Gamma-aminobutyric Acid Secretion Research Articles

Abstract 3858: Peristalsis-like mechanical stimuli in the intestinal milieu promotes colorectal cancer invasion through GABAergic signaling changes in an organ-on-chip platform

Abstract Mechanical forces in the tumor microenvironmental milieu are often understudied due to a lack of relevant preclinical model systems. Here we describe a microfluidic organ-on-chip platform that incorporates tissue-tissue interfaces and physical forces to aid in the examination of colorectal cancer (CRC) progression. A major advantage of this model is the ability to mimic peristalsis, a physiological process occurring in the gut. Live-cell imaging with a 3D printed organ-on-chip cradle was used to quantify the number of CRC cells that have invaded from the epithelial channel into the vascular channel over time. We determined that peristalsis-like motions in our organ-on-chip model enhanced the invasion capacity of CRC cells, mimicking intravasation. We subsequently examined the effluent media in stretched compared to non-stretched conditions using mass spectrometry based metabolomics and discovered an increase in the secretion of gamma-aminobutyric acid (GABA) by CRC cells, implicating peristalsis-mediated tumor cell invasion with neurotransmitter release. Inhibitors targeting the GABA-A receptor reversed the observed tumor cell invasion phenotype when cells were subjected to peristalsis-like motions. Interestingly, even in the absence of peristalsis, tumor cell invasion was promoted when GABA agonists were introduced into the organ-on-chip system. This work reveals important interactions between CRC cells and their microenvironment, and that disrupting GABAergic signaling might be an approach to prevent or delay cancer progression. Citation Format: Carly Strelez, Sujatha Chilakala, Kimya Ghaffarian, Ah Young Yoon, Jonathan Katz, Shannon M. Mumenthaler. Peristalsis-like mechanical stimuli in the intestinal milieu promotes colorectal cancer invasion through GABAergic signaling changes in an organ-on-chip platform [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3858.

Endogenous Levels of Gamma Amino-Butyric Acid Are Correlated to Glutamic-Acid Decarboxylase Antibody Levels in Type 1 Diabetes.

Gamma-aminobutyric acid (GABA) is an important inhibitory neurotransmitter in the central nervous system (CNS) and outside of the CNS, found in the highest concentrations in immune cells and pancreatic beta-cells. GABA is gaining increasing interest in diabetes research due to its immune-modulatory and beta-cell stimulatory effects and is a highly interesting drug candidate for the treatment of type 1 diabetes (T1D). GABA is synthesized from glutamate by glutamic acid decarboxylase (GAD), one of the targets for autoantibodies linked to T1D. Using mass spectrometry, we have quantified the endogenous circulating levels of GABA in patients with new-onset and long-standing T1D and found that the levels are unaltered when compared to healthy controls, i.e., T1D patients do not have a deficit of systemic GABA levels. In T1D, GABA levels were negatively correlated with IL-1 beta, IL-12, and IL-15 15 and positively correlated to levels of IL-36 beta and IL-37. Interestingly, GABA levels were also correlated to the levels of GAD-autoantibodies. The unaltered levels of GABA in T1D patients suggest that the GABA secretion from beta-cells only has a minor impact on the circulating systemic levels. However, the local levels of GABA could be altered within pancreatic islets in the presence of GAD-autoantibodies.

Transcriptomics reveal different metabolic strategies for acid resistance and gamma-aminobutyric acid (GABA) production in select Levilactobacillus brevis strains

BackgroundOf the many neurotransmitters in humans, gamma-aminobutyric acid (GABA) shows potential for improving several mental health indications such as stress and anxiety. The microbiota-gut-brain axis is an important pathway for GABAergic effects, as microbially-secreted GABA within the gut can affect host mental health outcomes. Understanding the molecular characteristics of GABA production by microbes within the gut can offer insight to novel therapies for mental health.ResultsThree strains of Levilactobacillus brevis with syntenous glutamate decarboxylase (GAD) operons were evaluated for overall growth, glutamate utilization, and GABA production in typical synthetic growth media supplemented with monosodium glutamate (MSG). Levilactobacillus brevis Lbr-6108™ (Lbr-6108), formerly known as L. brevis DPC 6108, and Levilactobacillus brevis Lbr-35 ™ (Lbr-35) had similar growth profiles but differed significantly in GABA secretion and acid resistance. Lbr-6108 produced GABA early within the growth phase and produced significantly more GABA than Lbr-35 and the type strain Levilactobacillus brevis ATCC 14869 after the stationary phase. The global gene expression during GABA production at several timepoints was determined by RNA sequencing. The GAD operon, responsible for GABA production and secretion, activated in Lbr-6108 after only 6 h of fermentation and continued throughout the stationary phase. Furthermore, Lbr-6108 activated many different acid resistance mechanisms concurrently, which contribute to acid tolerance and energy production. In contrast, Lbr-35, which has a genetically similar GAD operon, including two copies of the GAD gene, showed no upregulation of the GAD operon, even when cultured with MSG.ConclusionsThis study is the first to evaluate whole transcriptome changes in Levilactobacillus brevis during GABA production in different growth phases. The concurrent expression of multiple acid-resistance mechanisms reveals niche-specific metabolic functionality between common human commensals and highlights the complex regulation of GABA metabolism in this important microbial species. Furthermore, the increased and rapid GABA production of Lbr-6108 highlights the strain’s potential as a therapeutic and the overall value of screening microbes for effector molecule output.

