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α5 subunit GABA type A receptor (GABA A R) preferring negative allosteric modulators (NAMs) are cognitive enhancers with antidepressant-like effects. α5-NAM success in treating mouse models of neurodevelopmental disorders with excessive inhibition have led to Phase 2 clinical trials for Down syndrome. Despite in vivo efficacy, no study has examined the effects of continued α5-NAM treatment on inhibitory and excitatory synapse plasticity to identify mechanisms of action. Here we used L-655,708, an imidazobenzodiazepine that acts as a highly selective but weak α5-NAM, to investigate the impact of sustained treatment on hippocampal neuron synapse and dendrite development. We show that 2-day pharmacological reduction of α5-GABA A R signaling from DIV12-14, when GABA A Rs contribute to depolarization, delays dendritic spine maturation and the NMDA receptor (NMDAR) GluN2B/GluN2A developmental shift. In contrast, α5-NAM treatment from DIV19-21, when hyperpolarizing GABA A R signaling predominates, enhances surface synaptic GluN2A while decreasing GluN2B. Despite changes in NMDAR subtype surface levels and localization, total levels of key excitatory synapse proteins were largely unchanged, and mEPSCs were unaltered. Importantly, 2-day α5-NAM treatment does not alter the total surface levels or distribution of α5-GABA A Rs, reduce the gephyrin inhibitory synaptic scaffold, or impair phasic or tonic inhibition. Furthermore, α5-NAM inhibition of the GABA A R tonic current in mature neurons is maintained after 2-day α5-NAM treatment, suggesting reduced tolerance liability, in contrast to other clinically relevant GABA A R-targeting drugs such as benzodiazepines. Together, these results show that α5-GABA A Rs contribute to dendritic spine maturation and excitatory synapse development via a NMDAR dependent mechanism without perturbing overall neuronal excitability., (Copyright © 2021. Published by Elsevier Ltd.)

γ-aminobutyric acid type-A receptors (GABA A Rs) are inhibitory ligand-gated ion channels in the brain that are crucial for controlling neuronal excitation. To explore their physiological roles in cellular and neural network activity, it is important to understand why specific GABA A R isoforms are distributed not only to various brain regions and cell types, but also to specific areas of the membrane in individual neurons. To address this aim we have developed a novel photosensitive compound, azogabazine, that targets and reversibly inhibits GABA A Rs. The receptor selectivity of the compound is based on the competitive antagonist, gabazine, and photosensitivity is conferred by a photoisomerisable azobenzene group. Azogabazine can exist in either cis or trans conformations that are controlled by UV and blue light respectively, to affect receptor inhibition. We report that the trans-isomer preferentially binds and inhibits GABA A R function, whilst promotion of the cis-isomer caused unbinding of azogabazine from GABA A Rs. Using cultured cerebellar granule cells, azogabazine in conjunction with UV light applied to defined membrane domains, revealed higher densities of GABA A Rs at somatic inhibitory synapses compared to those populating proximal dendritic zones, even though the latter displayed a higher number of synapses per unit area of membrane. Azogabazine also revealed more pronounced GABA-mediated inhibition of action potential firing in proximal dendrites compared to the soma. Overall, azogabazine is a valuable addition to the photochemical toolkit that can be used to interrogate GABA A R function and inhibition., (Copyright © 2020 The Authors. Published by Elsevier Ltd.. All rights reserved.)

The flexibility of neuronal networks is believed to rely mainly on the plasticity of excitatory synapses. However, like their excitatory counterparts, inhibitory synapses also undergo several forms of synaptic plasticity. This review examines recent advances in the understanding of the molecular mechanisms leading to postsynaptic GABAergic plasticity. Specifically, modulation of GABAA receptor (GABAAR) number at postsynaptic sites plays a key role, with the interaction of GABAARs with the scaffold protein gephyrin and other postsynaptic scaffold/regulatory proteins having particular importance. Our understanding of these molecular interactions are progressing, based on recent insights into the processes of GABAAR lateral diffusion, gephyrin dynamics, and gephyrin nanoscale organization. This article is part of the special issue entitled 'Mobility and trafficking of neuronal membrane proteins'., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Delayed secondary degeneration in the non-ischemic sites such as ipsilateral thalamus would occur after cortical infarction. Hence, alleviating secondary damage is considered to be a promising novel target for acute stroke therapy. In the current study, the neuroprotective effects of bis(propyl)-cognitin (B3C), a multifunctional dimer, against secondary damage in the VPN of ipsilateral thalamus were investigated in a distal middle cerebral artery occlusion (dMCAO) stroke model in adult rats. It was found that B3C (0.5 and 1 mg/kg, ip) effectively improved neurological function of rats at day 7 and day 14 after dMCAO. Additionally, the treatment with B3C alleviated neuronal loss and gliosis in ipsilateral VPN after dMCAO, as evidenced by the higher immunoreactivity of neuron-specific nuclear-binding protein (NeuN) as well as lower immunostaining intensity of glial fibrillary acidic protein (GFAP) and cluster of differentiation 68 (CD68). Most encouragingly, immunohistochemistry and western blotting further revealed that B3C treatment greatly reduced Aβ deposits and cathepsin B expression in the VPN of ipsilateral thalamus at day 7 and day 14 after dMCAO. In parallel, we demonstrated herein that the neuroprotective effects of B3C in dMCAO model were similar to L-3-trans-(Propyl-carbamoyloxirane-2-carbonyl)- L-isoleucyl-l-proline methyl ester (CA-074Me), a specific inhibitor of cathepsin B, suggesting that B3C attenuated secondary damage and Aβ deposits in the VPN of ipsilateral thalamus after dMCAO possibly through the reduction of cathepsin B. These findings taken together provide novel molecular sights into the potential application of B3C for the treatment of secondary degeneration after cortical infarction., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

