Gastrointestinal microbiome and cholelithiasis: Prospect in the nervous system.

Dan and colleagues recently published research suggesting that the gastrointestinal microbiome (microorganisms and metabolites) in cholelithiasis. They reviewed gallbladder stones, choledocholithiasis, and asymptomatic gallstones. Finally, their discussion was on the gastrointestinal. We focused on complementing the effect of the S1 protein and neuroinflammatory changes caused by severe acute respiratory syndrome coronavirus 2. Our contribution was about to involve the microbiota and the nervous system. They can have similar functions because they have similar pathways and advantages, bearing in mind γ-aminobutyric acid in schizophrenia and serotonin in Parkinson's disease. Therefore in the next few years, more research should be encouraged on the microbiota consequences for development, and mobility.

Probiotic Properties, Prebiotic Fermentability, and GABA-Producing Capacity of Microorganisms Isolated from Mexican Milk Kefir Grains: A Clustering Evaluation for Functional Dairy Food Applications.

Isolation and functional characterization of microorganisms are relevant steps for generating starter cultures with functional properties, and more recently, those related to improving mental health. Milk kefir grains have been recently investigated as a source of health-related strains. This study focused on the evaluation of microorganisms from artisanal Mexican milk kefir grains regarding probiotic properties, in vitro fermentability with commercial prebiotics (lactulose, inulin, and citrus pectin), and γ-aminobutyric acid (GABA)-producing capacity. Microorganisms were identified belonging to genera Lactococcus, Lactobacillus, Leuconostoc, and Kluyveromyces. The probiotic properties were assessed by aggregation abilities, antimicrobial activity, antibiotic susceptibility, and resistance to in vitro gastrointestinal digestion, showing a good performance compared with commercial probiotics. Most of isolates maintained a concentration above 6 log colony forming units/mL after the intestinal phase. Specific isolates of Kluyveromyces (BIOTEC009 and BIOTEC010), Leuconostoc (BIOTEC011 and BIOTEC012), and Lactobacillus (BIOTEC014 and BIOTEC15) showed a high fermentability in media supplemented with commercial prebiotics. The capacity to produce GABA was classified as medium for L. lactis BIOTEC006, BIOTEC007, and BIOTEC008; K. lactis BIOTEC009; L. pseudomesenteroides BIOTEC012; and L. kefiri BIOTEC014, and comparable to that obtained for commercial probiotics. Finally, a multivariate approach was performed, allowing the grouping of 2-5 clusters of microorganisms that could be further considered new promising cultures for functional dairy food applications.

Myorelaxation, respiratory depression and electrocardiographic changes caused by the administration of extract of açai (Euterpe oleracea Mart.) stone in rats.

The biological and pharmacological properties of natural polyphenols of the extract of Euterpe oleracea stone (EEOS) are associated with the central nervous system (CNS). To investigate the sedative and myorelaxant activity of EEOS in vivo, this study aimed to present the myorelaxant and sedative effects of EEOS in Wistar rats using spontaneous locomotor activity and motor electrophysiology. A total of 108 animals were used in the following experiments: a) behavioral tests (n = 27); b) electromyographic recordings of skeletal muscle (n = 27); c) respiratory muscle activity recordings (n = 27); d) cardiac muscle activity recordings (n = 27). The behavioral characteristics were measured according to the latency time of onset, the transient loss of posture reflex and maximum muscle relaxation. Electrodes were implanted in the gastrocnemius muscle and in the tenth intercostal space for electromyographic (EMG) signal capture to record muscle contraction, and in the D2 lead for electrocardiogram acquisition. After using the 300 mg/kg dose of EEOS intraperitoneally, a myorelaxant activity exhibited a lower frequency of contractility with an amplitude pattern of low and short duration at gastrocnemius muscle and intercostal muscle, which clearly describes a myorelaxant activity and changes in cardiac activity. The present report is so far the first study to demonstrate the myorelaxant activity of this extract, indicating an alternative route for açai stone valorization and its application in pharmaceutical fields.

