suggest that in the pgACC, but not in the mid-ACC or OC, GABA and Glx concentrations play a distinct role in regulating impulsiveness and risk probability during decision-making behavior under conditions of risk, respectively.

  9. Functional consequences of sulfhydryl modification of the γ-aminobutyric acid transporter 1 at a single solvent-exposed cysteine residue.

    PubMed

    Omoto, Jaison J; Maestas, Matthew J; Rahnama-Vaghef, Ali; Choi, Ye E; Salto, Gerardo; Sanchez, Rachel V; Anderson, Cynthia M; Eskandari, Sepehr

    2012-12-01

    The aims of this study were to optimize the experimental conditions for labeling extracellularly oriented, solvent-exposed cysteine residues of γ-aminobutyric acid transporter 1 (GAT1) with the membrane-impermeant sulfhydryl reagent [2-(trimethylammonium)ethyl]methanethiosulfonate (MTSET) and to characterize the functional and pharmacological consequences of labeling on transporter steady-state and presteady-state kinetic properties. We expressed human GAT1 in Xenopus laevis oocytes and used radiotracer and electrophysiological methods to assay transporter function before and after sulfhydryl modification with MTSET. In the presence of NaCl, transporter exposure to MTSET (1-2.5 mM for 5-20 min) led to partial inhibition of GAT1-mediated transport, and this loss of function was completely reversed by the reducing reagent dithiothreitol. MTSET treatment had no functional effect on the mutant GAT1 C74A, whereas the membrane-permeant reagents N-ethylmaleimide and tetramethylrhodamine-6-maleimide inhibited GABA transport mediated by GAT1 C74A. Ion replacement experiments indicated that MTSET labeling of GAT1 could be driven to completion when valproate replaced chloride in the labeling buffer, suggesting that valproate induces a GAT1 conformation that significantly increases C74 accessibility to the extracellular fluid. Following partial inhibition by MTSET, there was a proportional reduction in both the presteady-state and steady-state macroscopic signals, and the functional and pharmacological properties of the remaining signals were indistinguishable from those of unlabeled GAT1. Therefore, covalent modification of GAT1 at C74 results in completely nonfunctional as well as electrically silent transporters.

  10. Positive regulation by γ-aminobutyric acid B receptor subunit-1 of chondrogenesis through acceleration of nuclear translocation of activating transcription factor-4.

    PubMed

    Takahata, Yoshifumi; Hinoi, Eiichi; Takarada, Takeshi; Nakamura, Yukari; Ogawa, Shinya; Yoneda, Yukio

    2012-09-28

    A view that signaling machineries for the neurotransmitter γ-aminobutyric acid (GABA) are functionally expressed by cells outside the central nervous system is now prevailing. In this study, we attempted to demonstrate functional expression of GABAergic signaling molecules by chondrocytes. In cultured murine costal chondrocytes, mRNA was constitutively expressed for metabotropic GABA(B) receptor subunit-1 (GABA(B)R1), but not for GABA(B)R2. Immunohistochemical analysis revealed the predominant expression of GABA(B)R1 by prehypertrophic to hypertrophic chondrocytes in tibial sections of newborn mice. The GABA(B)R agonist baclofen failed to significantly affect chondrocytic differentiation determined by Alcian blue staining and alkaline phosphatase activity in cultured chondrocytes, whereas newborn mice knocked out of GABA(B)R1 (KO) showed a decreased body size and delayed calcification in hyoid bone and forelimb and hindlimb digits. Delayed calcification was also seen in cultured metatarsals from KO mice with a marked reduction of Indian hedgehog gene (Ihh) expression. Introduction of GABA(B)R1 led to synergistic promotion of the transcriptional activity of activating transcription factor-4 (ATF4) essential for normal chondrogenesis, in addition to facilitating ATF4-dependent Ihh promoter activation. Although immunoreactive ATF4 was negligibly detected in the nucleus of chondrocytes from KO mice, ATF4 expression was again seen in the nucleus and cytoplasm after the retroviral introduction of GABA(B)R1 into cultured chondrocytes from KO mice. In nuclear extracts of KO chondrocytes, a marked decrease was seen in ATF4 DNA binding. These results suggest that GABA(B)R1 positively regulates chondrogenesis through a mechanism relevant to the acceleration of nuclear translocation of ATF4 for Ihh expression in chondrocytes. PMID:22879594

  11. Neurotoxins from Snake Venoms and α-Conotoxin ImI Inhibit Functionally Active Ionotropic γ-Aminobutyric Acid (GABA) Receptors*

    PubMed Central

    Kudryavtsev, Denis S.; Shelukhina, Irina V.; Son, Lina V.; Ojomoko, Lucy O.; Kryukova, Elena V.; Lyukmanova, Ekaterina N.; Zhmak, Maxim N.; Dolgikh, Dmitry A.; Ivanov, Igor A.; Kasheverov, Igor E.; Starkov, Vladislav G.; Ramerstorfer, Joachim; Sieghart, Werner; Tsetlin, Victor I.; Utkin, Yuri N.

    2015-01-01

    Ionotropic receptors of γ-aminobutyric acid (GABAAR) regulate neuronal inhibition and are targeted by benzodiazepines and general anesthetics. We show that a fluorescent derivative of α-cobratoxin (α-Ctx), belonging to the family of three-finger toxins from snake venoms, specifically stained the α1β3γ2 receptor; and at 10 μm α-Ctx completely blocked GABA-induced currents in this receptor expressed in Xenopus oocytes (IC50 = 236 nm) and less potently inhibited α1β2γ2 ≈ α2β2γ2 > α5β2γ2 > α2β3γ2 and α1β3δ GABAARs. The α1β3γ2 receptor was also inhibited by some other three-finger toxins, long α-neurotoxin Ls III and nonconventional toxin WTX. α-Conotoxin ImI displayed inhibitory activity as well. Electrophysiology experiments showed mixed competitive and noncompetitive α-Ctx action. Fluorescent α-Ctx, however, could be displaced by muscimol indicating that most of the α-Ctx-binding sites overlap with the orthosteric sites at the β/α subunit interface. Modeling and molecular dynamic studies indicated that α-Ctx or α-bungarotoxin seem to interact with GABAAR in a way similar to their interaction with the acetylcholine-binding protein or the ligand-binding domain of nicotinic receptors. This was supported by mutagenesis studies and experiments with α-conotoxin ImI and a chimeric Naja oxiana α-neurotoxin indicating that the major role in α-Ctx binding to GABAAR is played by the tip of its central loop II accommodating under loop C of the receptors. PMID:26221036

  12. Quantification of induced resistance against Phytophthora species expressing GFP as a vital marker: beta-aminobutyric acid but not BTH protects potato and Arabidopsis from infection.

    PubMed

    Si-Ammour, Azeddine; Mauch-Mani, Brigitte; Mauch, Felix

    2003-07-01

    SUMMARY Induced resistance was studied in the model pathosystem Arabidopsis-Phytophthora brassicae (formerly P. porri) in comparison with the agronomically important late blight disease of potato caused by Phytophthora infestans. For the quantification of disease progress, both Phytophthora species were transformed with the vector p34GFN carrying the selectable marker gene neomycine phosphotransferase (nptII) and the reporter gene green fluorescent protein (gfp). Eighty five per cent of the transformants of P. brassicae and P. infestans constitutively expressed GFP at high levels at all developmental stages both in vitro and in planta. Transformants with high GFP expression and normal in vitro growth and virulence were selected to quantify pathogen growth by measuring the in planta emitted GFP fluorescence. This non-destructive monitoring of the infection process was applied to analyse the efficacy of two chemical inducers of disease resistance, a functional SA-analogue, benzothiadiazole (BTH), and beta-aminobutyric acid (BABA) which is involved in priming mechanisms of unknown nature. BABA pre-treatment (300 microm) via soil drench applied 24 h before inoculation completely protected the susceptible Arabidopsis accession Landsberg erecta (Ler) from infection with P. brassicae. A similar treatment with BTH (330 microm) did not induce resistance. Spraying the susceptible potato cultivar Bintje with BABA (1 mm) 2 days before inoculation resulted in a phenocopy of the incompatible interaction shown by the resistant potato cultivar Matilda while BTH (1.5 mm) did not protect Bintje from severe infection. Thus, in both pathosystems, the mechanisms of induced resistance appeared to be similar, suggesting that the Arabidopsis-P. brassicae pathosystem is a promising model for the molecular analysis of induced resistance mechanisms of potato against the late blight disease.

  13. Investigation of Cortical Glutamate–Glutamine and γ-Aminobutyric Acid in Obsessive–Compulsive Disorder by Proton Magnetic Resonance Spectroscopy

    PubMed Central

    Simpson, Helen B; Shungu, Dikoma C; Bender, James; Mao, Xiangling; Xu, Xiaoyan; Slifstein, Mark; Kegeles, Lawrence S

    2012-01-01

    Glutamatergic abnormalities in corticostriatal brain circuits are thought to underlie obsessive–compulsive disorder (OCD). Whether these abnormalities exist in adults with OCD is not clear. We used proton magnetic resonance spectroscopy (1H MRS) to test our hypothesis that unmedicated adults with OCD have reduced glutamate plus glutamine (Glx) levels in the medial prefrontal cortex (MPFC) compared with healthy controls. Levels of γ-aminobutyric acid (GABA) were also explored. Twenty-four unmedicated adults with OCD and 22 matched healthy control subjects underwent 1H MRS scans at 3.0 T. Resonances of both Glx and GABA were obtained using the standard J-editing technique and assessed as ratios relative to voxel tissue water (W) in the MPFC (the region of interest) and the dorsolateral prefrontal cortex (DLPFC) to explore the regional specificity of any finding. In the MPFC, Glx/W did not differ by diagnostic group (p=0.98) or sex (p=0.57). However, GABA/W was decreased in OCD (2.16±0.46 × 10−3) compared with healthy controls (2.43±0.45 × 10−3, p=0.045); moreover, age of OCD onset was inversely correlated with MPFC GABA/W (r=−0.50, p=0.015). MPFC GABA/W was higher in females than in males. In the DLPFC, there were no main effects of diagnosis or gender on Glx/W or GABA/W. These data indicate that unmedicated adults with OCD do not have Glx abnormalities in a MPFC voxel that includes the pregenual anterior cingulate cortex. However, they may have decreased MPFC GABA levels. How GABA abnormalities might contribute to corticostriatal dysfunction in OCD deserves further study. PMID:22850733

  14. The novel isoxazoline ectoparasiticide fluralaner: selective inhibition of arthropod γ-aminobutyric acid- and L-glutamate-gated chloride channels and insecticidal/acaricidal activity.

    PubMed

    Gassel, Michael; Wolf, Christian; Noack, Sandra; Williams, Heike; Ilg, Thomas

    2014-02-01

    Isoxazolines are a novel class of parasiticides that are potent inhibitors of γ-aminobutyric acid (GABA)-gated chloride channels (GABACls) and L-glutamate-gated chloride channels (GluCls). In this study, the effects of the isoxazoline drug fluralaner on insect and acarid GABACl (RDL) and GluCl and its parasiticidal potency were investigated. We report the identification and cDNA cloning of Rhipicephalus (R.) microplus RDL and GluCl genes, and their functional expression in Xenopus laevis oocytes. The generation of six clonal HEK293 cell lines expressing Rhipicephalus microplus RDL and GluCl, Ctenocephalides felis RDL-A285 and RDL-S285, as well as Drosophila melanogaster RDLCl-A302 and RDL-S302, combined with the development of a membrane potential fluorescence dye assay allowed the comparison of ion channel inhibition by fluralaner with that of established insecticides addressing RDL and GluCl as targets. In these assays fluralaner was several orders of magnitude more potent than picrotoxinin and dieldrin, and performed 5-236 fold better than fipronil on the arthropod RDLs, while a rat GABACl remained unaffected. Comparative studies showed that R. microplus RDL is 52-fold more sensitive than R. microplus GluCl to fluralaner inhibition, confirming that the GABA-gated chloride channel is the primary target of this new parasiticide. In agreement with the superior RDL on-target activity, fluralaner outperformed dieldrin and fipronil in insecticidal screens on cat fleas (Ctenocephalides felis), yellow fever mosquito larvae (Aedes aegypti) and sheep blowfly larvae (Lucilia cuprina), as well as in acaricidal screens on cattle tick (R. microplus) adult females, brown dog tick (Rhipicephalus sanguineus) adult females and Ornithodoros moubata nymphs. These findings highlight the potential of fluralaner as a novel ectoparasiticide.

  15. Mutations in γ-aminobutyric acid (GABA) transaminase genes in plants or Pseudomonas syringae reduce bacterial virulence.

    PubMed

    Park, Duck Hwan; Mirabella, Rossana; Bronstein, Philip A; Preston, Gail M; Haring, Michel A; Lim, Chun Keun; Collmer, Alan; Schuurink, Robert C

    2010-10-01

    Pseudomonas syringae pv. tomato DC3000 is a bacterial pathogen of Arabidopsis and tomato that grows in the apoplast. The non-protein amino acid γ-amino butyric acid (GABA) is produced by Arabidopsis and tomato and is the most abundant amino acid in the apoplastic fluid of tomato. The DC3000 genome harbors three genes annotated as gabT GABA transaminases. A DC3000 mutant lacking all three gabT genes was constructed and found to be unable to utilize GABA as a sole carbon and nitrogen source. In complete minimal media supplemented with GABA, the mutant grew less well than wild-type DC3000 and showed strongly reduced expression of hrpL and avrPto, which encode an alternative sigma factor and effector, respectively, associated with the type III secretion system. The growth of the gabT triple mutant was weakly reduced in Arabidopsis ecotype Landberg erecta (Ler) and strongly reduced in the Ler pop2-1 GABA transaminase-deficient mutant that accumulates higher levels of GABA. Much of the ability to grow on GABA-amended minimal media or in Arabidopsis pop2-1 leaves could be restored to the gabT triple mutant by expression in trans of just gabT2. The ability of DC3000 to elicit the hypersensitive response (HR) in tobacco leaves is dependent upon deployment of the type III secretion system, and the gabT triple mutant was less able than wild-type DC3000 to elicit this HR when bacteria were infiltrated along with GABA at levels of 1 mm or more. GABA may have multiple effects on P. syringae-plant interactions, with elevated levels increasing disease resistance.

  16. Activation of γ-aminobutyric Acid (A) Receptor Protects Hippocampus from Intense Exercise-induced Synapses Damage and Apoptosis in Rats

    PubMed Central

    Ding, Yi; Xie, Lan; Chang, Cun-Qing; Chen, Zhi-Min; Ai, Hua

    2015-01-01

    Background: Our previous study has confirmed that one bout of exhaustion (Ex) can cause hippocampus neurocyte damage, excessive apoptosis, and dysfunction. Its initial reason is intracellular calcium overload in hippocampus triggered by N-methyl-D-aspartic acid receptor (NMDAR) over-activation. NMDAR activation can be suppressed by γ-aminobutyric acid (A) receptor (GABAAR). Whether GABAAR can prevent intense exercise-induced hippocampus apoptosis, damage, or dysfunction will be studied in this study. Methods: According to dose test, rats were randomly divided into control (Con), Ex, muscimol (MUS, 0.1 mg/kg) and bicuculline (BIC, 0.5 mg/kg) groups, then all rats underwent once swimming Ex except ones in Con group only underwent training. Intracellular free calcium concentration ([Ca2+]i) was measured by Fura-2-acetoxymethyl ester; glial fibrillary acidic protein (GFAP) and synaptophysin (SYP) immunofluorescence were also performed; apoptosis were displayed by dUTP nick end labeling (TUNEL) stain; endoplasmic reticulum stress-induced apoptosis pathway was detected by Western blotting analysis; Morris water maze was used to detect learning ability and spatial memory. Results: The appropriate dose was 0.1 mg/kg for MUS and 0.5 mg/kg for BIC. Ex group showed significantly increased [Ca2+]i and astrogliosis; TUNEL positive cells and levels of GFAP, B cell lymphoma-2 (Bcl-2) associated X protein (Bax), caspase-3, caspase-12 cleavage, CCAAT/enhancer binding protein homologous protein (CHOP), and p-Jun amino-terminal kinase (p-JNK) in Ex group also raised significantly compared to Con group, while SYP, synapse plasticity, and Bcl-2 levels in Ex group were significantly lower than those in Con group. These indexes were back to normal in MUS group. BIC group had the highest levels of [Ca2+]i, astrogliosis, TUNEL positive cell, GFAP, Bax, caspase-3, caspase-12 cleavage, CHOP, and p-JNK, it also gained the lowest SYP, synapse plasticity, and Bcl-2 levels among all groups

  17. Electrospray ionization tandem mass spectrometric study on the effect of N-terminal beta- and gamma-carbo amino acids on fragmentation of GABA-hybrid peptides.

    PubMed

    Ramesh, V; Ramesh, M; Srinivas, R; Sharma, G V M; Jayaprakash, P

    2008-11-01

    The fragmentations of protonated and deprotonated ions of a new class of N-blocked hybrid Boc-carbopeptides containing repeats of gamma-Caa/gammaAbu- and beta-Caa/gammaAbu- (Caa==C-linked carbo gamma(4)-/beta(3)- amino acids derived from D-xylose, gammaAbu = gamma-aminobutyric acid) have been studied using electrospray ionization (ESI) ion-trap tandem mass spectrometry (MS/MS). MS/MS of a pair of these protonated diastereomers produces distinct fragmentation of the Boc group. The formation of [M + H-56](+) corresponding to loss of isobutylene is more pronounced for Boc-NH-(R)-gamma-Caa-gammaAbu-OH (2) whereas it is of low abundance for Boc-NH-(S)-gamma-Caa-gammaAbu--OH (1). Similarly, MS(2) of [M--H](-) of 2 produces an abundant [M--H--C(CH(3))(3)OH--CO(2)](-) ion, which is absent for its diastereomeric isomer 1. From this, it can be suggested that MS/MS of N-blocked Boc-protected carbopeptides may be helpful in distinguishing the stereochemistry of the N-terminus Caa. MS(3) of [M + H-Boc + H](+) ions of peptides with a gamma-amino acid (gamma-Caa/gammaAbu) at the N-terminus produces only abundant y(n) (+) ions. On the other hand, characteristic fragmentations involving the peptide backbone (b(n) (+) and y(n) (+)) and the side chain are seen when beta-Caa is at the N-terminus of the peptides. MS(3) of the [M--H--C(CH(3))(3)OH](-) ion of peptides containing gamma-Caa/gammaAbu at the N-terminus gave y(n) (-) and [M--H--C(CH(3))(3)OH--CO(2)](-) ions, whereas the presence of beta-Caa at the N-terminus yielded predominantly [M--H--C(CH(3))(3)OH--HNCO](-). Thus, on the basis of our previous study and that presented here we propose that the fragmentation of these hybrid carbopeptides is highly influenced by the type of carbo amino acid present at the N-terminus. PMID:18837002

  18. Oral administration of γ-aminobutyric acid affects heat production in a hot environment in resting humans

    PubMed Central

    2012-01-01

    Background Central administration of γ-amino butyric acid (GABA) induces lower body temperature in animals in hot ambient air. However, it is still unknown whether oral GABA administration affects temperature regulation at rest in a hot environment in humans. Therefore, in the present study, we specifically hypothesized that systemic administration of GABA in humans would induce hypothermia in a hot environment and that this response would be observed in association with decreased heat production. Methods Eight male participants drank a 200-ml sports drink with 1 g of GABA (trial G) or without GABA (trial C), then rested for 30 minutes in a sitting position in a hot environment (ambient air temperature 33°C, relative humidity 50%). Results We found that changes in esophageal temperature from before drinking the sports drink were lower in trial G than in trial C (-0.046 ± 0.079°C vs 0.001 ± 0.063°C; P < 0.05), with lower heat production calculated by oxygen consumption (41 ± 5 W/m2 vs 47 ± 8 W/m2; P < 0.05). Conclusions In this study, we have demonstrated that a single oral administration of GABA induced a larger decrease in body core temperature compared to a control condition during rest in a hot environment and that this response was concomitant with a decrease in total heat production. PMID:22738209

  19. A phase transition from monoclinic C2 with Z' = 1 to triclinic P1 with Z' = 4 for the quasiracemate L-2-aminobutyric acid-D-methionine (1/1).

    PubMed

    Görbitz, Carl Henrik; Wragg, David S; Bakke, Ingrid Marie Bergh; Fleischer, Christian; Grønnevik, Gaute; Mykland, Maria; Park, Yoomin; Trovik, Kristian Wiedicke; Serigstad, Halvard; Sundsli, Bård Edgar Vestheim

    2016-07-01

    Racemates of hydrophobic amino acids with linear side chains are known to undergo a unique series of solid-state phase transitions that involve sliding of molecular bilayers upon heating or cooling. Recently, this behaviour was shown to extend also to quasiracemates of two different amino acids with opposite handedness [Görbitz & Karen (2015). J. Phys. Chem. B, 119, 4975-4984]. Previous investigations are here extended to an L-2-aminobutyric acid-D-methionine (1/1) co-crystal, C4H9NO2·C5H11NO2S. The significant difference in size between the -CH2CH3 and -CH2CH2SCH3 side chains leads to extensive disorder at room temperature, which is essentially resolved after a phase transition at 229 K to an unprecedented triclinic form where all four D-methionine molecules in the asymmetric unit have different side-chain conformations and all three side-chain rotamers are used for the four partner L-2-aminobutyric acid molecules. PMID:27377274

  20. Comparative proteomic analysis of gamma-aminobutyric acid responses in hypoxia-treated and untreated melon roots.

    PubMed

    Fan, Longquan; Wu, Xiaolei; Tian, Zhen; Jia, Kaizhi; Pan, Yinghong; Li, Jingrui; Gao, Hongbo

    2015-08-01

    Hypoxia is one of the main environmental stresses that accounts for decreasing crop yield. To further investigate the mechanisms whereby exogenous GABA alleviates hypoxia injury to melon seedlings, a comparative proteomic analysis was performed using roots subjected to normal aeration and hypoxia conditions with or without GABA (5mM). The results indicated that protein spots on gels after hypoxia and hypoxia+GABA treatment were significantly changed. Three "matched sets" were analyzed from four treatments, and 13 protein spots with large significant differences in expression were identified by MALDI-TOF/TOF mass spectrometry. Exogenous GABA treatment enhanced the expression of protein in cytosolic phosphoglycerate kinase 1, exaA2 gene product, dnaJ and myb-like DNA-binding domain-containing proteins, as well as elongation factor-1 alpha and hypothetical proteins in hypoxia-induced roots. However, the hypoxia+GABA treated roots had a significantly lower expression of proteins including malate dehydrogenase, nucleoside diphosphate kinase, disease resistance-like protein, disulfide isomerase, actin, ferrodoxin NADP oxidoreductase, glutathione transferase, netting associated peroxidase. This paper describes the effect of GABA on melon plants under hypoxia-induced stress using proteomics, and supports the alleviating function of GABA in melon plants grown under hypoxic conditions.

  1. Neuronal gamma-aminobutyric acid (GABA) type A receptors undergo cognate ligand chaperoning in the endoplasmic reticulum by endogenous GABA

    PubMed Central

    Wang, Ping; Eshaq, Randa S.; Meshul, Charles K.; Moore, Cynthia; Hood, Rebecca L.; Leidenheimer, Nancy J.

    2015-01-01

    GABAA receptors mediate fast inhibitory neurotransmission in the brain. Dysfunction of these receptors is associated with various psychiatric/neurological disorders and drugs targeting this receptor are widely used therapeutic agents. Both the efficacy and plasticity of GABAA receptor-mediated neurotransmission depends on the number of surface GABAA receptors. An understudied aspect of receptor cell surface expression is the post-translational regulation of receptor biogenesis within the endoplasmic reticulum (ER). We have previously shown that exogenous GABA can act as a ligand chaperone of recombinant GABAA receptors in the early secretory pathway leading us to now investigate whether endogenous GABA facilitates the biogenesis of GABAA receptors in primary cerebral cortical cultures. In immunofluorescence labeling experiments, we have determined that neurons expressing surface GABAA receptors contain both GABA and its degradative enzyme GABA transaminase (GABA-T). Treatment of neurons with GABA-T inhibitors, a treatment known to increase intracellular GABA levels, decreases the interaction of the receptor with the ER quality control protein calnexin, concomittantly increasing receptor forward-trafficking and plasma membrane insertion. The effect of GABA-T inhibition on the receptor/calnexin interaction is not due to the activation of surface GABAA or GABAB receptors. Consistent with our hypothesis that GABA acts as a cognate ligand chaperone in the ER, immunogold-labeling of rodent brain slices reveals the presence of GABA within the rough ER. The density of this labeling is similar to that present in mitochondria, the organelle in which GABA is degraded. Lastly, the effect of GABA-T inhibition on the receptor/calnexin interaction was prevented by pretreatment with a GABA transporter inhibitor. Together, these data indicate that endogenous GABA acts in the rough ER as a cognate ligand chaperone to facilitate the biogenesis of neuronal GABAA receptors. PMID:26041994

  2. Transmembrane topology, subcellular distribution and turnover of the gamma-aminobutyric acid/benzodizaepine receptor in chick brain cell cultures

    SciTech Connect

    Czajkowski, C.M.

    1987-01-01

    Experiments were performed utilizing trypsinization of the GABA/BZD-R in intact cells to determine (1) the subcellular distribution of membrane-associated GABA/BZD-Rs and (2) aspects of the transmembrane topology of the BZD-R. Additionally, R07-0213, a positively charged benzodiazepine, was used to distinguish between cell surface and intracellular BZD-Rs. Following trypsin treatment of intact cells a cleaved receptor fragment of M{sub r} = 24,000 (xRF24) is generated. It remains anchored in the plasma membrane and not only retains the ability to bind ({sup 3}H)flunitrazepan reversibly and irreversibly but also retains the ability to be modulated by GABA. xRF24 is not observed following trypsinization of saponin-treated cells or cell homogenates, indicating that it has a cytoplasmic domain as well as a cell surface domain, as expected for a transmembrane fragment of the BZD-R. By utilizing ({sup 3}H)flunitrazepam as an irreversible photoaffinity label, BZD-R turnover was also investigated.

