Glutamic Acid Decarboxylase Activity Research Articles

Augmentation of endogenous GABA pool size induced by Magainin II peptide

BackgroundGamma aminobutyric acid (GABA), an inhibitory neurotransmitter, is produced via decarboxylation of l-glutamate through the glutamic acid decarboxylase (GAD) enzyme. The synchronic action of GABA-transaminase (GABA-T) and succinic semialdehyde dehydrogenase (SSADH) enzymes convert the GABA metabolite into succinate. Given this background, our research was aimed at probing the effect of Magainin II, on the activity of GABA shunt metabolizing enzymes. MethodsMale NIH mice were administered peripherally by Magainin II (50 μg/kg body weight) and saline solution (%0.9 (w/v)) as the control vehicle. At different time intervals, the mice were sacrificed to evaluate the effect of Magainin II injection on the GABA shunt pathway. The activity of hypothalamic GAD, GABA-T and SSADH enzymes were determined using relevant enzyme assays. ResultsMagainin II effectively enhanced the activity of GAD, by %90, 24 h after injection, while quenching the activities of GABA-T and SSADH by %43 and %71, respectively. In vitro models also revealed the direct but reversible interaction between the peptide and each of the individual enzymes of GABA shunt pathway. ConclusionThis study confirms the probable role of Magainin II in increasing the GABA content of the mouse hypothalamus. This property might candidate the peptide as a novel agent for improving the symptoms of many GABA dependent psychiatric disorders.

PB11. Long term GABA alterations induced by anodal Transcranial Direct Current Stimulation (tDCS) in motor cortex: A proton magnetic resonance spectroscopy study

Background Transcranial direct current stimulation (tDCS) is a non-invasive technique that modulates excitability of neurons in the motor cortex. In particular, anodal tDCS leads to cortical facilitation and overall increase in excitability ( Nitsche et al., 2000 ). Objective Although the effects of anodal tDCS on modulating motor cortex activation have been reported, there is little evidence of long term reduction of GABA concentration following anodal stimulation. Therefore, to characterize the duration of after effects of primary motor cortex (M1) excitability following anodal stimulation, we performed repeated magnetic resonance spectroscopy (MRS) measurements of GABA for 60 min after the stimulation. Materials and methods Thirty-two right-handed healthy volunteers were recruited in RWTH Aachen University. GABA-edited spectra were acquired using 3 T Siemens scanner from a volume of interest of 3 × 3 × 3 cm (27 mL) carefully placed on the hand motor area of the left primary motor cortex [ Fig. 1 ]. MEGA-PRESS J-editing ( Mescher et al., 1998 ), was used for GABA detection. Data processing and quantification of spectra was performed with TARQUIN ( Wilson et al., 2011 ). Results An ANOVA was performed on percentage of GABA concentrations with stimulation (sham vs. anodal) and measurement (12 measurements: 2 pre-stimulation and 10 post-stimulation measurements taken every 6 min) as between- and within-participants factors. The main effects of stimulation was not significant F(1, 30) = 1.157, p = .291. However, main effect of measurement was significant F(11, 330) = 2.378 p = .008. Remarkably, the interaction of stimulation x measurement was significant F(11, 330) = 2.249, p = .0128 [ Fig. 2 ]. GABA concentration decreased significantly in the anodal stimulation group at first (20th minute) post-stimulation measurement, and then stayed the same at measurements eighth, ninth and tenth ts (3) Discussion and conclusion Our results revealed a significant early drop of GABA concentration within 20 min following anodal stimulation and late drop of GABA after 66th minutes until the last post stimulation measurement (86th min). Early and late decrease in GABA could be due to a decrease of activity of glutamic acid decarboxylase (GAD) 67 and long term depression like plasticity, respectively. This technique may open a new field of application for long-lasting neuroplastic induction, especially in the rehabilitation of stroke patients.

S-nitrosylation of GAD65 is implicated in decreased GAD activity and oxygen-induced seizures.