The role of allopregnanolone in depressive-like behaviors: Focus on neurotrophic proteins.

Allopregnanolone (3α,5α-tetrahydroprogesterone; pharmaceutical formulation: brexanolone) is a neurosteroid that has recently been approved for the treatment of postpartum depression, promising to fill part of a long-lasting gap in the effectiveness of pharmacotherapies for depressive disorders. In this review, we explore the experimental research that characterized the antidepressant-like effects of allopregnanolone, with a particular focus on the neurotrophic adaptations induced by this neurosteroid in preclinical studies. We demonstrate that there is a consistent decrease in allopregnanolone levels in limbic brain areas in rodents submitted to stress-induced models of depression, such as social isolation and chronic unpredictable stress. Further, both the drug-induced upregulation of allopregnanolone or its direct administration reduce depressive-like behaviors in models such as the forced swim test. The main drugs of interest that upregulate allopregnanolone levels are selective serotonin reuptake inhibitors (SSRIs), which present the neurosteroidogenic property even in lower, non-SSRI doses. Finally, we explore how these antidepressant-like behaviors are related to neurogenesis, particularly in the hippocampus. The protagonist in this mechanism is likely the brain-derived neurotrophic factor (BFNF), which is decreased in animal models of depression and may be restored by the normalization of allopregnanolone levels. The role of an interaction between GABA and the neurotrophic mechanisms needs to be further investigated.

Renal Sensory Activity Regulates the γ-Aminobutyric Acidergic Inputs to the Paraventricular Nucleus of the Hypothalamus in Goldblatt Hypertension.

Renal sensory activity is centrally integrated within brain nuclei involved in the control of cardiovascular function, suggesting that renal afferents regulate basal and reflex sympathetic vasomotor activity. Evidence has shown that renal deafferentation (DAx) evokes a hypotensive and sympathoinhibitory effect in experimental models of cardiovascular diseases; however, the underlying mechanisms involved in this phenomenon need to be clarified, especially those related to central aspects. We aimed to investigate the role of renal afferents in the control of γ-aminobutyric acid (GABA)ergic inputs to the paraventricular nucleus (PVN) of the hypothalamus in renovascular hypertensive (2K1C) rats and their influence in the regulation of cardiovascular function. Hypertension was induced by clipping the left renal artery. After 4 weeks, renal DAx was performed by exposing the left renal nerve to a 33 mM capsaicin solution for 15 min. After 2 weeks of DAx, microinjection of muscimol into the PVN was performed in order to evaluate the influence of GABAergic activity in the PVN and its contribution to the control of renal sympathetic nerve activity (rSNA) and blood pressure (BP). Muscimol microinjected into the PVN triggered a higher drop in BP and rSNA in the 2K1C rats and renal DAx mitigated these responses. These results suggest that renal afferents are involved in the GABAergic changes found in the PVN of 2K1C rats. Although the functional significance of this phenomenon needs to be clarified, it is reasonable to speculate that GABAergic alterations occur to mitigate microglia activation-induced sympathoexcitation in the PVN of 2K1C rats.

Strawberry Intake Ameliorates Oxidative Stress and Decreases GABA Levels Induced by High-Fat Diet in Frontal Cortex of Rats.

It has been proposed that there is a correlation between high-fat diet (HFD), oxidative stress and decreased γ-aminobutyric acid (GABA) levels, but this has not been thoroughly demonstrated. In the present study, we determined the effects of strawberry extract intake on the oxidative stress and GABA levels in the frontal cortex (FC) of obese rats. We observed that an HFD increased lipid and protein oxidation, and decreased GABA levels. Moreover, UV-irradiated strawberry extract (UViSE) decreased lipid peroxidation but not protein oxidation, whereas non-irradiated strawberry extract (NSE) reduced protein oxidation but not lipid peroxidation. Interestingly, NSE increased GABA concentration, whereas UViSE was not as effective. In conclusion, our results suggest that an HFD increases oxidative damage in the FC, whereas strawberry extract intake may ameliorate the disturbances associated with HFD-induced oxidative damage.