Modulation of neurosecretion and approaches for its multistep analysis

BackgroundNeurosecretion is the multistep process occurring in separate spatial and temporal cellular boundaries which complicates its comprehensive analysis. Most of the research are focused on one distinct stage of synaptic vesicle recycling. Here, we describe approaches for complex analysis of synaptic vesicle (SV) endocytosis and separate steps of exocytosis at the level of presynaptic bouton and highly purified SVs. MethodsProposed fluorescence-based strategies and analysis of neurotransmitter transport provided the advantages in studies of exocytosis steps. We evaluated SV docking/tethering, their Ca2+-dependent fusion and release of neurotransmitters gamma-aminobutyric acid (GABA) and glutamate in two animal models. ResultsApproaches enabled us to study: 1) endocytosis/Ca2+-dependent release of fluorescent carbon nanodots (CNDs) during stimulation of nerve terminals; 2) the action of levetiracetam, modulator of SV glycoprotein SV2, on fusion competence of SVs and stimulated release of GABA and glutamate; 3) impairments of several steps of neurosecretion under vitamin D3 deficiency. ConclusionsOur algorithm enabled us to verify the method validity for multidimensional analysis of SV turnover. By increasing SV docking and the size of readily releasable pool (RRP), levetiracetam is able to selectively enhance the stimulated GABA secretion in hippocampal neurons. Findings suggest that SV2 regulates RRP through impact on the number of docked/primed SVs. General significanceMethodology can be widely applied to study the stimulated neurosecretion in presynapse, regulation of SV docking, their Ca2+-dependent fusion with target membranes, quantitative analysis of expression of neuron-specific proteins, as well as for testing the efficiency of pre-selected designed neuroactive substances.

Different effects of etomidate and propofol on memory in immature rats

This study is to investigate the effects of etomidate and propofol on memory and possible involved mechanisms using immature rats. Forty-eight rats randomly received intraperitoneal injection of 5 mg/kg etomidate ( n = 16), 50 mg/kg propofol ( n = 16) or normal saline (control, n = 16). Three hours after awakening, memory was assessed by Y-maze test using 10 rats in each drug group. Gamma-aminobutyric acid (GABA) content in hippocampal tissue was measured using six rats in each group. Etomidate group had more total reaction time (TRT) compared with the control group in Y-maze test ( p < 0.05). No other difference between these two groups was observed. Propofol group had less number of correct response ( p < 0.01) and more TRT ( p < 0.05) in Y-maze test, as well as more GABA concentration detected in hippocampal tissue ( p < 0.01) than the control group. Propofol group also showed less number of correct response ( p < 0.05) and more hippocampal GABA concentration ( p < 0.01) compared with etomidate group. Etomidate does not show significant effects on memory in rat and further investigation is required. Propofol can affect memory in rat possibly via increasing the synthesis and/or secretion of GABA as one of the factors.

Spinal cord stimulation: panacea for incurable diseases?