  3. Effect of Functional Bread Rich in Potassium, γ-Aminobutyric Acid and Angiotensin-Converting Enzyme Inhibitors on Blood Pressure, Glucose Metabolism and Endothelial Function

    PubMed Central

    Becerra-Tomás, Nerea; Guasch-Ferré, Marta; Quilez, Joan; Merino, Jordi; Ferré, Raimon; Díaz-López, Andrés; Bulló, Mònica; Hernández-Alonso, Pablo; Palau-Galindo, Antoni; Salas-Salvadó, Jordi

    2015-01-01

    Abstract Because it has been suggested that food rich in γ-aminobutyric acid (GABA) or angiotensin-converting enzyme inhibitor (ACEI) peptides have beneficial effects on blood pressure (BP) and other cardiovascular risk factors, we tested the effects of low-sodium bread, but rich in potassium, GABA, and ACEI peptides on 24-hour BP, glucose metabolism, and endothelial function. A randomized, double-blind, crossover trial was conducted in 30 patients with pre or mild-to-moderate hypertension, comparing three 4-week nutritional interventions separated by 2-week washout periods. Patients were randomly assigned to consume 120 g/day of 1 of the 3 types of bread for each nutritional intervention: conventional wheat bread (CB), low-sodium wheat bread enriched in potassium (LSB), and low-sodium wheat bread rich in potassium, GABA, and ACEI peptides (LSB + G). For each period, 24-hour BP measurements, in vivo endothelial function, and biochemical samples were obtained. After LSB + G consumption, 24-hour ambulatory BP underwent a nonsignificant greater reduction than after the consumption of CB and LSB (0.26 mm Hg in systolic BP and −0.63 mm Hg in diastolic BP for CB; −0.71 mm Hg in systolic BP and −1.08 mm Hg in diastolic BP for LSB; and −0.75 mm Hg in systolic BP and −2.12 mm Hg in diastolic BP for LSB + G, respectively). Diastolic BP at rest decreased significantly during the LSB + G intervention, although there were no significant differences in changes between interventions. There were no significant differences between interventions in terms of changes in in vivo endothelial function, glucose metabolism, and peripheral inflammatory parameters. Compared with the consumption of CB or LSB, no greater beneficial effects on 24-hour BP, endothelial function, or glucose metabolism were demonstrated after the consumption of LSB + G in a population with pre or mild-to-moderate hypertension. Further studies are warranted to clarify the

  4. Dietary supplementation with glutamine and γ-aminobutyric acid improves growth performance and serum parameters in 22- to 35-day-old broilers exposed to hot environment.

    PubMed

    Hu, H; Bai, X; Shah, A A; Wen, A Y; Hua, J L; Che, C Y; He, S J; Jiang, J P; Cai, Z H; Dai, S F

    2016-04-01

    This study was designed using 360 21-day-old chicks to determine the influences of diet supplementation with glutamine (5 g/kg), γ-aminobutyric acid (GABA, 100 mg/kg) or their combinations on performance and serum parameters exposed to cycling high temperatures. From 22 to 35 days, the experimental groups (2 × 2) were subjected to circular heat stress by exposing them to 30-34 °C cycling, while the positive control group was exposed to 23 °C constant. The blood of broilers was collected to detect serum parameters on days 28 and 35. Compared with the positive control group, the cycling high temperature decreased (p < 0.05) the feed consumption, weight gain and serum total protein (TP), glucose, thyroxine (T4), insulin, alkaline phosphatase (ALP), glutamine, GABA and glutamate levels, while increased (p < 0.05) the serum triglyceride (TG), corticosterone (CS), glucagon (GN), creatine kinase (CK), glutamic oxaloacetic transaminase (GOT), nitric oxide synthase (NOS), glutamate pyruvate transaminase (GPT) and lactate dehydrogenase (LDH) levels during 22-35 days. However, dietary glutamine (5 g/kg) increased (p < 0.05) the feed consumption, weight gain and serum levels of glutamine, TP, insulin and ALP, but decreased (p < 0.05) the serum TG, CK, GOT, NOS and GPT levels. Diet supplemented with GABA also increased (p < 0.05) weight gain and the serum levels of TP, T4, ALP, GABA and glutamine. In addition, the significant interactions (p < 0.05) between glutamine and GABA were found in the feed consumption, weight gain and the serum ALP, CK, LDH, GABA, T3 and T4 levels of heat-stressed chickens. This research indicated that dietary glutamine and GABA improved the antistress ability in performance and serum parameters of broilers under hot environment. PMID:25980810

  5. Effect of γ-Aminobutyric Acid-producing Lactobacillus Strain on Laying Performance, Egg Quality and Serum Enzyme Activity in Hy-Line Brown Hens under Heat Stress

    PubMed Central

    Zhu, Y. Z.; Cheng, J. L.; Ren, M.; Yin, L.; Piao, X. S.

    2015-01-01

    Heat-stress remains a costly issue for animal production, especially for poultry as they lack sweat glands, and alleviating heat-stress is necessary for ensuring animal production in hot environment. A high γ-aminobutyric acid (GABA)-producer Lactobacillus strain was used to investigate the effect of dietary GABA-producer on laying performance and egg quality in heat-stressed Hy-line brown hens. Hy-Line brown hens (n = 1,164) at 280 days of age were randomly divided into 4 groups based on the amount of freeze-dried GABA-producer added to the basal diet as follows: i) 0 mg/kg, ii) 25 mg/kg, iii) 50 mg/kg, and iv) 100 mg/kg. All hens were subjected to heat-stress treatment through maintaining the temperature and the relative humidity at 28.83±3.85°C and 37% to 53.9%, respectively. During the experiment, laying rate, egg weight and feed intake of hens were recorded daily. At the 30th and 60th day after the start of the experiment, biochemical parameters, enzyme activity and immune activity in serum were measured. Egg production, average egg weight, average daily feed intake, feed conversion ratio and percentage of speckled egg, soft shell egg and misshaped egg were significantly improved (p<0.05) by the increasing supplementation of the dietary GABA-producer. Shape index, eggshell thickness, strength and weight were increased linearly with increasing GABA-producer supplementation. The level of calcium, phosphorus, glucose, total protein and albumin in serum of the hens fed GABA-producing strain supplemented diet was significantly higher (p<0.05) than that of the hens fed the basal diet, whereas cholesterol level was decreased. Compared with the basal diet, GABA-producer strain supplementation increased serum level of glutathione peroxidase (p = 0.009) and superoxide dismutase. In conclusion, GABA-producer played an important role in alleviating heat-stress, the isolated GABA-producer strain might be a potential natural and safe probiotic to use to improve laying

  6. γ-Aminobutyric acid type B (GABAB) receptor expression is needed for inhibition of N-type (Cav2.2) calcium channels by analgesic α-conotoxins.

    PubMed

    Cuny, Hartmut; de Faoite, Andrew; Huynh, Thuan G; Yasuda, Takahiro; Berecki, Géza; Adams, David J

    2012-07-01

    α-Conotoxins Vc1.1 and RgIA are small peptides isolated from the venom of marine cone snails. They have effective anti-nociceptive actions in rat models of neuropathic pain. Pharmacological studies in rodent dorsal root ganglion (DRG) show their analgesic effect is mediated by inhibition of N-type (Ca(v)2.2) calcium channels via a pathway involving γ-aminobutyric acid type B (GABA(B)) receptor. However, there is no direct demonstration that functional GABA(B) receptors are needed for inhibition of the Ca(v)2.2 channel by analgesic α-conotoxins. This study examined the effect of the GABA(B) agonist baclofen and α-conotoxins Vc1.1 and RgIA on calcium channel currents after transient knockdown of the GABA(B) receptor using RNA interference. Isolated rat DRG neurons were transfected with small interfering RNAs (siRNA) targeting GABA(B) subunits R1 and R2. Efficient knockdown of GABA(B) receptor expression at mRNA and protein levels was confirmed by quantitative real time PCR (qRT-PCR) and immunocytochemical analysis, respectively. Whole-cell patch clamp recordings conducted 2-4 days after transfection showed that inhibition of N-type calcium channels in response to baclofen, Vc1.1 and RgIA was significantly reduced in GABA(B) receptor knockdown DRG neurons. In contrast, neurons transfected with a scrambled nontargeting siRNA were indistinguishable from untransfected neurons. In the HEK 293 cell heterologous expression system, Vc1.1 and RgIA inhibition of Ca(v)2.2 channels needed functional expression of both human GABA(B) receptor subunits. Together, these results confirm that GABA(B) receptors must be activated for the modulation of N-type (Ca(v)2.2) calcium channels by analgesic α-conotoxins Vc1.1 and RgIA.

  7. Frontal Glutamate and γ-Aminobutyric Acid Levels and Their Associations With Mismatch Negativity and Digit Sequencing Task Performance in Schizophrenia

    PubMed Central

    Rowland, Laura M.; Summerfelt, Ann; Wijtenburg, S. Andrea; Du, Xiaoming; Chiappelli, Joshua J.; Krishna, Nithin; West, Jeffrey; Muellerklein, Florian; Kochunov, Peter; Hong, L. Elliot

    2016-01-01

    IMPORTANCE Auditory mismatch negativity (MMN) is a biomarker for schizophrenia thought to reflect glutamatergic N-methyl-d-aspartate receptor function and excitatory-inhibitory neurotransmission balance. However, the association of glutamate level with MMN has not been directly examined in patients with schizophrenia, to our knowledge. OBJECTIVE To investigate the contributions of glutamate and γ-aminobutyric acid (GABA) to MMN and digit sequencing task (DST) performance, an assessment of verbal working memory, in schizophrenia. DESIGN, SETTING, AND PARTICIPANTS Fifty-three control participants from the community and 45 persons with schizophrenia from outpatient clinics completed an electroencephalographic session for MMN, magnetic resonance spectroscopy for glutamate and GABA, and a DST. The study dates were July 2011 to May 2014, and the dates of our analysis were May 2014 to August 2015. MAIN OUTCOMES AND MEASURES Glutamate, GABA, the ratio of glutamine to glutamate, MMN amplitude, and DST. Structural equation modeling was used to test the effects of neurochemistry and MMN amplitude on DST performance. RESULTS The 45 persons with schizophrenia were a mean (SD) of 37.7 (12.8) years and the control participants were 37.1 (13.1) years. The schizophrenia group had a mean (SD) of 14.7 (12.1) years of illness. Mismatch negativity amplitude (F = 4.39, P = .04) and glutamate (F = 9.69, P = .002) were reduced in the schizophrenia group. Smaller MMN amplitude was significantly associated with lower GABA level (P = .008), lower glutamate level (P = .05), and higher ratio of glutamine to glutamate (P = .003). Reduced MMN amplitude was linked to poor verbal working memory in schizophrenia (P = .002). Modeling revealed that a proxy of glutamatergic function, indexed by the ratio of glutamine to glutamate, influenced a path from the ratio of glutamine to glutamate to MMN to verbal working memory (P = .38 [root-mean-square error of approximation, P < .001] by χ2 test

  8. Effect of γ-Aminobutyric Acid-producing Lactobacillus Strain on Laying Performance, Egg Quality and Serum Enzyme Activity in Hy-Line Brown Hens under Heat Stress.

    PubMed

    Zhu, Y Z; Cheng, J L; Ren, M; Yin, L; Piao, X S

    2015-07-01

    Heat-stress remains a costly issue for animal production, especially for poultry as they lack sweat glands, and alleviating heat-stress is necessary for ensuring animal production in hot environment. A high γ-aminobutyric acid (GABA)-producer Lactobacillus strain was used to investigate the effect of dietary GABA-producer on laying performance and egg quality in heat-stressed Hy-line brown hens. Hy-Line brown hens (n = 1,164) at 280 days of age were randomly divided into 4 groups based on the amount of freeze-dried GABA-producer added to the basal diet as follows: i) 0 mg/kg, ii) 25 mg/kg, iii) 50 mg/kg, and iv) 100 mg/kg. All hens were subjected to heat-stress treatment through maintaining the temperature and the relative humidity at 28.83±3.85°C and 37% to 53.9%, respectively. During the experiment, laying rate, egg weight and feed intake of hens were recorded daily. At the 30th and 60th day after the start of the experiment, biochemical parameters, enzyme activity and immune activity in serum were measured. Egg production, average egg weight, average daily feed intake, feed conversion ratio and percentage of speckled egg, soft shell egg and misshaped egg were significantly improved (p<0.05) by the increasing supplementation of the dietary GABA-producer. Shape index, eggshell thickness, strength and weight were increased linearly with increasing GABA-producer supplementation. The level of calcium, phosphorus, glucose, total protein and albumin in serum of the hens fed GABA-producing strain supplemented diet was significantly higher (p<0.05) than that of the hens fed the basal diet, whereas cholesterol level was decreased. Compared with the basal diet, GABA-producer strain supplementation increased serum level of glutathione peroxidase (p = 0.009) and superoxide dismutase. In conclusion, GABA-producer played an important role in alleviating heat-stress, the isolated GABA-producer strain might be a potential natural and safe probiotic to use to improve laying

  9. Effect of Jian-Pi-Zhi-Dong Decoction on striatal glutamate and γ-aminobutyric acid levels detected using microdialysis in a rat model of Tourette syndrome

    PubMed Central

    Zhang, Wen; Wei, Li; Yu, Wenjing; Cui, Xia; Liu, Xiaofang; Wang, Qian; Wang, Sumei

    2016-01-01

    Background Jian-Pi-Zhi-Dong Decoction (JPZDD) is a dedicated treatment of Tourette syndrome (TS). The balance of neurotransmitters in the cortico-striato-pallido-thalamo-cortical network is crucial to the occurrence of TS and related to its severity. This study evaluated the effect of JPZDD on glutamate (Glu) and γ-aminobutyric acid (GABA) and their receptors in a TS rat model. Materials and methods Rats were divided into four groups (n=12 each). TS was induced in three of the groups by injecting them with 3,3′-iminodipropionitrile for 7 consecutive days. Two model groups were treated with tiapride (Tia) or JPZDD, while the control and the remaining model group were gavaged with saline. Behavior was assessed by stereotypic score and autonomic activity. Striatal Glu and GABA contents were detected using microdialysis. Expressions of N-methyl-D-aspartate receptor 1 and GABAA receptor (GABAAR) were observed using Western blot and real-time polymerase chain reaction. Results Tia and JPZDD groups had decreased stereotypy compared with model rats; however, the JPZDD group showed a larger decrease in stereotypy than the Tia group at a 4-week time point. In a spontaneous activity test, the total distance of the JPZDD and Tia groups was significantly decreased compared with the model group. The Glu levels of the model group were higher than the control group and decreased with Tia or JPZDD treatment. The GABA level was higher in the model group than the control group. Expressions of GABAAR protein in the model group were higher than in the control group. Treatment with Tia or JPZDD reduced the expression of GABAAR protein. In the case of the mRNA expression, only Tia reduced the expression of N-methyl-D-aspartate receptor 1, compared with the model group. Conclusion JPZDD could alleviate impairments in behavior and dysfunctional signaling by downregulating GABAAR in the striatum. We suggest that this acts to maintain the balance of Glu and GABA. PMID:27279743

  10. β-Aminobutyric acid increases abscisic acid accumulation and desiccation tolerance and decreases water use but fails to improve grain yield in two spring wheat cultivars under soil drying.

    PubMed

    Du, Yan-Lei; Wang, Zhen-Yu; Fan, Jing-Wei; Turner, Neil C; Wang, Tao; Li, Feng-Min

    2012-08-01

    A pot experiment was conducted to investigate the effect of the non-protein amino acid, β-aminobutyric acid (BABA), on the homeostasis between reactive oxygen species (ROS) and antioxidant defence during progressive soil drying, and its relationship with the accumulation of abscisic acid (ABA), water use, grain yield, and desiccation tolerance in two spring wheat (Triticum aestivum L.) cultivars released in different decades and with different yields under drought. Drenching the soil with 100 µM BABA increased drought-induced ABA production, leading to a decrease in the lethal leaf water potential (Ψ) used to measure desiccation tolerance, decreased water use, and increased water use efficiency for grain (WUEG) under moderate water stress. In addition, at severe water stress levels, drenching the soil with BABA reduced ROS production, increased antioxidant enzyme activity, and reduced the oxidative damage to lipid membranes. The data suggest that the addition of BABA triggers ABA accumulation that acts as a non-hydraulic root signal, thereby closing stomata, and reducing water use at moderate stress levels, and also reduces the production of ROS and increases the antioxidant defence enzymes at severe stress levels, thus increasing the desiccation tolerance. However, BABA treatment had no effect on grain yield of wheat when water availability was limited. The results suggest that there are ways of effectively priming the pre-existing defence pathways, in addition to genetic means, to improve the desiccation tolerance and WUEG of wheat.

  11. Formation of biologically relevant carboxylic acids during the gamma irradiation of acetic acid

    NASA Technical Reports Server (NTRS)

    Negron-Mendoza, A.; Ponnamperuma, C.

    1976-01-01

    Irradiation of aqueous solutions of acetic acid with gamma rays produced several carboxylic acids in small yield. Their identification was based on the technique of gas chromatography combined with mass spectrometry. Some of these acids are Krebs Cycle intermediates. Their simultaneous formation in experiments simulating the primitive conditions on the earth suggests that metabolic pathways may have had their origin in prebiotic chemical processes.

  12. Development and Validation of a HPTLC Method for Simultaneous Estimation of L-Glutamic Acid and γ-Aminobutyric Acid in Mice Brain.

    PubMed

    Sancheti, J S; Shaikh, M F; Khatwani, P F; Kulkarni, Savita R; Sathaye, Sadhana

    2013-11-01

    A new robust, simple and economic high performance thin layer chromatographic method was developed for simultaneous estimation of L-glutamic acid and γ-amino butyric acid in brain homogenate. The high performance thin layer chromatographic separation of these amino acid was achieved using n-butanol:glacial acetic acid:water (22:3:5 v/v/v) as mobile phase and ninhydrin as a derivatising agent. Quantitation of the method was achieved by densitometric method at 550 nm over the concentration range of 10-100 ng/spot. This method showed good separation of amino acids in the brain homogenate with Rf value of L-glutamic acid and γ-amino butyric acid as 21.67±0.58 and 33.67±0.58, respectively. The limit of detection and limit of quantification for L-glutamic acid was found to be 10 and 20 ng and for γ-amino butyric acid it was 4 and 10 ng, respectively. The method was also validated in terms of accuracy, precision and repeatability. The developed method was found to be precise and accurate with good reproducibility and shows promising applicability for studying pathological status of disease and therapeutic significance of drug treatment. PMID:24591747

  13. Mechanism of Inactivation of γ-Aminobutyric Acid Aminotransferase by (1S ,3S)-3-Amino-4-difluoromethylene-1-cyclopentanoic Acid (CPP-115)

    SciTech Connect

    Lee, Hyunbeom; Doud, Emma H.; Wu, Rui; Sanishvili, Ruslan; Juncosa, Jose I.; Liu, Dali; Kelleher, Neil L.; Silverman, Richard B.

    2015-01-23

    γ-Aminobutyric acid aminotransferase (GABA-AT) is a pyridoxal 5'-phosphate (PLP)-dependent enzyme that degrades GABA, the principal inhibitory neurotransmitter in mammalian cells. When the concentration of GABA falls below a threshold level, convulsions can occur. Inhibition of GABA-AT raises GABA levels in the brain, which can terminate seizures as well as have potential therapeutic applications in treating other neurological disorders, including drug addiction. Among the analogues that we previously developed, (1S,3S)-3-amino-4-difluoromethylene-1-cyclopentanoic acid (CPP-115) showed 187 times greater potency than that of vigabatrin, a known inactivator of GABA-AT and approved drug (Sabril) for the treatment of infantile spasms and refractory adult epilepsy. Recently, CPP-115 was shown to have no adverse effects in a Phase I clinical trial. Here we report a novel inactivation mechanism for CPP-115, a mechanism-based inactivator that undergoes GABA-AT-catalyzed hydrolysis of the difluoromethylene group to a carboxylic acid with concomitant loss of two fluoride ions and coenzyme conversion to pyridoxamine 5'-phosphate (PMP). The partition ratio for CPP-115 with GABA-AT is about 2000, releasing cyclopentanone-2,4-dicarboxylate (22) and two other precursors of this compound (20 and 21). Time-dependent inactivation occurs by a conformational change induced by the formation of the aldimine of 4-aminocyclopentane-1,3-dicarboxylic acid and PMP (20), which disrupts an electrostatic interaction between Glu270 and Arg445 to form an electrostatic interaction between Arg445 and the newly formed carboxylate produced by hydrolysis of the difluoromethylene group in CPP-115, resulting in a noncovalent, tightly bound complex. Ultimately, this represents a novel mechanism for inactivation of GABA-AT and a new approach for the design of mechanism-based inactivators in general.

  14. Manganese accumulation in membrane fractions of primary astrocytes is associated with decreased γ-aminobutyric acid (GABA) uptake, and is exacerbated by oleic acid and palmitate.

    PubMed

    Fordahl, Steve C; Erikson, Keith M

    2014-05-01

    Manganese (Mn) exposure interferes with GABA uptake; however, the effects of Mn on GABA transport proteins (GATs) have not been identified. We sought to characterize how Mn impairs GAT function in primary rat astrocytes. Astrocytes exposed to Mn (500 μM) had significantly reduced (3)H-GABA uptake despite no change in membrane or cytosolic GAT3 protein levels. Co-treatment with 100 μM oleic or palmitic acids (both known to be elevated in Mn neurotoxicity), exacerbated the Mn-induced decline in (3)H-GABA uptake. Mn accumulation in the membrane fraction of astrocytes was enhanced with fatty acid administration, and was negatively correlated with (3)H-GABA uptake. Furthermore, control cells exposed to Mn only during the experimental uptake had significantly reduced (3)H-GABA uptake, and the addition of GABA (50 μM) blunted cytosolic Mn accumulation. These data indicate that reduced GAT function in astrocytes is influenced by Mn and fatty acids accumulating at or interacting with the plasma membrane.

  15. Intoxications due to ingestion of gamma-butyrolactone: organ distribution of gamma-hydroxybutyric acid and gamma-butyrolactone.

    PubMed

    Lenz, Daniel; Rothschild, Markus A; Kröner, Lars

    2008-12-01

    In Europe, the misuse of gamma-hydroxybutyric acid (GHB) and its analogues has increased within the recent years. Here, 2 fatalities and 1 nonfatal intoxication resulting from ingestion of gamma-butyrolactone (GBL), a precursor of GHB, are presented. GHB was quantified involving the conversion to GBL by application of a gas chromatography-mass spectrometry (GC/MS) method. Besides quantitation of GHB equivalents ("total GBL"), all specimens of case 1 were analyzed for the metabolic precursor GBL itself (absolute GBL). The cause of death in each case was attributed to GHB intoxication; the manner of death was suicide in the first case and accidental in the second one. Another yet nonfatal GHB intoxication was reported by an emergency department concerning a 36-year-old woman who was hospitalized due to her comatose state and loss of adverse effects reflexes. Here nail polish remover pads were used as source for GBL. PMID:18824955

  16. The study of gamma irradiation effects on poly (glycolic acid)

    NASA Astrophysics Data System (ADS)

    Rao Nakka, Rajeswara; Rao Thumu, Venkatappa; Reddy SVS, Ramana; Rao Buddhiraju, Sanjeeva

    2015-05-01

    We have investigated the effects of gamma irradiation on chemical structure, thermal and morphological properties of biodegradable semi-crystalline poly (glycolic acid) (PGA). PGA samples were subjected to irradiation treatment using a 60Co gamma source with a delivered dose of 30, 60 and 90 kGy, respectively. Gamma irradiation induces cleavage of PGA main chains forming ∼OĊH2 and ĊH2COO∼ radicals in both amorphous and crystalline regions. The free radicals formed in the amorphous region abstract atmospheric oxygen and convert them to peroxy radicals. The peroxy radical causes chain scission at the crystal interface through hydrogen abstraction from methylene groups forming the ∼ĊHCOO∼ (I) radical. Consequently, the observed electron spin resonance (ESR) doublet of irradiated PGA is assigned to (I). The disappearance of the ESR signal above 190°C indicates that free radicals are formed in the amorphous region and decay below the melting temperature of PGA. Fourier transform infrared and optical absorption studies confirm that the ? groups are not influenced by gamma irradiation. Differential scanning calorimetry (DSC) studies showed that the melting temperature of PGA decreased from 212°C to 202°C upon irradiation. Degree of crystallinity increased initially and then decreased with an increase in radiation as per DSC and X-ray diffraction studies. Irradiation produced changes in the physical properties of PGA as well as affecting the morphology of the material.

  17. γ-Aminobutyric Acid (GABA) Production and Angiotensin-I Converting Enzyme (ACE) Inhibitory Activity of Fermented Soybean Containing Sea Tangle by the Co-Culture of Lactobacillus brevis with Aspergillus oryzae.

    PubMed

    Jang, Eun Kyeong; Kim, Nam Yeun; Ahn, Hyung Jin; Ji, Geun Eog

    2015-08-01

    To enhance the γ-aminobutyric acid (GABA) content, the optimized fermentation of soybean with added sea tangle extract was evaluated at 30°C and pH 5.0. The medium was first inoculated with Aspergillus oryzae strain FMB S46471 and fermented for 3 days, followed by the subsequent inoculation with Lactobacillus brevis GABA 100. After fermentation for 7 days, the fermented soybean showed approximately 1.9 g/kg GABA and exhibited higher ACE inhibitory activity than the traditional soybean product. Furthermore, several peptides in the fraction containing the highest ACE inhibitory activity were identified. The novel fermented soybean enriched with GABA and ACE inhibitory components has great pharmaceutical and functional food values.

  18. Eskimo plasma constituents, dihomo-gamma-linolenic acid, eicosapentaenoic acid and docosahexaenoic acid inhibit the release of atherogenic mitogens.