Breathing oxygen at partial pressures ≥2.5 atmospheres absolute, which can occur in diving and hyperbaric oxygen (HBO2) therapy, can rapidly become toxic to the central nervous system (CNS). This neurotoxicity culminates in generalized EEG epileptiform discharges, tonic-clonic convulsions and ultimately death. Increased production of neuronal nitric oxide (NO) has been implicated in eliciting hyperoxic seizures by altering the equilibrium between glutamatergic and GABAergic synaptic transmission. Inhibition of glutamic acid decarboxylase (GAD) activity in HBO2 promotes this imbalance; however, the mechanisms by which this occurs is unknown. Therefore, we conducted a series of experiments using mice, a species that is highly susceptible to CNS oxygen toxicity, to explore the possibility that NO modulates GABA metabolism. Mice were exposed to 100% oxygen at 4 ATA for various durations, and brain GAD and GABA transaminase (GABA-T) activity, as well as S-nitrosylation of GAD65 and GAD67 were determined. HBO2 inhibited GAD activity by 50% and this was negatively correlated with S-nitrosylation of GAD65, whereas GABA-T activity and S-nitrosylation of GAD67 were unaltered. These results suggest a new mechanism by which NO alters GABA metabolism, leading to neuroexcitation and seizures in HBO2.

Protective roles of hepatic GABA signaling in acute liver injury of rats.

γ-Aminobutyric acid (GABA) is produced by various cells through the catalytic activity of glutamic acid decarboxylase (GAD). Activation of type-A GABA receptor (GABAAR) inhibits stem cell proliferation but protects differentiated cells from injures. The present study investigated hepatic GABA signaling system and the role of this system in liver physiology and pathophysiology. RT-PCR and immunoblot assays identified GAD and GABAAR subunits in rat livers and in HepG2 and Clone 9 hepatocytes. Patch-clamp recording detected GABA-induced currents in Clone 9 hepatocytes and depolarization in WITT cholangiocytes. The function of hepatic GABA signaling system in rats was examined using models of d-galactosamine (GalN)-induced acute hepatocytic injury in vivo and in vitro. The expression of GAD increased whereas GABAAR subunits decreased in the liver of GalN-treated rats. Remarkably, treating rats with GABA or the GABAAR agonist muscimol, but not the GABABR agonist baclofen, protected hepatocytes against GalN toxicity and improved liver function. In addition, muscimol treatment decreased the formation of pseudobile ductules and the enlargement of hepatocytic canaliculi in GalN-treated rats. Our results revealed that a complex GABA signaling system exists in the rat liver. Activation of this intrahepatic GABAergic system protected the liver against toxic injury.NEW & NOTEWORTHY Auto- and paracrine GABAergic signaling systems exist in the rat hepatocytes and cholangiocytes. Activation of GABA signaling protects liver function from d-galactosamine injury by reducing toxic impairment of hepatocytes and by decreasing cholangiocyte proliferation.

Enzyme activities involved in the glutamate-glutamine cycle are altered to reduce glutamate after spinal cord injury in rats.

Glutamate (Glu) neurotransmitter is involved in the excitotoxic damage after spinal cord injury (SCI). Glu is transformed into glutamine (Gln) by glutamine synthetase (GS) enzyme in glial cells. Once into the neurons, Gln is transformed back into Glu by phosphate-activated glutaminase (PAG). Glu is also a precursor for the synthesis of γ-aminobutyric acid through the action of the glutamic acid decarboxylase (GAD) enzyme. The contribution of all these Glu biotransformations after SCI has not been determined. The aim of this work is to characterize the role of GS, PAG, and GAD in the acute phase after SCI. Female Wistar rats were subjected to SCI by contusion and killed 2, 4, 8, and 12 h after surgery. Sham-injury animals, killed at the same time points served as controls. PAG and GAD activities were analyzed by high-performance liquid chromatography, whereas GS activity was determined by ultraviolet-visible spectroscopy. GS activity showed a significant decrease in animals with SCI at all time points evaluated versus the sham group. Similarly, the activity of the PAG was decreased at all time points compared with the control group. Finally, GAD activity was significantly increased in the SCI group when measured at 2, 4, and 8 h after lesion. The results of this study suggest that excitotoxicity is highly regulated through Glu/Gln and Glu/γ-aminobutyric acid cycles as an important mechanism to prevent further damage in the acute phase after lesion.