In preclinical models, it has been reported that social defeat stress activates microglial cells in the CNS. Translocator protein 18 kDa (TSPO) is a mitochondrial protein expressed on microglia in the CNS that has been proposed to be a useful biomarker for brain injury and inflammation. We hypothesized that a TSPO antagonist, ONO-2952, would inhibit the neuroinflammation induced by microglial hyperactivation and associated depressive-like behaviors. An in vitro analysis showed that ONO-2952 suppressed the release of pro-inflammatory cytokines and mitochondrial reactive oxygen species in cultured microglia stimulated by lipopolysaccharide. In mice submitted to chronic social defeat stress, microglia predominantly expressed TSPO in limbic areas implicated in depressive-like behaviors, including the amygdala, ventral hippocampus and nucleus accumbens, in which an increase in the production of pro-inflammatory cytokines in vivo were associated. Treating animals with ONO-2952 during chronic social defeat stress ameliorated impairments in social avoidance and anxiety-like behaviors and suppressed pro-inflammatory cytokine production, suggesting that ONO-2952 exerted an anti-stress effect in this animal model of depression. Thus, targeting TSPO as a candidate for the development of antidepressants that reduce susceptibility to chronic stress could pave the way toward therapeutic interventions for relapse prophylaxis in depression., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Deficits in prefrontal cortex (PFC) GABAergic neurotransmission are linked to cognitive impairments seen in schizophrenia and other disorders, and pharmacological reduction of PFC GABA A transmission disrupts processes including working and spatial memory. This provides an opportunity to examine whether compounds capable of neutralizing GABAergic dysfunction may ameliorate these cognitive deficits. PFC dopamine (DA) D 1 receptor activation enhances GABA transmission, raising the possibly that direct or indirect agonists of DA D 1 receptors would be effective in reversing working memory and other forms of cognitive deficits. To test this, male rats were pre-treated with two drugs that augment PFC D 1 signalling before PFC infusion of the GABA A antagonist, bicuculline (50 ng) and assessment of spatial working and reference memory function. A moderate dose of the full D 1 agonist SKF-81297 (0.1 mg/kg) completely reversed PFC GABA hypofunction-induced working memory deficits assessed in an delayed-response task, whereas lower and higher doses (0.05 and 0.3 mg/kg respectively) were associated with mild improvements or deleterious effects. Treatment with the tetrahydroprotoberberine d-govadine (0.5 or 1.0 mg/kg), a synthetic compound known to enhance DA release selectively in the PFC, also significantly improved delayed-response working memory function induced by PFC GABA A antagonism. Furthermore, administration of the optimal dose of both drugs led to a partial rescue of PFC GABA hypofunction-induced reference and short-term spatial memory impairments assessed on a radial maze task. These findings suggest that modulation of PFC DA signalling via actions on the DA D 1 receptor represents a promising therapeutic strategy for working memory and other cognitive impairments observed in psychiatric disorders, including those with causes that extend beyond DA dysfunction., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Fenamates mefanamic and niflumic acids (MFA and NFA) induced dual potentiating and inhibitory effects on GABA currents recorded in isolated cerebellar Purkinje cells using the whole-cell patch-clamp and fast-application techniques. Regardless of the concentration, both drugs induced a pronounced prolongation of the current response. We demonstrated that the same concentration of drugs can produce both potentiating and inhibitory effects, depending on the GABA concentration, which indicates that both processes take place simultaneously and the net effect depends on the concentrations of both the agonist and fenamate. We found that the NFA-induced block is strongly voltage-dependent. The Woodhull analysis of the block suggests that NFA has two binding sites in the pore - shallow and deep. We built a homology model of the open GABA A R based on the cryo-EM structure of the open α1 GlyR and applied Monte-Carlo energy minimization to optimize the ligand-receptor complexes. A systematic search for MFA/NFA binding sites in the GABA A R pore revealed the existence of two sites, the location of which coincides well with predictions of the Woodhull model. In silico docking suggests that two fenamate molecules are necessary to occlude the pore. We showed that MFA, acting as a PAM, competes with an intravenous anesthetic etomidate for a common binding site. We built structural models of MFA and NFA binding at the transmembrane β(+)/α(-) intersubunit interface. We suggested a hypothesis on the molecular mechanism underlying the prolongation of the receptor lifetime in open state after MFA/NFA binding and β subunit specificity of the fenamate potentiation., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

HIV-Associated Neurocognitive disorder (HAND) affects nearly half of infected patients. The HIV envelope protein gp120 is shed by infected cells and is a potent neurotoxin in vitro that reproduces many aspects of HAND when expressed in vivo. Here, we show that HIV gp120 increases the amplitude of a tonic current mediated by γ-aminobutyric acid type-A receptors (GABA A Rs). Treating rat hippocampal cultures with 600 pM gp120 IIIB for 4 h increased a tonic bicuculline-sensitive current, which remained elevated for 24 h. The increased current resulted from upregulation of extrasynaptic α5-containing GABA A Rs, as indicated by inhibition with the selective inverse agonist basmisanil. Treatment with gp120 increased α5-GABA A R immunoreactivity on the cell surface without new protein synthesis. The increase in tonic inhibition was prevented by a C-X-C chemokine receptor type 4 (CXCR4) antagonist or elimination of microglia from the culture. Treatment with interleukin-1β (IL-1β) increased the tonic current and an IL-1 receptor antagonist blocked the gp120-evoked response. Pharmacological or genetic inhibition of p38 mitogen-activated protein kinase (MAPK) prevented the gp120-evoked increase in tonic current and direct activation of a mutant form of p38 MAPK expressed in neurons increased the