Updates in the Diagnosis and Treatment of Paraneoplastic Neurologic Syndromes.

The disorders of the central nervous system associated with cancer by remote immune-mediated mechanisms are a heterogeneous group. These disorders encompass the classic paraneoplastic disorders and the recently recognized autoimmune encephalitis associated with antibodies against neuronal cell surface or synaptic proteins that occur with or without cancer association. In the last decade, the new surge of interest in neuronal diseases associated with anti-neuronal antibodies led to the rapid discovery of new forms of disease that have different manifestations and were not previously suspected to be immune mediated. The recognition of these syndromes is important because it may lead to early detection of an underlying malignancy and prompt initiation of treatment, improving chances for a better outcome.

Molecular cloning, expression, and immobilization of glutamate decarboxylase from Lactobacillus fermentum YS2

Background: GABA (γ-aminobutyric acid) is a four-carbon nonprotein amino acid that has hypotensive, diuretic, and tranquilizing properties. Glutamate decarboxylase (GAD) is the key enzyme to generate GABA. A simple and economical method of preparing and immobilizing GAD would be helpful for GABA production. In this study, the GAD from Lactobacillus fermentum YS2 was expressed under the control of a stress-inducible promoter and was purified and immobilized in a fusion form, and its reusability was investigated. Results: The fusion protein CBM-GAD was expressed in Escherichia coli DH5α carrying pCROCB-gadB, which contained promoter PrpoS, cbm3 (family 3 carbohydrate-binding module from Clostridium thermocellum) coding sequence, the gadB gene from L. fermentum YS2 coding for GAD, and the T7 terminator. After a one-step purification of CBM-GAD using regenerated amorphous cellulose (RAC) as an adsorbent, SDS-PAGE analysis revealed a clear band of 71 kDa; the specific activity of the purified fusion protein CBM-GAD reached 83.6 ± 0.7 U·mg-1. After adsorption onto RAC, the immobilized GAD with CBM3 tag was repeatedly used for GABA synthesis. The protein-binding capacity of RAC was 174 ± 8 mg·g-1. The immobilized CBM-GAD could repeatedly catalyze GABA synthesis, and 8% of the initial activities was retained after 10 uses. We tested the conversion of monosodium glutamate to GABA by the immobilized enzyme; the yield reached 5.15 g/L and the productivity reached 3.09 g/L·h. Conclusions: RAC could be used as an adsorbent in one-step purification and immobilization of CBM-GAD, and the immobilized enzyme could be repeatedly used to catalyze the conversion of glutamate to GABA.

Ghrelin activates hypophysiotropic corticotropin-releasing factor neurons independently of the arcuate nucleus.

Previous work has established that the hormone ghrelin engages the hypothalamic-pituitary-adrenal neuroendocrine axis via activation of corticotropin-releasing factor (CRF) neurons of the hypothalamic paraventricular nucleus (PVN). The neuronal circuitry that mediates this effect of ghrelin is currently unknown. Here, we show that ghrelin-induced activation of PVN CRF neurons involved inhibition of γ-aminobutyric acid (GABA) inputs, likely via ghrelin binding sites that were localized at GABAergic terminals within the PVN. While ghrelin activated PVN CRF neurons in the presence of neuropeptide Y (NPY) receptor antagonists or in arcuate nucleus (ARC)-ablated mice, it failed to do it so in mice with ghrelin receptor expression limited to ARC agouti gene related protein (AgRP)/NPY neurons. These data support the notion that ghrelin activates PVN CRF neurons via inhibition of local GABAergic tone, in an ARC-independent manner. Furthermore, these data suggest that the neuronal circuits mediating ghrelin's orexigenic action vs. its role as a stress signal are anatomically dissociated.

Neonatal low-protein diet reduces the masticatory efficiency in rats.