Renal Denervation Improves the Baroreflex and GABA System in Chronic Kidney Disease-induced Hypertension.

Hypertensive rats with chronic kidney disease (CKD) exhibit enhanced gamma-aminobutyric acid (GABA)B receptor function and regulation within the nucleus tractus solitarii (NTS). For CKD with hypertension, renal denervation (RD) interrupts the afferent renal sympathetic nerves, which are connecting to the NTS. The objective of the present study was to investigate how RD improves CKD-induced hypertension. Rats underwent 5/6 nephrectomy for 8 weeks, which induced CKD and hypertension. RD was induced by applying phenol to surround the renal artery in CKD. RD improved blood pressure (BP) by lowering sympathetic nerve activity and markedly restored the baroreflex response in CKD. The GABAB receptor expression was increased in the NTS of CKD; moreover, the central GABA levels were reduced in the cerebrospinal fluid, and the peripheral GABA levels were increased in the serum. RD restored the glutamic acid decarboxylase activity in the NTS in CKD, similar to the effect observed for central treatment with baclofen, and the systemic administration of gabapentin reduced BP. RD slightly improved renal function and cardiac load in CKD. RD may improve CKD-induced hypertension by modulating the baroreflex response, improving GABA system dysfunction and preventing the development and reducing the severity of cardiorenal syndrome type 4 in CKD rats.

Detection, isolation and characterization of Fusobacterium gastrosuis sp. nov. colonizing the stomach of pigs

Nine strains of a novel Fusobacterium sp. were isolated from the stomach of 6–8 months old and adult pigs. The isolates were obligately anaerobic, although they endured 2h exposure to air. Phylogenetic analysis based on 16S rRNA and gyrase B genes demonstrated that the isolates showed high sequence similarity with Fusobacterium mortiferum, Fusobacterium ulcerans, Fusobacterium varium, Fusobacterium russii and Fusobacterium necrogenes, but formed a distinct lineage in the genus Fusobacterium. Comparative analysis of the genome of the type strain of this novel Fusobacterium sp. confirmed that it is different from other recognized Fusobacterium spp. DNA-DNA hybridization, fingerprinting and genomic %GC determination further supported the conclusion that the isolates belong to a new, distinct species. The isolates were also distinguishable from these and other Fusobacterium spp. by phenotypical characterization. The strains produced indole and exhibited proline arylamidase and glutamic acid decarboxylase activity. They did not hydrolyse esculin, did not exhibit pyroglutamic acid arylamidase, valine arylamidase, α-galactosidase, β-galactosidase, β-galactosidase-6-phosphate or α-glucosidase activity nor produced acid from cellobiose, glucose, lactose, mannitol, mannose, maltose, raffinose, saccharose, salicin or trehalose. The major fatty acids were C16:0 and C18:1ω9c. The name Fusobacterium gastrosuis sp. nov. is proposed for the novel isolates with the type strain CDW1(T) (=DSM 101753(T)=LMG 29236(T)). We also demonstrated that Clostridium rectum and mortiferum Fusobacterium represent the same species, with nomenclatural priority for the latter.

A genetic context for the study of audiogenic seizures.

Here, the genetic context for the study of audiogenic seizures is four single-gene, spontaneous mutations that occurred in the Behavior Genetics Laboratory at the University of Chicago from 1959 to 1969. Three of these increased the incidence of audiogenic seizures, and one of these decreased the incidence of audiogenic seizures. The genetics of one of these mutants is described in detail, and the effect of diet on the same mutant is also described in detail. Research on genetic and environmental effects on the cortical EEG of audiogenic seizures is reviewed; this research included two of these mutants. The cortical EEG associated with audiogenic seizures in this study was consistent with audiogenic seizures being a type of brain stem epilepsy as had been proposed by others. Also, I proposed that brain stem pathophysiology is the same regardless of the genetic or environmental pathway to audiogenic seizure susceptibility. Research is also reviewed using these mutants to determine whether or not a strain association between glutamic acid decarboxylase (GAD) activity in whole brain and susceptibility to audiogenic seizures is pleiotropic and whether or not a strain association between nucleoside triphosphatase (NTPase) activity in the granule cell layer of the dentate fascia of the hippocampus and susceptibility to audiogenic seizures is a lineal or collateral pleiotropy. Lastly, pleiotropy as an explanation for strain comorbidities in aggressive behavior and audiogenic seizures is considered. This article is part of a Special Issue entitled "Genetic and Reflex Epilepsies, Audiogenic Seizures and Strains: From Experimental Models to the Clinic".

