GABA Receptor Antagonist Research Articles

Acute deep brain stimulation of the paraventricular nucleus of the hypothalamus increases brown adipose tissue thermogenesis in rats

Brown adipose tissue (BAT) activity is controlled by the sympathetic nervous system. Activation of BAT has shown significant promise in preclinical studies to elicit weight loss. Since the hypothalamic paraventricular nucleus (PVN) contributes to the regulation of BAT thermogenic activity, we sought to determine the effects of electrical stimulation of the PVN as a model of deep brain stimulation (DBS) for increasing BAT sympathetic nerve activity (SNA). The rostral raphe pallidus area (rRPa) was also chosen as a target for DBS since it contains the sympathetic premotor neurons for BAT. Electrical stimulation (100 µA, 100 µs, 100 Hz, for 5 min at a 50 % duty cycle) of the PVN increased BAT SNA and BAT thermogenesis. These effects were prevented by a local nanoinjection of bicuculline, a GABAA receptor antagonist. We suggest that electrical stimulation of the PVN elicited local release of GABA, which inhibited BAT sympathoinhibitory neurons in PVN, thereby releasing a restraint on BAT SNA. Electrical stimulation of the rRPa inhibited BAT thermogenesis and this was prevented by a local nanoinjection of bicuculline, suggesting that local release of GABA suppressed BAT SNA. Electrical stimulation of the PVN activates BAT metabolism via a mechanism that may include activation of local GABAA receptors. These findings contribute to our understanding of the mechanisms underlying the effects of DBS in the regulation of fat metabolism and provide a foundation for further DBS studies targeting hypothalamic circuits regulating BAT thermogenesis as a therapy for obesity.

Gamma-aminobutyric acid (GABA) suppresses hemocyte phagocytosis by binding to GABA receptors and modulating corresponding downstream pathways in blood clam, Tegillarca granosa

Although accumulating data demonstrated that gamma-aminobutyric acid (GABA), an inhibitory neurotransmitter, plays an important regulatory role in immunity of vertebrates, its immunomodulatory function and mechanisms of action remain poorly understood in invertebrates such as bivalve mollusks. In this study, the effect of GABA on phagocytic activity of hemocytes was evaluated in a commercial bivalve species, Tegillarca granosa. Furthermore, the potential regulatory mechanism underpinning was investigated by assessing potential downstream targets. Data obtained demonstrated that in vitro GABA incubation significantly constrained the phagocytic activity of hemocytes. In addition, the GABA-induced suppression of phagocytosis was markedly relieved by blocking of GABAA and GABAB receptors using corresponding antagonists. Hemocytes incubated with lipopolysaccharides (LPS) and GABA had significant higher K+-Cl- cotransporter 2 (KCC2) content compared to the control. In addition, GABA treatment led to an elevation in intracellular Cl−, which was shown to be leveled down to normal by blocking the ionotropic GABAA receptor. Treatment with GABAA receptor antagonist also rescued the suppression of GABAA receptor-associated protein (GABARAP), KCC, TNF receptor associated factor 6 (TRAF6), inhibitor of nuclear factor kappa-B kinase subunit alpha (IKKα), and nuclear factor kappa B subunit 1 (NFκB) caused by GABA incubation. Furthermore, incubation of hemocytes with GABA resulted in a decrease in cAMP content, an increase in intracellular Ca2+, and downregulation of cAMP-dependent protein kinase (PKA), calmodulin kinase II (CAMK2), calmodulin (CaM), calcineurin (CaN), TRAF6, IKKα, and NFκB. All these above-mentioned changes were found to be evidently relieved by blocking the metabotropic G-protein-coupled GABAB receptor. Our results suggest GABA may play an inhibitory role on phagocytosis through binding to both GABAA and GABAB receptors, and subsequently regulating corresponding downstream pathways in bivalve invertebrates.

Cortical interneuron development is affected in 4H leukodystrophy.