A spinal cord stimulator is usually used as an electro-modulator for pain treatment. Shealy et al. [1] first reported the use of a spinal cord stimulator in an elderly male suffering with terminal bronchogenic carcinoma and right-sided chest pain in 1967. Since then, spinal cord stimulation (SCS) has been applied to many disease entities, such as neuropathic pain, failed back surgery syndrome, complex regional pain syndrome, and ischemic peripheral vascular disease [2-4]. The indications for SCS have been extended to include intractable angina pain, interstitial cystitis, intractable pain, headache, and non-painrelated applications such as refractory congestive heart failure, intractable spasticity, or treatment of cerebral vasospasm after subarachnoid hemorrhage [5-9]. SCS systems are composed of three components: leads/electrodes, a generator/power source, and a programmer/controller [5]. Leads can be divided into percutaneous leads and paddle leads. Small-sized rechargeable implanted batteries are becoming increasingly common. An electrical current is applied to the dorsal columns, creating a tingling sensation in the dermatomes whose afferent fibers traverse the regions being stimulated. The physician can control the degree, range of current, pulse width, and rate of the stimulation wave to optimize pain control. In this issue of the Korean Journal of Anesthesiology, Ryu et al. [10] report the effect of cervical SCS for treatment of digit ulcers in a patient with Buerger’s disease. Thromboangiitis obliterans is a type of segmental inflammatory vasculitis asso ciated with severe pain and ulcers, which involves the small or medium-sized arteries and veins. Treatment of this painful ulcer is difficult and often involves amputation of the affected limb. This case report suggests that cervical SCS is a good choice for reducing pain and healing of an ulcer in patients with vasculitis. There are some reports of the effectiveness of SCS for patients with Raynaud’s disease and Buerger’s disease [11-13]. Cook et al. [14] reported using electronic stimulation of spinal nerves and the dorsal root in patients with a vascular disease of the limbs. Since then, SCS has been successfully used to treat atherosclerotic and vasospastic peripheral arterial disease. SCS improves microcirculation by inhibiting sympathetic vasoconstriction. SCS reduces pain in Buerger’s disease by stimulating the secretion of a number of inhibitory neurotransmitters and promoting the secretion of gamma aminobutyric acid, substance P, and serotonin at the spinal nerve dorsal horn [15,16]. Wolter and Kieselbach [17] reported the long-term outcomes of patients with cervical SCS. Treatment indications included patients with causalgia, cervicobrachialgia, complex regional pain syndrome, phantom pain, and Raynaud’s syndrome [18]. Pain reduction and paresthesia coverage were effective in almost all patients, and treatment satisfaction was high. No severe complications, such as neurological deficits or infection, were reported. They concluded that cervical SCS appears to be a safe and efficacious treatment option for upper limb neuropathic pain. Nevertheless, SCS has some disadvantages. The procedure for implanting SCS is invasive, so it is associated with complications such as infection, bleeding, and dural puncture [19]. The effectiveness of SCS is limited, and regular checks and continuous follow-ups are needed. Furthermore, SCS is expensive because of the limited medical insurance coverage in Korea. However, SCS is cost effective as compared with conservative management alone in cases with medical insurance coverage. Advances in SCS technology and understanding of the

Global Versus Local Perspectives on Schizophrenia

Back to table of contents Previous article Next article LETTERFull AccessGlobal Versus Local Perspectives on SchizophreniaSareh ZendehrouhFatemeh BakouieShahriar Gharibzadeh,Sareh ZendehrouhSearch for more papers by this authorFatemeh BakouieSearch for more papers by this authorShahriar GharibzadehSearch for more papers by this author,Published Online:1 Apr 2009AboutSectionsPDF/EPUB ToolsAdd to favoritesDownload CitationsTrack Citations ShareShare onFacebookTwitterLinked InEmail To the Editor: Recent neurobiological studies indicate that schizophrenia may be a neurodevelopmental and progressive disorder with multiple biochemical abnormalities involving dopamine, serotonin, glutamate, and gamma-aminobutyric acid secretion. In postmortem tissue studies, structural abnormalities and alterations in synaptic connectivity have been observed in the intracortical circuitry of the prefrontal dorsal cortex of schizophrenia patients. These morphological changes could be the sequelae of earlier environmental insults and genetic processes. There are probably multiple susceptibility genes, each of small effect, which act in conjunction with environmental factors as obstetric abnormalities, intrauterine infection, and abnormal nutrition. Candidate identified genes could influence neurodevelopment, synaptic plasticity, and neurotransmission. 1 The recent findings are consistent with notions that epigenetic factors play a major role in the disease process and epigenetic factors may continue to influence the expression of the affected genes in adulthood. 2 It is also demonstrated that schizophrenia is a dynamical disease—i.e., that important aspects of schizophrenia can be understood on the basis of concepts of the theory of nonlinear dynamical systems. 3 Chaotic dynamical systems are characterized by a lawful but delicate sensitivity to initial conditions. This leads to the observation that initially similar behaviors evolve into a striking divergence of behavioral patterns over time. Therefore, small differences in the input can result in an entirely different sequence of outputs. 4 We believe that genes act as initial conditions and environmental factors are the control parameters of a chaotic human brain system, which can force the system to special states with particular characteristics (e.g., schizophrenia). Since in chaotic dynamical systems, control parameters can direct the system to special states, environmental factors play a key role in aggravation or decline of schizophrenia symptoms. This complies with some reports that showed high relapse rate of schizophrenia in families that expressed high emotion, 5 which can be considered as a positive control factor. Based on our hypothesis, we propose that schizophrenia could be considered a chaotic model and the global features of the disease could be extracted instead of paying attention to local detailed features. In such a perspective, the manner of managing the disease will be changed. For example, we think that it is not necessary to include the effects of each part of the brain circuits and to correct any change in the amount of neurotransmitters; instead it is better to model the disease state as a chaotic system and recognize the interaction of different environmental factors, such as maternal health, birth complications, emotional states, familial relationships, etc., on it and try to minimize the pathological effects of them. We stress that it is not the compartments but the global interactions of different elements that play major roles in the disease and deserve attention. Modeling the behavior of schizophrenia on the basis of chaos theory to control the disease seems to be a good place to start.Biomedical Engineering, Amirkabir University of Technology, Tehran, Iran