current. Collectively, these data show that gp120 activates CXCR4 to stimulate microglia to release IL-1β. Subsequent stimulation of IL-1 receptors activates p38 MAPK in neurons leading to the upregulation of α5-containing GABA A Rs. Increased tonic inhibition impairs neuroplasticity and inhibition of α5-containing GABA A Rs improves cognitive function in disease models. Thus, gp120-induced upregulation of α5-containing GABA A Rs presents a novel therapeutic target for HAND., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Zinc is an abundant trace metal in the hippocampus nerve terminals. Previous studies demonstrate the ability of zinc to selectively block neurosteroid-sensitive, extrasynaptic GABA-A receptors in the hippocampus (Carver et al, 2016). Here we report that zinc prevents the seizure protective effects of the synthetic neurosteroid ganaxolone (GX) in an experimental model of epilepsy. GABA-gated and tonic currents were recorded from dissociated dentate gyrus granule cells (DGGCs), CA1 pyramidal cells (CA1PCs), and hippocampal slices from adult mice. Antiseizure effects of GX and the reversal of these effects by zinc were evaluated in fully-kindled mice expressing generalized (stage 5) seizures. In electrophysiological studies, zinc blocked the GABA-evoked and GX-potentiated GABA-gated chloride currents in DGGCs and CA1PCs in a concentration-dependent fashion similar to the competitive GABA-A receptor antagonists bicuculline and gabazine. Zinc completely blocked GX potentiation of extrasynaptic tonic currents, but not synaptic phasic currents. In hippocampus kindling studies, systemic administration of GX produced a dose-dependent suppression of behavioral and electrographic seizures in fully-kindled mice with complete seizure protection at the 10 mg/kg dose. However, the antiseizure effects of GX were significantly prevented by intrahippocampal administration of zinc (ED 50 , 150 μM). The zinc antagonistic response was reversible as animals responded normally to GX administration 24 h post-zinc blockade. These results demonstrate that zinc reduces the antiseizure effects of GX by selectively blocking extrasynaptic δGABA-A receptors in the hippocampus. These pharmacodynamic interactions have clinical implications in neurosteroid therapy for brain conditions associated with zinc fluctuations., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Feeding can be inhibited by satiety, sickness, or food unpalatability. The central nucleus of the amygdala (CeA) has been considered the key region for processing multiple anorexigenic signals, although the detailed cellular and molecular mechanisms remain largely unclear. Here we identify that methyleugenol (ME), a novel agonist of A type ionotropic γ-aminobutyric acid receptors (GABA A Rs), significantly counteracts the anorexigenic effects caused by satiety or sickness in association with GABAergic inhibition in the CeA. Electrophysiologically, ME enhanced GABAergic transmission and repressed neuronal excitability of the CeA. Behaviorally, ME increased feeding but not affect locomotor activity and basal anxiety in naïve mice. Notably, both systemic and CeA-specific delivery of ME significantly rescued satiety- or sickness-induced inhibition of feeding. The effects of ME were mainly dependent on the GABA A Rs in the CeA. Indeed, viral-mediated, the CeA region-specific genetic knockdown of the γ2 subunit of GABA A Rs largely abolished the above pharmacological effects, while its re-expression in a subpopulation of GABAergic neurons in the CeA, that produce protein kinase C-δ (PKC-δ), recovered the effects of ME on anorexigenic signals. Taken together, these results reveal a novel molecular mechanism for counter-anorexigenic signals dependent on GABAergic inhibition in the CeA, suggesting the possibility of ME as a leading compound for anorexia treatment., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Novel treatments against migraine are an urgent medical requirement. The α6 subunit-containing GABA A receptors (α6GABA A Rs) are expressed in trigeminal ganglia (TG), the hub of the trigeminal vascular system (TGVS) that is involved in the pathogenesis of migraine. Here we reveal an unprecedented role of α6GABA A Rs in ameliorating TGVS activation using several pharmacological approaches in an animal model mimicking pathological changes in migraine. TGVS activation was induced by intra-cisternal (i.c.) instillation of capsaicin in Wistar rats. Centrally, i.c. capsaicin activated the trigeminal cervical complex (TCC) measured by the increased number of c-Fos-immunoreactive (c-Fos-ir) TCC neurons. Peripherally, it elevated calcitonin gene-related peptide immunoreactivity (CGRP-ir) in TG and depleted CGRP-ir in the dura mater. Pharmacological approaches included a recently identified α6GABA A R-selective positive allosteric modulator (PAM), the pyrazoloquinolinone Compound 6, two α6GABA A R-active PAMs (Ro15-4513 and loreclezole), an α6GABA A R-inactive benzodiazepine (diazepam), an α6GABA A R-selective antagonist (furosemide), and a clinically effective antimigraine agent (topiramate). We examined effects of these compounds on both central and peripheral TGVS responses induced by i.c. capsaicin. Compound 6 (3-10 mg/kg, i.p.) significantly attenuated the TCC neuronal activation and TG CGRP-ir elevation, and dural CGRP depletion induced by capsaicin. All these effects of Compound 6 were mimicked by topiramate, Ro15-4513 and loreclezole, but not by diazepam. The brain-impermeable furosemide antagonized the peripheral, but not central, effects of Compound 6. These results suggest that the α6GABA A R in TG is a novel drug target for TGVS activation and that α6GABA A R-selective PAMs have the potential to be developed as a novel pharmacotherapy for migraine., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Chronic activity perturbations in neurons induce homeostatic plasticity through modulation of synaptic strength or other intrinsic properties to maintain the correct physiological range of excitability. Although similar plasticity can also occur at the population level, what molecular mechanisms are involved remain unclear. In the current study, we utilized a multielectrode array (MEA) recording system to evaluate homeostatic neural network activity of primary mouse cortical neuron cultures. We demonstrated that chronic elevation of neuronal activity through the inhibition of GABA(A) receptors elicits