4H leukodystrophy is a rare genetic disorder classically characterized by hypomyelination, hypodontia and hypogonadotropic hypogonadism. With the discovery that 4H is caused by mutations that affect RNA polymerase III, mainly involved in the transcription of small non-coding RNAs, patients with atypical presentations with mainly a neuronal phenotype were also identified. Pathomechanisms of 4H brain abnormalities are still unknown and research is hampered by a lack of preclinical models. We aimed to identify cells and pathways that are affected by 4H mutations using induced pluripotent stem cell models. RNA sequencing analysis on induced pluripotent stem cell-derived cerebellar cells revealed several differentially expressed genes between 4H patients and control samples, including reduced ARX expression. As ARX is involved in early brain and interneuron development, we studied and confirmed interneuron changes in primary tissue of 4H patients. Subsequently, we studied interneuron changes in more depth and analysed induced pluripotent stem cell-derived cortical neuron cultures for changes in neuronal morphology, synaptic balance, network activity and myelination. We showed a decreased percentage of GABAergic synapses in 4H, which correlated to increased neuronal network activity. Treatment of cultures with GABA antagonists led to a significant increase in neuronal network activity in control cells but not in 4H cells, also pointing to lack of inhibitory activity in 4H. Myelination and oligodendrocyte maturation in cultures with 4H neurons was normal, and treatment with sonic hedgehog agonist SAG did not improve 4H related neuronal phenotypes. Quantitative PCR analysis revealed increased expression of parvalbumin interneuron marker ERBB4, suggesting that the development rather than generation of interneurons may be affected in 4H. Together, these results indicate that interneurons are involved, possibly parvalbumin interneurons, in disease mechanisms of 4H leukodystrophy.

GABAergic Signaling during Spinal Cord Stimulation Reduces Cardiac Arrhythmias in a Porcine Model.

Neuraxial modulation, including spinal cord stimulation, reduces cardiac sympathoexcitation and ventricular arrhythmogenesis. There is an incomplete understanding of the molecular mechanisms through which spinal cord stimulation modulates cardiospinal neural pathways. The authors hypothesize that spinal cord stimulation reduces myocardial ischemia-reperfusion-induced sympathetic excitation and ventricular arrhythmias through γ-aminobutyric acid (GABA)-mediated pathways in the thoracic spinal cord. Yorkshire pigs were randomized to control (n = 11), ischemia-reperfusion (n = 16), ischemia-reperfusion plus spinal cord stimulation (n = 17), ischemia-reperfusion plus spinal cord stimulation plus γ-aminobutyric acid type A (GABAA) or γ-aminobutyric acid type B (GABAB) receptor antagonist (GABAA, n = 8; GABAB, n = 8), and ischemia-reperfusion plus GABA transaminase inhibitor (GABAculine, n = 8). A four-pole spinal cord stimulation lead was placed epidurally (T1 to T4). GABA modulating pharmacologic agents were administered intrathecally. Spinal cord stimulation at 50 Hz was applied 30 min before ischemia. A 56-electrode epicardial mesh was used for high-resolution electrophysiologic recordings, including activation recovery intervals and ventricular arrhythmia scores. Immunohistochemistry and Western blots were performed to measure GABA receptor expression in the thoracic spinal cord. Cardiac ischemia led to myocardial sympathoexcitation with reduction in activation recovery interval (mean ± SD, -42 ± 11%), which was attenuated by spinal cord stimulation (-21 ± 17%, P = 0.001). GABAA and GABAB receptor antagonists abolished spinal cord stimulation attenuation of sympathoexcitation (GABAA, -9.7 ± 9.7%, P = 0.043 vs. ischemia-reperfusion plus spinal cord stimulation; GABAB, -13 ± 14%, P = 0.012 vs. ischemia-reperfusion plus spinal cord stimulation), while GABAculine alone caused a therapeutic effect similar to spinal cord stimulation (-4.1 ± 3.7%, P = 0.038 vs. ischemia-reperfusion). The ventricular arrhythmia score supported these findings. Spinal cord stimulation during ischemia-reperfusion increased GABAA receptor expression with no change in GABAB receptor expression. Thoracic spinal cord stimulation reduces ischemia-reperfusion-induced sympathoexcitation and ventricular arrhythmias through activation of GABA signaling pathways. These data support the hypothesis that spinal cord stimulation-induced release of GABA activates inhibitory interneurons to decrease primary afferent signaling from superficial dorsal horn to sympathetic output neurons in the intermediolateral nucleus.

γ-Aminobutyric acid promotes the inhibition of hair growth induced by chronic restraint stress

Stress plays a critical role in hair loss, although the underlying mechanisms are largely unknown. γ-aminobutyric acid (GABA) has been reported to be associated with stress; however, whether it affects stress-induced hair growth inhibition is unclear. This study aimed to investigate the potential roles and mechanisms of action of GABA in chronic restraint stress (CRS)-induced hair growth inhibition. We performed RNA-seq analysis and found that differentially expressed genes (DEGs) associated with neuroactive ligand-receptor interaction, including genes related to GABA receptors, significantly changed after mice were treated with CRS. Targeted metabolomics analysis and enzyme-linked immunosorbent assay (ELISA) also showed that GABA levels in back skin tissues and serum significantly elevated in the CRS group. Notably, CRS-induced hair growth inhibition got aggravated by GABA and alleviated through GABAA antagonists, such as picrotoxin and ginkgolide A. RNA sequencing analysis revealed that DEGs related to the cell cycle, DNA replication, purine metabolism, and pyrimidine metabolism pathways were significantly downregulated in dermal papilla (DP) cells after GABA treatment. Moreover, ginkgolide A, a GABAA antagonist extracted from the leaves of Ginkgo biloba, promoted the cell cycle of DP cells. Therefore, the present study demonstrated that the increase in GABA could promote CRS-induced hair growth inhibition by downregulating the cell cycle of DP cells and suggested that ginkgolide A may be a promising therapeutic drug for hair loss.