γ-aminobutyric acid secreted from islet β-cells modulates exocrine secretion in rat pancreas

To investigate the role of endogenous gamma-amino-butyric acid (GABA) in pancreatic exocrine secretion. The isolated, vascularly perfused rat pancreas was employed in this study to eliminate the possible influences of extrinsic nerves and hormones. Cholecystokinin (CCK; 10 pmol/L) was intra-arterially given to stimulate exocrine secretion of the pancreas. Glutamine, a major precursor of GABA, which was given intra-arterially at concentrations of 1, 4 and 10 mmol/L, dose-dependently elevated the CCK-stimulated secretions of fluid and amylase in the normal pancreas. Bicuculline (10 micromol/L), a GABA(A) receptor antagonist, blocked the enhancing effect of glutamine (4 mmol/L) on the CCK-stimulated exocrine secretions. Glutamine, at concentrations of 1, 4 and 10 mmol/L, dose-dependently increased the GABA concentration in portal effluent of the normal pancreas. The effects of glutamine on the CCK-stimulated exocrine secretion as well as the GABA secretion were markedly reduced in the streptozotocin-treated pancreas. GABA could be secreted from beta-cells into the islet-acinar portal system after administration of glutainine, and could enhance the CCK-stimulated exocrine secretion through GABA(A) receptors. Thus, GABA in islet beta-cells is a hormone modulating pancreatic exocrine secretion.

A gamma-aminobutyric acid transporter driven by a proton pump is present in synaptic-like microvesicles of pancreatic beta cells.

A variety of peptide-secreting endocrine cells contain a population of recycling microvesicles that share several major membrane polypeptides with neuronal synaptic vesicles (SVs). The function of these synaptic-like microvesicles (SLMVs) remains to be elucidated. It was previously suggested that SLMVs of pancreatic beta cells may store and secrete gamma-aminobutyric acid (GABA). GABA, the major nonpeptide inhibitory neurotransmitter of the central nervous system, is stored in and secreted from SVs. GABA uptake into SVs is mediated by a transporter that is driven by a vacuolar proton ATPase. GABA is also present at high concentration in the endocrine pancreas where it is selectively localized in insulin-secreting beta cells, the core cells of pancreatic islets. GABA is not present in peripheral islet cells (mantle cells), represented primarily by glucagon-secreting alpha cells. In this study, an immunoisolation procedure was used to purify SLMVs from cell lines derived from mouse beta cells and alpha cells. SLMVs obtained from the beta-cell line, but not those obtained from the alpha-cell line, displayed a GABA-transport activity dependent upon a proton electrochemical gradient generated by a vacuolar proton ATPase. These data support the hypotheses that (i) SLMVs have a secretory function similar to that of SVs and (ii) beta-cell SLMVs are involved in the secretion of GABA, which in turn may have a paracrine function on mantle cells of the islet.

Gamma-aminobutyric acid-glutamate interaction in the control of somatostatin release from hypothalamic neurons in primary culture: in vivo corroboration.