    PubMed

    Smith, D L; Willis, A L; Nguyen, N; Conner, D; Zahedi, S; Fulks, J

    1989-01-01

    Studies in man and laboratory animals suggest that omega 3 polyunsaturated fatty acid constituents of fish oils have antiatherosclerotic properties. We have studied the effects of several such polyunsaturated fatty acids for ability to modify the in vitro release of mitogens from human platelets. Such mitogens may produce the fibro-proliferative component of atherosclerotic plaques. Both 5,8,11,14,17-eicosapentaenoic acid (20:5 omega 3) and 4,7,10,13,16,19-docosahexaenoic acid (22:6 omega 3), major constituents of fish oils, inhibited adenosine diphosphate-induced aggregation of platelets and the accompanying release of mitogens. These effects are dose dependent. Linolenic acid (18:3 omega 3), the biosynthetic precursor of eicosapentaenoic acid, also inhibited platelet aggregation and mitogen release. Eicosapentaenoic acid also inhibited mitogen release from human monocyte-derived macrophages, which, in vivo, are an additional source of mitogens during atherogenesis. Potent inhibition of human platelet aggregation and mitogen release was also seen with dihomo-gamma-linolenic acid (8,11,14-eicosatrienoic acid 20:3 omega 6), whose levels are reportedly elevated in Eskimos subsisting on marine diets. We conclude that diets that elevate plasma and/or tissue levels of eicosapentaenoic acid, docosahexaenoic acid and dihomo-gamma-linolenic acid precursor gamma-linolenic acid (18:3 omega 6) may exert antiatherosclerotic effects by inhibiting the release of mitogens from platelets and other cells.

  19. Incorporation and distribution of dihomo-gamma-linolenic acid, arachidonic acid, and eicosapentaenoic acid in cultured human keratinocytes

    SciTech Connect

    Punnonen, K.; Puustinen, T.; Jansen, C.T.

    1986-02-01

    Human keratinocytes in culture were labelled with /sup 14/C-dihomo-gamma-linolenic acid, /sup 14/C-arachidonic acid or /sup 14/C-eicosapentaenoic acid. All three eicosanoid precursor fatty acids were effectively incorporated into the cells. In phospholipids most of the radioactivity was recovered, in neutral lipids a substantial amount, and as free unesterified fatty acids only a minor amount. Most of the radioactivity was found in phosphatidylethanolamine which was also the major phospholipid as measured by phosphorous assay. The incorporation of dihomo-gamma-linolenic acid and arachidonic acid into lipid subfractions was essentially similar. Eicosapentaenoic acid was, however, much less effectively incorporated into phosphatidylinositol + phosphatidylserine and, correspondingly, more effectively into triacylglycerols as compared to the two other precursor fatty acids. Once incorporated, the distribution of all three precursor fatty acids was relatively stable, and only minor amounts of fatty acids were released into the culture medium during short term culture (two days). Our study demonstrates that eicosanoid precursor fatty acids are avidly taken up by human keratinocytes and esterified into membrane lipids. The clinical implication of this finding is that dietary manipulations might be employed to cause changes in the fatty acid composition of keratinocytes.

  20. Novel high-affinity and selective biaromatic 4-substituted gamma-hydroxybutyric acid (GHB) analogues as GHB ligands: design, synthesis, and binding studies.

    PubMed

    Høg, Signe; Wellendorph, Petrine; Nielsen, Birgitte; Frydenvang, Karla; Dahl, Ivar F; Bräuner-Osborne, Hans; Brehm, Lotte; Frølund, Bente; Clausen, Rasmus P

    2008-12-25

    Gamma-hydroxybutyrate (GHB) is a metabolite of gamma-aminobutyric acid (GABA) and has been proposed to function as a neurotransmitter or neuromodulator. GHB is used in the treatment of narcolepsy and is a drug of abuse. GHB binds to both GABA(B) receptors and specific high-affinity GHB sites in brain, of which the latter have not been linked unequivocally to function, but are speculated to be GHB receptors. In this study, a series of biaromatic 4-substituted GHB analogues, including 4'-phenethylphenyl, 4'-styrylphenyl, and 4'-benzyloxyphenyl GHB analogues, were synthesized and characterized pharmacologically in a [3H](E,RS)-(6,7,8,9-tetrahydro-5-hydroxy-5H-benzocyclohept-6-ylidene)acetic acid ([3H]NCS-382) binding assay and in GABA(A) and GABA(B) receptor binding assays. The compounds were selective for the high-affinity GHB binding sites and several displayed Ki values below 100 nM. The affinity of the 4-[4'-(2-iodobenzyloxy)phenyl] GHB analogue 17b was shown to reside predominantly with the R-enantiomer (Ki = 22 nM), which has higher affinity than previously reported GHB ligands.

  1. Gamma amino butyric acid accumulation in medicinal plants without stress

    PubMed Central

    Anju, P.; Moothedath, Ismail; Rema Shree, Azhimala Bhaskaranpillai

    2014-01-01

    Introduction: Gamma amino butyric acid (GABA) is an important ubiquitous four carbon nonprotein amino acid with an amino group attached to gamma carbon instead of beta carbon. It exists in different organisms including bacteria, plants, and animals and plays a crucial role in humans by regulating neuronal excitability throughout the nervous system. It is directly responsible for the regulation of muscle tone and also effective in lowering stress, blood pressure, and hypertension. Aim and Objective: The aim of the study was to develop the fingerprint profile of selected medicinally and economically important plants having central nervous system (CNS) activity and to determine the quantity of GABA in the selected plants grown under natural conditions without any added stress. Materials and Methods: The high-performance thin layer chromatography analysis was performed on precoated silica gel plate 60F–254 plate (20 cm × 10 cm) in the form of bands with width 8 mm using Hamilton syringe (100 μl) using n-butanol, acetic acid, and water in the proportion 5:2:2 as mobile phase in a CAMAG chamber which was previously saturated for 30 min. CAMAG TLC scanner 3 was used for the densitometric scanning at 550 nm. Specific marker compounds were used for the quantification. Results and Conclusion: Among the screened medicinal plants, Zingiber officinale and Solanum torvum were found to have GABA. The percentage of GABA present in Z. officinale and S. torvum were found to be 0.0114% and 0.0119%, respectively. The present work confirmed that among the selected CNS active medicinal plants, only two plants contain GABA. We found a negative correlation with plant having CNS activity and accumulation of GABA. The GABA shunt is a conserved pathway in eukaryotes and prokaryotes but, although the role of GABA as a neurotransmitter in mammals is clearly established, its role in plants is still vague. PMID:25861139

  2. Drug-facilitated sexual assault involving gamma-hydroxybutyric acid.

    PubMed

    Stillwell, Matthew E

    2002-09-01

    The first case involving an alleged sexual assault linked to the use of gamma-hydroxybutyric acid (GHB) in Oklahoma is reported. A-48-year-old Caucasian woman taking amitriptyline was known to have voluntarily ingested a sports drink containing a relaxing health product. She purportedly experienced unconsciousness that persisted for approximately 4 h. The toxicological testing on urine identified GHB, amitriptyline, and nortriptyline using a capillary Hewlett-Packard 6890 gas chromatograph coupled to a Hewlett-Packard 5973 mass selective detector (MSD). The GHB concentration in urine was 26.9 microg/mL. Urine concentrations of amitriptyline and nortriptyline were not determined. The analytical method used for identifying and quantitating GHB can be applied to matters of forensic interests.

  3. Characterization of cDNAs encoding the chick retinoic acid receptor gamma 2 and preferential distribution of retinoic acid receptor gamma transcripts during chick skin development.

    PubMed

    Michaille, J J; Blanchet, S; Kanzler, B; Garnier, J M; Dhouailly, D

    1994-12-01

    Retinoic acid receptors alpha, beta and gamma (RAR alpha, beta and gamma) are ligand-inductible transcriptional activators which belong to the steroid/thyroid hormone receptor superfamily. At least two major isoforms (1 and 2) of each RAR arise by differential use of two promoters and alternative splicing. In mouse, the three RAR genes are expressed in stage- and tissue-specific patterns during embryonic development. In order to understand the role of the different RARs in chick, RAR gamma 2 cDNAs were isolated from an 8.5-day (stage 35 of Hamburger and Hamilton) chick embryo skin library. The deduced chick RAR gamma 2 amino acid sequence displays uncommon features such as 21 specific amino acid replacements, 12 of them being clustered in the amino-terminal region (domains A2 and B), and a truncated acidic carboxy-terminal region (F domain). However, the pattern of RAR gamma expression in chick embryo resembles that reported in mouse, particularly in skin where RAR gamma expression occurs in both the dermal and epidermal layers at the beginning of feather formation, and is subsequently restricted to the differentiating epidermal cells. Northern blot analysis suggests that different RAR gamma isoforms could be successively required during chick development.

  4. Insect γ-aminobutyric acid receptors and isoxazoline insecticides: toxicological profiles relative to the binding sites of [³H]fluralaner, [³H]-4'-ethynyl-4-n-propylbicycloorthobenzoate, and [³H]avermectin.

    PubMed

    Zhao, Chunqing; Casida, John E

    2014-02-01

    Isoxazoline insecticides, such as fluralaner (formerly A1443), are noncompetitive γ-aminobutyric acid (GABA) receptor (GABA-R) antagonists with selective toxicity for insects versus mammals. The isoxazoline target in house fly ( Musca domestica ) brain has subnanomolar affinity for [³H]fluralaner and a unique pattern of sensitivity to isoxazolines and avermectin B(1a) (AVE) but not to fipronil and α-endosulfan. Inhibitor specificity profiles for 15 isoxazolines examined with Musca GABA-R and [³H]fluralaner, [³H]-4'-ethynyl-4-n-propylbicycloorthobenzoate ([³H]EBOB), and [³H]AVE binding follow the same structure-activity trends although without high correlation. The 3 most potent of the 15 isoxazolines tested in Musca [³H]fluralaner, [³H]EBOB, and [³H]AVE binding assays and in honeybee (Apis mellifera) brain [³H]fluralaner assays are generally those most toxic to Musca and four agricultural pests. Fluralaner does not inhibit [³H]EBOB binding to the human GABA-R recombinant β₃ homopentamer, which is highly sensitive to all of the commercial GABAergic insecticides. The unique isoxazoline binding site may resurrect the GABA-R as a major insecticide target.

  5. Synthesis and characterization of Co(II), Ni(II), Cu(II) and Zn(II) complexes of tridentate Schiff base derived from vanillin and DL-α-aminobutyric acid

    NASA Astrophysics Data System (ADS)

    Nair, M. Sivasankaran; Joseyphus, R. Selwin

    2008-09-01

    Co(II), Ni(II), Cu(II) and Zn(II) complexes of the Schiff base derived from vanillin and DL-α-aminobutyric acid were synthesized and characterized by elemental analysis, IR, electronic spectra, conductance measurements, magnetic measurements, powder XRD and biological activity. The analytical data show the composition of the metal complex to be [ML(H 2O)], where L is the Schiff base ligand. The conductance data indicate that all the complexes are non-electrolytes. IR results demonstrate the tridentate binding of the Schiff base ligand involving azomethine nitrogen, phenolic oxygen and carboxylato oxygen atoms. The IR data also indicate the coordination of a water molecule with the metal ion in the complex. The electronic spectral measurements show that Co(II) and Ni(II) complexes have tetrahedral geometry, while Cu(II) complex has square planar geometry. The powder XRD studies indicate that Co(II) and Cu(II) complexes are amorphous, whereas Ni(II) and Zn(II) complexes are crystalline in nature. Magnetic measurements show that Co(II), Ni(II) and Cu(II) complexes have paramagnetic behaviour. Antibacterial results indicated that the metal complexes are more active than the ligand.

  6. Orexin a phosphorylates the γ-Aminobutyric acid type A receptor β2 subunit on a serine residue and changes the surface expression of the receptor in SH-SY5Y cells exposed to propofol.

    PubMed

    Andersson, Henrik; Björnström, Karin; Eintrei, Christina; Sundqvist, Tommy

    2015-11-01

    Propofol activates the γ-aminobutyric acid type A receptor (GABAA R) and causes a reversible neurite retraction, leaving a thin, thread-like structure behind; it also reverses the transport of vesicles in rat cortical neurons. The awakening peptide orexin A (OA) inhibits this retraction via phospholipase D (PLD) and protein kinase Cɛ (PKCɛ). The human SH-SY5Y cells express both GABAA Rs and orexin 1 and 2 receptors. These cells are used to examine the interaction between OA and the GABAA R. The effects of OA are studied with flow cytometry and immunoblotting. This study shows that OA stimulates phosphorylation on the serine residues of the GABAA R β2 subunit and that the phosphorylation is caused by the activation of PLD and PKCɛ. OA administration followed by propofol reduces the cell surface expression of the GABAA R, whereas propofol stimulation before OA increases the surface expression. The GABAA R β2 subunit is important for receptor recirculation, and the effect of OA on propofol-stimulated cells may be due to a disturbed recirculation of the GABAA R.

  7. Determination of γ-Aminobutyric Acid (GABA) in Rambutan Fruit cv. Rongrian by HPLC-ELSD and Separation of GABA from Rambutan Fruit Using Dowex 50W-X8 Column.

    PubMed

    Meeploy, Maneerat; Deewatthanawong, Rujira

    2016-03-01

    A high-performance liquid chromatography method coupled with an evaporative light scattering detector (ELSD) was validated for the determination of γ-aminobutyric acid (GABA) in rambutan fruit without any sample pretreatment or derivatization. In the concentration range of 0.05-1.0 mg/mL GABA, the ELSD response was linear with a correlation coefficient (r) >0.999. Limit of detection and limit of quantitation were found to be 0.7 and 2.0 µg/mL, respectively. The method enabled the complete separation of GABA in the aqueous extract of rambutan flesh from the impurity peaks at 45.7 min. The recoveries of sample added GABA were obtained in the range of 92.0-99.3%. Intraday and interday relative standard deviations were <5.3%. Repeatability of the extraction process showed the acceptable precision. From the analysis of GABA content in rambutan flesh, 0.71 ± 0.23 mg of GABA was found in 1 g fresh weight. The recovery of GABA after passing through the Dowex 50W-X8 column was 96.65%. The analytical methodology could be potentially applied to the detection and quantification of GABA in other fruits and complex matrices when a sufficient quantity is available. PMID:26590236

  8. Gamma-hydroxybutyric acid (GHB) measurement by GC-MS in blood, urine and gastric contents, following an acute intoxication in Belgium.

    PubMed

    Bodson, Q; Denooz, R; Serpe, P; Charlier, C

    2008-01-01

    Gamma-hydroxybutyrate (GHB, sodium oxybate) is a compound related to neuromodulator gamma-aminobutyric acid (GABA), emerging as a recreational drug of abuse and as a rape drug. GHB-related emergencies have dramatically increased in the 1990s, but a decrease is observed since 2000. We describe the case of an acute GHB intoxication in a 28-year-old male who fell unconscious after ingestion of a mouthful of an unknown beverage, and required medical support for 2 days. A cocaine abuse was also detected by preliminary toxicological screening, but the clinical presentation was not typical of cocaine intoxication. A simple liquid-liquid extraction was used for quantitation of GHB, followed by disilyl-derivatization and analysis in selective ion monitoring (SIM) mode by gas chromatography-mass spectrometry (GC-MS), using GHB-d6 as internal standard. High concentrations of GHB were detected in urine (3020 mg/L) and gastric contents (71487 mg/L) at admission. After a 6-hours delay, GHB was still present in urine at 2324 mg/L and in blood at 43 mg/L. The clinical symptoms of cocaine intoxication were diminished by GHB consumption, and the cerebral scan was modified. Attention must thus be paid to acute intoxications with surprising clinical symptoms, and GHB has probably to be added to the preliminary toxicological screening. Data available regarding GHB are briefly reviewed, and our results are compared with previously published reports of non-fatal GHB intoxication.

  9. [Effect of gamma-linolenic acid on microsomal oxidation in the rat liver following gamma-irradiation].

    PubMed

    Zavodnik, L B; Sushko, L I; Tarasov, Iu A; Ignatenko, K V; Chumachenko, S S; Ovchinnikov, V A; Brzosko, V; Buko, V U

    2001-01-01

    The antioxidant and radioprotector properties of gamma-linolenic acid isolated from the seeds of Borago officialis were studied on rats gamma-irradiated to a dose of 1 Gy. The irradiation caused an increase in the content of malonaldehyde in microsomal liver fraction and disturbed the metabolism of xenobiotics. The administration of gamma-linolenic acid in the form of a commercial drug Neoglandin (daily dose, 150 mg/kg, p.o.; over 1, 3, or 7 days after irradiation reduced the level of lipid peroxidation (for all treatment schedules), normalized the activity of NADPH-oxidase, NADH-oxidase, and NADPH-reductase, and increased the content of cytochromes P-450 and b5 as compared to bothirradiated and control animals. PMID:11589114

  10. Mapping general anesthetic binding site(s) in human α1β3 γ-aminobutyric acid type A receptors with [³H]TDBzl-etomidate, a photoreactive etomidate analogue.

    PubMed

    Chiara, David C; Dostalova, Zuzana; Jayakar, Selwyn S; Zhou, Xiaojuan; Miller, Keith W; Cohen, Jonathan B

    2012-01-31

    The γ-aminobutyric acid type A receptor (GABA(A)R) is a target for general anesthetics of diverse chemical structures, which act as positive allosteric modulators at clinical doses. Previously, in a heterogeneous mixture of GABA(A)Rs purified from bovine brain, [³H]azietomidate photolabeling of αMet-236 and βMet-286 in the αM1 and βM3 transmembrane helices identified an etomidate binding site in the GABA(A)R transmembrane domain at the interface between the β and α subunits [Li, G. D., et.al. (2006) J. Neurosci. 26, 11599-11605]. To further define GABA(A)R etomidate binding sites, we now use [³H]TDBzl-etomidate, an aryl diazirine with broader amino acid side chain reactivity than azietomidate, to photolabel purified human FLAG-α1β3 GABA(A)Rs and more extensively identify photolabeled GABA(A)R amino acids. [³H]TDBzl-etomidate photolabeled in an etomidate-inhibitable manner β3Val-290, in the β3M3 transmembrane helix, as well as α1Met-236 in α1M1, a residue photolabeled by [³H]azietomidate, while no photolabeling of amino acids in the αM2 and βM2 helices that also border the etomidate binding site was detected. The location of these photolabeled amino acids in GABA(A)R homology models derived from the recently determined structures of prokaryote (GLIC) or invertebrate (GluCl) homologues and the results of computational docking studies predict the orientation of [³H]TDBzl-etomidate bound in that site and the other amino acids contributing to this GABA(A)R intersubunit etomidate binding site. Etomidate-inhibitable photolabeling of β3Met-227 in βM1 by [³H]TDBzl-etomidate and [³H]azietomidate also provides evidence of a homologous etomidate binding site at the β3-β3 subunit interface in the α1β3 GABA(A)R.

  11. A potential new metabolite of gamma-hydroxybutyrate: sulfonated gamma-hydroxybutyric acid.

    PubMed

    Hanisch, Stephanie; Stachel, Nicole; Skopp, Gisela

    2016-03-01

    Detection of gamma-hydroxybutyric acid (GHB) became crucial in many clinical and forensic settings due to its increasing use for recreational purposes and drug-facilitated sexual assault. Its narrow window of detection of about 3-12 h in urine represents a major problem. Analogous to ethyl glucuronide, the recently identified GHB-glucuronide exhibits a longer window of detection than the parent drug. It appeared reasonable that a sulfonated metabolite of GHB (GHB-SUL) will also be formed. Due to the lack of an appropriate standard, GHB was incubated with a human liver cytosolic fraction to produce GHB-SUL. Following development of a liquid chromatography/tandem mass spectrometry (LC-MS/MS) assay to measure GHB and GHB-SUL, authentic urine samples (n = 5) were tested for GHB-SUL. These investigations revealed detectable signals of both GHB and GHB-SUL, strongly indicating that GHB is not only glucuronidated but also sulfonated. Given that sulfonated metabolites generally have longer half-life times than the corresponding free drugs, GHB-SUL may serve as a biomarker of GHB misuse along with its glucuronide. PMID:26210636

  12. Gamma-polyglutamic acid (gamma-PGA) produced by Bacillus amyloliquefaciens C06 promoting its colonization on fruit surface.

    PubMed

    Liu, Jun; He, Dan; Li, Xiu-zhen; Gao, Shengfeng; Wu, Huijun; Liu, Wenzhe; Gao, Xuewen; Zhou, Ting

    2010-08-15

    Bacillus amyloliquefaciens C06, an effective biological agent in controlling brown rot of stone fruit caused by Monilinia fructicola, was also found to produce extra-cellular mucilage and form mucoid colonies on semi-solid surfaces. This study aimed to characterize the extra-cellular mucilage produced by B. amyloliquefaciens C06 using transposon mutagenesis and biochemical and physical analyses. The mucilage production in B. amyloliquefaciens C06 was demonstrated to be associated with ywsC gene expression and characterized to be of high molecular weight, consisted of only glutamic acid and linked with non-peptide bonds, thus identified as gamma-polyglutamic acid (gamma-PGA). Compared with wild type B. amyloliquefaciens C06, its mutants deficient in producing gamma-PGA, e.g. M106 and C06DeltaywsC showed less efficiency in biofilm formation, surface adhesion and swarming ability. It was also demonstrated that gamma-PGA was not essential for C06 to form colony on semi-solid surfaces, but was able to improve its colony structure. In vivo evaluation showed that disruption of gamma-PGA production in C06DeltaywsC impaired its efficiency of colonizing apple surfaces.

  13. Bio-derived poly(gamma-glutamic acid) nanogels as controlled anticancer drug delivery carriers.

    PubMed

    Bae, Hee Ho; Cho, Mi Young; Hong, Ji Hyeon; Poo, Haryoung; Sung, Moon-Hee; Lim, Yong Taik

    2012-12-01

    We have developed a novel type of polymer nanogel loaded with anticancer drug based on bio-derived poly(gamma- glutamic acid) (gamma-PGA). gamma-PGA is a highly anionic polymer that is synthesized naturally by microbial species, most prominently in various bacilli, and has been shown to have excellent biocompatibility. Thiolated gamma-PGA was synthesized by covalent coupling between the carboxyl groups of gamma-PGA and the primary amine group of cysteamine. Doxorubicin (Dox)-loaded gamma-PGA nanogels were fabricated using the following steps: (1) an ionic nanocomplex was formed between thiolated gamma-PGA as the negative charge component, and Dox as the positive charge component; (2) addition of poly(ethylene glycol) (PEG) induced hydrogen-bond interactions between thiol groups of thiolated gamma-PGA and hydroxyl groups of PEG, resulting in the nanocomplex; and (3) disulfide crosslinked gamma-PGA nanogels were fabricated by ultrasonication. The average size and surface charge of Dox-loaded disulfide cross-linked gamma-PGA nanogels in aqueous solution were 136.3 +/- 37.6 nm and -32.5 +/- 5.3 mV, respectively. The loading amount of Dox was approximately 38.7 microgram per mg of gamma-PGA nanogel. The Dox-loaded disulfide cross-linked gamma-PGA nanogels showed controlled drug release behavior in the presence of reducing agents, glutathione (GSH) (1- 10 mM). Through fluorescence microscopy and FACS, the cellular uptake of gamma-PGA nanogels into breast cancer cells (MCF-7) was analyzed. The cytotoxic effect was evaluated using the MTT assay and was determined to be dependent on both the concentration and treatment time of gamma-PGA nanogels. The bio-derived gamma-PGA nanogels are expected to be a well-designed delivery carrier for controlled drug delivery applications. PMID:23221543

  14. Cellular localization of retinoic acid receptor-gamma expression in normal and neoplastic skin.

    PubMed Central

    Finzi, E.; Blake, M. J.; Celano, P.; Skouge, J.; Diwan, R.

    1992-01-01

    Retinoids profoundly affect the normal growth and differentiation of epithelial tissues. Retinoic acid receptor-gamma (RAR-gamma) is a member of a family of retinoid receptors, and has been shown to be expressed almost exclusively in skin. However, little is known about the cellular localization of this receptor in human skin. The authors studied the expression of RAR-gamma in normal skin and human skin tumors by Northern blot analysis and in situ hybridization. RAR-gamma mRNA was detected in normal skin as well as in cultures of neonatal keratinocytes. Using an oligonucleotide specific for the RAR-gamma cDNA isoform 1 (RAR-gamma 1), RAR-gamma 1 mRNA was localized to all layers of the epidermis, the outer root sheath of hair follicles, follicular hair bulbs, eccrine and sebaceous glands. Basal cell carcinoma constitutively expressed gamma-1 mRNA and one of seven squamous cell carcinomas showed loss of gamma-1 mRNA expression, relative to adjacent epithelium. By contrast, normal melanocytic nevi and tumor-associated lymphocytes expressed little or no RAR-gamma mRNA. These results suggest that RAR-gamma 1 may play an important role in the maintenance and differentiation of normal epidermis and skin appendages. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 PMID:1318641

  15. Epileptiform activity in the CA1 region of the hippocampus becomes refractory to attenuation by cannabinoids in part because of endogenous γ-aminobutyric acid type B receptor activity.

    PubMed

    Messer, Ricka D; Levine, Eric S

    2012-07-01

    The anticonvulsant properties of marijuana have been known for centuries. The recently characterized endogenous cannabinoid system thus represents a promising target for novel anticonvulsant agents; however, administration of exogenous cannabinoids has shown mixed results in both human epilepsy and animal models. The ability of cannabinoids to attenuate release of both excitatory and inhibitory neurotransmitters may explain the variable effects of cannabinoids in different models of epilepsy, but this has not been well explored. Using acute mouse brain slices, we monitored field potentials in the CA1 region of the hippocampus to characterize systematically the effects of the cannabinoid agonist WIN55212-2 (WIN) on evoked basal and epileptiform activity. WIN, acting presynaptically, significantly reduced the amplitude and slope of basal field excitatory postsynaptic potentials as well as stimulus-evoked epileptiform responses induced by omission of magnesium from the extracellular solution. In contrast, the combination of omission of magnesium plus elevation of potassium induced an epileptiform response that was refractory to attenuation by WIN. The effect of WIN in this model was partially restored by blocking γ-aminobutyric acid type B (GABA(B) ), but not GABA(A) , receptors. Subtle differences in models of epileptiform activity can profoundly alter the efficacy of cannabinoids. Endogenous GABA(B) receptor activation played a role in the decreased cannabinoid sensitivity observed for epileptiform activity induced by omission of magnesium plus elevation of potassium. These results suggest that interplay between presynaptic G protein-coupled receptors with overlapping downstream targets may underlie the variable efficacy of cannabinoids in different models of epilepsy.