Alpha-synuclein oligomers alter the spontaneous firing discharge of cultured midbrain neurons.

The aim of this work was to monitor the effects of extracellular α-synuclein on the firing activity of midbrain neurons dissociated from substantia nigra TH-GFP mice embryos and cultured on microelectrode arrays (MEA). We monitored the spontaneous firing discharge of the network for 21 days after plating and the role of glutamatergic and GABAergic inputs in regulating burst generation and network synchronism. Addition of GABA A , AMPA and NMDA antagonists did not suppress the spontaneous activity but allowed to identify three types of neurons that exhibited different modalities of firing and response to applied L-DOPA: high-rate (HR) neurons, low-rate pacemaking (LR-p), and low-rate non-pacemaking (LR-np) neurons. Most HR neurons were insensitive to L-DOPA, while the majority of LR-p neurons responded with a decrease of the firing discharge; less defined was the response of LR-np neurons. The effect of exogenous α-synuclein (α-syn) on the firing discharge of midbrain neurons was then studied by varying the exposure time (0-48 h) and the α-syn concentration (0.3-70 μM), while the formation of α-syn oligomers was monitored by means of AFM. Independently of the applied concentration, acute exposure to α-syn monomers did not exert any effect on the spontaneous firing rate of HR, LR-p, and LR-np neurons. On the contrary, after 48 h exposure, the firing activity was drastically altered at late developmental stages (14 days in vitro, DIV, neurons): α-syn oligomers progressively reduced the spontaneous firing discharge (IC50 = 1.03 μM), impaired burst generation and network synchronism, proportionally to the increased oligomer/monomer ratio. Different effects were found on early-stage developed neurons (9 DIV), whose firing discharge remained unaltered, regardless of the applied α-syn concentration and the exposure time. Our findings unravel, for the first time, the variable effects of exogenous α-syn at different stages of midbrain network development and provide new evidence for the early detection of neuronal function impairment associated to aggregated forms of α-syn.

Reversibility of alfaxalone with intravenous flumazenil in cats: a prospective study.

Introduction: This study assessed the GABAA receptor antagonist flumazenil effect on the anesthetic activity of the GABAA receptor modulator and anesthetic, alfaxalone, when administered intravenously in healthy cats.

Dihydromyricetin Protects Against Salsolinol-Induced Toxicity in Dopaminergic Cell Line: Implication for Parkinson's Disease.

Parkinson's disease (PD) is a progressive neurodegenerative disease associated with loss of dopaminergic neurons in the substantia nigra pars compacta. Although aging is the primary cause, environmental and genetic factors have also been implicated in its etiology. In fact, the sporadic nature of PD (i.e., unknown etiology) renders the uncovering of the exact pathogenic mechanism(s) or development of effective pharmacotherapies challenging. In search of novel neuroprotectants, we showed that butyrate (BUT), a short-chain fatty acid, protects against salsolinol (SALS)-induced toxicity in human neuroblastoma-derived SH-SY5Y cells, which are considered an in-vitro model of PD. Dihydromyricetin (DHM), a flavonoid derived from Asian medicinal plant, has also shown effectiveness against oxidative damage and neuroinflammation, hallmarks of neurodegenerative diseases. Here we show that pretreatment of SH-SY5Y cells with DHM concentration-dependently prevented SALS-induced toxicity and that a combination of DHM and BUT resulted in a synergistic protection. The effects of both DHM and BUT in turn could be completely blocked by flumazenil (FLU), a GABAA antagonist acting at benzodiazepine receptor site, and by bicuculline (BIC), a GABAA antagonist acting at orthosteric site. Beta-hydroxybutyrate (BHB), a free fatty acid 3 (FA3) receptor antagonist, also fully blocked the protective effect of DHM. BHB was shown previously to only partially block the protective effect of BUT. Thus, there are some overlaps and some distinct differences in protective mechanisms of DHM and BUT against SALS-induced toxicity. It is suggested that a combination of DHM and BUT may have therapeutic potential in PD. However, further in-vivo verifications are necessary.