synchronization of neural network activity and homeostatic reduction of the amplitude of spontaneous neural network spikes. We subsequently showed that this phenomenon is mediated by the ubiquitination of tumor suppressor p53, which is triggered by murine double minute-2 (Mdm2). Using a mouse model of fragile X syndrome, in which fragile X mental retardation protein (FMRP) is absent (Fmr1 knockout), we found that Mdm2-p53 signaling, network synchronization, and the reduction of network spike amplitude upon chronic activity stimulation were all impaired. Pharmacologically inhibiting p53 with Pifithrin-α or genetically employing p53 heterozygous mice to enforce the inactivation of p53 in Fmr1 knockout cultures restored the synchronization of neural network activity after chronic activity stimulation and partially corrects the homeostatic reduction of neural network spike amplitude. Together, our findings reveal the roles of both Fmr1 and Mdm2-p53 signaling in the homeostatic regulation of neural network activity and provide insight into the deficits of excitability homeostasis seen when Fmr1 is compromised, such as occurs with fragile X syndrome., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

gamma-Aminobutyric acid (GABA)-mediated inhibitory neurotransmission and the gene products involved were discovered during the mid-twentieth century. Historically, myriad existing nervous system drugs act as positive and negative allosteric modulators of these proteins, making GABA a major component of modern neuropharmacology, and suggesting that many potential drugs will be found that share these targets. Although some of these drugs act on proteins involved in synthesis, degradation, and membrane transport of GABA, the GABA receptors Type A (GABA A R) and Type B (GABA B R) are the targets of the great majority of GABAergic drugs. This discovery is due in no small part to Professor Norman Bowery. Whereas the topic of GABA B R is appropriately emphasized in this special issue, Norman Bowery also made many insights into GABA A R pharmacology, the topic of this article. GABA A R are members of the ligand-gated ion channel receptor superfamily, a chloride channel family of a dozen or more heteropentameric subtypes containing 19 possible different subunits. These subtypes show different brain regional and subcellular localization, age-dependent expression, and potential for plastic changes with experience including drug exposure. Not only are GABA A R the targets of agonist depressants and antagonist convulsants, but most GABA A R drugs act at other (allosteric) binding sites on the GABA A R proteins. Some anxiolytic and sedative drugs, like benzodiazepine and related drugs, act on GABA A R subtype-dependent extracellular domain sites. General anesthetics including alcohols and neurosteroids act at GABA A R subunit-interface trans-membrane sites. Ethanol at high anesthetic doses acts on GABA A R subtype-dependent trans-membrane domain sites. Ethanol at low intoxicating doses acts at GABA A R subtype-dependent extracellular domain sites. Thus GABA A R subtypes possess pharmacologically specific receptor binding sites for a large group of different chemical classes of clinically important neuropharmacological agents. This article is part of the "Special Issue Dedicated to Norman G. Bowery"., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Unlike in the central nervous system (CNS), in the adult peripheral nervous system (PNS), activation of GABA A receptors (GABA A R) is excitatory because of the relatively high concentration of intracellular chloride in these neurons. Indeed, exogenous GABA and muscimol, a GABA A R agonist, exacerbate acute inflammatory hypersensitivity in rodents. However, it remains unclear whether peripheral GABA A R and the endogenous GABA play an important role in persistent inflammatory hypersensitivity. In this study, we thus investigated how peripheral GABA A R affects pain hypersensitivity by using the complete Freund's adjuvant (CFA)-induced persistent inflammatory pain mouse model. We found that intraplantar (i.pl.) administration of GABA A R antagonists, picrotoxin, and 1(S),9(R)-(-)-bicuculline methiodide significantly inhibited both spontaneous nociceptive (paw licking and flinching) behavior and mechanical hypersensitivity in CFA-injected mice at day 3 (D3), but not in naïve mice. Interestingly, CFA-induced mechanical hypersensitivity was significantly reversed by anti-GABA antibody (anti-GABA, i.pl.). In addition, RT-qPCR revealed that glutamate decarboxylase Gad1 (GAD 67) and Gad2 (GAD 65) mRNA expression was also upregulated in the ipsilateral hind paw of CFA-injected mice at D3. Finally, 5α-pregnan-3α-ol-20-one (3α,5α-THP), a selective positive allosteric modulator of GABA A R, produced mechanical hypersensitivity in naïve mice in a dose-dependent manner. Taken together, our results indicate that peripheral GABA A R and endogenous GABA, possibly produced by the inflamed tissue, potentiate CFA-induced persistent inflammatory hypersensitivity, suggesting that they can be used as a therapeutic target for alleviating inflammatory pain., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

In the mammalian central nervous system (CNS) GABA A receptors (GABA A Rs) mediate neuronal inhibition and are important therapeutic targets. GABA A Rs are composed of 5 subunits, drawn from 19 proteins, underpinning expression of 20-30 GABA A R subtypes. In the CNS these isoforms are heterogeneously expressed and exhibit distinct physiological and pharmacological properties. We report the discovery of S44819, a novel tricyclic oxazolo-2,3-benzodiazepine-derivative, that selectively inhibits α5-subunit-containing GABA A Rs (α5-GABA A Rs). Current α5-GABA A R inhibitors bind to the "benzodiazepine site". However, in HEK293 cells expressing recombinant α5-GABA A Rs, S44819 had no effect on 3 H-flumazenil binding, but displaced the GABA A R agonist 3 H-muscimol and competitively inhibited the GABA-induced responses. Importantly, we reveal that the α5-subunit selectivity is uniquely governed by amino acid residues within the α-subunit F-loop, a region associated with GABA binding. In mouse hippocampal CA1 neurons, S44819 enhanced long-term potentiation (LTP), blocked a tonic current mediated by extrasynaptic α5-GABA A Rs, but had no effect on synaptic GABA A Rs. In mouse thalamic neurons, S44819 had no effect on the tonic current mediated by δ-GABA A Rs, or