Recent studies have provided new data on the neuroendocrine role of glutamate (the major excitatory neurotransmitter) on somatostatin release. The neuroendocrine role of gamma-aminobutyric acid (GABA) (the major inhibitory neurotransmitter) on this same secretion, is also well established. Our objective was thus to investigate whether GABA and glutamate, which have opposite neurotransmission signals, could interact in the control of hypothalamic somatostatin release. Pharmacological manipulations of the two types of receptors were performed in vitro on primary cultures of hypothalamic neurons secreting somatostatin. We found that tonic release of somatostatin was reduced by 76% in the presence of tetrodotoxin (TTX) and was regulated by endogenous secretion of glutamate and GABA. CGS 19755, a highly selective N-methyl-D-aspartate (NMDA) receptor antagonist, significantly reduced tonic somatostatin secretion whereas it was strongly increased by picrotoxin and bicuculline, two GABAA antagonists. When CGS 19755 was applied with picrotoxin, somatostatin release was the same as levels obtained in the control group with TTX. GABA reduced tonic somatostatin release (in the presence or absence of TTX), and glutamate-stimulated secretion in a dose-dependent manner. Picrotoxin stimulation of tonic somatostatin release was additive with that obtained after glutamate stimulation and was also dose-dependent. This interaction was also studied in vivo in unanesthetized rats bearing a push-pull cannula stereotaxically implanted into the median eminence. Ip injected CGS 19755 (an antagonist that can freely permeate the blood-brain barrier) completely blocked the peak secretion of somatostatin observed after ip picrotoxin administration, whereas there was no significant effect when it was injected alone. These findings corroborated our in vitro data and allow us to postulate that GABA and glutamate interact in the control of somatostatin.

Gamma-Aminobutyric Acid Neurons in the Preoptic/Anterior Hypothalamic Area Synchronize the Phasic Activity of the Gonadotropin-Releasing Hormone Pulse Generator in Ovariectomized Rats

To achieve a bolus-type release of gonadotropin-releasing hormone (GnRH) into the portal vessels it is required that GnRH neurons exert phasic and synchronous activity. The activity of GnRH neurons appears to be under an inhibitory influence of the amino acid neurotransmitter gamma-aminobutyric acid (GABA). Preoptic GABA concentrations in ovariectomized (OVX) rats decrease prior to a luteinizing hormone (LH) episode. This reduction of GABAergic activity in the preoptic/anterior hypothalamic area (PO/AH) may be the synchronizing signal for the simultaneous release of GnRH from the hypothalamus. To further study the role of GABA in controlling the GnRH pulse generator we applied GABA, 3-mercaptopropionic acid (MPA) or bicuculline (BIC) locally into the PO/AH by means of push-pull cannulae (PPC). PPC were implanted into the PO/AH of OVX rats and the contralateral, not PPC-implanted PO/AH was lesioned electrochemically. The effects of GABA, MPA or BIC on the GnRH pulse generator were determined by measuring LH levels in blood samples collected in 5-min intervals. Local application of GABA into the PO/AH caused a pronounced reduction of average LH secretion and abolished LH pulsatility. This inhibitory effect was completely reversible. Results of intrapreoptic MPA application on GABA secretion were variable. In only 45% of treated rats MPA caused a reduction of GABA secretion which was associated with a cessation of pulsatile LH release. A pronounced reduction of LH secretion and pulsatility was observed upon local application of the GABA antagonist BIC. Based on these data we propose that oscillating GABA levels in the PO/AH may be the synchronizing signal which triggers bolus release of GnRH into the portal vessels.(ABSTRACT TRUNCATED AT 250 WORDS)

Gamma amino butyric acid (GABA) levels in hypophyseal stalk plasma of rats

To determine the possible physiologic contribution of GABA to the tonic hypothalamic inhibition of adenohypophyseal prolactin secretion, we compared GABA levels in hypophyseal stalk plasma with those found in the peripheral circulation. Hypophyseal stalk blood was collected via a parapharyngeal approach from 8 urethane anesthetized diestrus rats. Peripheral blood was collected simultaneously from the external jugular vein of the same rat at a rate similar to hypophyseal stalk blood flow. Blood samples resulting from a single 4 hr collection per animal were centrifuged, and the plasma stored frozen before ethanol extraction and assay using a radioreceptor method. GABA levels in hypophyseal stalk plasma ( 909±171 pmol/ml; X ± S.E.M. ) were not significantly higher than levels in peripheral plasma (845±182; p>0.05), indicating little or no secretion of GABA by the median eminence.