Corticostriatal Projections Relying on GABA Levels Mediate Exercise-Induced Functional Recovery in Cerebral Ischemic Mice.

Stroke is a neurological disorder characterized by high disability and death worldwide. The occlusion of the middle cerebral artery (MCAO) supplying the cortical motor regions and its projection pathway regions can either kill the cortical neurons or block their projections to the spinal cord and subcortical structure. The cerebral cortex is the primary striatal afferent, and the medium spiny neurons of the striatum have been identified as the major output neurons projecting to the substantia nigra and pallidum. Thus, disconnection of the corticostriatal circuit often occurs in the model of MCAO. In this study, we hypothesize that striatal network dysfunction in cerebral ischemic mice ultimately modulates the activity of striatal projections from cortical neurons to improve dysfunction during exercise training. In this study, we observed that the corticostriatal circuit originating from glutamatergic neurons could partially medicate the improvement of motor and anxiety-like behavior in mice with exercise. Furthermore, exercising or activating a single optogenetic corticostriatal circuit can increase the striatal gamma-aminobutyric acid (GABA) level. Using the GABA-A receptor antagonist, bicuculline, we further identified that the striatal glutamatergic projection from the cortical neurons relies on the GABAergic synapse's activity to modulate exercise-induced functional recovery. Overall, those results reveal that the dorsal striatum-projecting subpopulation of cortical glutamatergic neurons can influence GABA levels in the striatum, playing a critical role in modulating exercise-induced improvement of motor and anxiety-like behavior.

A role of GABAA receptor α1 subunit in the hippocampus for rapid-acting antidepressant-like effects of ketamine

Ketamine can produce rapid-acting antidepressant effects in treatment-resistant patients with depression. Although alterations in glutamatergic and GABAergic neurotransmission in the brain play a role in depression, the precise molecular mechanisms in these neurotransmission underlying ketamine's antidepressant actions remain largely unknown. Mice exposed to FSS (forced swimming stress) showed depression-like behavior and decreased levels of GABA (γ-aminobutyric acid), but not glutamate, in the hippocampus. Ketamine increased GABA levels and decreased glutamate levels in the hippocampus of mice exposed to FSS. There was a correlation between GABA levels and depression-like behavior. Furthermore, ketamine increased the levels of enzymes and transporters on the GABAergic neurons (SAT1, GAD67, GAD65, VGAT and GAT1) and astrocytes (EAAT2 and GAT3), without affecting the levels of enzymes and transporters (SAT2, VGluT1 and GABAAR γ2) on glutamatergic neurons. Moreover, ketamine caused a decreased expression of GABAAR α1 subunit, which was specifically expressed on GABAergic neurons and astrocytes, an increased GABA synthesis and metabolism in GABAergic neurons, a plasticity change in astrocytes, and an increase in ATP (adenosine triphosphate) contents. Finally, GABAAR antagonist bicuculline or ATP exerted a rapid antidepressant-like effect whereas pretreatment with GABAAR agonist muscimol blocked the antidepressant-like effects of ketamine. In addition, pharmacological activation and inhibition of GABAAR modulated the synthesis and metabolism of GABA, and the plasticity of astrocytes in the hippocampus. The present data suggest that ketamine could increase GABA synthesis and astrocyte plasticity through downregulation of GABAAR α1, increases in GABA, and conversion of GABA into ATP, resulting in a rapid-acting antidepressant-like action.This article is part of the Special Issue on ‘Ketamine and its Metabolites’.

Histamine Release in the Prefrontal Cortex Excites Fast-Spiking Interneurons while GABA Released from the Same Axons Inhibits Pyramidal Cells.