on synaptic (α1β2γ2) GABA A Rs. In rats, S44819 enhanced object recognition memory and reversed scopolamine-induced impairment of working memory in the eight-arm radial maze. In conclusion, S44819 is a first in class compound that uniquely acts as a potent, competitive, selective antagonist of recombinant and native α5-GABA A Rs. Consequently, S44819 enhances hippocampal synaptic plasticity and exhibits pro-cognitive efficacy. Given this profile, S44819 may improve cognitive function in neurodegenerative disorders and facilitate post-stroke recovery., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Previous work has shown that S44819 is a novel GABAA receptor (GABA A R) antagonist, which is selective for extrasynaptic GABA A Rs incorporating the α5 subunit (α5-GABA A Rs). The present study reports on the preclinical neuropsychopharmacological profile of S44819. Significantly, no sedative or pro-convulsive side effects of S44819 were found at doses up to 30 mg/kg i.p. Object recognition (OR) memory in intact mice was enhanced by S44819 (0.3 mg/kg p.o.) given before the acquisition trial. Mice treated with phencyclidine for two weeks and tested six days after the cessation of treatment failed to show OR memory. This deficit was corrected by a single administration of S44819 (0.1, 0.3 or 1 mg/kg p.o.) prior to the acquisition trial. The amnestic effect of ketamine in rats tested in the eight-arm radial maze (reference and working memory versions) was blocked by S44819 (3 mg/kg p.o.). Extinction of cued fear was preserved during treatment with S44819 (3 mg/kg/diem i.p.). Administration of S44819 had no significant effect in the Vogel-conflict test, the elevated plus maze, the forced swim, the marble-burying and the tail-suspension tests. In contrast, anxiolytic/antidepressant-like effects of the compound were found in paradigms that have mnemonic components, such as social interaction, fear-potentiated startle and social avoidance induced by negative life experience. In summary, S44819 enhanced intact recognition memory and ameliorated memory deficits induced by inhibition of NMDA receptors. Anxiolytic/antidepressant efficacy was limited to paradigms involving cognitive function. In conclusion, S44819 is a novel psychoactive pro-cognitive compound with potential as a therapeutic agent in dementia., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Numerous studies in animals and humans have established that oxytocin (OT) reduces anxiety. In rats, the prelimbic (PL) subregion of the medial prefrontal cortex (mPFC) is among the brain areas implicated in the anxiolytic actions of OT. However, questions remain about the anatomical and receptor specificity of OT and its mechanism of action. Here we assessed whether the regulation of anxiety by mPFC OT is restricted to the PL subregion and evaluated whether oxytocin receptor (OTR) activation is required for OT to have an anxiolytic effect. We also examined whether OT interacts with GABA in the mPFC to reduce anxiety and investigated the extent to which OT in the mPFC affects activation of mPFC GABA neurons as well as neuronal activation in the amygdala, a primary target of the mPFC which is part of the neural network regulating anxiety. We found that OT reduced anxiety-like behavior when delivered to the PL, but not infralimbic or anterior cingulate subregions of the mPFC. The anxiolytic effect of OT in the PL mPFC was blocked by pretreatment with an OTR, but not a vasopressin receptor, antagonist as well as with a GABA A receptor antagonist. Lastly, administration of OT to the PL mPFC was accompanied by increased activation of GABA neurons in the PL mPFC and altered neuronal activation of the amygdala following anxiety testing. These results demonstrate that OT in the PL mPFC attenuates anxiety-related behavior and may do so by engaging GABAergic neurons which ultimately modulate downstream brain regions implicated in anxiety., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

α5-containing GABA A Rs are potential therapeutic targets for clinical conditions including age-related dementia, stroke, schizophrenia, Down syndrome, anaesthetic-induced amnesia, anxiety and pain. α5-containing GABA A Rs are expressed in layer 5 cortical neurons and hippocampal pyramidal neurons where they mediate both tonic currents and slow inhibitory postsynaptic currents (IPSCs). A range of drugs has been developed to specifically modulate these receptors. The main α5-containing GABA A Rs that are likely to exist in vivo are the α5β1γ2, α5β2γ2 and α5β3γ2 isoforms. We currently have few clues as to how these isoforms are distributed between synaptic and extrasynaptic compartments or their relative roles in controlling neuronal excitability. Accordingly, the aim of this study was to define the basic biophysical and pharmacological properties of IPSCs mediated by the three isoforms in a hippocampal neuron-HEK293 cell co-culture assay. The IPSC decay time constants were slow (α5β1γ2L: 45 ms; α5β1γ2L: 80 ms; α5β3γ2L: 184 ms) and were largely dominated by the intrinsic channel deactivation rates. By comparing IPSC rise times, we inferred that α5β1γ2L GABA A Rs are located postsynaptically whereas the other two are predominantly perisynaptic. α5β3γ2L GABA A Rs alone mediated tonic currents. We quantified the effects of four α5-specific inverse agonists (TB-21007, MRK-016, α5IA and L-655708) on IPSCs mediated by the three isoforms. All compounds selectively inhibited IPSC amplitudes and accelerated IPSC decay rates, albeit with distinct isoform specificities. MRK-016 also significantly accelerated IPSC rise times. These results provide a reference for future studies seeking to identify and characterize the properties of IPSCs mediated by α5-containing GABA A R isoforms in neurons., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