  16. A functional role for both γ-aminobutyric acid (GABA) transporter-1 and GABA transporter-3 in the modulation of extracellular GABA and GABAergic tonic conductances in the rat hippocampus

    PubMed Central

    Kersanté, Flavie; Rowley, Samuel C S; Pavlov, Ivan; Gutièrrez-Mecinas, María; Semyanov, Alexey; Reul, Johannes M H M; Walker, Matthew C; Linthorst, Astrid C E

    2013-01-01

    Tonic γ-aminobutyric acid (GABA)A receptor-mediated signalling controls neuronal network excitability in the hippocampus. Although the extracellular concentration of GABA (e[GABA]) is critical in determining tonic conductances, knowledge on how e[GABA] is regulated by different GABA transporters (GATs) in vivo is limited. Therefore, we studied the role of GATs in the regulation of hippocampal e[GABA] using in vivo microdialysis in freely moving rats. Here we show that GAT-1, which is predominantly presynaptically located, is the major GABA transporter under baseline, quiescent conditions. Furthermore, a significant contribution of GAT-3 in regulating e[GABA] was revealed by administration of the GAT-3 inhibitor SNAP-5114 during simultaneous blockade of GAT-1 by NNC-711. Thus, the GABA transporting activity of GAT-3 (the expression of which is confined to astrocytes) is apparent under conditions in which GAT-1 is blocked. However, sustained neuronal activation by K+-induced depolarization caused a profound spillover of GABA into the extrasynaptic space and this increase in e[GABA] was significantly potentiated by sole blockade of GAT-3 (i.e. even when uptake of GAT-1 is intact). Furthermore, experiments using tetrodotoxin to block action potentials revealed that GAT-3 regulates extrasynaptic GABA levels from action potential-independent sources when GAT-1 is blocked. Importantly, changes in e[GABA] resulting from both GAT-1 and GAT-3 inhibition directly precipitate changes in tonic conductances in dentate granule cells as measured by whole-cell patch-clamp recording. Thus, astrocytic GAT-3 contributes to the regulation of e[GABA] in the hippocampus in vivo and may play an important role in controlling the excitability of hippocampal cells when network activity is increased. PMID:23381899

  17. Positive and Negative Allosteric Modulation of an α1β3γ2 γ-Aminobutyric Acid Type A (GABAA) Receptor by Binding to a Site in the Transmembrane Domain at the γ+-β- Interface.

    PubMed

    Jayakar, Selwyn S; Zhou, Xiaojuan; Savechenkov, Pavel Y; Chiara, David C; Desai, Rooma; Bruzik, Karol S; Miller, Keith W; Cohen, Jonathan B

    2015-09-18

    In the process of developing safer general anesthetics, isomers of anesthetic ethers and barbiturates have been discovered that act as convulsants and inhibitors of γ-aminobutyric acid type A receptors (GABAARs) rather than potentiators. It is unknown whether these convulsants act as negative allosteric modulators by binding to the intersubunit anesthetic-binding sites in the GABAAR transmembrane domain (Chiara, D. C., Jayakar, S. S., Zhou, X., Zhang, X., Savechenkov, P. Y., Bruzik, K. S., Miller, K. W., and Cohen, J. B. (2013) J. Biol. Chem. 288, 19343-19357) or to known convulsant sites in the ion channel or extracellular domains. Here, we show that S-1-methyl-5-propyl-5-(m-trifluoromethyl-diazirynylphenyl) barbituric acid (S-mTFD-MPPB), a photoreactive analog of the convulsant barbiturate S-MPPB, inhibits α1β3γ2 but potentiates α1β3 GABAAR responses. In the α1β3γ2 GABAAR, S-mTFD-MPPB binds in the transmembrane domain with high affinity to the γ(+)-β(-) subunit interface site with negative energetic coupling to GABA binding in the extracellular domain at the β(+)-α(-) subunit interfaces. GABA inhibits S-[(3)H]mTFD-MPPB photolabeling of γ2Ser-280 (γM2-15') in this site. In contrast, within the same site GABA enhances photolabeling of β3Met-227 in βM1 by an anesthetic barbiturate, R-[(3)H]methyl-5-allyl-5-(m-trifluoromethyl-diazirynylphenyl)barbituric acid (mTFD-MPAB), which differs from S-mTFD-MPPB in structure only by chirality and two hydrogens (propyl versus allyl). S-mTFD-MPPB and R-mTFD-MPAB are predicted to bind in different orientations at the γ(+)-β(-) site, based upon the distance in GABAAR homology models between γ2Ser-280 and β3Met-227. These results provide an explanation for S-mTFD-MPPB inhibition of α1β3γ2 GABAAR function and provide a first demonstration that an intersubunit-binding site in the GABAAR transmembrane domain binds negative and positive allosteric modulators.

  18. Positive and Negative Allosteric Modulation of an α1β3γ2 γ-Aminobutyric Acid Type A (GABAA) Receptor by Binding to a Site in the Transmembrane Domain at the γ+-β− Interface*

    PubMed Central

    Jayakar, Selwyn S.; Zhou, Xiaojuan; Savechenkov, Pavel Y.; Chiara, David C.; Desai, Rooma; Bruzik, Karol S.; Miller, Keith W.; Cohen, Jonathan B.

    2015-01-01

    In the process of developing safer general anesthetics, isomers of anesthetic ethers and barbiturates have been discovered that act as convulsants and inhibitors of γ-aminobutyric acid type A receptors (GABAARs) rather than potentiators. It is unknown whether these convulsants act as negative allosteric modulators by binding to the intersubunit anesthetic-binding sites in the GABAAR transmembrane domain (Chiara, D. C., Jayakar, S. S., Zhou, X., Zhang, X., Savechenkov, P. Y., Bruzik, K. S., Miller, K. W., and Cohen, J. B. (2013) J. Biol. Chem. 288, 19343–19357) or to known convulsant sites in the ion channel or extracellular domains. Here, we show that S-1-methyl-5-propyl-5-(m-trifluoromethyl-diazirynylphenyl) barbituric acid (S-mTFD-MPPB), a photoreactive analog of the convulsant barbiturate S-MPPB, inhibits α1β3γ2 but potentiates α1β3 GABAAR responses. In the α1β3γ2 GABAAR, S-mTFD-MPPB binds in the transmembrane domain with high affinity to the γ+-β− subunit interface site with negative energetic coupling to GABA binding in the extracellular domain at the β+-α− subunit interfaces. GABA inhibits S-[3H]mTFD-MPPB photolabeling of γ2Ser-280 (γM2–15′) in this site. In contrast, within the same site GABA enhances photolabeling of β3Met-227 in βM1 by an anesthetic barbiturate, R-[3H]methyl-5-allyl-5-(m-trifluoromethyl-diazirynylphenyl)barbituric acid (mTFD-MPAB), which differs from S-mTFD-MPPB in structure only by chirality and two hydrogens (propyl versus allyl). S-mTFD-MPPB and R-mTFD-MPAB are predicted to bind in different orientations at the γ+-β− site, based upon the distance in GABAAR homology models between γ2Ser-280 and β3Met-227. These results provide an explanation for S-mTFD-MPPB inhibition of α1β3γ2 GABAAR function and provide a first demonstration that an intersubunit-binding site in the GABAAR transmembrane domain binds negative and positive allosteric modulators. PMID:26229099

  19. Kinetic studies on the inhibition of GABA-T by gamma-vinyl GABA and taurine.

    PubMed

    Sulaiman, Saba A J; Suliman, Fakhr Eldin O; Barghouthi, Samira

    2003-08-01

    Gamma-aminobutyric acid transaminase (GABA-T, EC 2.6.1.19) is a pyridoxal phosphate (PLP) dependent enzyme that catalyzes the degradation of gamma-aminobutyric acid. The kinetics of this reaction are studied in vitro, both in the absence, and in the presence of two inhibitors: gamma-vinyl GABA (4-aminohex-5-enoic acid), and a natural product, taurine (ethylamine-2-sulfonic acid). A kinetic model that describes the transamination process is proposed. GABA-T from Pseudomonas fluorescens is inhibited by gamma-vinyl GABA and taurine at concentrations of 51.0 and 78.5 mM. Both inhibitors show competitive inhibition behavior when GABA is the substrate and the inhibition constant (Ki) values for gamma-vinyl GABA and taurine were found to be 26 +/- 3 mM and 68 +/- 7 mM respectively. The transamination process of alpha-ketoglutarate was not affected by the presence of gamma-vinyl GABA, whereas, taurine was a noncompetitive inhibitor of GABA-T when alpha-ketoglutarate was the substrate. The inhibition dissociation constant (Kii) for this system was found to be 96 +/- 10 mM. The Michaelis-Menten constant (Km) in the absence of inhibition, was found to be 0.79 +/- 0.11 mM, and 0.47 +/- 0.10 mM for GABA and alpha-ketoglutarate respectively.

  20. Effect of γ-Aminobutyric Acid (GABA) Producing Bacteria on In vitro Rumen Fermentation, Biogenic Amine Production and Anti-oxidation Using Corn Meal as Substrate.

    PubMed

    Ku, Bum Seung; Mamuad, Lovelia L; Kim, Seon-Ho; Jeong, Chang Dae; Soriano, Alvin P; Lee, Ho-Il; Nam, Ki-Chang; Ha, Jong K; Lee, Sang Suk

    2013-06-01

    The effects and significance of γ-amino butyric acid (GABA) producing bacteria (GPB) on in vitro rumen fermentation and reduction of biogenic amines (histamine, methylamine, ethylamine, and tyramine) using corn meal as a substrate were determined. Ruminal samples collected from ruminally fistulated Holstein cows served as inoculum and corn was used as substrate at 2% dry matter (DM). Different inclusion rates of GPB and GABA were evaluated. After incubation, addition of GPB had no significant effect on in vitro fermentation pH and total gas production, but significantly increased the ammonia nitrogen (NH3-N) concentration and reduced the total biogenic amines production (p<0.05). Furthermore, antioxidation activity was improved as indicated by the significantly higher concentration of superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) among treated samples when compared to the control (p<0.05). Additionally, 0.2% GPB was established as the optimum inclusion level. Taken together, these results suggest the potential of utilizing GPB as feed additives to improve growth performance in ruminants by reducing biogenic amines and increasing anti-oxidation.

  1. Effect of γ-Aminobutyric Acid (GABA) Producing Bacteria on In vitro Rumen Fermentation, Biogenic Amine Production and Anti-oxidation Using Corn Meal as Substrate

    PubMed Central

    Ku, Bum Seung; Mamuad, Lovelia L.; Kim, Seon-Ho; Jeong, Chang Dae; Soriano, Alvin P.; Lee, Ho-Il; Nam, Ki-Chang; Ha, Jong K.; Lee, Sang Suk

    2013-01-01

    The effects and significance of γ-amino butyric acid (GABA) producing bacteria (GPB) on in vitro rumen fermentation and reduction of biogenic amines (histamine, methylamine, ethylamine, and tyramine) using corn meal as a substrate were determined. Ruminal samples collected from ruminally fistulated Holstein cows served as inoculum and corn was used as substrate at 2% dry matter (DM). Different inclusion rates of GPB and GABA were evaluated. After incubation, addition of GPB had no significant effect on in vitro fermentation pH and total gas production, but significantly increased the ammonia nitrogen (NH3-N) concentration and reduced the total biogenic amines production (p<0.05). Furthermore, antioxidation activity was improved as indicated by the significantly higher concentration of superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) among treated samples when compared to the control (p<0.05). Additionally, 0.2% GPB was established as the optimum inclusion level. Taken together, these results suggest the potential of utilizing GPB as feed additives to improve growth performance in ruminants by reducing biogenic amines and increasing anti-oxidation. PMID:25049853

  2. Evaluation of radiological data of some saturated fatty acids using gamma ray spectrometry

    NASA Astrophysics Data System (ADS)

    Kore, Prashant S.; Pawar, Pravina P.; Palani Selvam, T.

    2016-02-01

    Radiological parameters such as mass attenuation coefficients (μm), total attenuation cross section (σtot), molar extinction coefficient (ε), mass energy absorption coefficient (μen/ρ) and effective electronic cross section (σt, el) of saturated fatty acids, namely butyric acid (C4H8O2), caproic acid (C6H12O2), enanthic acid (C7H14O2), caprylic acid (C8H16O2), pelargonic acid (C9H18O2) and valeric acid (C5H10O2) were measured using NaI(Tl)-based gamma spectrometry. Radioactive sources used in the study are 57Co, 133Ba, 137Cs, 54Mn, 60Co and 22Na. Gamma ray transmission method in a narrow beam good geometry set up was used in the study. The measured data were compared against Win-XCOM-based data. The agreement is within 1%.

  3. GC-MS Analysis of [gamma]-Hydroxybutyric Acid Analogs: A Forensic Chemistry Experiment

    ERIC Educational Resources Information Center

    Henck, Colin; Nally, Luke

    2007-01-01

    An upper-division forensic chemistry experiment is described. It involves using glycolic acid and sodium glycolate as analogs of [gamma]-hydroxybutyric acid and its sodium salt. The experiment shows the use of silylation in GC-MS analysis and gives students the opportunity to work with a commonly used silylating reagent,…

  4. Induction of renal cytochrome P450 arachidonic acid epoxygenase activity by dietary gamma-linolenic acid.

    PubMed

    Yu, Zhigang; Ng, Valerie Y; Su, Ping; Engler, Marguerite M; Engler, Mary B; Huang, Yong; Lin, Emil; Kroetz, Deanna L

    2006-05-01

    Dietary gamma-linolenic acid (GLA), a omega-6 polyunsaturated fatty acid found in borage oil (BOR), lowers systolic blood pressure in spontaneously hypertensive rats (SHRs). GLA is converted into arachidonic acid (AA) by elongation and desaturation steps. Epoxyeicosatrienoic acids (EETs) and 20-hydroxyeicosatetraenoic acid (20-HETE) are cytochrome P450 (P450)-derived AA eicosanoids with important roles in regulating blood pressure. This study tested the hypothesis that the blood pressure-lowering effect of a GLA-enriched diet involves alteration of P450-catalyzed AA metabolism. Microsomes and RNA were isolated from the renal cortex of male SHRs fed a basal fat-free diet for 5 weeks to which 11% by weight of sesame oil (SES) or BOR was added. There was a 2.6- to 3.5-fold increase in P450 epoxygenase activity in renal microsomes isolated from the BOR-fed SHRs compared with the SES-fed rats. Epoxygenase activity accounted for 58% of the total AA metabolism in the BOR-treated kidney microsomes compared with 33% in the SES-treated rats. More importantly, renal 14,15- and 8,9-EET levels increased 1.6- to 2.5-fold after dietary BOR treatment. The increase in EET formation is consistent with increases in CYP2C23, CYP2C11, and CYP2J protein levels. There were no differences in the level of renal P450 epoxygenase mRNA between the SES- and BOR-treated rats. Enhanced synthesis of the vasodilatory EETs and decreased formation of the vasoconstrictive 20-HETE suggests that changes in P450-mediated AA metabolism may contribute, at least in part, to the blood pressure-lowering effect of a BOR-enriched diet. PMID:16421287

  5. Marked resistance of RAR gamma-deficient mice to the toxic effects of retinoic acid.

    PubMed

    Look, J; Landwehr, J; Bauer, F; Hoffmann, A S; Bluethmann, H; LeMotte, P

    1995-07-01

    Excessive intake of retinol or of retinoic acid causes a syndrome of characteristic toxic effects known as hypervitaminosis A. To test the role of the nuclear retinoic acid receptor (RAR gamma) in this process we produced mice with a targeted disruption of the RAR gamma gene and examined toxic effects of repeated doses of retinoic acid and two other synthetic retinoids, Ro 15-1570 and Ro 40-6055. Surprisingly, homozygous mutant mice were resistant to fourfold higher doses of retinoic acid than wild-type mice as well as to elevated doses of the synthetic retinoids, indicating that RAR gamma may have a major role in mediating retinoid toxicity, a finding that possibly has practical implications for reducing the toxicity of synthetic retinoids in clinical use.

  6. Two isoforms of Xenopus retinoic acid receptor gamma 2 (B) exhibit differential expression and sensitivity to retinoic acid during embryogenesis.

    PubMed

    Crawford, M J; Liversage, R A; Varmuza, S L

    1995-01-01

    We report the isolation of two retinoic acid receptor isoforms (RAR gamma), which differ only in the 5'untranslated and putative N-terminus A regions. The two isoforms appear to serve as early markers for the presumptive neural axis; however, their expression patterns differ. RAR-gamma 2.1 is first expressed at gastrulation at the dorsal lip and subsequently along the presumptive neural axis. RAR- gamma 2.2 represents the full-length sequence of a receptor cDNA already partially characterized and present as a maternal transcript [Ellinger-Ziegelbauer and Dreyer (1991); Genes Dev 5:94-104, (1993): Mech Dev 41:31-46; Pfeffer and DeRobertis, (1994) Mech Dev: 45:147-153]. Unlike RAR-gamma 2.2, the 2.1 variant is not expressed either in pre-somitic mesoderm or notochord. RAR-gamma 2.1 is strongly expressed in branchial arches and to a lesser extent in the neural floor plate. The two isoforms also exhibit differential sensitivity to retinoic acid. Constitutive expression of RAR gamma 2.2 following neurulation appears to be depressed by treatment with retinoic acid, but domains of highest expression, namely, the head and tail, remain relatively unaffected, as do patterns of expression prior to late neurulation. By contrast, RAR-gamma 2.1 is not transcribed in retinoid-inhibited structures. Using microinjection techniques, we show that changes of RAR-gamma 2.1 expression in presumptive head structures occur as an early and local consequence of retinoic acid administration. Since RAR-gamma 2.1 expression is inhibited by retinoic acid, we tested to see if other treatments that perturb axis formation had any effect. Surprisingly, UV irradiation did not suppress that its inhibition by retinoic acid is not due solely to inhibition of anterior neural development. These experiments demonstrate a new subdivision of isoforms that undergo differential expression during development and that exhibit differential sensitivity to retinoic acid and to UV. This sensitivity and the presence

  7. Effect of Functional Bread Rich in Potassium, γ-Aminobutyric Acid and Angiotensin-Converting Enzyme Inhibitors on Blood Pressure, Glucose Metabolism and Endothelial Function: A Double-blind Randomized Crossover Clinical Trial.

    PubMed

    Becerra-Tomás, Nerea; Guasch-Ferré, Marta; Quilez, Joan; Merino, Jordi; Ferré, Raimon; Díaz-López, Andrés; Bulló, Mònica; Hernández-Alonso, Pablo; Palau-Galindo, Antoni; Salas-Salvadó, Jordi

    2015-11-01

    Because it has been suggested that food rich in γ-aminobutyric acid (GABA) or angiotensin-converting enzyme inhibitor (ACEI) peptides have beneficial effects on blood pressure (BP) and other cardiovascular risk factors, we tested the effects of low-sodium bread, but rich in potassium, GABA, and ACEI peptides on 24-hour BP, glucose metabolism, and endothelial function.A randomized, double-blind, crossover trial was conducted in 30 patients with pre or mild-to-moderate hypertension, comparing three 4-week nutritional interventions separated by 2-week washout periods. Patients were randomly assigned to consume 120 g/day of 1 of the 3 types of bread for each nutritional intervention: conventional wheat bread (CB), low-sodium wheat bread enriched in potassium (LSB), and low-sodium wheat bread rich in potassium, GABA, and ACEI peptides (LSB + G). For each period, 24-hour BP measurements, in vivo endothelial function, and biochemical samples were obtained.After LSB + G consumption, 24-hour ambulatory BP underwent a nonsignificant greater reduction than after the consumption of CB and LSB (0.26 mm Hg in systolic BP and -0.63 mm Hg in diastolic BP for CB; -0.71 mm Hg in systolic BP and -1.08 mm Hg in diastolic BP for LSB; and -0.75 mm Hg in systolic BP and -2.12 mm Hg in diastolic BP for LSB + G, respectively). Diastolic BP at rest decreased significantly during the LSB + G intervention, although there were no significant differences in changes between interventions. There were no significant differences between interventions in terms of changes in in vivo endothelial function, glucose metabolism, and peripheral inflammatory parameters.Compared with the consumption of CB or LSB, no greater beneficial effects on 24-hour BP, endothelial function, or glucose metabolism were demonstrated after the consumption of LSB + G in a population with pre or mild-to-moderate hypertension. Further studies are warranted to clarify the effect of GABA on BP

  8. Effect of Functional Bread Rich in Potassium, γ-Aminobutyric Acid and Angiotensin-Converting Enzyme Inhibitors on Blood Pressure, Glucose Metabolism and Endothelial Function: A Double-blind Randomized Crossover Clinical Trial.

    PubMed

    Becerra-Tomás, Nerea; Guasch-Ferré, Marta; Quilez, Joan; Merino, Jordi; Ferré, Raimon; Díaz-López, Andrés; Bulló, Mònica; Hernández-Alonso, Pablo; Palau-Galindo, Antoni; Salas-Salvadó, Jordi

    2015-11-01

    Because it has been suggested that food rich in γ-aminobutyric acid (GABA) or angiotensin-converting enzyme inhibitor (ACEI) peptides have beneficial effects on blood pressure (BP) and other cardiovascular risk factors, we tested the effects of low-sodium bread, but rich in potassium, GABA, and ACEI peptides on 24-hour BP, glucose metabolism, and endothelial function.A randomized, double-blind, crossover trial was conducted in 30 patients with pre or mild-to-moderate hypertension, comparing three 4-week nutritional interventions separated by 2-week washout periods. Patients were randomly assigned to consume 120 g/day of 1 of the 3 types of bread for each nutritional intervention: conventional wheat bread (CB), low-sodium wheat bread enriched in potassium (LSB), and low-sodium wheat bread rich in potassium, GABA, and ACEI peptides (LSB + G). For each period, 24-hour BP measurements, in vivo endothelial function, and biochemical samples were obtained.After LSB + G consumption, 24-hour ambulatory BP underwent a nonsignificant greater reduction than after the consumption of CB and LSB (0.26 mm Hg in systolic BP and -0.63 mm Hg in diastolic BP for CB; -0.71 mm Hg in systolic BP and -1.08 mm Hg in diastolic BP for LSB; and -0.75 mm Hg in systolic BP and -2.12 mm Hg in diastolic BP for LSB + G, respectively). Diastolic BP at rest decreased significantly during the LSB + G intervention, although there were no significant differences in changes between interventions. There were no significant differences between interventions in terms of changes in in vivo endothelial function, glucose metabolism, and peripheral inflammatory parameters.Compared with the consumption of CB or LSB, no greater beneficial effects on 24-hour BP, endothelial function, or glucose metabolism were demonstrated after the consumption of LSB + G in a population with pre or mild-to-moderate hypertension. Further studies are warranted to clarify the effect of GABA on BP

  9. Differential effects of methylmercury on gamma-aminobutyric acid type A receptor currents in rat cerebellar granule and cerebral cortical neurons in culture.

    PubMed

    Herden, Christina J; Pardo, Nicole E; Hajela, Ravindra K; Yuan, Yukun; Atchison, William D

    2008-02-01

    Cerebellar granule cells are particularly sensitive to inhibition by methylmercury (MeHg) on GABA(A) receptor function. This is manifested as a more rapid block of inhibitory postsynaptic currents/inhibitory postsynaptic potentials than for Purkinje cells. The underlying mechanism(s) for differential sensitivity of GABAergic transmission to MeHg in cerebellar neurons is unknown. Differential expression of alpha(6) subunit-containing GABA(A) receptors in cerebellar granule and Purkinje neurons could partially explain this. GABA-evoked currents (I(GABA)) were recorded in response to MeHg in alpha(6) subunit-containing cerebellar granule cells and alpha(6) subunit-deficient cerebral cortical cells in culture. Cortical cells were substituted for Purkinje cells, which do not express alpha(6) subunits. They express the same alpha(1)-containing GABA(A) receptor as Purkinje cells but lack characteristics that enhance Purkinje cell resistance to MeHg. I(GABA) were obtained using whole-cell recording and symmetrical [Cl(-)]. MeHg reduced I(GABA) to complete block in both cell types in a time- and concentration-dependent manner. This effect was faster in granule cells than cortical cells. Effects of MeHg on I(GABA) were recorded in granule cells at various developmental stages (days in vitro 4, 6, and 8) to alter the expression level of alpha(6) subunit-containing GABA(A) receptors. Effects of MeHg on I(GABA) were similar in cells at all days. In human embryonic kidney 293 cells expressing either alpha(6) or alpha(1) subunit-containing GABA(A) receptors, time to block of I(GABA) by MeHg was comparable. Thus, the presence of the alpha(6) subunit alone may not underlie the differential effects of MeHg on I(GABA) observed in cerebellar granule and cortical neurons; other factors are likely to be involved as well. PMID:17977981

  10. Polyamine biosynthesis and degradation are modulated by exogenous gamma-aminobutyric acid in root-zone hypoxia-stressed melon roots.

    PubMed

    Wang, Chunyan; Fan, Longquan; Gao, Hongbo; Wu, Xiaolei; Li, Jingrui; Lv, Guiyun; Gong, Binbin

    2014-09-01

    We detected physiological change and gene expression related to PA metabolism in melon roots under controlled and hypoxic conditions with or without 5 mM GABA. Roots with hypoxia treatment showed a significant increase in glutamate decarboxylase (GAD) activity and endogenous GABA concentration. Concurrently, PA biosynthesis and degradation accelerated with higher gene expression and enzymes activity. However, endogenous GABA concentrations showed a large and rapid increase in Hypoxia + GABA treated roots. This led to a marked increase in Glu concentration by feedback inhibition of GAD activity. Hypoxia + GABA treatment enhanced arginine (Arg), ornithine (Orn) and methionine (Met) levels, promoting enzyme gene expression levels and arginine decarboxylase (ADC), ornithine decarboxylase (ODC) and S-adenosylmethionine decarboxylase (SAMDC) activities in roots. Hypoxia + GABA treatment significantly increased concentrations of free putrescine (Put), spermidine (Spd) and spermine (Spm) from day two to eight, promoting the PA conversion to soluble conjugated and insoluble bound forms. However, PA degradation was significantly inhibited in hypoxia + GABA treated roots by significantly decreasing gene expression and activity of diamine oxidase (DAO) and polyamine oxidase (PAO). However, exogenous GABA showed a reduced effect in control compared with hypoxic conditions. Our data suggest that alleviating effect of exogenous GABA to hypoxia is closely associated with physiological regulation of PA metabolism. We propose a potential negative feedback mechanism of higher endogenous GABA levels from combined effects of hypoxia and exogenous GABA, which alleviate the hypoxia damage by accelerating PA biosynthesis and conversion as well as preventing PA degradation in melon plants.