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

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 20.3mg/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 739.9mg/100g 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.

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.

The present experiment was conducted to investigate the interactive effects between dietary glutamine (Gln, 0 and 5g/kg) and gamma-aminobutyric acid (GABA, 0 and 100mg/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 42days, while it increased (P < 0.05) the activities of glutamine synthetase (GS) and gamma-aminobutyric acid transaminase (GABA-T) at 28, 35, and 42days. Dietary Gln and GABA improved (P < 0.05) DWG and DFC of broilers under cyclic HS during 28-42days. In breast muscle, the Gln supplementation increased (P < 0.05) the concentrations of Gln (28, 35, and 42days), Glu (28, 35, and 42days), and GABA (42days) and the activities of glutaminase (28, 35, and 42days) and GAD (28, 35, and 42days) but decreased (P < 0.05) GS activities at 28, 35, and 42days and GABA-T activities at 28days. 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 42days. Significant interactions (P < 0.05) between Gln and GABA were found on breast skeletal muscle Gln concentrations, glutaminase activities, GS activities at 28 and 35days, and DWG, GABA concentrations, and GABA-T activities at 28, 35, and 42days 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 GABA metabolism of broilers.

S‐Nitrosoglutathione Reductase Null Mice Display Increased Brain Glutamic Acid Decarboxylase Activity and Seizure Resistance in Hyperbaric Oxygen

An imbalance between glutamatergic and g‐aminobutyric acid (GABA)ergic synaptic transmission is associated with hyperbaric oxygen (HBO2) induced seizures. Here we explored the impact of S‐nitrosylation in preventing oxygen toxicity in S‐nitrosoglutathione reductase (GSNOR) null mice and tested whether S‐nitrosylation of brain glutamic acid decarboxylase (GAD) may alter its activity, thus the synthesis of inhibitory GABA. Wild type (WT) and GSNOR ‐/‐ mice were exposed to HBO2 at 4 ATA for up to 100 min and seizure latency was recorded. The medulla, cerebellum and forebrain were harvested from HBO2 exposed and control mice for determination of GAD activity (fluorometrically) and S‐nitrosylated GAD‐65 (Biotin‐switch). The mean (± SE) seizure latency was significantly delayed in the GSNOR ‐/‐ (63 ± 7 min) compared to WT (34 ± 6 min) mice, and the proportion of GSNOR ‐/‐ mice that experienced seizures (56%) was significantly less than WT mice (87%). Relative to controls, GAD activity in the medulla and cerebellum was significantly increased in the GSNOR ‐/‐ compared to a reduction in WT mice: medulla (GSNOR ‐/‐: +22% and WT: ‐13%) and cerebellum (GSNOR‐/‐: +23% and WT: ‐12%). A significant fall in the degree of GAD‐65 S‐nitrosylation was observed in GSNOR ‐/‐ (‐63%) compared to WT (‐17%) mice suggesting augmented denitrosylation after GSNOR deletion. Dysregulated S‐nitrosylation increased seizure latency and reduced the percentage of mice that developed seizures. The protection afforded by reduced S‐nitrosylation may be explained in part by increased GAD activity that would favor GABA synthesis, thus inhibit neuronal excitation and seizures.