Little is known about the effects of undernutrition on the specific muscles and neuronal circuits involved in mastication. The aim of this study was to document the effects of neonatal low-protein diet on masticatory efficiency. Newborn rats whose mothers were fed 17% (nourished (N), n 60) or 8% (undernourished (U), n 56) protein were compared. Their weight was monitored and their masticatory jaw movements were video-recorded. Whole-cell patch-clamp recordings were performed in brainstem slice preparations to investigate the intrinsic membrane properties and N-methyl-d-aspartate-induced bursting characteristics of the rhythmogenic neurons (N, n 43; U, n 39) within the trigeminal main sensory nucleus (NVsnpr). Morphometric analysis (N, n 4; U, n 5) were conducted on masseteric muscles serial cross-sections. Our results showed that undernourished animals had lower numbers of masticatory sequences (P=0·049) and cycles (P=0·045) and slower chewing frequencies (P=0·004) (N, n 32; U, n 28). Undernutrition reduced body weight but had little effect on many basic NVsnpr neuronal electrophysiological parameters. It did, however, affect sag potentials (P<0·001) and rebound firing (P=0·005) that influence firing pattern. Undernutrition delayed the appearance of bursting and reduced the propensity to burst (P=0·002), as well as the bursting frequency (P=0·032). Undernourished animals showed increased and reduced proportions of fibre type IIA (P<0·0001) and IIB (P<0·0001), respectively. In addition, their fibre areas (IIA, P<0·001; IIB, P<0·001) and perimeters (IIA, P<0·001; IIB, P<0·001) were smaller. The changes observed at the behavioural, neuronal and muscular levels suggest that undernutrition reduces chewing efficiency by slowing, weakening and delaying maturation of the masticatory muscles and the associated neuronal circuitry.

Maximising the phytochemical content and antioxidant activity of Ecuadorian brown rice sprouts through optimal germination conditions.

Germinated brown rice (GBR) is considered a healthy alternative to white rice in the fight against chronic diseases. As the functional quality of GBR depends on genotype and germination conditions, the objectives were to identify suitable Ecuadorian brown rice cultivars and optimal germination time and temperature to maximise γ-aminobutyric acid (GABA), total phenolics compounds (TPC) and antioxidant activity of GBR. Regression models for the prediction of phytochemical composition and antioxidant activity in GBR were also obtained. Germination improved GABA, TPC and antioxidant activity in all cultivars. Maximum GABA and antioxidant activity were attained at 34 °C for 96 h, while the highest TPC was found at 28 °C for 96 h in all cultivars. GBR cv. GO displayed the highest antioxidant activity and cv. 15 was the most effective at accumulating GABA and TPC in the optimal germination conditions. Therefore, Ecuadorian GBR could be used for the preparation of functional foods serving as preventative strategies in combating chronic diseases.

Submerged fermentation of Lactobacillus rhamnosus YS9 for γ-aminobutyric acid (GABA) production.

γ-Aminobutyric acid (GABA) is a major inhibitory neurotransmitter in central nervous system, and its application in drugs and functional foods has attracted great attention. To enhance production of γ-aminobutyric acid, Lactobacillus rhamnosus YS9, a strain isolated from Chinese traditional fermented food pickled vegetable, was grown under submerged fermentation. Its cultivation conditions were investigated. When culture pH condition was adjusted to the optimal pH of glutamate decarboxylase activity, culture of Lb. rhamnosus YS9 in medium supplemented with 200 mM of monosodium glutamate and 200 µM of pyridoxal phosphate (PLP), produced 187 mM of GABA.

Quercetin antagonism of GABAAρ1 receptors is prevented by ascorbic acid through a redox-independent mechanism.