  11. Effects of quercetin on the sleep-wake cycle in rats: involvement of gamma-aminobutyric acid receptor type A in regulation of rapid eye movement sleep.

    PubMed

    Kambe, Daiji; Kotani, Makiko; Yoshimoto, Makoto; Kaku, Shinsuke; Chaki, Shigeyuki; Honda, Kazuki

    2010-05-12

    The bioflavonoid quercetin is widely found in plants and exerts a large number of biological activities such as anti-hypertensive and anti-inflammatory properties. However, the effect of quercetin on the sleep-wake cycle has not been investigated. In the present study, we investigated the effect of quercetin on sleep-wake regulation. Intraperitoneal administration of quercetin (200mg/kg) significantly increased non-rapid eye movement (non-REM) sleep during dark period in rats, while it significantly decreased REM sleep. The decrease in REM sleep induced by quercetin was blocked by intracerebroventricular (i.c.v.) injection of bicuculline, a GABA(A) receptor antagonist. In contrast, the increase in non-REM sleep induced by quercetin was not affected by i.c.v. injection of bicuculline. Therefore, the present results suggest that quercetin alters the sleep-wake cycle partly through activation of GABA(A) receptors.

  12. Interaction of carvacrol with the Ascaris suum nicotinic acetylcholine receptors and gamma-aminobutyric acid receptors, potential mechanism of antinematodal action

    PubMed Central

    Marjanović, Djordje S.; Trailović, Jelena Nedeljković; Robertson, Alan P.; Martin, Richard J.

    2015-01-01

    Essential plant oils (or their active principles) are safe to use and a potentially attractive alternative to current antiparasitic drugs. In the present study, we tested the effects of carvacrol on the isolated tissues of Ascaris suum and investigated potential interactions with other antiparasitic drugs. We used somatic muscle flaps for contraction assays, as well as for electrophysiological investigations. Carvacrol 300 μM highly significantly inhibited contractions caused by 1, 3, 10, 30, and 100 μM of ACh (p=0.0023, p=0.0002, p=0.0002, p<0.0001, and p<0.0001). The control EC50 for acetylcholine was 8.87 μM (log EC50=0.95±0.26), while Rmax was 2.53±0.24 g. The EC50 of acetylcholine in the presence of 300 μM of carvacrol was 27.71 μM (log EC50=1.44±0.28) and the Rmax decreased to 1.63±0.32 g. Furthermore, carvacrol highly significant potentiates inhibitory effect of GABA and piperazine on the contractions induced by ACh. However, carvacrol (100 and 300 μM), did not produce any changes in the membrane potential or conductance of the A. suum muscle cell. While, 300 μM of carvacrol showed a significant inhibitory effect on ACh-induced depolarization response. The mean control depolarization was 13.58±0.66 mV and decreased in presence of carvacrol to 4.50±1.02 mV (p<0.0001). Mean control Δg was 0.168±0.017 μS, while in the presence of 300 μM of carvacrol, Δg significantly decreased to 0.060±0.018 ΔS (p=0.0017). The inhibitory effect on contractions may be the explanation of the antinematodal potential of carvacrol. Moreover, inhibition of depolarizations caused by ACh and reduction of conductance changes directly points to an interaction with the nAChR in A. suum. PMID:25944741

  13. Gamma-irradiation of malic acid in aqueous solutions. [prebiotic significance

    NASA Technical Reports Server (NTRS)

    Negron-Mendoza, A.; Graff, R. L.; Ponnamperuma, C.

    1980-01-01

    The gamma-irradiation of malic acid in aqueous solutions was studied under initially oxygenated and oxygen-free conditions in an attempt to determine the possible interconversion of malic acid into other carboxylic acids, specifically those associated with Krebs cycle. The effect of dose on product formation of the system was investigated. Gas-liquid chromatography combined with mass spectrometry was used as the principal means of identification of the nonvolatile products. Thin layer chromatography and direct probe mass spectroscopy were also employed. The findings show that a variety of carboxylic acids are formed, with malonic and succinic acids in greatest abundance. These products have all been identified as being formed in the gamma-irradiation of acetic acid, suggesting a common intermediary. Since these molecules fit into a metabolic cycle, it is strongly suggestive that prebiotic pathways provided the basis for biological systems.

  14. Cortical and subcortical gamma amino acid butyric acid deficits in anxiety and stress disorders: Clinical implications

    PubMed Central

    Goddard, Andrew W

    2016-01-01

    Anxiety and stress disorders are a major public health issue. However, their pathophysiology is still unclear. The gamma amino acid butyric acid (GABA) neurochemical system has been strongly implicated in their pathogenesis and treatment by numerous preclinical and clinical studies, the most recent of which have been highlighted and critical review in this paper. Changes in cortical GABA appear related to normal personality styles and responses to stress. While there is accumulating animal and human neuroimaging evidence of cortical and subcortical GABA deficits across a number of anxiety conditions, a clear pattern of findings in specific brain regions for a given disorder is yet to emerge. Neuropsychiatric conditions with anxiety as a clinical feature may have GABA deficits as an underlying feature. Different classes of anxiolytic therapies support GABA function, and this may be an area in which newer GABA neuroimaging techniques could soon offer more personalized therapy. Novel GABAergic pharmacotherapies in development offer potential improvements over current therapies in reducing sedative and physiologic dependency effects, while offering rapid anxiolysis. PMID:27014597

  15. Impact evaluation of α-lipoic acid in gamma-irradiated erythrocytes

    NASA Astrophysics Data System (ADS)

    Desouky, Omar S.; Selim, Nabila S.; Elbakrawy, Eman M.; Rezk, Rezk A.

    2011-03-01

    This work is intended to study in vitro the ability of lipoic acid to protect erythrocytes against the oxidative damage resulting from exposure to gamma radiation through measurement of their rheological properties and to study the effects of detergent on their membrane solubility and permeability. Different doses of gamma radiation were applied: the most recommended and applied dose (25 Gy), and two higher doses, namely 50 and 100 Gy. The effect of addition of lipoic acid as well as its effect as a radioprotector was tested. The obtained results show changes in structural integrity of the erythrocyte cell membrane components as a result of oxidative damage due to gamma radiation that could be improved by pre-treatment with the antioxidant lipoic acid.

  16. Safety and tolerability of gamma-hydroxybutyric acid in the treatment of alcohol-dependent patients.

    PubMed

    Beghè, F; Carpanini, M T

    2000-04-01

    Gamma-hydroxybutyric acid (GHB) has been in clinical use in Italy since 1991 for treatment of alcohol dependence. Results of phase III and phase IV studies have shown that the drug is effective and well tolerated in the treatment of alcohol withdrawal syndrome and in reducing alcohol consumption and alcohol craving. Pharmacosurveillance indicates that abuse of gamma-hydroxybutyric acid is a limited phenomenon in clinical settings when the drug is dispensed under strict medical surveillance and entrusted to a referring familiar member of the patient.

  17. Adsorption of benzoic acid on [alpha]-alumina and [gamma]-boehmite

    SciTech Connect

    Madsen, L. . Dept. of Geology and Geotechnical Engineering); Blokhus, A.M. . Dept. of Chemistry)

    1994-08-01

    The adsorption of benzoic acid (BzCOOH) on [alpha]-alumina ([alpha]-Al[sub 2]O[sub 3]) and [gamma]-boehmite ([gamma]-AlOOH) from the aqueous phase has been studied. The adsorption experiments were carried out in 0 and 0.1 M NaCl solutions, with pH adjusted to 4 or 6. For both [alpha]-alumina and [gamma]-boehmite, increasing ionic strength decreases the maximum adsorption. Increasing the pH to 6 at the same ionic strength also reduces the maximum adsorption markedly. This suggests that both the anion and the corresponding acid participate in the adsorption process. The results show that benzoic acid has a greater affinity for [alpha]-alumina than for [gamma]-boehmite. Under the same experimental conditions (0.1 M NaCl, pH 4) the maximum adsorption capacities are 5.0 and 1.5 [mu]mol/m[sup 2] for [alpha]-alumina and [gamma]-boehmite, respectively. This difference in adsorption capacities is probably due to the mineralogical difference. These results illustrate the importance of knowing the mineralogical composition of the solid phase.

  18. Radiolysis of aqueous solutions of 2-aminoethanethiosulfuric acid. [Gamma radiation

    SciTech Connect

    Grachev, S.A.; Koroleva, I.K.; Kropachev, E.V.; Litvyakova, G.I.

    1982-07-10

    In the radiolysis products of aerated and deaerated solutions of the 2-aminoethanethiosulfuric acid the authors have identified cystamine monoxide, cystamine, taurine, mercamine, the sulfate ion, the sulfite ion, and the dithionate ion. The yields of these products under different conditions have been determined. Results indicated that the sulfate ion is formed both from the divalent and the hexavalent sulfur atom of the 2-aminoethanethiosulfuric acid moelcule. A possible radiolysis mechanism is discussed.

  19. Solute-enhanced production of gamma-valerolactone (GVL) from aqueous solutions of levulinic acid

    DOEpatents

    Dumesic, James A; Wettstein, Stephanie G; Alonso, David Martin; Gurbuz, Elif Ispir

    2015-02-24

    A method to produce levulinic acid (LA) and gamma-valerolactone (GVL) from biomass-derived cellulose or lignocellulose by selective extraction of LA using GVL and optionally converting the LA so isolated into GVL, with no purifications steps required to yield the GVL.

  20. Degradation of 3-chloro-4-hydroxybenzoic acid in biological treated effluent by gamma irradiation

    NASA Astrophysics Data System (ADS)

    Chu, Libing; Wang, Jianlong

    2016-02-01

    Gamma irradiation-induced degradation of a chlorinated aromatic compound, 3-chloro-4-hydroxybenzoic acid (CHBA) in biological treated effluent was studied and the results were compared with those obtained in deionized water. Gamma irradiation led to a complete decomposition of CHBA and a partial mineralization in the treated effluent. The removal of CHBA followed the pseudo first-order reaction kinetic model and the rate constant in the treated effluent was 1.7-3.5 times lower than that in deionized water. The CHBA degradation rate was higher at acidic condition than at neutral and alkaline conditions. The radiolytic yield, G-value for CHBA degradation was lower in the treated effluent, which decreased with increase in absorbed doses and increased with increase in initial concentrations of CHBA. The degradation mechanism of CHBA using gamma irradiation was proposed through the oxidation by -OH and reduction by eaq- and H- radicals. As exposed to gamma irradiation, dechlorination takes place rapidly and combines with the oxidation and cleavage of the aromatic ring, producing chloride ions, small carboxylic acids, acetaldehyde and other intermediates into the solution.

  1. Monitoring of the interconversion of gamma-butyrolactone (GBL) to gamma hydroxybutyric acid (GHB) by Raman spectroscopy.

    PubMed

    Munshi, Tasnim; Brewster, Victoria L; Edwards, Howell G M; Hargreaves, Michael D; Jilani, Shelina K; Scowen, Ian J

    2013-08-01

    Gamma-hydroxybutyric acid (GHB) is a drug-of-abuse that has recently become associated with drug-facilitated sexual assault, known as date rape. For this reason the drug is commonly found 'spiked' in alcoholic beverages. When GHB is in solution it may undergo conversion into the corresponding lactone, Gamma-butyrolactone (GBL). Studies have been carried out to determine the detection limits of GHB and GBL in various solutions by Raman spectroscopy and to monitor the interconversion of GHB and GBL in solution with different pH conditions and temperature. In this study, a portable Raman spectrometer was used to study the interconversion of GHB and GBL in water and ethanol solutions as a function of pH, time, and temperature. The aim of this was to determine the optimum pH range for conversion in order to relate this to the pH ranges that the drug is likely to be subjected to, first in spiked beverages and secondly after ingestion in the digestive system. The aim was also to identify a timescale for this conversion in relation to possible scenarios, for example if GHB takes a number of hours to convert to GBL, it is likely for the beverage to be ingested before esterification can take place. GHB and GBL were then spiked into a selection of beverages of known pH in order to study the stability of GHB and GBL in real systems.

  2. [Current knowledge on gamma-hydroxybutyric acid (GHB), gamma-butyrolactone (GBL) and 1 ,4-butanediol (1,4-BD)].

    PubMed

    Dematteis, Maurice; Pennel, Lucie; Mallaret, Michel

    2012-05-01

    Gamma-hydroxybutyric acid (GHB) is an old anaesthetic drug which was misused in the 80-90's as an anabolic agent (bodybuilding), recreational drug (drunkenness, euphoric, disinhibiting and aphrodisiac effects) and as a date rape drug (disinhibiting, hypnotic and amnesic effects). Its use in the general population is low, and mainly concerns gay population in nightclubs and young people in parties. The intoxications, above all with alcohol combination, can be severe, with coma and breathing depression, or even fatal. Chronic use leads to psychic and physical dependence; withdrawal syndrome can be severe, with agitation and delirium. In 1999, GHB classification as a narcotic resulted in the increased use of GHB prodrugs gamma-butyrolactone (GBL) and 1,4-butanediol (1,4-BD), which were easily commercially available as solvent and cleaning products. Like GHB, they have a narrow window of use, and share similar toxicity. Their increased cases of recreational use and of severe drug intoxication, abuse and dependence, led the French Ministry of Health in 2011 to prohibit their sale and transfer to the public. PMID:22730800

  3. Monitoring of the interconversion of gamma-butyrolactone (GBL) to gamma hydroxybutyric acid (GHB) by Raman spectroscopy.

    PubMed

    Munshi, Tasnim; Brewster, Victoria L; Edwards, Howell G M; Hargreaves, Michael D; Jilani, Shelina K; Scowen, Ian J

    2013-08-01

    Gamma-hydroxybutyric acid (GHB) is a drug-of-abuse that has recently become associated with drug-facilitated sexual assault, known as date rape. For this reason the drug is commonly found 'spiked' in alcoholic beverages. When GHB is in solution it may undergo conversion into the corresponding lactone, Gamma-butyrolactone (GBL). Studies have been carried out to determine the detection limits of GHB and GBL in various solutions by Raman spectroscopy and to monitor the interconversion of GHB and GBL in solution with different pH conditions and temperature. In this study, a portable Raman spectrometer was used to study the interconversion of GHB and GBL in water and ethanol solutions as a function of pH, time, and temperature. The aim of this was to determine the optimum pH range for conversion in order to relate this to the pH ranges that the drug is likely to be subjected to, first in spiked beverages and secondly after ingestion in the digestive system. The aim was also to identify a timescale for this conversion in relation to possible scenarios, for example if GHB takes a number of hours to convert to GBL, it is likely for the beverage to be ingested before esterification can take place. GHB and GBL were then spiked into a selection of beverages of known pH in order to study the stability of GHB and GBL in real systems. PMID:23225646

  4. Ultraviolet B irradiation induces changes in the distribution and release of arachidonic acid, dihomo-gamma-linolenic acid, and eicosapentaenoic acid in human keratinocytes in culture

    SciTech Connect

    Punnonen, K.; Puustinen, T.; Jansen, C.T.

    1987-05-01

    There is increasing evidence that derivatives of 20-carbon polyunsaturated fatty acids, the eicosanoids, play an important role in the inflammatory responses of the human skin. To better understand the metabolic fate of fatty acids in the skin, the effect of ultraviolet B (UVB) irradiation (280-320 nm) on the distribution and release of /sup 14/C-labeled arachidonic acid, dihomo-gamma-linolenic acid, and eicosapentaenoic acid in human keratinocytes in culture was investigated. Ultraviolet B irradiation induced the release of all three /sup 14/C-labeled fatty acids from the phospholipids, especially from phosphatidylethanolamine, and this was accompanied by increased labeling of the nonphosphorus lipids. This finding suggests that UVB induces a significant liberation of eicosanoid precursor fatty acids from cellular phospholipids, but the liberated fatty acids are largely reincorporated into the nonphosphorus lipids. In conclusion, the present study suggests that not only arachidonic acid but also dihomo-gamma-linolenic acid, and eicosapentaenoic acid might be involved in the UVB irradiation-induced inflammatory reactions of human skin.

  5. Gamma-irradiation stability of saturated and unsaturated aliphatic polyanhydrides--ricinoleic acid based polymers.

    PubMed

    Teomim, D; Mäder, K; Bentolila, A; Magora, A; Domb, A J

    2001-01-01

    The effect of terminal sterilization by gamma-irradiation on several ricinoleic acid based polyanhydrides was investigated. The following polymers were used: poly(ricinoleic acid maleate) [P(RAM)], poly(ricinoleic acid succinate) [P(RAS)], poly(hydroxy stearic acid succinate) [P(HSAS)], poly(hydroxy stearic acid maleate) [P(HSAM)], and their copolymers with sebacic acid. The polymers were irradiated with an absorbed dose of 2.5 or 10 Mrad by means of a 60Co source under dry ice or at room temperature. No differences were found between samples irradiated under dry ice and at room temperature. Polymers prepared from monomers containing maleate residues, which contain double bonds adjusted to the anhydride linkage along the polymer chain, decreased in molecular weight, became insoluble, and showed fast hydrolytic degradation. For example, p(RAM), p(HSAM), and their copolymers with sebacic acid decreased in Mw from about 10,000 to about 2000, and from about 30,000 to about 5000, respectively, while polymers based on RAS and HSAS remained stable. This phenomenon was explained by an anhydride interchange-self-depolymerization process of the unsaturated anhydride bonds induced by gamma-irradiation. This explanation was supported by the depolymerization of another class of polymers having an anhydride bond between two double bonds, fumaric acid anhydride polymers. The anhydride bond that lies between two double bonds was found to be more sensitive to gamma-irradiation. This anhydride bond may be cleaved to form two radicals that further react with aliphatic anhydride bonds along the polymer chain to form inter- and/or intracyclization products. PMID:11710004

  6. Gamma-amino butyric acid inhibits the nicotine-imposed stimulatory challenge in xenograft models of non-small cell lung carcinoma.

    PubMed

    Al-Wadei, H A N; Al-Wadei, M H; Ullah, M F; Schuller, H M

    2012-02-01

    Non-small cell lung carcinoma (NSCLC) is the leading type of lung cancer; smoking is a documented risk factor. Nicotinic acetylcholine receptor (nAChR)-mediated intracellular signaling in response to nicotine has recently been implicated in the growth regulation of NSCLC. In the current study nude mice carrying xenografts of the human lung NSCLC cell lines NCI-H322 or NCI-H441 were used as animal models. Nicotine administration and gamma aminobutyric acid (GABA) treatment lasted for 30 days. Catecholamines, cortisol, GABA, and cAMP were analyzed in blood and tumor tissues by immunoassays. Expression of nicotinic receptors and effector proteins in the xenografts was assessed by Western blotting. Our data indicate that nicotine stimulated the growth of NSCLC xenografts via modulation of nAChR upregulation and activation of cAMP signaling. The nicotine-treated group showed an enhanced level of stress neurotransmitters and second messenger cAMP in serum, blood cellular fraction, and xenograft tissues. Activation of critical proteins in the oncogenic pathway, including CREB, ERK, Akt, and Src, and upregulation of α-4 and α-7 subunits of nAChR provided mechanistic insight for the observed stimulatory effect of nicotine. Interestingly, GABA, being an antagonist to cAMP signaling, showed a promising intervention by reversing the stimulatory effect of nicotine on cancer growth and all signaling pathways. GABA has potential to lower the risk of NSCLC among smokers and could be used to enhance the clinical outcome of standard cancer intervention strategies.

  7. Peptide nucleic acid (PNA) binding-mediated induction of human gamma-globin gene expression.

    PubMed

    Wang, G; Xu, X; Pace, B; Dean, D A; Glazer, P M; Chan, P; Goodman, S R; Shokolenko, I

    1999-07-01

    Peptide nucleic acids (PNAs) can bind to homopurine/homopyrimidine sequences of double-stranded DNA targets in a sequence-specific manner and form [PNA]2/DNA triplexes with single-stranded DNA D-loop structures at the PNA binding sites. These D-loop structures have been found to have a capacity to initiate transcription in vitro. If this strategy can be used to induce transcription of endogenous genes, it may provide a novel approach for gene therapy of many human diseases. Human [beta] globin disorders such as sickle cell anemia and beta-thalassemia are very common genetic diseases that are caused by mutations in the beta-globin gene. When gamma-globin genes are highly expressed in sickle cell patients, the presence of high levels of fetal hemoglobin (HbF, alpha2gamma2) can compensate for the defective beta-globin gene product and such patients have much improved symptoms or are free of disease. However, the gamma-globin genes are developmentally regulated and normally expressed at very low levels (>1%) in adult blood cells. We have investigated the possibility of inducing gamma-globin gene expression with PNAs. Using PNAs designed to bind to the 5' flanking region of the gamma-globin gene, induction of expression of a reporter gene construct was demonstrated both in vitro and in vivo. More importantly, PNA-mediated induction of endogenous gamma-globin gene expression was also demonstrated in K562 human erythroleukemia cells. This result suggests that induction of gamma-globin gene expression with PNAs might provide a new approach for the treatment of sickle cell disease. PNA-induced gene expression strategy also may have implications in gene therapy of other diseases such as genetic diseases, cancer and infectious diseases.

  8. Effect of gamma-linolenic acid and dihomo-gamma-linolenic acid 7,12-dimethylbenz(a)anthracene-induced mammary tumorigenesis

    SciTech Connect

    Ramchurren, N.; Karmali, R.A. )

    1991-03-15

    Gamma-linolenic acid (GLA) and its sequential metabolite, dihomogamma-linolenic acid (DHLA) have been reported to influence growth of neoplastic cells in culture. The pure forms of these fatty acids have not been tested in vivo. The authors have studied the effect of GLA and DHLA on mammary tumors induced by 7,12-dimethylbenz(a) anthracene (DMBA) (7.5 mg/rat) in female Sprague-Dawley rats. Rats received either 0.15 g of GLA or DHLA or corn oil (CO) orally, twice weekly for a period of 12 weeks. All three groups of rats were maintained on a diet containing 5% (w/w) corn oil as fat. Tumor incidence and multiplicity were recorded. The group receiving 0.15 g Co had higher tumor yields than those receiving GLA or DHLA. At the end of the experiment, tumor incidence was the lowest in the group receiving DHLA. Tumor multiplicity was consistently lowest with GLA. Fatty acid composition of mammary tissue and liver reflected that of fatty acid treatment. These results suggest that oral administration of GLA or DHLA retards the development of DMBA-induced mammary tumors in rats receiving a diet containing 5% (w/w) corn oil.

  9. Antioxidant activities of fucoidan degraded by gamma irradiation and acidic hydrolysis

    NASA Astrophysics Data System (ADS)

    Lim, Sangyong; Choi, Jong-il; Park, Hyun

    2015-04-01

    Low molecular weight fucoidan, prepared by radical degradation using gamma ray was investigated for its antioxidant activities with different assay methods. As the molecular weight of fucoidan decreased with a higher absorbed dose, ferric-reducing antioxidant power values increased, but β-carotene bleaching inhibition did not change significantly. The antioxidant activity of acid-degraded fucoidan was also examined to investigate the effect of different degradation methods. At the same molecular weight, fucoidan degraded by gamma irradiation showed higher 1,1-diphenyl-2-picrylhydrazyl radical scavenging activity than that observed with the acidic method. This result reveals that in addition to molecular weight, the degradation method affects the antioxidant activity of fucoidan.

  10. Discovery of Azetidinone Acids as Conformationally-Constrained Dual PPARalpha/gamma Agonists

    SciTech Connect

    Wang, W.; Devasthale, P; Farrelly, D; Gu, L; Harrity, T; Cap, M; Chu, C; Kunselman, L; Morgan, N; et. al.

    2008-01-01

    A novel class of azetidinone acid-derived dual PPAR{alpha}/{gamma} agonists has been synthesized for the treatment of diabetes and dyslipidemia. The preferred stereochemistry in this series for binding and functional agonist activity against both PPARa and PPAR? receptors was shown to be 3S,4S. Synthesis, in vitro and in vivo activities of compounds in this series are described. A high-yielding method for N-arylation of azetidinone esters is also described.

  11. A heavy metal biotrap for wastewater remediation using poly-gamma-glutamic acid.

    PubMed

    Mark, Sonny S; Crusberg, Theodore C; Dacunha, Christopher M; Di Iorio, Alexander A

    2006-01-01

    Poly-gamma-glutamic acid (gamma-PGA) obtained from Bacillus licheniformis ATCC 9945 was evaluated as a potential biosorbent material for use in the removal of heavy metals from aqueous solution. Copper (Cu(2+)) was chosen as the model heavy metal used in these studies since it is extensively used by electroplating and other industries, has been the model for many other similar studies, and can be easily assayed through a number of convenient methods. Cu(2+)-gamma-PGA binding parameters under varying conditions of pH, temperature, ionic strength, and in the presence of other heavy metal ions were determined for the purified biopolymer using a specially designed dialysis apparatus. Applying the Langmuir adsorption isotherm model showed that gamma-PGA had a copper capacity approaching 77.9 mg/g and a binding constant of 32 mg/L (0.5 mM) at pH 4.0 and 25 degrees C. Cu(2+)-gamma-PGA adsorption was relatively temperature independent between 7 and 40 degrees C, while an increase in ionic strength led to a decrease in metal ion binding. Cd(2+) and Zn(2+) ions compete with Cu(2+) for binding sites on the gamma-PGA biopolymer. Metal uptake by gamma-PGA was further tested using a tangential flow filtration apparatus in a diafiltration mode in which metal was continually processed through a dilute solution of gamma-PGA without allowing for equilibrium to be established. The circulating polymer solution was able to complex metal as well as successfully prevent passage of unbound copper ions present in solution through the membrane. Using 500 mL of a 0.2% gamma-PGA solution, up to 97% of a 50 mg/L copper sulfate solution processed at a flow rate of 115 mL/min was retained by the polymer. For a 10 mg/L solution of Cu(2+) as copper sulfate, filtrate concentrations of Cu(2+) never rose above 0.6 mg/L while processing 2.5 L of dilute copper sulfate.