We studied how histamine and GABA release from axons originating from the hypothalamic tuberomammillary nucleus (TMN) and projecting to the prefrontal cortex (PFC) influence circuit processing. We optostimulated histamine/GABA from genetically defined TMN axons that express the histidine decarboxylase gene (TMNHDC axons). Whole-cell recordings from PFC neurons in layer 2/3 of prelimbic, anterior cingulate, and infralimbic regions were used to monitor excitability before and after optostimulated histamine/GABA release in male and female mice. We found that histamine-GABA release influences the PFC through actions on distinct neuronal types: the histamine stimulates fast-spiking interneurons; and the released GABA enhances tonic (extrasynaptic) inhibition on pyramidal cells (PyrNs). For fast-spiking nonaccommodating interneurons, histamine released from TMNHDC axons induced additive gain changes, which were blocked by histamine H1 and H2 receptor antagonists. The excitability of other fast-spiking interneurons in the PFC was not altered. In contrast, the GABA released from TMNHDC axons predominantly produced divisive gain changes in PyrNs, increasing their resting input conductance, and decreasing the slope of the input-output relationship. This inhibitory effect on PyrNs was not blocked by histamine receptor antagonists but was blocked by GABAA receptor antagonists. Across the adult life span (from 3 to 18 months of age), the GABA released from TMNHDC axons in the PFC inhibited PyrN excitability significantly more in older mice. For individuals who maintain cognitive performance into later life, the increases in TMNHDC GABA modulation of PyrNs during aging could enhance information processing and be an adaptive mechanism to buttress cognition.SIGNIFICANCE STATEMENT The hypothalamus controls arousal state by releasing chemical neurotransmitters throughout the brain to modulate neuronal excitability. Evidence is emerging that the release of multiple types of neurotransmitters may have opposing actions on neuronal populations in key cortical regions. This study demonstrates for the first time that the neurotransmitters histamine and GABA are released in the prefrontal cortex from axons originating from the tuberomammillary nucleus of the hypothalamus. This work demonstrates how hypothalamic modulation of neuronal excitability is maintained throughout adult life, highlighting an unexpected aspect of the aging process that may help maintain cognitive abilities.

Impulsive and compulsive behaviors can be induced by opposite GABAergic dysfunctions inside the primate ventral pallidum.

Introduction: The ventral pallidum (VP) is central in the limbic Basal Ganglia circuit, controlling both appetitive (approach) and aversive (avoidance) motivated behaviors. Nevertheless, VP involvement in pathological aspects remains unclear, especially in the behavioral expression of different motivational dysfunctions. This study aimed to investigate how the VP contributes to the expression of abnormal behaviors via opposite GABAergic dysfunctions. Methods: Opposite GABAergic dysfunctions were induced by injecting muscimol (a GABAA agonist) and bicuculline (a GABAA antagonist) into monkeys. We determined the effects of both substances on self-initiated behaviors in lab-chair and in free-moving home-cage contexts in six monkeys, and in two animals performing an approach-avoidance task in appetitive and aversive contexts. Results: While the self-initiated behaviors induced by bicuculline injections in VP were characterized by compulsive behaviors such as repetitive grooming and self-biting, muscimol injections induced impulsive behaviors including limb movements in a lab-chair context and exploration behaviors in a free-moving context. More specific behavioral effects were observed in the approach-avoidance task. The muscimol injections induced premature responses and erroneous screen touches, which characterize impulsive and attention disorders, while the bicuculline injections into the VP increased passive avoidance (non-initiated action) and task-escape in an aversive context, suggesting an anxiety disorder. Conclusions: These results show that activating or blocking GABAergic transmission in the VP impairs motivated behaviors. Furthermore, the behavioral expressions produced by these opposite disturbances show that the VP could be involved in anxiety-driven compulsive disorders, such as OCD, as well as in impulsive disorders motivated by attention deficits or reward-seeking, as seen in ADHD or impulse control disorders.

Dexmedetomidine Attenuates Lipopolysaccharide-Induced Sympathetic Activation and Sepsis via Suppressing Superoxide Signaling in Paraventricular Nucleus.

Sympathetic overactivity contributes to the pathogenesis of sepsis. The selective α2-adrenergic receptor agonist dexmedetomidine (DEX) is widely used for perioperative sedation and analgesia. We aimed to determine the central roles and mechanisms of DEX in attenuating sympathetic activity and inflammation in sepsis. Sepsis was induced by a single intraperitoneal injection of lipopolysaccharide (LPS) in rats. Effects of DEX were investigated 24 h after injection of LPS. Bilateral microinjection of DEX in the paraventricular nucleus (PVN) attenuated LPS-induced sympathetic overactivity, which was attenuated by the superoxide dismutase inhibitor DETC, cAMP analog db-cAMP or GABAA receptor antagonist gabazine. Superoxide scavenger tempol, NADPH oxidase inhibitor apocynin, adenylate cyclase inhibitor SQ22536 or PKA inhibitor Rp-cAMP caused similar effects to DEX in attenuating LPS-induced sympathetic activation. DEX inhibited LPS-induced superoxide and cAMP production, as well as NADPH oxidase, adenylate cyclase and PKA activation. The roles of DEX in reducing superoxide production and NADPH oxidase activation were attenuated by db-cAMP or gabazine. Intravenous infusion of DEX inhibited LPS-induced sympathetic overactivity, NOX activation, superoxide production, TNF-α and IL-1β upregulation in the PVN and plasma, as well as lung and renal injury, which were attenuated by the PVN microinjection of yohimbine and DETC. We conclude that activation of α2-adrenergic receptors with DEX in the PVN attenuated LPS-induced sympathetic overactivity by reducing NADPH oxidase-dependent superoxide production via both inhibiting adenylate cyclase-cAMP-PKA signaling and activating GABAA receptors. The inhibition of NADPH oxidase-dependent superoxide production in the PVN partially contributes to the roles of intravenous infusion of DEX in attenuating LPS-induced sympathetic activation, oxidative stress and inflammation.