GABA circuitry, cells and molecular regulation in schizophrenia: Life in the graveyard

Post mortem studies of the GABA system in schizophrenia had an inauspicious start, when a high-profile report (Bird et al., 1977) of decreased activity of glutamic acid decarboxylase (GAD), the GABA synthetic enzyme, was later shown to reflect the mode of death (agonal state) not the underlying diagnosis (Spokes, 1979). Fortunately, subsequent studies have been much more successful in measuring and controlling for this and other confounders (Berretta et al., 2015–in this issue a), and they have also been of increasing quality in terms of design, rigour, and technical sophistication. Indeed, the cumulative post mortem evidence for a ‘GABAergic involvement’ in schizophrenia is now robust and compelling. The findings are complex, with an intriguing – and still incompletely understood – regional, cellular, and molecular pattern. The field has largely moved on from the initial, essential, descriptive phase of research, and is now attempting to characterise better the profile of alterations, explain their genetic anddevelopmental origins, understand the pathophysiological consequences, and explore the therapeutic potential. That is, the question is no longer ‘Is the GABA system affected in schizophrenia?’ but why, how, when, and so what? As part of this shift in focus, there is an increasing iteration and integration between post mortem studies and those using othermodalities (e.g. functional neuroimaging, electrophysiology) and approaches (e.g. animal models, computational modelling). This special issue is therefore timely, and provides a range of reviews and original submissions. It has been expertly brought together by FrancineBenes, a pioneer and still a leader of the field. The articles provide a valuable, balanced,

Screening and kinetics of glutaminase and glutamate decarboxylase producing lactic acid bacteria from fermented Thai foods

L-glutaminase and glutamic acid decarboxylase (GAD) catalyzes the hydrolysis of L-glutamine and glutamate, respectively. L-glutaminase widely used in cancer therapy along with a combination of other enzymes and most importantly these enzymes were used in food industries, as a major catalyst of bioconversion. The current investigation was aimed to screen and select L-glutaminase, and GAD producing lactic acid bacteria (LAB). A total of 338 LAB were isolated from fermented meat, fermented fish, fermented soya bean, fermented vegetables and fruits. Among 338 isolates, 22 and 237 LAB has been found to be positive for L-glutaminase and GAD, respectively. We found that 30 days of incubation at 35 oC and pH 6.0 was the optimum condition for glutaminase activity by G507/1. G254/2 was found to be the best for GAD activity with the optimum condition of pH 6.5, temperature 40 oC and ten days of incubation. These LAB strains, G507/1 and G254/2, were identified as close relative of Lactobacillus brevis ATCC 14869 and Lactobacillus fermentum NBRC 3956, respectively by 16S rRNA sequencing. Further, improvements in up-stream of the fermentation process with these LAB strains are currently under development.

The role of neurotransmitter systems separate the hemispheres of the brain in action the new antiepileptic compounds AGB-31

Compound AGB-31, a monocarbamate derivative, is shown to possess a high antiepileptic activity. The mechanisms of antiepileptic action are connected with significant increase in glutamic acid decarboxylase activity in the left hemisphere of the brain, with trend of the glutamate content decrease in the left hemisphere and the tendency to increase GABA in both hemispheres. AGB-31 significantly (more than 3-fold) increases syntase nitric oxide activity in the left hemisphere and has a tendency to reduce the NO content in both hemispheres. AGB-31 significantly (by 63.4%), reduced glutathione peroxydase activity in the right hemisphere without changing it in the left, with a tendency to increase the activity of glutathione reductase in both hemispheres.

Synthesis and Relaxometric Characterization of a MRI Gd-Based Probe Responsive to Glutamic Acid Decarboxylase Enzymatic Activity

Novel contrast agent based systems, which selectively visualize specific cells, e.g., neurons in the brain, would be of substantial importance for the fast developing field of molecular magnetic resonance imaging (MRI). We report here the synthesis and in vitro validation of a Gd(III)-based contrast agent designed to act as an MRI responsive probe for imaging the activity of the enzyme glutamic acid decarboxylase (GAD) present in neurons. Upon the action of the enzyme, the Gd(III) complex increases its hydration sphere and takes on a residual positive charge that promotes its binding to endogenous macromolecules. Both effects contribute in a synergic way to generate a marked relaxation enhancement, which directly reports enzyme activity and will allow activity detection of GAD positive cells in vitro and in vivo selectively.