Quercetin is a natural flavonoid widely distributed in plants that acts as a neuroprotective agent and modulates the activity of different synaptic receptors and ion channels, including the ionotropic GABA receptors. GABA(Aρ1) receptors were shown to be antagonized by quercetin, but the mechanisms underlying these antagonistic actions are still unknown. We have analyzed here if the antagonistic action produced by quercetin on GABA(Aρ1) receptors was related to its redox activity or due to alternative mechanism/s. Homomeric GABA(Aρ1) receptors were expressed in frog oocytes and GABA-evoked responses electrophysiologically recorded. Quercetin effects on GABA(Aρ1) receptors were examined in the absence or presence of ascorbic acid. Chemical protection of cysteines by selective sulfhydryl reagents and site directed mutagenesis experiments were also used to determine ρ1 subunit residues involved in quercetin actions. Quercetin antagonized GABA(Aρ1) receptor responses in a dose-dependent, fast and reversible manner. Quercetin inhibition was prevented in the presence of ascorbic acid, but not by thiol reagents that modify the extracellular Cys-loop of these receptors. H141, an aminoacidic residue located near to the ρ1 subunit GABA binding site, was involved in the allosteric modulation of GABA(Aρ1) receptors by several agents including ascorbic acid. Quercetin similarly antagonized GABA-evoked responses mediated by mutant (H141D)GABA(Aρ1) and wild-type receptors, but prevention exerted by ascorbic acid on quercetin effects was impaired in mutant receptors. Taken together the present results suggest that quercetin antagonistic actions on GABA(Aρ1) receptors are mediated through a redox-independent allosteric mechanism.

Changes on auditory physiology in response to the inactivation of amygdala nuclei in high anxiety rats expressing learned fear.

The inferior colliculus (IC) is primarily involved in the processing of acoustic stimuli, including those emitted by prey and predators. The role of the central nucleus of the IC (CIC) in fear and anxiety has been suggested based on electrophysiological, behavioral and immunohistochemical studies. The reactivity of high-anxiety rats (HA) to diverse challenges is different from low-anxiety ones (LA). In humans and laboratory animals, pathological anxiety is often accompanied by heightened vigilance and alertness, hyperactivity of the amygdala (AM), and increased amplitude of the auditory evoked potentials (AEP) from the IC. This study aims to evaluate the influence of the inactivation of the central (CEA) and basolateral (BLA) nuclei of the amygdala, after local infusions of the full GABAA agonist muscimol (1nmol/0.2µl), on the AEP elicited in the CIC of rats tested under a learned fear state. Our results showed that both BLA and CEA inactivation change the expression of conditioned fear, in a paradigm using the context as the conditioned stimulus (CS). These changes are correlated to the innate anxiety levels of the animals. It is supposed that this shortcoming is in addition to the imbalance between the regulatory role of the top-down and bottom-up processes in the control of anxiety.

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

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.

Enhancement of gamma-aminobutyric acid in a fermented red seaweed beverage by starter culture Lactobacillus plantarum DW12

Lactobacillus plantarum DW12, a gamma-aminobutyric acid (GABA) producing strain, was used as a starter culture to produce a functional fermented red seaweed beverage (FSB). Optimal conditions for producing FSB were established using Central Composite Design by varying the amounts of monosodium glutamate (MSG), sucrose and the initial pH in MRS medium. After a verification test, 1 percent MSG, 6 percent sucrose and an initial pH of 6 were selected. Four treatments were tested: traditional formula (A), red seaweed-cane sugar-potable water = 3:1:10, w/w/v, initial pH 6; the traditional formula with a 5 percent starter culture consisting of 4.1 x 10(9) CFU of DW 12/ml (B); formula A modified by changing the amounts of cane sugar and MSG to 6 percent and 1 percent, respectively (C); formula C with a 5 percent starter culture added (D). Comparison among the 4 treatments showed that the treatment D produced the highest amount of GABA (4000 mg/L) during days 45-60 while the GABA content of A, B and C treatments was 340, 730 and 1690 mg/L, respectively. However, the results of the sensory test for the treatments C and D showed that the presence of MSG produced an unsatisfactory salty taste. All finished products from the 4 treatments met Thai standard guidelines for chemical and microbiological qualities after 120 days. The results indicated that enrichment of the GABA content in FSB is possible by adding MSG and the GABA producing strain DW12; however, the appropriate amount of MSG addition should be further studied.