In Huntington's disease (HD), dysfunctional affective processes emerge as key symptoms of disturbances. In human HD and transgenic rat models of the disease, the amygdala was previously shown to have a reduced volume and to carry a high load of mutant huntingtin (mHTT) aggregates. In search of the pathophysiology of affective dysregulation in HD, we hypothesized a specific role of the central amygdala (CeA), known to be particularly involved in emotional regulation. Using transgenic BACHD rats carrying full-length human mHTT, we compared behavioral consequences of pharmacological modulation of CeA function by infusing GABA A receptor (GABA A R) antagonist picrotoxin into ∼4.5 month old BACHD and WT rats before confronting them to potentially threatening situations. Our results show that disinhibition of the CeA induced differential behaviors in WT and BACHD rats in our tasks: it increased social contacts and responses to the threatening warning signal in an avoidance task in BACHD rats but not in WT animals. At the cellular level, analyzes of amygdala alteration/dysfunction showed (1) an age-dependent increase in number and size of mHTT aggregates specifically in the CeA of BACHD rats; (2) no alteration of GABA and GABA A R expression level, but (3) an increased neuronal reactivity (Arc labelling) to a threatening stimulus in the medial part of this nucleus in 4.5 months old BACHD rats. These results suggest a basal pathological hyper-reactivity in the CeA (in particular its medial part) in the transgenic animals. Such amygdala dysfunction could account, at least in part, for affective symptoms in HD patients., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Many clinical and molecular features of the fragile X syndrome, a common form of intellectual disability and autism, can be modeled by deletion of the Fmr1 protein (Fmrp) in mice. Previous studies showed a decreased expression of several components of the GABAergic system in Fmr1 knockout mice. Here, we used this mouse model to investigate the functional consequences of Fmrp deletion on hippocampal GABAergic inhibition in the CA1-region of the hippocampus. Whole-cell patch-clamp recordings demonstrated a significantly reduced amplitude of evoked inhibitory postsynaptic currents (eIPSCs) and a decrease in the amplitude and frequency of spontaneous IPSCs. In addition, miniature IPSCs were reduced in amplitude and frequency and decayed significantly slower than mIPSCs in controls. Quantitative real-time PCR revealed a significantly lower expression of α2, β1 and δ GABA A receptor subunits in the hippocampus of the juvenile mice (P22) compared to wild-type littermates. Correspondingly, we found also at the protein level reduced amounts of α2, β1 and δ subunits in Fmr1 knockout mice. Overall, these results demonstrate that the reduction in several components of the GABAergic system is already present at young age and that this reduction results in measurable abnormalities on GABA A receptor-mediated phasic inhibition. These abnormalities might contribute to the behavioral and cognitive deficits of this fragile X mouse model., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

NMDA glutamatergic and GABAergic transmission have both been implicated in regulating working memory functions mediated by the prefrontal cortex (PFC), and perturbations in these neurotransmitter systems have been proposed to underlie deficits in these functions observed in schizophrenia. Here, we examined the consequence of disrupting GABAergic or NMDA glutamatergic transmission within the medial PFC of rats on a delayed-response paradigm with translational relevance to working memory tasks used with humans. The operant delayed non-match to position task consisted of a sample phase (one lever extended) and a choice phase wherein rats were required to choose the opposite lever, separated by a variable delay (1-24 s). In well-trained rats, inactivation of the PFC via infusions of GABA agonists baclofen/muscimol (100 ng each) induced delay-independent deficits. Reducing PFC GABA transmission with the GABA-A receptor antagonist bicuculline (12.5-50 ng) also caused delay-independent impairments and increased trial omissions and response latencies during the sample and end-of-delay phases. On the other hand, non-selective blockade of PFC NMDA receptors with MK-801 (3-6 μg) disrupted performance, but these effects more closely resembled delay-dependent impairments. However, selective blockade of GluN2B-containing NMDA receptors with Ro-25-6981 (2.5 μg) did not affect any measures of performance. These results demonstrate that both intact PFC GABA and NMDA receptor signalling are integral for accurate delayed-responding, although they may differentially regulate encoding vs maintenance of information within working memory. Furthermore they suggest that perturbations of both of these neurochemical signals within the PFC may contribute differentially to impairments in working memory observed in schizophrenia., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Despite the highest expression in striatum, physiological function of cylindromatosis (CYLD), a deubiquitinating enzyme, remains unexplored. We found, in the present study, that the duration of spontaneous up-states in the striatum is shorter and membrane potential fluctuation preceding action potential and firing rate are increased in Cyld(-/-) mice. Excess striatal GABAergic inhibition likely plays the major role in this alteration as supported by the findings: (1) the levels of striatal GABAA and GABAB receptors in Cyld(-/-) mice are increased, (2) pharmacological blockade of GABAB receptors rescues the shortened up-state phenotype, and (3) pharmacological blockade of GABAA receptors rescues the power of beta frequency oscillations. Our results indicate that CYLD alters striatal network function by regulating the protein expression levels of GABA receptors., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Stress-induced reinstatement of cocaine seeking requires corticotropin releasing factor (CRF) actions in the ventral tegmental area (VTA). However the mechanisms through which CRF regulates VTA function to promote cocaine use are not fully understood. Here we examined the role of GABAergic neurotransmission in the VTA mediated by GABA-A or GABA-B receptors in the reinstatement of extinguished cocaine seeking by a stressor, uncontrollable intermittent footshock, or bilateral intra-VTA administration of CRF. Rats underwent repeated daily cocaine self-administration (1.0 mg/kg/ing; 14 × 6 h/day) and extinction and were tested for reinstatement in response to footshock (0.5 mA, 0.5" duration, average every 40 s; range 10-70 s) or intra-VTA CRF delivery (500 ng/side) following intra-VTA pretreatment with the GABA-A antagonist, bicuculline, the GABA-B antagonist, 2-hydroxysaclofen or vehicle. Intra-VTA bicuculline (1, 10 or 20 ng/side) failed to block footshock- or CRF-induced cocaine seeking at either dose tested. By contrast, 2-hydroxysaclofen (0.2 or 2 μg/side) prevented reinstatement by both footshock and intra-VTA CRF at a concentration that failed to attenuate food-reinforced lever pressing (45 mg sucrose-sweetened pellets; FR4 schedule) in a separate group of rats. These data suggest that GABA-B receptor-dependent CRF actions in the VTA mediate stress-induced cocaine seeking and that GABA-B receptor antagonists may have utility for the management of stress-induced relapse in cocaine addicts., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