Postmortem stability of brain GABAergic and glutamatergic receptors and enzymes under ecological conditions

Neurochemical biomarkers have emerged as useful tools for assessing the subclinical neurological impacts of environmental toxicants in birds and other wildlife. Careful consideration of biomarker stability is necessary before implementing their use on tissues from ecological studies, as receptors and enzymes in the brain may be affected by postmortem conditions. The goal of this study was to evaluate the postmortem stability of key GABAergic and glutamatergic receptors (N-methyl-d-aspartate (NMDA), gamma-aminobutyric acid (GABAA-benzodiazepine)) and enzymes (glutamine synthetase (GS), glutamic acid decarboxylase (GAD)) under environmentally relevant field and storage conditions to determine their suitability as biomarkers. We exposed chicken embryo brains to postmortem environmental and storage conditions typical for ecological studies (12, 24, and 48h at 7°C or 25°C; 1, 4, and 8 weeks at −80°C or −20°C; 1 or 2 freeze thaw cycles), and measured [3H] MK-801 binding to the NMDA receptor, [3H] flunitrazipam binding to the GABAA-benzodiazepine receptor, GS activity, and GAD activity.We found that [3H] MK-801 binding is stable under all conditions studied. GAD activity was fairly stable under each storage and environmental temperatures for all durations, but was significantly less stable when stored at −20°C than at −80°C. [3H] flunitrazipam binding and GS activity were both impacted by environmental and storage temperature and duration, and might best be utilized in studies of samples with similar histories. Our findings here demonstrate that caution is warranted when comparing samples with different collection and storage histories, but that some biomarkers are fairly stable under various conditions.

The influence of NMDA and GABA(A) receptors and glutamic acid decarboxylase (GAD) activity on attention.

Attention dysfunction is the hallmark of cognitive deficits associated with major psychiatric illnesses including schizophrenia. Cognitive deficits of schizophrenia have been attributed to reduced function of the N-methyl-D-aspartate (NMDA) receptor or reduced expression of the gamma-aminobutyric acid (GABA)-synthesizing enzyme glutamic acid decarboxylase-67, which presumably leads to attenuated neurotransmission at GABA(A) receptors. The present study used a rodent model to compare the inhibition of NMDA and GABA(A) receptors, and GAD activity on attention. We tested the impact of inhibiting these proteins brain wide or in the anterior cingulate cortex (ACC), a prefrontal cortex region critical for attentional processing. Rats were trained on the three choice serial reaction time task (3-CSRT), an attention test. The impact of systemic or intra-ACC injection of drugs on performance was measured in well-trained rats. Reducing GABA(A) receptor function within the ACC with the direct antagonist SR95531 (1 or 3 ng/side) or brain wide using systemic injection of the benzodiazepine inverse agonist FG7142 (5 mg/kg) impaired accuracy and increased omissions. Systemic or intra-ACC inhibition of NMDA receptors using MK-801 (at 3 mg/kg or 3 μg, respectively) also impaired performance. Inhibition of GAD with 3-mercaptopropionic acid, even at high doses, had no effect on 3-CSRT accuracy or omissions when administered systemically or within the ACC. These data demonstrate that, while tonic stimulation of NMDA and GABA(A) receptors within the ACC are critical for attentional performance, reduction in GAD activity may have little functional significance and is not indicative of reduced GABA neurotransmission.

Effects of tutin and its derivatives on GAD and GABA-T in Pseudelatia separata (Lepidoptera: Noctuidae)