Posterodorsal Medial Amygdala Urocortin-3, GABA, and Glutamate Mediate Suppression of LH Pulsatility in Female Mice.

The posterodorsal subnucleus of the medial amygdala (MePD) is an upstream modulator of the hypothalamic-pituitary-gonadal (HPG) and hypothalamic-pituitary-adrenal (HPA) axes. Inhibition of MePD urocortin-3 (Ucn3) neurons prevents psychological stress-induced suppression of luteinizing hormone (LH) pulsatility while blocking the stress-induced elevations in corticosterone (CORT) secretion in female mice. We explore the neurotransmission and neural circuitry suppressing the gonadotropin-releasing hormone (GnRH) pulse generator by MePD Ucn3 neurons and we further investigate whether MePD Ucn3 efferent projections to the hypothalamic paraventricular nucleus (PVN) control CORT secretion and LH pulsatility. Ucn3-cre-tdTomato female ovariectomized (OVX) mice were unilaterally injected with adeno-associated virus (AAV)-channelrhodopsin 2 (ChR2) and implanted with optofluid cannulae targeting the MePD. We optically activated Ucn3 neurons in the MePD with blue light at 10Hz and monitored the effect on LH pulses. Next, we combined optogenetic stimulation of MePD Ucn3 neurons with pharmacological antagonism of GABAA or GABAB receptors with bicuculline or CGP-35348, respectively, as well as a combination of NMDA and AMPA receptor antagonists, AP5 and CNQX, respectively, and observed the effect on pulsatile LH secretion. A separate group of Ucn3-cre-tdTomato OVX mice with 17β-estradiol replacement were unilaterally injected with AAV-ChR2 in the MePD and implanted with fiber-optic cannulae targeting the PVN. We optically stimulated the MePD Ucn3 efferent projections in the PVN with blue light at 20Hz and monitored the effect on CORT secretion and LH pulses. We reveal for the first time that activation of Ucn3 neurons in the MePD inhibits GnRH pulse generator frequency via GABA and glutamate signaling within the MePD, while MePD Ucn3 projections to the PVN modulate the HPG and HPA axes.

Regulation of the central amygdala on intestinal motility and behavior via the lateral hypothalamus in irritable bowel syndrome model mice.

Impaired bidirectional communication between the gastrointestinal tract and the central nervous system (CNS) is closely related to the development of irritable bowel syndrome (IBS). Studies in patients with IBS have also shown significant activation of the hypothalamus and amygdala. However, how neural circuits of the CNS participate in and process the emotional and intestinal disorders of IBS remains unclear. The GABAergic neural pathway projecting from the central amygdala (CeA) to the lateral hypothalamus (LHA) in mice was investigated by retrograde tracking combined with fluorescence immunohistochemistry. Anxiety, depression-like behavior, and intestinal motility were observed in the water-immersion restraint stress group and the control group. Furthermore, the effects of the chemogenetic activation of the GABAergic neural pathway of CeA-LHA on behavior and intestinal motility, as well as the co-expression of orexin-A and c-Fos in the LHA, were explored. In our study, Fluoro-Gold retrograde tracking combined with fluorescence immunohistochemistry showed that GABAergic neurons in the CeA were projected to the LHA. The microinjection of the gamma-aminobutyric acid (GABA) receptor antagonist into the LHA relieved anxiety, depression-like behavior, and intestinal motility disorder in the IBS mice. The chemogenetic activation of GABAergic neurons in the CeA-LHA pathway led to anxiety, depression-like behavior, and intestinal motility disorder. In addition, GABAergic neurons in the CeA-LHA pathway inhibited the expression of orexin-A in the LHA, and orexin-A was co-expressed with GABAA receptors. The CeA-LHA GABAergic pathway might participate in the occurrence and development of IBS by regulating orexin-A neurons.

Attenuation of Epileptogenesis and Cognitive Deficits by a Selective and Potent Kv7 Channel Opener in Rodent Models of Seizures.