  12. Fatty acid profile of gamma-irradiated and cooked African oil bean seed (Pentaclethra macrophylla Benth)

    PubMed Central

    Olotu, Ifeoluwa; Enujiugha, Victor; Obadina, Adewale; Owolabi, Kikelomo

    2014-01-01

    The safety and shelf-life of food products can be, respectively, ensured and extended with important food-processing technologies such as irradiation. The joint effect of cooking and 10 kGy gamma irradiation on the fatty acid composition of the oil of Pentaclethra macrophylla Benth was evaluated. Oils from the raw seed, cooked seeds, irradiated seeds (10 kGy), cooked, and irradiated seeds (10 kGy) were extracted and analyzed for their fatty acid content. An omega-6-fatty acid (linoleic acid) was the principal unsaturated fatty acid in the bean seed oil (24.6%). Cooking significantly (P < 0.05) increased Erucic acid by 3.3% and Linolenic acid by 23.0%. Combined treatment significantly (P < 0.05) increased C18:2, C6:0, C20:2, C18:3, C20:3, C24:0, and C22:6 being linoleic, caproic, eicosadienoic, linolenic, eicosatrienoic, ligoceric, and docosahexaenoic acid, respectively, and this increase made the oil sample to have the highest total fatty acid content (154.9%), unsaturated to saturated fatty acid ratio (109.6), and unsaturated fatty acid content (153.9%). 10 kGy irradiation induces the formation of C20:5 (eicosapentaenoic), while cooking induced the formation of C20:4 (arachidic acid), C22:6 (Heneicosanoic acid), and C22:2 (docosadienoic acid). Combined 10 kGy cooking and irradiation increased the susceptibility of the oil of the African oil bean to rancidity. PMID:25493197

  13. Fatty acid profile of gamma-irradiated and cooked African oil bean seed (Pentaclethra macrophylla Benth).

    PubMed

    Olotu, Ifeoluwa; Enujiugha, Victor; Obadina, Adewale; Owolabi, Kikelomo

    2014-11-01

    The safety and shelf-life of food products can be, respectively, ensured and extended with important food-processing technologies such as irradiation. The joint effect of cooking and 10 kGy gamma irradiation on the fatty acid composition of the oil of Pentaclethra macrophylla Benth was evaluated. Oils from the raw seed, cooked seeds, irradiated seeds (10 kGy), cooked, and irradiated seeds (10 kGy) were extracted and analyzed for their fatty acid content. An omega-6-fatty acid (linoleic acid) was the principal unsaturated fatty acid in the bean seed oil (24.6%). Cooking significantly (P < 0.05) increased Erucic acid by 3.3% and Linolenic acid by 23.0%. Combined treatment significantly (P < 0.05) increased C18:2, C6:0, C20:2, C18:3, C20:3, C24:0, and C22:6 being linoleic, caproic, eicosadienoic, linolenic, eicosatrienoic, ligoceric, and docosahexaenoic acid, respectively, and this increase made the oil sample to have the highest total fatty acid content (154.9%), unsaturated to saturated fatty acid ratio (109.6), and unsaturated fatty acid content (153.9%). 10 kGy irradiation induces the formation of C20:5 (eicosapentaenoic), while cooking induced the formation of C20:4 (arachidic acid), C22:6 (Heneicosanoic acid), and C22:2 (docosadienoic acid). Combined 10 kGy cooking and irradiation increased the susceptibility of the oil of the African oil bean to rancidity.

  14. Gamma-hydroxybutyric acid affects the acquisition and reinstatement of cocaine-induced conditioned place preference in mice.

    PubMed

    Maldonado, Concepción; Rodríguez-Arias, Marta; Castillo, Ana; Aguilar, María A; Miñarro, José

    2006-03-01

    Cocaine addicts very often use different combinations of cocaine and other drugs of abuse such as gamma-hydroxybutyric acid. The objective of the present work was to evaluate the impact of gamma-hydroxybutyric acid administration on the rewarding actions of cocaine, using the conditioned place preference procedure. Cocaine-induced conditioned place preference (50 mg/kg) was studied after pairing this drug with different gamma-hydroxybutyric acid doses (6.25, 12.5, 25, 50 and 100 mg/kg) during either the acquisition or the expression phase of the procedure. After conditioned place preference had been established, and the preference was extinguished, a reinstatement was induced by a dose of cocaine half of that used to produce conditioning, or by gamma-hydroxybutyric acid alone or by both drugs together. The doses of 12.5 and 100 mg/kg of gamma-hydroxybutyric acid blocked the acquisition of cocaine-induced conditioned place preference, and no dose affected the expression of this conditioning. Reinstatement was abolished only with the dose of 25 mg/kg gamma-hydroxybutyric acid, which did not reinstate the preference by itself. This is the first study evaluating the effects of gamma-hydroxybutyric acid on the rewarding properties of cocaine using the conditioned place preference procedure. The principal conclusion of the study is that gamma-hydroxybutyric acid does not enhance the rewarding effect of cocaine, and within a narrow margin of effective doses, blocks the acquisition and reinstatement of cocaine-induced preference.

  15. Two proteins with gamma-carboxyglutamic acid in frog bone: isolation and comparative characterization.

    PubMed

    Dohi, Y; Iwami, K; Yonemasu, K; Moriyama, T

    1987-10-15

    Two gamma-carboxyglutamic acid-containing proteins were purified from neutral (pH 7.5) EDTA-extract of frog, Rana catesbiana, cortical bone by Sephadex G-75 gel filtration, DEAE-Sephadex A-25 chromatography and successive hydroxyapatite column chromatography. These two bone gamma-carboxyglutamic acid-containing proteins, termed osteocalcin, P-1 and P-2, had molecular weights of about 5100 and 4900, respectively, based on their amino-acid composition. Both species of osteocalcin have two gamma-carboxyglutamic acid residues, one disulfide bond, but there was no 4-hydroxyproline in either molecule. Each N-terminus of both proteins was acetylated and each C-terminal amino acid was lysine. The isoelectric points of P-1 and P-2 are 4.02 and 3.91, respectively, and their pI values shifted to more neutral pH in the presence of calcium ions. Equilibrium dialysis has indicated that each of these two proteins binds specifically 2 mol Ca2+, and nonspecifically more, 4-5 mol, Ca2+ in 0.02 M Tris-HCl/0.15 M NaCl (pH 7.4), at 4 degrees C. By the best-fitted calculation, P-1 had one high affinity Ca2+-binding site (Kd1 = 0.17 mM) and one lower affinity site (Kd2 = 0.29 mM), and P-2 contained one high affinity site (Kd1 = 0.154 mM) and one lower affinity site (Kd2 = 0.67 mM). PMID:2443180

  16. Synthesis and characterization of zinc chloride containing poly(acrylic acid) hydrogel by gamma irradiation

    NASA Astrophysics Data System (ADS)

    Park, Jong-Seok; Kuang, Jia; Gwon, Hui-Jeong; Lim, Youn-Mook; Jeong, Sung-In; Shin, Young-Min; Seob Khil, Myung; Nho, Young-Chang

    2013-07-01

    In this study, the characterization of zinc chloride incorporated into a poly(acrylic acid) (PAAc) hydrogel prepared by gamma-ray irradiation was investigated. Zinc chloride powder with different concentrations was dissolved in the PAAc solution, and it was crosslinked with gamma-ray irradiation. The effects of various parameters such as zinc ion concentration and irradiation doses on characteristics of the hydrogel formed were investigated in detail for obtaining an antibacterial wound dressing. In addition, the gel content, pH-sensitive (pH 4 or 7) swelling ratio, and UV-vis absorption spectra of the zinc particles in the hydrogels were characterized. Moreover, antibacterial properties of these new materials against Staphylococcus aureus and Escherichia coli strains were observed on solid growth media. The antibacterial tests indicated that the zinc chloride containing PAAc hydrogels have good antibacterial activity.

  17. [Effect of synthetic cyclopentane beta,beta'-triketones on amino acid metabolism in roots of buckwheat (Fagopyrum esculentum Moench.) seedlings].

    PubMed

    Demina, E A; Tishchenko, L Ia; Shestak, O P; Novikov, V L; Anisimov, M M

    2009-01-01

    Germination of buckwheat seeds in solutions of synthetic mono- and tricyclic cyclopentane-containing beta,beta'-triketones of various concentrations was accompanied by inhibition of seedling root growth and changes in the contents of glutamate, gamma-aminobutyric acid, proline, glutamine, and alanine. The monocyclic triketone also affected the amount of isoleucine. It is likely that the increase in proline content is a nonspecific response significant for enhancing stress tolerance in seedlings.

  18. Gamma-linolenic acid enrichment from Borago officinalis and Echium fastuosum seed oils and fatty acids by low temperature crystallization.

    PubMed

    López-Martínez, Juan Carlos; Campra-Madrid, Pablo; Guil-Guerrero, José Luis

    2004-01-01

    Solvent winterization of seed oil and free fatty acids (FFAs) was employed to obtain gamma-linolenic acid (GLA; 18:3omega6) concentrates from seed oils of two Boraginaceae species, Echium fastuosum and Borago officinalis. Different solutions of seed oils and FFAs from these two oils at 10%, 20% and 40% (w/w) were crystallized at 4 degrees C, -24 degrees C and -70 degrees C, respectively, using hexane, acetone, diethyl ether, isobutanol and ethanol as solvents. Best results were obtained for B. officinalis FFAs in hexane, reaching a maximum GLA concentration of 58.8% in the liquid fraction (LF). In E. fastuosum, the highest GLA concentration (39.9%) was also achieved with FFAs in hexane. PMID:16233632

  19. Gamma-linolenic acid enrichment from Borago officinalis and Echium fastuosum seed oils and fatty acids by low temperature crystallization.

    PubMed

    López-Martínez, Juan Carlos; Campra-Madrid, Pablo; Guil-Guerrero, José Luis

    2004-01-01

    Solvent winterization of seed oil and free fatty acids (FFAs) was employed to obtain gamma-linolenic acid (GLA; 18:3omega6) concentrates from seed oils of two Boraginaceae species, Echium fastuosum and Borago officinalis. Different solutions of seed oils and FFAs from these two oils at 10%, 20% and 40% (w/w) were crystallized at 4 degrees C, -24 degrees C and -70 degrees C, respectively, using hexane, acetone, diethyl ether, isobutanol and ethanol as solvents. Best results were obtained for B. officinalis FFAs in hexane, reaching a maximum GLA concentration of 58.8% in the liquid fraction (LF). In E. fastuosum, the highest GLA concentration (39.9%) was also achieved with FFAs in hexane.

  20. Delta-6 desaturase from borage converts linoleic acid to gamma-linolenic acid in HEK293 cells.

    PubMed

    Chen, Qing; Nimal, Jonathan; Li, Wanli; Liu, Xia; Cao, Wenguang

    2011-07-01

    Gamma-linolenic acid (GLA, 18:3 n6) is an essential polyunsaturated fatty acid of the omega-6 family and is found to be effective in prevention and/or treatment of various health problems. In this study, we evaluated the possibility of increasing γ-linolenic acid contents in mammalian cells using the delta-6 gene from Borago officinalis. The borage Δ6-desaturase gene (sDelta-6) was codon-optimized and introduced into HEK293 cells by lipofectin transfection. Co-expression of GFP with sDelta-6 and RT-PCR analysis indicated that sDelta-6 could be expressed in mammalian cells. Subsequently, the heterologous expression of borage Δ6-desaturase was evaluated by fatty acid analysis. Total cellular lipid analysis of transformed cells fed with linoleic acid (LA 18:2 n6) as a substrate showed that the expression of sDelta-6 resulted in an 228-483% (p<0.05) increase of GLA when compared with that in the control cells. The highest conversion efficiency of LA into GLA in sDelta-6(+) cells was 6.9 times higher than that in the control group (11.59% vs. 1.69%; p<0.05). Our present work demonstrated that the sDelta-6 gene from borage could be functionally expressed in mammalian cells, and could convert LA into GLA. Furthermore, this study may pave the way to generate transgenic livestock that can synthesise GLA.

  1. Effect of gamma irradiation on hyaluronic acid and dipalmitoylphosphatidylcholine (DPPC) interaction

    SciTech Connect

    Ahmad, Ainee Fatimah; Mohd, Hur Munawar Kabir; Taqiyuddin Mawardi bin Ayob, Muhammad; Rosli, Nur Ratasha Alia Md; Mohamed, Faizal; Radiman, Shahidan; Rahman, Irman Abdul

    2014-09-03

    DPPC lipids are the major component constituting the biological membrane, and their importances in various physiological functions are well documented. Hyaluronic acid (HA) in the synovial joint fluid functions as a lubricant, shock absorber and a nutrient carrier. Gamma irradiation has also been found to be effective in depolymerizing and cleaving molecular chains related to free radicals, thus extends with changes in chemical composition as well as its physiological functions. This research are conducted to investigate the hyaluronic acid (HA) and 1,2-dipalmitoyl-sn-glycero-3-phosphatidylcholine (DPPC) interaction in form of vesicles and its effect to gamma radiation. The size of DPPC vesicles formed via gentle hydration method is between 100 to 200 nm in diameter. HA (0.1, 0.5 and 1.0 mg/ml) was added into the vesicles and characterized by using TEM to determine vesicle size distributions, fusion and rupture of DPPC structure. The results demonstrated that the size of the vesicles approximately between 200 to 300 nm which caused by vesicles fusion with HA and formed even larger vesicles. After being irradiated by 0 to 200 Gy, the size of vesicles decreased as HA was degraded. To elucidate the mechanism of these effects, FTIR spectra were carried out and have shown that at absorption bands at 1700–1750 cm{sup −1} due to formation of carboxylic acid and leads to alteration of HA structure.

  2. Effect of gamma irradiation on hyaluronic acid and dipalmitoylphosphatidylcholine (DPPC) interaction

    NASA Astrophysics Data System (ADS)

    Ahmad, Ainee Fatimah; Mohd, Hur Munawar Kabir; bin Ayob, Muhammad Taqiyuddin Mawardi; Rosli, Nur Ratasha Alia Md; Mohamed, Faizal; Radiman, Shahidan; Rahman, Irman Abdul

    2014-09-01

    DPPC lipids are the major component constituting the biological membrane, and their importances in various physiological functions are well documented. Hyaluronic acid (HA) in the synovial joint fluid functions as a lubricant, shock absorber and a nutrient carrier. Gamma irradiation has also been found to be effective in depolymerizing and cleaving molecular chains related to free radicals, thus extends with changes in chemical composition as well as its physiological functions. This research are conducted to investigate the hyaluronic acid (HA) and 1,2-dipalmitoyl-sn-glycero-3-phosphatidylcholine (DPPC) interaction in form of vesicles and its effect to gamma radiation. The size of DPPC vesicles formed via gentle hydration method is between 100 to 200 nm in diameter. HA (0.1, 0.5 and 1.0 mg/ml) was added into the vesicles and characterized by using TEM to determine vesicle size distributions, fusion and rupture of DPPC structure. The results demonstrated that the size of the vesicles approximately between 200 to 300 nm which caused by vesicles fusion with HA and formed even larger vesicles. After being irradiated by 0 to 200 Gy, the size of vesicles decreased as HA was degraded. To elucidate the mechanism of these effects, FTIR spectra were carried out and have shown that at absorption bands at 1700-1750 cm-1 due to formation of carboxylic acid and leads to alteration of HA structure.

  3. 14C dating of bone using (gamma) Carboxyglutamic Acid and Carboxyglycine (Aminomalonate)

    SciTech Connect

    Southon, J R; Burky, R T; Kirner, D L; Taylor, R E; Hare, P E

    1999-04-27

    Radiocarbon determinations have been obtained on {gamma}-carboxyglutamic acid [Gla] and {alpha}-carboxyglycine (aminomalonate) [Am] as well as acid- and base-hydrolyzed total amino acids isolated from a series of fossil bones. As far as they are aware, Am has not been reported previously in fossil bone and neither Gla nor Am {sup 14}C values have been measured previously. Interest in Gla, an amino acid found in the non-collagen proteins osteocalcin and matrix Gla-protein (MGP), proceeds from the suggestion that it may be preferentially retained and more resistant to diagenetic contamination affecting {sup 14}C values in bones exhibiting low and trace amounts of collagen. The data do not support these suggestions. The suite of bones examined showed a general tendency for total amino acid and Gla concentrations to decrease in concert. Even for bones retaining significant amounts of collagen, Gla (and Am extracts) can yield {sup 14}C values discordant with their expected age and with {sup 14}C values obtained on total amino-acid fractions isolated from the same bone sample.

  4. Biosynthesis of gamma-linolenic acid and beta-carotene by Zygomycetes fungi.

    PubMed

    Klempova, Tatiana; Basil, Eva; Kubatova, Alena; Certik, Milan

    2013-07-01

    Due to increasing demand for natural sources of both polyunsaturated fatty acids (PUFAs) and beta-carotene, 28 Zygomycetes fungal soil isolates were screened for their potential to synthesize these biologically active compounds. Although all fungi produced C18 PUFAs, only nine strains also formed beta-carotene. Although Actinomucor elegans CCF 3218 was the best producer of gamma-linolenic acid (GLA) (251 mg/L), Umbelopsis isabellina CCF 2412 was found to be the most valuable fungus because of the dual production of GLA (217 mg/L) and beta-carotene (40.7 mg/L). The calculated ratio of formed PUFAs provided new insight into activities of individual fatty acid desaturases involved in biosynthetic pathways for various types of PUFAs. The maximal activity of delta-9 desaturase was accompanied by high accumulation of storage lipids in fungal cells. On the other hand, maximal activity of delta-15 desaturase was found in strains synthesizing low amounts of oleic acid due to diminished delta-9 desaturase. Activities of delta-6 desaturase showed competition for fatty acids engaged in n3, n6, and n9 biosynthetic pathways. Such knowledge about fatty acid desaturase activities provides new challenges for the regulation of biotechnological production of PUFAs by Zygomycetes fungi. PMID:23625863

  5. Metal ion blockage of tritium incorporation into gamma-carboxyglutamic acid of prothrombin. Stoichiometry of gamma-carboxyglutamic acid to Gd3+ for the high affinity sites

    SciTech Connect

    Bajaj, S.P.; Saini, R.; Katz, A.; Cai, G.Z.; Maki, S.L.; Brodsky, G.L.

    1988-07-15

    Prothrombin possesses two high affinity and four low affinity gamma-carboxyglutamic acid (Gla)-dependent gadolinium binding sites. Earlier work has shown that tritium can be specifically incorporated at the gamma-carbon of Gla in proteins at pH 5. In the present work we show that inclusion of saturating concentrations of Ca2+ in nondenaturing buffer systems ranging from pH 5.5 to 8.5 prevents the exchange of tritium into all 10 Gla residues of prothrombin. Similarly, saturating concentrations of Gd3+ prevent tritium incorporation into Gla at pH 5.5. Positive cooperativity was observed for the binding of Gd3+ to human prothrombin (at pH 5.5) for the two high affinity sites (Kd congruent to 35 nM). The four low affinity sites bind Gd3+ with a Kd congruent to 5 microM. Incubation of prothrombin ranging in concentrations from 10 to 40 microM with 2 eq of Gd3+ at pH 5.5 prevents 5.7 (average of seven determinations) Gla residues from tritium incorporation. Sedimentation velocity experiments conducted at pH 5.5 indicate that prothrombin in the presence of saturating concentrations of Gd3+ polymerizes, most likely, to a trimer. Further, in the presence of 2 eq of Gd3+, calculated percent weight average concentration of monomer prothrombin is congruent to 100% at 10 microM, approximately equal to 95% at 20 microM, and congruento to 80% at 40 microM protein concentration. Thus, it appears that under conditions in which prothrombin primarily exists as a monomer, occupancy of the initial two metal binding sites by Gd3+ involves six Gla residues.

  6. Furan formation from fatty acids as a result of storage, gamma irradiation, UV-C and heat treatments

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Furan is a possible human carcinogen that has been found in many thermally processed foods. The effects of thermal processing, gamma and UV-C irradiation on formation of furan from different fatty acids was studied. In addition, formation of furan from fatty acid emulsions during storage at 25C and...

  7. Biosynthesis of gamma-linolenic acid in cotyledons and microsomal preparations of the developing seeds of common borage (Borago officinalis).

    PubMed

    Stymne, S; Stobart, A K

    1986-12-01

    The developing seeds of Borago officinalis (common borage) accumulate a triacylglycerol oil that is relatively rich in the uncommon fatty acid gamma-linolenate (octadec-6,9,12-trienoic acid). Incubation of developing, whole, cotyledons with [14C]oleate and [14C]linoleate showed that the gamma-linolenate was synthesized by the sequential desaturation of oleate----linoleate----gamma-linolenate. Microsomal membrane preparations from the developing cotyledons contained an active delta 6-desaturase enzyme that catalysed the conversion of linoleate into gamma-linolenate. Experiments were designed to manipulate the [14C]linoleate content of the microsomal phosphatidylcholine. The [14C]linoleoyl phosphatidylcholine labelled in situ was converted into gamma-linolenoyl phosphatidylcholine in the presence of NADH. The substrate for the delta 6-desaturase in borage was, therefore, the linoleate in the complex microsomal lipid phosphatidylcholine, rather than, as in animals, the acyl-CoA. This was further confirmed in experiments that compared the specific radioactivity of the gamma-linolenate, in acyl-CoA and phosphatidylcholine, that was synthesized when [14C]linoleoyl-CoA was incubated with microsomal membranes, NADH and non-radioactive gamma-linolenoyl-CoA. The delta 6-desaturase was positionally specific and only utilized the linoleate in position 2 of sn-phosphatidylcholine. Analysis of the positional distribution of fatty acids in the endogenous microsomal sn-phosphatidylcholine showed that, whereas position 1 contained substantial linoleate, only small amounts of gamma-linolenate were present. The results shed further light on the synthesis of C18 polyunsaturated fatty acids in plants and in particular its relationship to the regulation of the acyl quality of the triacylglycerols in oilseeds.

  8. Direct analysis reveals an absence of gamma-carboxyglutamic acid in cancer procoagulant from human tissues.

    PubMed

    Kaplinska, Katarzyna; Mielicki, Wojciech P

    2009-07-01

    Additional carboxylation of glutamic acid by vitamin K-dependent gamma-carboxylase is a common posttranslational modification of many proteins, including some of blood clotting factors. Vitamin K-antagonists, such as warfarin, are often included in the therapy of malignant disease, decreasing the blood coagulation potential. Cancer procoagulant, a direct blood coagulation factor X activator from malignant tissue, is considered as a vitamin K-dependent protein, so it could serve as one of possible targets for the therapy with warfarin. However, there is still no experimental data demonstrating directly the presence of gamma-carboxyglutamic acid (Gla) in a cancer procoagulant molecule. The presence of Gla in cancer procoagulant isolated from human amnion-chorion membranes and from human malignant melanoma WM 115 cell line was analyzed directly, using specific anti-Gla monoclonal antibodies. There was no detectable amount of Gla in cancer procoagulant isolated from fetal or malignant tissue. Cancer procoagulant from human tissues does not contain Gla-rich domain. The finding indicates that cancer procoagulant is rather a poor target for warfarin therapy of malignant disease.

  9. Gamma-irradiation effect on a commercial composite anticorrosive pigment and acidity-to-alkalinity conversion

    NASA Astrophysics Data System (ADS)

    Song, Weiqiang; Niu, Kaihui; Wu, Longchao

    2016-05-01

    A commercial composite anticorrosive pigment based on aluminum dihydrogen tripolyphosphate was studied after exposure to gamma irradiation (Co60, 0, 20, 50, 100 and 150 kGy) using FTIR, XRD, TGA and acid-base titration technologies. Although the FTIR spectra showed that the effect of the irradiation on functional groups in the pigments was not obvious, the decrease in the crystal lattice parameters of the irradiated pigments was observed in the XRD spectra compared to the non-irradiated sample. But the extent of the lattice parameter decrease monotonically with the increase of absorbed dose from 20 to 150 kGy, which was attributed to the decomposition of water and the simultaneous occurrence of lattice damage when the pigments were exposed to gamma rays. Of particular significance was the displayed basicity of the aqueous solutions of the irradiated pigments compared to the acidity of the solution of the non-irradiated pigment, which was attributed to the decomposition of P-OH groups (combined water).

  10. Genetic manipulation of gamma-linolenic acid (GLA) synthesis in a commercial variety of evening primrose (Oenothera sp.).

    PubMed

    de Gyves, Emilio Mendoza; Sparks, Caroline A; Sayanova, Olga; Lazzeri, Paul; Napier, Johnathan A; Jones, Huw D

    2004-07-01

    A robust Agrobacterium-mediated transformation procedure was developed for Rigel, a commercial cultivar of evening primrose, and used to deliver a cDNA encoding a Delta(6)-desaturase from borage under the control of a cauliflower mosaic virus (CaMV) 35S promoter. Analysis of the transformed plants demonstrated an altered profile of polyunsaturated fatty acids, with an increase in gamma-linolenic acid and octadecatetraenoic acid in leaf tissues when compared with control lines. PMID:17134396

  11. Formation of hydrocarbons from acid-Clay suspensions by gamma irradiation

    SciTech Connect

    Cruz-Castaneda, J.; Negron-Mendoza, A.; Ramos-Bernal, S.

    2013-07-03

    The adsorption of certain organic compounds by clays gives rise to the transformation of the adsorbate through the action of the clays. This phenomenon can be enhanced using ionizing radiation. In this context, these kinds of reactions play an important role in many natural and industrial processes. For example, in oil and gas exploration, the source and trap of petroleum hydrocarbons is frequently clay-rich rocks. Clay-water-based muds are also seen as environmentally friendly alternatives to toxic oil-based fluids. The principal processes that occur in sediments are usually held to be of bacterial action and thermal transformation, which may include thermally induced catalytic alteration of the organic debris. On the other hand, radioactive materials are widely distributed throughout Earth. They were more abundant in the past, but are present in petroleum reservoirs. Their presence induced radioactive bombardment, which may have altered these sediments. This important subject has not been extensively studied. The aim of this work is to study the behavior of fatty acids-like behenic acid-and dicarboxylic acids-like fumaric acid-as model compounds, which are adsorbed in a clay mineral (Na-montmorillonite) and exposed to gamma radiation. The results show that the radiation-induced decomposition of the clay-acid system goes along a definitive path (oxidation), rather than following several modes of simultaneous decomposition, as happens in radiolysis without clay or by heating the system. The main radiolytic products for fatty acids are their corresponding hydrocarbons, with one C-atom less than the original acid.

  12. [Expression of retinoic acid receptor gamma gene in ES cells and its effect on their differentiation and apoptosis].