Maternal cigarette smoking during pregnancy and maternal nicotine exposure in animal models are associated with cognitive impairments in offspring. However, the underlying mechanism remains unknown. Oriens-lacunosum moleculare (OLM) cells expressing α2* nicotinic acetylcholine receptors (nAChRs) are an important component of hippocampal circuitry, gating information flow and long-term potentiation (LTP) in the CA1 region. Here we investigated whether early postnatal nicotine exposure alters the normal role of α2*-nAChR-expressing OLM cells during adolescence in rats. We found that early postnatal nicotine exposure significantly decreased not only the number of α2-mRNA-expressing interneurons in the stratum oriens/alveus, but also α2*-nAChR-mediated responses in OLM cells. These effects of nicotine were prevented by co-administration with the nonselective nAChR antagonist mecamylamine, suggesting that nicotine-induced activation, but not desensitization, of nAChRs mediates the effects. α2*-nAChR-mediated depolarization of OLM cells normally triggers action potentials, causing an increase in spontaneous inhibitory postsynaptic currents in synaptically connected pyramidal cells. However, these α2*-nAChR-mediated effects were profoundly reduced after early postnatal nicotine exposure, suggesting altered control of CA1 circuits by α2*-nAChR-expressing OLM cells. Furthermore, these effects were associated with altered excitatory neural activity and LTP as well as the loss of normal α2*-nAChR-mediated control of excitatory neural activity and LTP. These findings suggest the altered function of α2*-nAChR-expressing OLM cells as an important target of further study for identifying the mechanisms underlying the cognitive impairment induced by maternal smoking during pregnancy., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

The prefrontal cortex (PFC) is critical for higher-order cognitive functions, including decision-making. In psychiatric conditions such as schizophrenia, prefrontal dysfunction co-occurs with pronounced alterations in decision-making ability. These alterations include a diminished ability to utilize probabilistic reinforcement in guiding future choice, and a reduced willingness to expend effort to receive reward. Among the neurochemical abnormalities observed in the PFC of individuals with schizophrenia are alterations in the production and function of the inhibitory neurotransmitter gamma-aminobutyric acid (GABA). To probe how PFC GABA hypofunction may contribute to alterations in cost/benefit decision-making, we assessed the effects GABAA-receptor antagonist bicuculline (BIC; 50 ng in 0.5 μl saline/hemisphere) infusion in the medial PFC of rats during performance on a series of well-validated cost/benefit decision-making tasks. Intra-PFC BIC reduced risky choice and reward sensitivity during probabilistic discounting and decreased the preference for larger rewards associated with a greater effort cost, similar to the behavioral sequelae observed in schizophrenia. Additional experiments revealed that these treatments did not alter instrumental responding on a progressive ratio schedule, nor did they impair the ability to discriminate between reward and no reward. However, BIC induced a subtle but consistent impairment in preference for larger vs. smaller rewards of equal cost. BIC infusion also increased decision latencies and impaired the ability to "stay on task" as indexed by reduced rates of instrumental responding. Collectively, these results implicate prefrontal GABAergic dysfunction as a key contributing factor to abnormal decision-making observed in schizophrenia and other neuropsychiatric conditions with similar neurobiological and behavioral alterations., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

The combination of the selective serotonin reuptake inhibitors (SSRIs) and atypical antipsychotic drugs shows better therapeutic efficacy than SSRI monotherapy in the treatment of depression. However, the underlying mechanisms responsible for the augmenting effects of olanzapine are not fully understood. Here, we report that olanzapine enhances the SSRI-induced increase in extracellular serotonin (5-HT) levels and antidepressant-like effects by inhibiting GABAergic neurons through 5-HT6 receptor antagonism in the dorsal raphe nucleus (DRN). In organotypic raphe slice cultures, treatment with olanzapine (1-100 μM) enhanced the increase in extracellular 5-HT levels in the presence of fluoxetine (10 μM) or citalopram (1 μM). The enhancing effect of olanzapine was not further augmented by the GABAA receptor antagonist bicuculline. Electrophysiological analysis revealed that olanzapine (50 μM) decreased the firing frequency of GABAergic neurons in acute DRN slices. Among many serotonergic agents, the 5-HT6 receptor antagonist SB399885 (1-100 μM) mimicked the effects of olanzapine by enhancing the SSRI-induced increase in extracellular 5-HT levels, which was not further augmented by bicuculline or olanzapine. SB399885 (50 μM) also decreased the firing frequency of GABAergic neurons in the DRN. In addition, an intraperitoneal administration of SB399885 (10 mg/kg) to mice significantly enhanced the antidepressant-like effect of a subeffective dose of citalopram (3 mg/kg) in the tail-suspension test. These results suggest that olanzapine decreases local inhibitory GABAergic tone in the DRN through antagonism of 5-HT6 receptors, thereby increasing the activity of at least part of serotonergic neurons, which may contribute to the augmentation of the efficacy of SSRIs., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