Targeting neuronal Kv7 channels by pharmacological activation has been proven to be an attractive therapeutic strategy for epilepsy. Here, we show that activation of Kv7 channels by an opener SCR2682 dose-dependently reduces seizure activity and severity in rodent models of epilepsy induced by a GABAa receptor antagonist pentylenetetrazole (PTZ), maximal electroshock, and a glutamate receptor agonist kainic acid (KA). Electroencephalographic recordings of rat cerebral cortex confirm that SCR2682 also decreases epileptiform discharges in KA-induced seizures. Nissl and neuronal nuclei staining further demonstrates that SCR2682 also protects neurons from injury induced by KA. In Morris water maze navigation and Y-maze tests, SCR2682 improves PTZ- and KA-induced cognitive impairment. Taken together, our findings demonstrate that pharmacological activation of Kv7 by novel opener SCR2682 may hold promise for therapy of epilepsy with cognitive impairment. SIGNIFICANCE STATEMENT: A neuronal Kv7 channel opener SCR2682 attenuates epileptogenesis and seizure-induced cognitive impairment in rodent models of seizures, thus possessing a developmental potential for effective therapy of epilepsy with cognitive impairment.

Claustrum modulates behavioral sensitivity and EEG activity of propofol anesthesia.

The claustrum has long been regarded as a vital center for conscious control. Electrical stimulation or damage to the claustrum can result in decreased awareness or loss of consciousness, suggesting that the claustrum may be a target for the action of general anesthetics. This study aimed to determine the role of the claustrum in propofol anesthesia. We first applied a fiber photometry calcium signal recording system to record the claustral neuronal activity during the entire process of propofol anesthesia. Chemogenetic activation of claustral neurones was then performed to verify their role in anesthesia. Finally, muscimol (GABAa receptor agonist) and gabazine (GABAa receptor antagonist) were microinjected into the claustrum to determine whether their GABAa receptors were involved in modulating propofol anesthesia. EEG and behavioral indicators, such as anesthetic sensitivity and efficacy, were recorded and analyzed. An evident anesthesia-related change in claustrum neuronal activity was suppressed during propofol-induced unconsciousness and restored following recovery from anesthesia. Chemogenetic activation of claustrum neuronsresults in attenuated propofol sensitivity, a shorter anesthesia duration, and an EEG shift toward wakefulness. Manipulation of GABAa receptors in the claustrum showed bidirectional control of propofol sensitivity, as activation decreases anesthesia efficiency while inactivation augments it. Additionally, inhibiting claustrum GABAa receptors increases cortical EEG slow waves. Claustrum neurones and their GABAa receptors are implicated in the modulation of propofol anesthesia in both behavioral and EEG assessments. Our findings create scope to reveal the brain targets of anesthetic action further and add to the existing evidence on the consciousness-modulating role of the claustrum.

GABAergic Regulation of Astroglial Gliotransmission through Cx43 Hemichannels

Gamma-Aminobutyric acid (GABA) is the primary inhibitory neurotransmitter in the brain. It is produced by interneurons and recycled by astrocytes. In neurons, GABA activates the influx of Cl- via the GABAA receptor or efflux or K+ via the GABAB receptor, inducing hyperpolarization and synaptic inhibition. In astrocytes, the activation of both GABAA and GABAB receptors induces an increase in intracellular Ca2+ and the release of glutamate and ATP. Connexin 43 (Cx43) hemichannels are among the main Ca2+-dependent cellular mechanisms for the astroglial release of glutamate and ATP. However, no study has evaluated the effect of GABA on astroglial Cx43 hemichannel activity and Cx43 hemichannel-mediated gliotransmission. Here we assessed the effects of GABA on Cx43 hemichannel activity in DI NCT1 rat astrocytes and hippocampal brain slices. We found that GABA induces a Ca2+-dependent increase in Cx43 hemichannel activity in astrocytes mediated by the GABAA receptor, as it was blunted by the GABAA receptor antagonist bicuculline but unaffected by GABAB receptor antagonist CGP55845. Moreover, GABA induced the Cx43 hemichannel-dependent release of glutamate and ATP, which was also prevented by bicuculline, but unaffected by CGP. Gliotransmission in response to GABA was also unaffected by pannexin 1 channel blockade. These results are discussed in terms of the possible role of astroglial Cx43 hemichannel-mediated glutamate and ATP release in regulating the excitatory/inhibitory balance in the brain and their possible contribution to psychiatric disorders.