    PubMed

    Du, Z W; Tsung, H C; Yao, Z

    1998-06-01

    We have constructed pSG5-RAR gamma-neo plasmid containing mouse retinoic acid receptor gamma (RAR gamma) gene and neo gene, and introduced it into embryonic stem ES-5 cells, by calcium phosphate mediated transfection. Some G418-resistant clones were isolated and from RNA dot blot analysis of these clones, a clone overexpressing RAR gamma gene was established, designated as ES-gamma cell line. Northern blot hydridization and Southern blot hydridization analysis of ES-gamma cells (Fig 3, 4) demonstrated that ES-gamma cells overexpressed exogenous RAR gamma mRNA and the exogenous RAR gamma cDNA integrated into the genome of ES cells. ES-gamma cells retained undifferentiated morphology and positive alkaline phosphatase activity (Plate I, Fig. 1, 2), so it resembled ES-5 cells in terms of stem cell characteristics. When ES-gamma cells were subcutaneously inoculated into nude mouse and differentiated in vivo, tumorous nodules containing various tissue structures were obtained, demonstrating their pluripotent properties just like parent ES-5 cells. Contrasting with ES-5 cells, the histological features of tumors showed no cartilage tissues, but abundant muscle tissues and keratinized cyst like structures constituted by stratified squamous epithelia (Plate I, Fig. 3). Differentiating in vitro by hanging drop culture methods, ES-gamma cells differentiated mostly into fibroblast-like cells, (Plate II, Fig. 1-5). The above results indicated that overexpression of RAR gamma gene changed the cell type of ES cells differentiating in vivo and in vitro. During the differentiation of ES-5 cells induced by RA, a large number of cells rounded up, detached from the dish and tended to die. We suspected that this phenomenon may be apoptosis. The ultrastructure appearance of the dying cells displayed typical apoptotic changes including chromatin condensation and nuclear fragmentation (Plate I, Fig. 4, 5). Detection of DNA fragments using agarose gel electrophoresis showed characteristic

  13. Delta-6 desaturase from borage converts linoleic acid to gamma-linolenic acid in HEK293 cells.

    PubMed

    Chen, Qing; Nimal, Jonathan; Li, Wanli; Liu, Xia; Cao, Wenguang

    2011-07-01

    Gamma-linolenic acid (GLA, 18:3 n6) is an essential polyunsaturated fatty acid of the omega-6 family and is found to be effective in prevention and/or treatment of various health problems. In this study, we evaluated the possibility of increasing γ-linolenic acid contents in mammalian cells using the delta-6 gene from Borago officinalis. The borage Δ6-desaturase gene (sDelta-6) was codon-optimized and introduced into HEK293 cells by lipofectin transfection. Co-expression of GFP with sDelta-6 and RT-PCR analysis indicated that sDelta-6 could be expressed in mammalian cells. Subsequently, the heterologous expression of borage Δ6-desaturase was evaluated by fatty acid analysis. Total cellular lipid analysis of transformed cells fed with linoleic acid (LA 18:2 n6) as a substrate showed that the expression of sDelta-6 resulted in an 228-483% (p<0.05) increase of GLA when compared with that in the control cells. The highest conversion efficiency of LA into GLA in sDelta-6(+) cells was 6.9 times higher than that in the control group (11.59% vs. 1.69%; p<0.05). Our present work demonstrated that the sDelta-6 gene from borage could be functionally expressed in mammalian cells, and could convert LA into GLA. Furthermore, this study may pave the way to generate transgenic livestock that can synthesise GLA. PMID:21679695

  14. Eicosapentaenoic and dihomo gamma linolenic acid metabolism by cultured rat mesangial cells

    SciTech Connect

    Scharschmidt, L.A.; Gibbons, N.B.; Neuwirth, R.

    1989-01-01

    To better understand the effects of dietary fatty acid manipulations on glomerular function, we compared mesangial incorporation, release, and metabolism of arachidonic (AA), eicosapentaenoic (EPA), and dihomo gamma linolenic (DHG) acids. We found marked differences in mesangial handling of these fatty acids. AA was incorporated into lipids of mesangial cells much more rapidly than EPA or DHG. Ionophore-induced stimulation of fatty acid release from mesangial cells prelabeled with (/sup 14/C)AA, (/sup 14/C)EPA, or (/sup 14/C)DHG caused a release of labeled AA greater than DHG much less than EPA, respectively. Preloading mesangial cells with DHG or EPA for 24 h reduced subsequent basal, ionophore-, and hormone-stimulated prostaglandin E2 (PGE2) synthesis. Finally, unlike AA, neither EPA nor DHG was converted to a significant extent by mesangial cyclooxygenase or lipoxygenase. Thus the mesangial metabolism of DHG and EPA differs both quantitatively and qualitatively from that of AA. Furthermore, EPA and DHG inhibit metabolism of AA at the level of mesangial cyclooxygenase.

  15. Synthesis of a novel acrylated abietic acid-g-bacterial cellulose hydrogel by gamma irradiation.

    PubMed

    Abeer, Muhammad Mustafa; Amin, Mohd Cairul Iqbal Mohd; Lazim, Azwan Mat; Pandey, Manisha; Martin, Claire

    2014-09-22

    Acrylated abietic acid (acrylated AbA) and acrylated abietic acid-grafted bacterial cellulose pH sensitive hydrogel (acrylated AbA-g-BC) were prepared by a one-pot synthesis. The successful dimerization of acrylic acid (AA) and abietic acid (AbA) and grafting of the dimer onto bacterial cellulose (BC) was confirmed by 13C solid state NMR as well as FT-IR. X-ray diffraction analysis showed characteristic peaks for AbA and BC; further, there was no effect of increasing amorphous AA content on the overall crystallinity of the hydrogel. Differential scanning calorimetry revealed a glass transition temperature of 80°C. Gel fraction and swelling studies gave insight into the features of the hydrogel, suggesting that it was suitable for future applications such as drug delivery. Scanning electron microscopy observations showed an interesting interpenetrating network within the walls of hydrogel samples with the lowest levels of AA and gamma radiation doses. Cell viability test revealed that the synthesized hydrogel is safe for future use in biomedical applications.

  16. Gamma Peptide Nucleic Acids: As Orthogonal Nucleic Acid Recognition Codes for Organizing Molecular Self-Assembly.

    PubMed

    Sacui, Iulia; Hsieh, Wei-Che; Manna, Arunava; Sahu, Bichismita; Ly, Danith H

    2015-07-01

    Nucleic acids are an attractive platform for organizing molecular self-assembly because of their specific nucleobase interactions and defined length scale. Routinely employed in the organization and assembly of materials in vitro, however, they have rarely been exploited in vivo, due to the concerns for enzymatic degradation and cross-hybridization with the host's genetic materials. Herein we report the development of a tight-binding, orthogonal, synthetically versatile, and informationally interfaced nucleic acid platform for programming molecular interactions, with implications for in vivo molecular assembly and computing. The system consists of three molecular entities: the right-handed and left-handed conformers and a nonhelical domain. The first two are orthogonal to each other in recognition, while the third is capable of binding to both, providing a means for interfacing the two conformers as well as the natural nucleic acid biopolymers (i.e., DNA and RNA). The three molecular entities are prepared from the same monomeric chemical scaffold, with the exception of the stereochemistry or lack thereof at the γ-backbone that determines if the corresponding oligo adopts a right-handed or left-handed helix, or a nonhelical motif. These conformers hybridize to each other with exquisite affinity, sequence selectivity, and level of orthogonality. Recognition modules as short as five nucleotides in length are capable of organizing molecular assembly.

  17. Gamma-linolenic acid in borage oil reverses epidermal hyperproliferation in guinea pigs.

    PubMed

    Chung, S; Kong, S; Seong, K; Cho, Y

    2002-10-01

    As dietary sources of gamma-linolenic acid [GLA; 18:3(n-6)], borage oil (BO; 24-25 g/100 g GLA) and evening primrose oil (PO; 8-10 g/100 g GLA) are efficacious in treating skin disorders. The triglycerol stereospecificity of these oils is distinct, with GLA being concentrated in the sn-2 position of BO and in the sn-3 position of PO. To determine whether the absolute level and/or the triglycerol stereospecificity of GLA in oils affect biological efficacy, epidermal hyperproliferation was induced in guinea pigs by a hydrogenated coconut oil (HCO) diet for 8 wk. Subsequently, guinea pigs were fed diets of PO, BO or a mixture of BO and safflower oil (SO) for 2 wk. The mixture of BO and SO (BS) diet had a similar level of GLA as PO but with sn-2 stereospecificity. As controls, two groups were fed SO and HCO for 10 wk. Epidermal hyperproliferation was reversed by all three oils in the order of BO > BS > PO. However, proliferation scores of group PO were higher than of the normal control group, SO. The accumulations of dihomo-gamma-linolenic acid [DGLA; 20:3(n-6)], an elongase product of GLA, into phospholipids and ceramides, of 15-hydroxyeicosatrienoic acid (15-HETrE), the potent antiproliferative metabolite of DGLA, and of ceramides, the major lipid maintaining epidermal barrier, in the epidermis of group BO were greater than of groups BS and PO. Group BS had higher levels of DGLA, 15-HETrE and ceramides than group PO. With primary dependence on absolute levels, our data demonstrate that the antiproliferative efficacy of GLA in the epidermis is preferably exerted from sn-2 stereospecificity of GLA in BO. PMID:12368400

  18. Gamma-linolenic acid in borage oil reverses epidermal hyperproliferation in guinea pigs.

    PubMed

    Chung, S; Kong, S; Seong, K; Cho, Y

    2002-10-01

    As dietary sources of gamma-linolenic acid [GLA; 18:3(n-6)], borage oil (BO; 24-25 g/100 g GLA) and evening primrose oil (PO; 8-10 g/100 g GLA) are efficacious in treating skin disorders. The triglycerol stereospecificity of these oils is distinct, with GLA being concentrated in the sn-2 position of BO and in the sn-3 position of PO. To determine whether the absolute level and/or the triglycerol stereospecificity of GLA in oils affect biological efficacy, epidermal hyperproliferation was induced in guinea pigs by a hydrogenated coconut oil (HCO) diet for 8 wk. Subsequently, guinea pigs were fed diets of PO, BO or a mixture of BO and safflower oil (SO) for 2 wk. The mixture of BO and SO (BS) diet had a similar level of GLA as PO but with sn-2 stereospecificity. As controls, two groups were fed SO and HCO for 10 wk. Epidermal hyperproliferation was reversed by all three oils in the order of BO > BS > PO. However, proliferation scores of group PO were higher than of the normal control group, SO. The accumulations of dihomo-gamma-linolenic acid [DGLA; 20:3(n-6)], an elongase product of GLA, into phospholipids and ceramides, of 15-hydroxyeicosatrienoic acid (15-HETrE), the potent antiproliferative metabolite of DGLA, and of ceramides, the major lipid maintaining epidermal barrier, in the epidermis of group BO were greater than of groups BS and PO. Group BS had higher levels of DGLA, 15-HETrE and ceramides than group PO. With primary dependence on absolute levels, our data demonstrate that the antiproliferative efficacy of GLA in the epidermis is preferably exerted from sn-2 stereospecificity of GLA in BO.

  19. Improved enzymatic hydrolysis of wheat straw by combined use of gamma ray and dilute acid for bioethanol production

    NASA Astrophysics Data System (ADS)

    Hyun Hong, Sung; Taek Lee, Jae; Lee, Sungbeom; Gon Wi, Seung; Ju Cho, Eun; Singh, Sudhir; Sik Lee, Seung; Yeoup Chung, Byung

    2014-01-01

    Pretreating wheat straw with a combination of dilute acid and gamma irradiation was performed in an attempt to enhance the enzymatic hydrolysis for bioethanol production. The glucose yield was significantly affected by combined pretreatment (3% sulfuric acid-gamma irradiation), compared with untreated wheat straw and individual pretreatment. The increasing enzymatic hydrolysis after combined pretreatment is resulting from decrease in crystallinity of cellulose, loss of hemicelluloses, and removal or modification of lignin. Therefore, combined pretreatment is one of the most effective methods for enhancing the enzymatic hydrolysis of wheat straw biomass.

  20. Amino acid composition of cerebrospinal fluid in actue neuroinfections in children.

    PubMed

    Buryakova, A V; Sytinsky, I A

    1975-01-01

    A survey of the cerebrospinal fluid (CSF) amino acids, glutamine, and glutamic and gamma-aminobutyric (GABA) acids was made in 168 children, aged 1 to 14 years, with various neurological infections. The glutamic acid and glutamine concentrations in the CSF of children with severe forms of acute serous and bacterial meningitis were about three to four times as great as in controls. The indices returned almost to normal during recovery. GABA is absent in normal CSF, but appeared in the CSF of patients with bacterial meningitis. Its determination may be used as an additional test to differentiate between serous and bacterial meningitis.

  1. Hepatic handling of a synthetic gamma-labeled bile acid (/sup 75/SeHCAT)

    SciTech Connect

    Galatola, G.; Jazrawi, R.P.; Bridges, C.; Joseph, A.E.; Northfield, T.C.

    1988-03-01

    /sup 75/Se-homocholic acid-taurine (/sup 75/SeHCAT) is the first available gamma-labeled bile acid, and should therefore be handled more efficiently and specifically by the liver than previous hepatoscintigraphic agents. We have measured serum and hepatic kinetics for /sup 75/SeHCAT, and compared them with those for the conventional hepatobiliary scintigraphic agent 99mTc-hepatoiminodiacetic acid, and with serum kinetics for the corresponding natural bile acid, (/sup 14/C)cholic acid-taurine. We used a dynamic scintigraphic technique and serial blood sampling in 8 subjects. Initial hepatic uptake rate was identical to initial serum disappearance rate (14% dose/min) for /sup 75/SeHCAT, but significantly lower for 99mTc-hepatoiminodiacetic acid (6% vs. 14% dose/min, p less than 0.001). Hepatic transit time was shorter for /sup 75/SeHCAT (13 min vs. 22 min, p less than 0.02), net hepatic excretory rate was more rapid (1.4% vs. 0.8% dose/min, p less than 0.001), and urinary excretion was lower (1.0% vs. 9.0% dose, p less than 0.001). Initial and late-plasma disappearance rates were significantly lower for /sup 75/SeHCAT (14.3% and 1.5% dose/min) than for (/sup 14/C)cholic acid-taurine (21.3% and 2.8% dose/min, respectively), and plasma clearance was also lower (2/sup 75/ vs. 670 ml/min). In vitro, /sup 75/SeHCAT was bound to serum proteins more completely than (/sup 14/C)cholic acid-taurine (90.4% vs. 86.5%, p less than 0.005). We conclude that /sup 75/SeHCAT provides a hepatoscintigraphic agent that is handled more efficiently and specifically by the liver than the conventionally used agent 99mTc-hepatoiminodiacetic acid. It is not cleared from the serum as rapidly as (/sup 14/C)cholic acid-taurine, probably due to its stronger protein binding. The clinical value of /sup 75/SeHCAT in assessing liver disease should be investigated.

  2. Preferential pi-pi complexation between tamoxifen and borage oil/gamma linolenic acid: transcutaneous delivery and NMR spectral modulation.

    PubMed

    Heard, Charles M; Gallagher, Simon J; Congiatu, Costantino; Harwood, John; Thomas, Christopher P; McGuigan, Christopher; Nemcová, Marta; Nouskova, Tereza

    2005-09-30

    The effect of different proportions of borage oil on the in vitro transcutaneous delivery of tamoxifen were studied, with the aim of developing a gel capable of the simultaneous delivery of tamoxifen and gamma linolenic acid across (breast) skin. Supplementary work probed 1H NMR spectral data for tamoxifen in the presence of different proportions of polyunsaturated or unsaturated fatty acids. Typical, non-aqueous gels were modified to contain 1% tamoxifen and three levels of borage oil ( approximately 25% gamma linolenic acid) and the transcutaneous delivery of both tamoxifen and GLA across full thickness skin determined in vitro. Both tamoxifen and gamma linolenic acid permeated the skin with the ratio of moles being consistent at approximately 4:1. This was irrespective of time, amount of borage oil contained in the formulation (above a minimum) and the presence of other (unsaturated) excipients: mineral oil, Miglyiol 810N, white soft paraffin, PEG400 and Cabosil M5. Dose-dependent downfield shifts of tamoxifen aromatic protons were observed in the presence of borage oil and linolenic acid (gamma and alpha), but not saturated triacyl glycerol. The permeation data suggested vehicular complexation between tamoxifen and polyunsaturated constituents of borage oil and that such complexes permeated the skin intact. The 1H NMR data supported the hypothesis that such complexation was a consequence of preferential pi-pi orbital interactions between the phenyl groups of tamoxifen and the multiple double bonds of GLA. The mechanism for the permeation of intact complexes across skin remains to be elucidated.

  3. Crystal structure of FAS thioesterase domain with polyunsaturated fatty acyl adduct and inhibition by dihomo-[gamma]-linolenic acid

    SciTech Connect

    Zhang, Wei; Chakravarty, Bornali; Zheng, Fei; Gu, Ziwei; Wu, Hongmei; Mao, Jianqiang; Wakil, Salih J.; Quiocho, Florante A.

    2012-05-29

    Human fatty acid synthase (hFAS) is a homodimeric multidomain enzyme that catalyzes a series of reactions leading to the de novo biosynthesis of long-chain fatty acids, mainly palmitate. The carboxy-terminal thioesterase (TE) domain determines the length of the fatty acyl chain and its ultimate release by hydrolysis. Because of the upregulation of hFAS in a variety of cancers, it is a target for antiproliferative agent development. Dietary long-chain polyunsaturated fatty acids (PUFAs) have been known to confer beneficial effects on many diseases and health conditions, including cancers, inflammations, diabetes, and heart diseases, but the precise molecular mechanisms involved have not been elucidated. We report the crystal structure of the hFAS TE domain covalently modified and inactivated by methyl {gamma}-linolenylfluorophosphonate. Whereas the structure confirmed the phosphorylation by the phosphonate head group of the active site serine, it also unexpectedly revealed the binding of the 18-carbon polyunsaturated {gamma}-linolenyl tail in a long groove-tunnel site, which itself is formed mainly by the emergence of an {alpha} helix (the 'helix flap'). We then found inhibition of the TE domain activity by the PUFA dihomo-{gamma}-linolenic acid; {gamma}- and {alpha}-linolenic acids, two popular dietary PUFAs, were less effective. Dihomo-{gamma}-linolenic acid also inhibited fatty acid biosynthesis in 3T3-L1 preadipocytes and selective human breast cancer cell lines, including SKBR3 and MDAMB231. In addition to revealing a novel mechanism for the molecular recognition of a polyunsaturated fatty acyl chain, our results offer a new framework for developing potent FAS inhibitors as therapeutics against cancers and other diseases.

  4. Degradation of cyanidin-3-rutinoside and formation of protocatechuic acid methyl ester in methanol solution by gamma irradiation.

    PubMed

    Lee, Seung Sik; Kim, Tae Hoon; Lee, Eun Mi; Lee, Min Hee; Lee, Ha Yeong; Chung, Byung Yeoup

    2014-08-01

    Anthocyanins are naturally occurring phenolic compounds having broad biological activities including anti-mutagenesis and anti-carcinogenesis. We studied the effects and the degradation mechanisms of the most common type of anthocyanins, cyanidin-3-rutinoside (cya-3-rut), by using gamma ray. Cya-3-rut in methanol (1mg/ml) was exposed to gamma-rays from 1 to 10kGy. We found that the reddish colour of cya-3-rut in methanol disappeared gradually in a dose-dependent manner and effectively disappeared (>97%) at 10kGy of gamma ray. Concomitantly, a new phenolic compound was generated and identified as a protocatechuic acid methyl ester by liquid chromatography, (1)H, and (13)C NMR. The formation of protocatechuic acid methyl ester increased with increasing irradiation and the amount of protocatechuic acid methyl ester formed by decomposition of cya-3-rut (20μg) at 10kGy of gamma ray was 1.95μg. In addition, the radical-scavenging activities were not affected by gamma irradiation. PMID:24629974

  5. Polyaniline/poly acid acrylic thin film composites: a new gamma radiation detector

    SciTech Connect

    Lima Pacheco, Ana P.; Araujo, Elmo S.; Azevedo, Walter M. de

    2003-03-15

    In this paper, we present a new and straightforward route to prepare polyaniline/poly acid acrylic (PAA) thin film composites in large areas and on almost any surface. This method was developed to improve the mechanical and adherence properties of polyaniline devices used as ionization radiation sensors. The route consists of the combination of the metal oxidant with polymer acid to form a highly homogeneous and viscous paste, which can be easily spread over any surface. In the second step, an aniline acid solution is brought in contact with the dried paste where polymerization occurs, yielding a high homogeneous and conducting polymer composite. The UV-visible absorption and infrared analysis confirm that a polyaniline/PAA complex is obtained. The four-point conductivity measurements show that the composite conductivity {rho} is the order of 5 {omega}{sup -1} cm{sup -1}. Preliminary gamma radiation interaction with the composite shows that the doped composite exhibits a linear response that can be used in the development of real-time radiation sensors for the dose range from 0 to 5000 Gy.

  6. [Gamma-hydroxybutyric acid (GHB) dependence and the GHB withdrawal syndrome: diagnosis and treatment].

    PubMed

    van Noorden, Martijn S; Kamal, Rama; de Jong, Cor A J; Vergouwen, A C M Ton; Zitman, Frans G

    2010-01-01

    Gamma-hydroxybutyric acid (GHB) is a neurotransmitter that occurs naturally in the brain and is increasingly being used as a 'party drug' because of its relaxing and euphoria-inducing effects. GHB has a limited medical use in the treatment of narcolepsy. GHB-intoxications occur often in non-medical use, and generally result in a coma of short duration. GHB use several times a day can lead to tolerance and dependence. After sudden cessation or reduction of intensive GHB use, a severe withdrawal syndrome may occur with symptoms varying from tremor, anxiety and agitation to autonomic instability, hallucinations and delirium. Treatment of the GHB withdrawal syndrome consists of supportive care and benzodiazepines, often in high doses. The controlled detoxification of GHB using pharmaceutical GHB in an adjusted dose is currently being investigated in the Netherlands. There is no literature concerning the treatment of patients following GHB intoxication or after detoxification.

  7. [The knowledge about gamma-hydroxybutyric acid as by students of Physical Education Academy].

    PubMed

    Chwaluk, Paweł; Chwaluk, Agnieszka; Parnicki, Florian

    2009-01-01

    Gamma-hydroxybutyric acid is a substance stealthily used by criminals to facilitate sexual assaults. It is also known as doping agent in sports. Physical Education Academies should prepare their graduates to be educators for young people, their trainers, organizers of sports and recreational events. Second year students of two majors: physical education and tourism and recreation were surveyed by means of questionnaire on "date-rape drug". As much as 320 among 327 students surveyed had heard about "date-rape drug". However their knowledge on it was shallow and unsystematic. None of the surveyed knew that the substance of "date-rape drug" could also be used as a doping agent. Only 31% of respondents were aware of existence of the test to detect "date-rape drug" in drinks. Physical Education Academy students should be thoroughly and relevantly educated on the matter of pharmacologic doping agents and drugs endangerment. PMID:19788135

  8. Articular-cartilage matrix gamma-carboxyglutamic acid-containing protein. Characterization and immunolocalization.

    PubMed Central

    Loeser, R; Carlson, C S; Tulli, H; Jerome, W G; Miller, L; Wallin, R

    1992-01-01

    Matrix gamma-carboxyglutamic acid (Gla)-containing protein (MGP) was found to be present in articular cartilage by Western-blot analysis of guanidinium chloride extracts of human and bovine cartilage and was further localized by immunohistochemical studies on human and monkey specimens. In newborn articular cartilage MGP was present diffusely throughout the matrix, whereas in growth-plate cartilage it was seen mainly in late hypertrophic and calcifying-zone chondrocytes. In adult articular cartilage MGP was present primarily in chondrocytes and the pericellular matrix. Immunoelectron microscopy studies revealed an association between MGP and vesicular structures with an appearance consistent with matrix vesicles. MGP may be an important regulator of cartilage calcification because of its localization in cartilage and the known affinity of Gla-containing proteins for Ca2+ and hydroxyapatite. Images Fig. 1. Fig. 2. Fig. 3. PMID:1540125

  9. [Expression of Mortierella isabellina delta6-fatty acid desaturase gene in gamma-linolenic acid production in transgenic tobacco].

    PubMed

    Li, Ming-Chun; Liu, Li; Hu, Guo-Wu; Xing, Lai-Jun

    2003-03-01

    Gamma-linolenic acid (GLA, C18:3delta6.9.12) is nutritional and important polyunsaturated fatty acid in human and animal diets. GLA play an important role in hormone regulation and fatty acid metabolization. Furthermore it is also the biological precursor of a group of molecules, including prostaglandins, leukotrienes and thromboxanes. Vast majority of oilseed crops do not produce GLA, but linoleic acid (LA, C18:2delta9.12) as its substrate. GLA is only produced by a small number of oilseed plants such as evening promrose ( Oenotheera spp.), borage (Borago officinalis) and etc. delta6-fatty acid desaturase (D6D) is the rate-limiting enzyme in the production of GLA. It can convert from linoleic acid to linolenic acid. To produce GLA in tobacco, plant expression vector was first constructed. To facilitate preparation of plant expression constructs, flanking Xba I and Bgl II restriction enzyme sites were added to the coding region of clone pTMICL6 by PCR amplification. pTMICL6 contains delta6-fatty acid desaturase gene cloned from Mortierella isabellina which is an oil-producing fugus. The PCR product was purified and subcloned into the plant expression vector pGA643 to generate the recombinant vector pGAMICL6 which contains the ORF of the D6D gene of Mortierella isabellina, together with regulatory elements consisting of the cauliflower mosaic virus 35S promoter and the nopaline synthase (nos) termination sequence. The plasmid pGAMICL6 was transformed into Agrobacterium tumefaciens strain LBA4404 by method of freeze thawing of liquid nitrogen. Transformants were selected by plating on YEB medium plates containing kanamycin and streptomycin and grown overnight at 28 degrees C, then transformants were further identified by PCR. The positive transformant containing the plant expression vector pGAMICL6 was transformed into tobacco ( Nicotiana tabacum cv. Xanthi) via Agrobacterium infection. Transgenic plants were selected on 100 microg/mL kanamycin. Plants were

  10. (+/-)-1-Tetralone-3-carboxylic acid and (+/-)-1-tetralone-2-acetic acid: hydrogen bonding in two gamma-keto acids.