Tutin inhibits the receptors of the neurotransmitter γ-aminobutyric acid (GABA). In this work, we compared tutin with its derivatives, 2-(4-methoxybenzoyloxy)-tutin (T1), 2-(3,5-dinitrobenzoyloxy)-tutin (T2), 2-(6-chloronicotinoyloxy)-tutin (T3), and 2-(4-nitrobenzoyloxy)-tutin (T4), in terms of their effects on feeding and the feedback regulation mechanism of GABA metabolism. Spectrophotometry was used to determine the glutamic acid decarboxylase (GAD) and γ-aminobutyric acid transaminase (GABA-T) activities of these compounds in 3rd instar larvae of Pseudelatia separata. Clear antifeedant activities were manifested by T1 and especially T3, whereas T4 stimulated feeding in P. separata. GAD and GABA-T activities in larvae treated with all five toxins differed significantly from the control and from one other (P &lt; 0.05) in terms of treatment types and exposure times. The effects of the toxins on GAD and GABA-T were time-dependent for 48 h. GAD activities were inhibited by tutin, T2, T3, and T4, and were enhanced by T1. GABA-T activities were increased by all the toxins in varying degrees. Variation of GABA content in P. separata larvae resulted from the disturbance of GAD and GABA-T by tutin and its derivatives. Results suggest that T3, with its nicotinoyl group, is the most promising novel active ingredient for pest control.

A novel role of intestine epithelial GABAergic signaling in regulating intestinal fluid secretion

γ-Aminobutyric acid (GABA) is the primary inhibitory neurotransmitter in the central nervous system, and it is produced via the enzymatic activity of glutamic acid decarboxylase (GAD). GABA generates fast biological signaling through type A receptors (GABA(A)R), an anionic channel. Intriguingly, GABA is found in the jejunum epithelium of rats. The present study intended to determine whether a functional GABA signaling system exists in the intestinal epithelium and if so whether the GABA signaling regulates intestinal epithelial functions. RT-PCR, Western blot, and immunohistochemical assays of small intestinal tissues of various species were performed to determine the expression of GABA-signaling proteins in intestinal epithelial cells. Perforated patch-clamp recording was used to measure GABA-induced transmembrane current in the small intestine epithelial cell line IEC-18. The fluid weight-to-intestine length ratio was measured in mice that were treated with GABA(A)R agonist and antagonist. The effect of GABA(A)R antagonist on allergic diarrhea was examined using a mouse model. GABA, GAD, and GABA(A)R subunits were identified in small intestine epithelial cells of mice, rats, pigs, and humans. GABA(A)R agonist induced an inward current and depolarized IEC-18. Both GABA and the GABA(A)R agonist muscimol increased intestinal fluid secretion of rats. The increased intestinal secretion was largely decreased by the GABA(A)R antagonist picrotoxin or gabazine, but not by tetrodotoxin. The expression levels of GABA-signaling proteins were increased in the intestinal epithelium of mice that were sensitized and challenged with ovalbumin (OVA). The OVA-treated mice exhibited diarrhea, which was alleviated by oral administration of gabazine or picrotoxin. An endogenous autocrine GABAergic signaling exists in the mammalian intestinal epithelium, which upregulates intestinal fluid secretion. The intestinal GABAergic signaling becomes intensified in allergic diarrhea, and inhibition of this GABA-signal system alleviates the allergic diarrhea.

In vivo imaging of inhibitory, GABAergic neurons by MRI

The unambiguous detection of specific neuronal subtypes is up to now only possible with invasive techniques or optical imaging after genetic modification. High field magnetic resonance imaging (MRI) has the ability to visualize the brain structure and anatomy noninvasively, with high resolution--but missing the cell specific and functional information. Here we present a new tool for neuroimaging with MRI, enabling the selective detection of GABAergic neurons under in vivo conditions. The specific imaging contrast is achieved by a novel paramagnetic contrast agent, which responds to the activity of the enzyme glutamic acid decarboxylase--expressed solely by inhibitory neurons. The relaxivity of the complex is increased upon decarboxylation of two glutamic acid moieties, thus allowing increased water access to the inner and outer coordination spheres of the paramagnetic ion. The mechanism and specificity of activation were proven with tissue lysates and further applied to a differentiation protocol for murine embryonic stem cells. The relaxation enhancement was studied quantitatively and revealed decreased longitudinal relaxation times in the inhibitory neuron samples compared to the naïve stem cells in vitro and in vivo. Furthermore, this approach offers not only the discrimination of inhibitory, GABAergic neurons in the brain but also may expand the usefulness of MRI for functional imaging on a cellular level.