RF01 | PMON45 Urocortin 3 Interaction with GABA and Glutamate in the Posterodorsal Subnucleus of The Medial Amygdala Mediates Suppression of GnRH Pulse Generator Frequency in Female Mice

Abstract Stress exerts a profound impact on reproductive function and disrupts pulsatile luteinizing hormone (LH) secretion. The posterodorsal subnucleus of the medial amygdala (MePD) is an upstream modulator of the hypothalamus pituitary gonadal axis. Stress alters neuronal activity within the MePD, increasing the expression of Urocortin3 (Ucn3) while enhancing the inhibitory output from the MePD to key hypothalamic reproductive centres. Designer receptor exclusively activated by designer drugs (DREADDs) inhibition of MePD Ucn3 neurons prevents psychological stress-induced suppression of LH pulses in female mice. The neurochemical pathways in the MePD responsible for regulating the GnRH pulse generator in the presence of stress remain unknown. We investigate the neurotransmission involved in the neural circuitry responsible for the suppression of GnRH pulse generator activity by MePD Ucn3 neuronal activation. Ucn3-Cre-tdTomato female ovariectomised (OVX) mice were unilaterally injected with AAV-ChR2 and implanted with optofluid cannulae targeting the MePD. Firstly, we optically stimulated MePD Ucn3 neurons with blue light at 10 Hz, 10 mW and monitored the effect on LH pulses. Next, we combined optogenetic activation of MePD Ucn3 neurons with pharmacological antagonism of GABAA or GABAB receptors with bicuculline or CGP, respectively, and observed the effect on LH pulsatility. Finally, during optical activation of MePD Ucn3 neurons we administered a combination of NMDA and AMPA receptor antagonists, AP5 and CNQX respectively, and monitored the effect on pulsatile LH secretion. Optogenetic stimulation of MePD Ucn3 neurons suppressed pulsatile LH secretion compared to controls and administration of aCSF had no effect on LH pulse interval (MePD Ucn3 activation: 36.04 ± 2.49 vs. pre-treatment control period: 17.50 ± 0.67 vs control virus: 21.67 ± 1.67 min; mean ± SEM; p<0.0001; n=12). Intra-MePD infusion of bicuculline (BIC) or CGP during optogenetic stimulation of MePD Ucn3 neurons completely blocked the suppressive effect of MePD Ucn3 neuronal activation on LH pulsatility (MePD Ucn3 activation and BIC: 20.50 ± 2.03 vs. pre-treatment control period: 17.58 ± 0.75 min; MePD Ucn3 activation and CGP: 16.30 ± 0.63 vs. pre-treatment control period: 17.50 ± 0.77 min; n=7-10). Intra-MePD infusion of AP5 + CNQX during optogenetic stimulation of MePD Ucn3 neurons similarly completely blocked the suppressive effect of MePD Ucn3 neuronal activation on LH pulsatility (MePD Ucn3 activation and AP5 + CNQX: 19.58 ± 1.42 min; n=6). Our findings show for the first time that optogenetic stimulation of MePD Ucn3 neurons inhibits GnRH pulse generator activity via GABA and glutamate signalling within the MePD. Presentation: Saturday, June 11, 2022 1:12 p.m. - 1:17 p.m., Monday, June 13, 2022 12:30 p.m. - 2:30 p.m.

Effects of Hyssopus Officinalis Hydroalcoholic Extract on Pentylenetetrazol-Induced Convulsive Seizures in Rat.

Hyssopus officinalis L. is one of the most important medicinal plants in traditional medicine used to treat seizures. In this study, we assessed the effects of H. officinalis hydroalcoholic extract against pentylenetetrazol (PTZ)-induced seizures in rat. The anti-seizure activity of the extract was assessed in three doses of 25, 50, and 100mg/kg. Kindling was induced by intraperitoneal injection of PTZ (35mg/kg) every 48h, and H. officinalis extract was administered daily and behavioral tests performed. The possible involvement of GABA receptors in the extract activity was investigated using flumazenil. Tonic seizure threshold and mortality rate were measured following intraperitoneal injection of 60mg/kg PTZ on the 14th day, following 14 days administration of H. officinalis hydroalcoholic extract. Blood and hippocampus samples were prepared to measure brain and serum antioxidant capacity, malondialdehyde (MDA), and nitric oxide (NO). Finally, the expression of GABA receptor gene in brain tissue was investigated. H. officinalis extract increased tonic seizure threshold and decreased mortality due to PTZ. Flumazenil, as a GABA receptor antagonist, reduced the tonic seizure threshold. Extract treatment significantly improved memory and learning, increased brain antioxidant capacity, decreased brain MDA and NO in kindled rats. It also increased GABA receptor gene expression in pre-treated groups compared to the negative control group. H. officinalis extract probably exerts potential antiepileptic effects through the GABAergic system. Also, H. officinalis extract has a supportive effect against hippocampal neuronal damage and improves memory and learning in kindled rats.