    PubMed

    Barcon, A; Brunskill, A P; Lalancette, R A; Thompson, H W; Miller, A J

    2001-03-01

    The crystal structure of (+/-)-4-oxo-1,2,3,4-tetrahydronaphthalene-2-carboxylic acid (C(11)H(10)O(3)) involves projection of the carboxyl group nearly orthogonal to the aromatic plane and hydrogen bonding of the acid groups by centrosymmetric pairing across the a edge and the center of the chosen cell [O...O = 2.705 (2) A]. Intermolecular C--H...O==C close contacts to translationally related molecules are found for both the ketone (2.55 A) and the acid (2.67 A). In (+/-)-1-oxo-1,2,3,4-tetrahydronaphthalene-2-acetic acid (C(12)H(12)O(3)), the aggregation involves centrosymmetric carboxyl dimers mutually hydrogen bonded across the bc face and the a edge of the chosen cell [O...O = 2.674 (2) A]. A 2.60 A close C--H...O==C contact is found to the carboxyl group of centrosymmetrically related molecule. PMID:11250596

  11. Enterohepatic circulation in man of a gamma-emitting bile-acid conjugate, 23-selena-25-homotaurocholic acid (SeHCAT).

    PubMed

    Merrick, M V; Eastwood, M A; Anderson, J R; Ross, H M

    1982-02-01

    A conjugated bile acid, 23-selena-25-homotaurocholic acid (SeHCAT), labeled with the gamma emitter Se-75, has been evaluated in man. Absorption and excretion were compared with that of simultaneously administered [23-14C]cholic acid. SeHCAT is absorbed quantitatively following oral administration, secreted into the bile at the same rate as cholic acid, reabsorbed from the small intestine, and resecreted. It is not absorbed when the terminal ileum has been excised or bypassed. SeHCAT is therefore the first of a new class of radiopharmaceuticals, namely, gamma-emitting tracers of the complete cycle of the enterohepatic circulation. Its use will simplify investigation of the functional state of the terminal ileum by eliminating the need to collect and process feces.

  12. Enterohepatic circulation in man of a gamma-emitting bile-acid conjugate, 23-selena-25-homotaurocholic acid (SeHCAT)

    SciTech Connect

    Merrick, M.V.; Eastwood, M.A.; Anderson, J.R.; Ross, H.M.

    1982-02-01

    A conjugated bile acid, 23-selena-25-homotaurocholic acid (SeHCAT), labeled with the gamma emitter Se-75, has been evaluated in man. Absorption and excretion were compared with that of simultaneously administered (23-14C)cholic acid. SeHCAT is absorbed quantitatively following oral administration, secreted into the bile at the same rate as cholic acid, reabsorbed from the small intestine, and resecreted. It is not absorbed when the terminal ileum has been excised or bypassed. SeHCAT is therefore the first of a new class of radiopharmaceuticals, namely, gamma-emitting tracers of the complete cycle of the enterohepatic circulation. Its use will simplify investigation of the functional state of the terminal ileum by eliminating the need to collect and process feces.

  13. Enterohepatic circulation in man of a gamma-emitting bile-acid conjugate, 23-selena-25-homotaurocholic acid (SeHCAT)

    SciTech Connect

    Merrick, M.V.; Eastwood, M.A.; Anderson, J.R.; Ross, H.M.

    1982-02-01

    A conjugated bile acid, 23-selena-25-homotaurocholic acid (SeHCAT), labeled with the gamma emitter Se-75, has been evaluated in man. Absorption and excretion were compared with that of simultaneously administered (23-/sup 14/C)cholic acid. SeHCAT is absorbed quantitatively following oral administration, secreted into the bile at the same rate as cholic acid, reabsorbed from the small intestine, and resecreted. It is not absorbed when the terminal ileum has been excised or bypassed. SeHCAT is therefore the first of a new class of radiopharmaceuticals, namely, gamma-emitting tracers of the complete cycle of the enterohepatic circulation. Its use will simplify investigation of the functional state of the terminal ileum by eliminating the need to collect and process feces.

  14. Gamma Frequency–Range Abnormalities to Auditory Stimulation in Schizophrenia

    PubMed Central

    Kwon, Jun Soo; O’Donnell, Brian F.; Wallenstein, Gene V.; Greene, Robert W.; Hirayasu, Yoshio; Nestor, Paul G.; Hasselmo, Michael E.; Potts, Geoffrey F.; Shenton, Martha E.; McCarley, Robert W.

    2010-01-01

    Background Basic science studies at the neuronal systems level have indicated that gamma-range (30–50 Hz) neural synchronization may be a key mechanism of information processing in neural networks, reflecting integration of various features of an object. Furthermore, gamma-range synchronization is thought to depend on the glutamatergically mediated interplay between excitatory projection neurons and inhibitory neurons utilizing g-aminobutyric acid (GABA), which postmortem studies suggest may be abnormal in schizophrenia. We therefore tested whether auditory neural networks in patients with schizophrenia could support gamma-range synchronization. Methods Synchronization of the electroencephalogram (EEG) to different rates (20–40 Hz) of auditory stimulation was recorded from 15 patients with schizophrenia and 15 sex-, age-, and handedness-matched control subjects. The EEG power at each stimulation frequency was compared between groups. The time course of the phase relationship between each stimulus and EEG peak was also evaluated for gamma-range (40 Hz) stimulation. Results Schizophrenic patients showed reduced EEG power at 40 Hz, but not at lower frequencies of stimulation. In addition, schizophrenic patients showed delayed onset of phase synchronization and delayed desynchronization to the click train. Conclusions These data provide new information on selective deficits in early-stage sensory processing in schizophrenia, a failure to support the entrainment of intrinsic gamma-frequency oscillators. The reduced EEG power at 40 Hz in schizophrenic patients may reflect a dysfunction of the recurrent inhibitory drive on auditory neural networks. PMID:10565499

  15. Abnormal essential fatty acid composition of tissue lipids in genetically diabetic mice is partially corrected by dietary linoleic and gamma-linolenic acids.

    PubMed

    Cunnane, S C; Manku, M S; Horrobin, D F

    1985-05-01

    Genetically diabetic mice (db/db) and their non-diabetic litter-mates were maintained for 15 weeks on diets supplemented with safflower oil or evening primrose (Oenothera bienis) oil, both essential fatty acid (EFA)-rich sources, or hydrogenated coconut oil (devoid of EFA). Plasma glucose was higher in the diabetic mice supplemented with the oils than in the unsupplemented diabetic mice. In the oil-supplemented non-diabetic mice, plasma glucose did not differ compared with the unsupplemented non-diabetic mice. The proportional content of arachidonic acid in the phospholipids of the pancreas was significantly decreased in diabetic mice, an effect which was completely prevented by supplementation with safflower or evening primrose oil but not hydrogenated coconut oil. In the liver phospholipids of the diabetic mice, dihomo-gamma-linolenic acid was proportionally increased, an effect reduced by supplementation with safflower oil but not evening primrose or hydrogenated coconut oils. In the liver triglycerides of the diabetic mice, gamma-linolenic acid, dihomo-gamma-linolenic acid and arachidonic acid were all proportionally decreased, effects which were also prevented by safflower or evening primrose oil but not hydrogenated coconut oil. Alopecia and dry scaly skin were prominent in the diabetic mice but less extensive in the diabetic mice supplemented with EFA.

  16. Surface modification of superparamagnetic iron nanoparticles with calcium salt of poly({gamma}-glutamic acid) as coating material

    SciTech Connect

    Kumar, Ramesh; Inbaraj, B. Stephen; Chen, B.H.

    2010-11-15

    Surface-modified magnetite nanoparticles (MNPs) were synthesized by co-precipitation of aqueous solution of ferrous and ferric salts (molar ratio 1:2) upon adding a base followed by calcium salt of poly({gamma}-glutamic acid) (Ca-{gamma}-PGA) for uniform coating on the surface of MNPs. Both uncoated and Ca-{gamma}-PGA-coated MNPs were characterized using various techniques including Fourier transform infrared spectroscopy (FTIR), X-ray diffraction (XRD), transmission electron microscopy (TEM), thermogravimetric analysis (TGA) and vibrating sample magnetometric (VSM) studies. Compared with bare MNPs, the IR spectra of coated MNPs showed characteristic peaks of {gamma}-PGA, implying the {gamma}-PGA coating on MNPs did occur. The TEM images depicted an average size of 8-10 nm for bare MNPs and 14 nm for coated MNPs, with their shape being spherical in nature. In the presence of applied magnetic field, a superparamagnetic behavior was observed at room temperature for both bare and Ca-{gamma}-PGA-coated MNPs, with no magnetism left upon magnetic-field removal.

  17. Effects of colistin on amino acid neurotransmitters and blood-brain barrier in the mouse brain.

    PubMed

    Wang, Jian; Yi, Meishuang; Chen, Xueping; Muhammad, Ishfaq; Liu, Fangping; Li, Rui; Li, Jian; Li, Jichang

    2016-01-01

    Neurotoxicity is one of the major potential side effects of colistin therapy. However, the mechanistic aspects of colistin-induced neurotoxicity remain largely unknown. The objective of this study was to examine the effects of colistin on the blood-brain barrier (BBB) and amino acid neurotransmitters in the cerebral cortex of mouse. Mice were divided into four groups (n=5) and were administrated intravenously with 15mg/kg/day of colistin sulfate for 1, 3 and 7days successively while the control group was administrated intravenously with saline solution. The permeability and ultrastructure of the BBB were detected using the Evans blue (EB) dye and transmission electron microscopy (TEM), and the expression of Claudin-5 were determined by real-time PCR examination and western blotting. The brain uptake of colistin was measured by high-performance liquid chromatography (HPLC). The effects of colistin on amino acid neurotransmitters and their receptors were also examined by HPLC and real-time PCR. The results of EB extravasation, TEM and expression of Claudin-5 showed that colistin treatment did not affect the BBB integrity. In addition, multiple doses of colistin could induce accumulation of this compound in the brain parenchyma although there was poor brain uptake of colistin. Moreover, colistin exposure significantly increased the contents of glutamate (Glu) and gamma aminobutyric acid (GABA), and enhanced the mRNA expression levels of gamma aminobutyric acid type A receptor (GABAAR), gamma aminobutyric acid type B receptor (GABABR), N-methyl-d-aspartate 1 receptor (NR1), N-methyl-d-aspartate 2A receptor (NR2A) and N-methyl-d-aspartate 2B receptor (NR2B) in the cerebral cortex. Our data demonstrate that colistin is able to accumulate in the mouse brain and elevate the levels of amino acid neurotransmitters. These findings may be associated with colistin-induced neurotoxicity.

  18. Effects of stearic acid and gamma irradiation, alone and in combination, on pasting properties of high amylose maize starch.

    PubMed

    Ocloo, Fidelis C K; Minnaar, Amanda; Emmambux, Naushad M

    2016-01-01

    The effects of stearic acid and gamma irradiation on pasting properties of high amylose maize starch (HAMS) were studied. Stearic acid (0%, 1.5%, and 5%) was added to HAMS, and then irradiated at 0, 30, and 60 kGy before pasting. Stearic acid increased the paste viscosity of un-irradiated HAMS from 420 mPas to 557 and 652 mPas for 1.5% and 5% stearic acid, respectively. This observation related well with the formation of type II amylose-lipid complexes, with melting temperatures of about 100-120 °C. Gamma irradiation (30 and 6 0kGy) reduced pasting viscosity of HAMS. Pasting of gamma irradiated HAMS resulted in the formation of type I amylose-lipid complexes, with melting temperatures and enthalpies ranging from 82 to 102 °C and 0.22 to 1.85 J/g, respectively. Stearic acid addition followed by irradiation creates means of producing different types of amylose-lipid complexes from HAMS for industrial utilization.

  19. Enhanced enzymatic hydrolysis of poplar bark by combined use of gamma ray and dilute acid for bioethanol production

    NASA Astrophysics Data System (ADS)

    Chung, Byung Yeoup; Lee, Jae Taek; Bai, Hyoung-Woo; Kim, Ung-Jin; Bae, Hyeun-Jong; Gon Wi, Seung; Cho, Jae-Young

    2012-08-01

    Pretreatment of poplar bark with a combination of sulfuric acid (3%, w/w, H2SO4) and gamma irradiation (0-1000 kGy) was performed in an attempt to enhance enzymatic hydrolysis for bioethanol production. The yields of reducing sugar were slightly increased with an increasing irradiation dose, ranging from 35.4% to 51.5%, with a 56.1% reducing sugar yield observed after dilute acid pretreatment. These results clearly showed that soluble sugars were released faster and to a greater extent in dilute acid-pretreated poplar bark than in gamma irradiation-pretreated bark. When combined pretreatment was carried out, a drastic increase in reducing sugar yield (83.1%) was found compared with individual pretreatment, indicating the possibility of increasing the convertibility of poplar bark following combined pretreatment. These findings are likely associated with cellulose crystallinity, lignin modification, and removal of hemicelluloses.

  20. Formation of hydrocarbons from acid-Clay suspensions by gamma irradiation

    NASA Astrophysics Data System (ADS)

    Cruz-Castañeda, J.; Negron-Mendoza, A.; Ramos-Bernal, S.

    2013-07-01

    The adsorption of certain organic compounds by clays gives rise to the transformation of the adsorbate through the action of the clays. This phenomenon can be enhanced using ionizing radiation. In this context, these kinds of reactions play an important role in many natural and industrial processes. For example, in oil and gas exploration, the source and trap of petroleum hydrocarbons is frequently clay-rich rocks. Clay-water-based muds are also seen as environmentally friendly alternatives to toxic oil-based fluids. The principal processes that occur in sediments are usually held to be of bacterial action and thermal transformation, which may include thermally induced catalytic alteration of the organic debris. On the other hand, radioactive materials are widely distributed throughout Earth. They were more abundant in the past, but are present in petroleum reservoirs. Their presence induced radioactive bombardment, which may have altered these sediments. This important subject has not been extensively studied. The aim of this work is to study the behavior of fatty acids—like behenic acid—and dicarboxylic acids—like fumaric acid—as model compounds, which are adsorbed in a clay mineral (Na-montmorillonite) and exposed to gamma radiation. The results show that the radiation-induced decomposition of the clay-acid system goes along a definitive path (oxidation), rather than following several modes of simultaneous decomposition, as happens in radiolysis without clay or by heating the system. The main radiolytic products for fatty acids are their corresponding hydrocarbons, with one C-atom less than the original acid.

  1. A widely used retinoic acid receptor antagonist induces peroxisome proliferator-activated receptor-gamma activity.

    PubMed

    Schupp, Michael; Curtin, Joshua C; Kim, Roy J; Billin, Andrew N; Lazar, Mitchell A

    2007-05-01

    Nuclear receptors (NRs) are transcription factors whose activity is regulated by the binding of small lipophilic ligands, including hormones, vitamins, and metabolites. Pharmacological NR ligands serve as important therapeutic agents; for example, all-trans retinoic acid, an activating ligand for retinoic acid receptor alpha (RARalpha), is used to treat leukemia. Another RARalpha ligand, (E)-S,S-dioxide-4-(2-(7-(heptyloxy)-3,4-dihydro-4,4-dimethyl-2H-1-benzothiopyran-6-yl)-1-propenyl)-benzoic acid (Ro 41-5253), is a potent antagonist that has been a useful and purportedly specific probe of RARalpha function. Here, we report that Ro 41-5253 also activates the peroxisome proliferator-activated receptor gamma (PPARgamma), a master regulator of adipocyte differentiation and target of widely prescribed antidiabetic thiazolidinediones (TZDs). Ro 41-5253 enhanced differentiation of mouse and human preadipocytes and activated PPARgamma target genes in mature adipocytes. Like the TZDs, Ro 41-5253 also down-regulated PPARgamma protein expression in adipocytes. In addition, Ro 41-5253 activated the PPARgamma-ligand binding domain in transiently transfected HEK293T cells. These effects were not prevented by a potent RARalpha agonist or by depleting cells of RARalpha, indicating that PPARgamma activation was not related to RARalpha antagonism. Indeed, Ro 41-5253 was able to compete with TZD ligands for binding to PPARgamma, suggesting that Ro 41-5253 directly affects PPAR activity. These results vividly demonstrate that pharmacological NR ligands may have "off-target" effects on other NRs. Ro 41-5253 is a PPARgamma agonist as well as an RARalpha antagonist whose pleiotropic effects on NRs may signify a unique spectrum of biological responses.

  2. A single gamma-carboxyglutamic acid residue in a novel cysteine-rich secretory protein without propeptide.

    PubMed

    Hansson, Karin; Thämlitz, Ann-Marie; Furie, Bruce; Furie, Barbara C; Stenflo, Johan

    2006-10-24

    Gamma-glutamyl carboxylase catalyzes the modification of specific glutamyl residues to gamma-carboxyglutamyl (Gla) residues in precursor proteins that possess the appropriate gamma-carboxylation recognition signal within the propeptide region. We describe the immunopurification and first biochemical characterization of an invertebrate high molecular weight Gla-containing protein with homologues in mammals. The protein, named GlaCrisp, was isolated from the venom of the marine cone snail Conus marmoreus. GlaCrisp gave intense signals in Western blot experiments employing the Gla-specific antibody M3B, and the presence of Gla was chemically confirmed by amino acid analysis after alkaline hydrolysis. Characterization of a full-length cDNA clone encoding GlaCrisp deduced a precursor containing an N-terminal signal peptide but, unlike other Gla-containing proteins, no apparent propeptide. The predicted mature protein of 265 amino acid residues showed considerable sequence similarity to the widely distributed cysteine-rich secretory protein family and closest similarity (65% identity) to the recently described substrate-specific protease Tex31. In addition, two cDNA clones encoding the precursors of two isoforms of GlaCrisp were identified. The predicted precursor isoforms differed at three amino acid positions (-6, 9, and 25). Analysis by Edman degradation and nanoelectrospray ionization mass spectrometry, before and after methyl esterfication, identified a Gla residue at amino acid position 9 in GlaCrisp. This is the first example of a Gla-containing protein without an obvious gamma-carboxylation recognition site. The results define a new class of Gla proteins and support the notion that gamma-carboxylation of glutamyl residues is phylogenetically older than blood coagulation and the vertebrate lineage.

  3. Transgenic manipulation of a single polyamine in poplar cells affects the accumulation of all amino acids.

    PubMed

    Mohapatra, Sridev; Minocha, Rakesh; Long, Stephanie; Minocha, Subhash C

    2010-04-01

    The polyamine metabolic pathway is intricately connected to metabolism of several amino acids. While ornithine and arginine are direct precursors of putrescine, they themselves are synthesized from glutamate in multiple steps involving several enzymes. Additionally, glutamate is an amino group donor for several other amino acids and acts as a substrate for biosynthesis of proline and gamma-aminobutyric acid, metabolites that play important roles in plant development and stress response. Suspension cultures of poplar (Populus nigra x maximowiczii), transformed with a constitutively expressing mouse ornithine decarboxylase gene, were used to study the effect of up-regulation of putrescine biosynthesis (and concomitantly its enhanced catabolism) on cellular contents of various protein and non-protein amino acids. It was observed that up-regulation of putrescine metabolism affected the steady state concentrations of most amino acids in the cells. While there was a decrease in the cellular contents of glutamine, glutamate, ornithine, arginine, histidine, serine, glycine, cysteine, phenylalanine, tryptophan, aspartate, lysine, leucine and methionine, an increase was seen in the contents of alanine, threonine, valine, isoleucine and gamma-aminobutyric acid. An overall increase in percent cellular nitrogen and carbon content was also observed in high putrescine metabolizing cells compared to control cells. It is concluded that genetic manipulation of putrescine biosynthesis affecting ornithine consumption caused a major change in the entire ornithine biosynthetic pathway and had pleiotropic effects on other amino acids and total cellular carbon and nitrogen, as well. We suggest that ornithine plays a key role in regulating this pathway.

  4. Studies of optimum conditions for covalent immobilization of Candida rugosa lipase on poly(gamma-glutamic acid) by RSM.

    PubMed

    Chang, S-W; Shaw, J-F; Yang, K-H; Chang, S-F; Shieh, C-J

    2008-05-01

    Poly(gamma-glutamic acid) (gamma-PGA) is a material of polymer. Immobilization of Candida rugosa lipase (Lipase AY-30) by covalent binding on gamma-PGA led to a markedly improved performance of the enzyme. Response surface methodology (RSM) and 3-level-3-factor fractional factorial design were employed to evaluate the effects of immobilization parameters, such as immobilization time (2-6h), immobilization temperature (0-26 degrees C), and enzyme/support ratio (0.1-0.5, w/w). Based on the analysis of ridge max, the optimum immobilization conditions were as follows: immobilization time 2.3h, immobilization temperature 13.3 degrees C, and enzyme/support ratio 0.41 (w/w); the highest lipase activity obtained was 1196 U/mg-protein.

  5. Preferential pi-pi complexation between tamoxifen and borage oil/gamma linolenic acid: transcutaneous delivery and NMR spectral modulation.

    PubMed

    Heard, Charles M; Gallagher, Simon J; Congiatu, Costantino; Harwood, John; Thomas, Christopher P; McGuigan, Christopher; Nemcová, Marta; Nouskova, Tereza

    2005-09-30

    The effect of different proportions of borage oil on the in vitro transcutaneous delivery of tamoxifen were studied, with the aim of developing a gel capable of the simultaneous delivery of tamoxifen and gamma linolenic acid across (breast) skin. Supplementary work probed 1H NMR spectral data for tamoxifen in the presence of different proportions of polyunsaturated or unsaturated fatty acids. Typical, non-aqueous gels were modified to contain 1% tamoxifen and three levels of borage oil ( approximately 25% gamma linolenic acid) and the transcutaneous delivery of both tamoxifen and GLA across full thickness skin determined in vitro. Both tamoxifen and gamma linolenic acid permeated the skin with the ratio of moles being consistent at approximately 4:1. This was irrespective of time, amount of borage oil contained in the formulation (above a minimum) and the presence of other (unsaturated) excipients: mineral oil, Miglyiol 810N, white soft paraffin, PEG400 and Cabosil M5. Dose-dependent downfield shifts of tamoxifen aromatic protons were observed in the presence of borage oil and linolenic acid (gamma and alpha), but not saturated triacyl glycerol. The permeation data suggested vehicular complexation between tamoxifen and polyunsaturated constituents of borage oil and that such complexes permeated the skin intact. The 1H NMR data supported the hypothesis that such complexation was a consequence of preferential pi-pi orbital interactions between the phenyl groups of tamoxifen and the multiple double bonds of GLA. The mechanism for the permeation of intact complexes across skin remains to be elucidated. PMID:16115741

  6. Glutamic acid gamma-monohydroxamate and hydroxylamine are alternate substrates for Escherichia coli asparagine synthetase B.

    PubMed

    Boehlein, S K; Schuster, S M; Richards, N G

    1996-03-01

    Escherichia coli asparagine synthetase B (AS-B) catalyzes the synthesis of asparagine from aspartic acid and glutamine in an ATP-dependent reaction. The ability of this enzyme to employ hydroxylamine and L-glutamic acid gamma-monohydroxamate (LGH) as alternative substrates in place of ammonia and L-glutamine, respectively, has been investigated. The enzyme is able to function as an amidohydrolase, liberating hydroxylamine from LGH with high catalytic efficiency, as measured by k(cat)/K(M). In addition, the kinetic parameters determined for hydroxylamine in AS-B synthetase activity are very similar to those of ammonia. Nitrogen transfer from LGH to yield aspartic acid beta-monohydroxamate is also catalyzed by AS-B. While such an observation has been made for a few members of the trpG amidotransferase family, our results appear to be the first demonstration that nitrogen transfer can occur from glutamine analogs in a purF amidotransferase. However, k(cat)/K(M) for the ATP-dependent transfer of hydroxylamine from LGH to aspartic acid is reduced 3-fold relative to that for glutamine-dependent asparagine synthesis. Further, the AS-B mutant in which asparagine is replaced by alanine (N74A) can also use hydroxylamine as an alternate substrate to ammonia and catalyze the hydrolysis of LGH. The catalytic efficiencies (k(cat)/K(M)) of nitrogen transfer from LGH and L-glutamine to beta-aspartyl-AMP are almost identical for the N74A AS-B mutant. These observations support the proposal that Asn-74 plays a role in catalyzing glutamine-dependent nitrogen transfer. We interpret our kinetic data as further evidence against ammonia-mediated nitrogen transfer from glutamine in the purF amidotransferase AS-B. These results are consistent with two alternate chemical mechanisms that have been proposed for this reaction [Boehlein, S. K., Richards, N. G. J., Walworth, E. S., & Schuster, S. M. (1994) J. Biol. Chem. 269, 26789-26795].

  7. Appearance of dentin gamma-carboxyglutamic acid-containing proteins in developing rat molars in vitro

    SciTech Connect

    Finkelman, R.D.; Butler, W.T.

    1985-07-01

    An in vitro model of mineralization was devised in order to study the developmental appearance of dentin gamma-carboxyglutamic acid-containing proteins (DGPs) in relation to the onset of mineralization. Maxillary third molars from 11-day-old rats were cultured with or without fetal calf serum (FCS) as modified from Navia et al. Molars were incubated without radiolabel, or with either /sup 45/CaCl/sub 2/ (5 microCi/ml) for 24 hr at various stages of a ten-day culture period or (/sup 3/H)-leucine (10 microCi/ml) for 24 hr at the eighth day of culture. Molars were lyophilized and extracted with 10% formic acid overnight at 4 degrees C. DGPs in extracts were detected by immunologic and chromatographic techniques; DGPs in molar sections were detected by immunolocalization using indirect immunofluorescence. Molar development was evaluated histologically using the Von Kossa staining technique. Molars cultured with FCS showed histologic evidence for mineralized dentin and enamel and a significant increase in /sup 45/Ca uptake after the sixth day in vitro. Eleven-day-old molars in vivo and molars cultured without FCS showed no evidence of the presence of mineralized tissues. (/sup 3/H)-Leucine-labeled DGPs were isolated and identified by affinity and reversed-phase high-performance liquid chromatography and by gel electrophoresis from both mineralized and unmineralized molars. DGP antigens were localized immunohistochemically using rabbit anti-rat antibodies raised against a highly purified DGP preparation. In the unmineralized molar, antigenicity was seen in odontoblasts but not in predentin matrix, preodontoblasts, or in any other cell type. Antigens in the mineralized molar were localized to odontoblasts and dentin.