Postictal refractoriness may be taken as an expression of lasting activity of inhibitory systems arresting seizures. We tested drugs interfering with GABAergic inhibitory system in pairs of cortical epileptic afterdischarges induced with 1-min interval in rats. Under control conditions the second stimulation failed to elicit an afterdischarge. This postictal refractoriness was not affected by antagonists of GABAA receptors acting at three binding sites (bicuculline, picrotoxin, benzodiazepine inverse agonist Ro 19-4603) as well as by a less specific antagonist pentetrazol. In contrast, antagonist of GABAB receptors CGP35348 partially blocked the refractoriness. Cooperation of different inhibitory systems is probably necessary to abolish postictal refractoriness in neocortex. This article is part of the Special Issue entitled 'GABAergic Signaling in Health and Disease'., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

N-acetylcysteine (NAC) has been reported to reverse the psychotomimetic effects in the rodent phencyclidine model of psychosis and shown beneficial effects in treating patients with schizophrenia. The effect of NAC has been associated with facilitating the activity of cystine-glutamate antiporters on glial cells concomitant with the release of non-vesicular glutamate, which mainly stimulates the presynaptic metabotropic glutamate receptor subtype 2 receptors (mGluR2). Recent evidence demonstrated that functional interactions between serotonin 5-HT2A receptor (5-HT(2A)R) and mGluR2 are responsible to unique cellular responses when targeted by hallucinogenic drugs. The present study determined the effects of NAC on hallucinogenic 5-HT(2A)R agonist (±)1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane (DOI)-elicited behavioral and molecular responses in mice and DOI-evoked field potentials in the mouse cortical slices. NAC significantly attenuated DOI-induced head twitch response and expression of c-Fos and Egr-2 in the infralimbic and motor cortex and suppressed the increase in the frequency of excitatory field potentials elicited by DOI in the medial prefrontal cortex. In addition, the cystine-glutamate antiporter inhibitor (S)-4-carboxyphenylglycine (CPG) and the mGluR2 antagonist LY341495 reversed the suppressing effects of NAC on DOI-induced head twitch and molecular responses and increased frequency of excitatory field potentials, supporting that NAC attenuates the 5-HT(2A)R-mediated hallucinogenic effects via increased activity of cystine-glutamate antiporter followed by activation of mGluR2 receptors. These findings implicate NAC as a potential therapeutic agent for hallucinations and psychosis associated with hallucinogen use and schizophrenia., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Corticotropin-releasing factor (CRF) is implicated in the pathogenesis of bipolar disorder, an illness associated with deficits in prepulse inhibition (PPI) of the acoustic startle response. Valproate is used in the treatment of bipolar disorder and may alter CRF activity via a GABA(A)-ergic mechanism. This study determined the effect of valproate on CRF-disrupted PPI and examined the role of the hypothalamic-pituitary-adrenal axis and GABA-ergic signaling in the effect of valproate. Valproate (60-240 mg/kg) dose-dependently reversed PPI deficits displayed by transgenic mice overexpressing CRF (CRFtg), and normalized PPI deficits induced by CRF i.c.v. infusion in 129Sv mice. Valproate enhanced corticosterone secretion more effectively in CRFtg than in wild-type mice. The effect of valproate on PPI was not blocked by the GABA(A) receptor antagonist bicuculline, the GABA(B) receptor antagonists phaclofen and SCH 50911 or combined administration of a GABA(A) and GABA(B) receptor antagonist. The beneficial effect of valproate on PPI was not mimicked by the GABA(A) receptor agonist muscimol, the GABA transaminase inhibitor vigabatrin, the histone deacetylase (HDAC) inhibitor sodium butyrate or by the mood stabilizers lithium, carbamazepine, lamotrigine or topiramate. Thus, we showed that valproate improves CRF-induced PPI deficits, albeit via a so far unknown mechanism. These marked beneficial effects of valproate on CRF-induced sensorimotor gating deficits suggest that valproate may be of particular value in specific subgroups of bipolar patients that are characterized by alterations in the CRF system., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Stimulation of cannabinoid CB1 receptors or inhibition of nitric oxide synthase (NOS) in the dorsolateral periaqueductal gray (dlPAG) decreases anxiety-like behavior. Moreover, activation of CB1 receptors attenuates flight responses induced by nitric oxide (NO) donors in the dlPAG, suggesting that endocannabinoids and NO could interact to control defensive responses such as anxiety-like behavior. To test this hypothesis male Wistar rats received intra-dlPAG microinjections of anandamide (AEA) or NO inhibitors and were tested in the elevated plus maze (EPM). Combined administration of low and ineffective doses of AEA and the NO scavenger (c-Ptio), the nNOS inhibitor (NPA) or the soluble guanylate cyclase inhibitor (ODQ) induced anxiolytic-like effects. The CB1 receptor antagonist AM251, but not the GABAA receptor antagonist bicuculline, attenuated the effect induced by AEA+c-Ptio combination. No effect, however, was found when anxiolytic doses of these same drugs were administered together. Combination of higher, ineffective doses of AEA and c-Ptio, NPA or ODQ was again anxiolytic. The effect of the former combination was prevented by low and ineffective doses of the GABAA receptor antagonist bicuculline or the GABA synthesis inhibitor L-allilglycine, suggesting that they depend on GABAA-mediated neurotransmission. AM251 was also able to attenuate this effect, indicating that in the presence of NO inhibition, the resultant anxiolytic-like effect could be due to AEA action on CB1 receptors. The present results suggest that the AEA and nitrergic systems exert a complex functional interaction in the dlPAG to modulate anxiety behavior, probably interfering, in addition to glutamate, also with GABAergic mechanisms., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Here we report that juxtacellular labeled GABA interneurons in the basolateral amygdaloid nucleus anterior part (BLA) of rats with 6-hydroxydopamine lesions of the substantia nigra pars compacta (SNc) showed a more burst-firing pattern, while having no change in the firing rate. In sham-operated and the lesioned rats, systemic administration of 5-HT(2A/2C) receptor agonist DOI produced excitation, inhibition and unchanged in the firing rate of the interneurons, and the mean response of DOI was excitatory. However, cumulative dose producing excitation in the lesioned rats was higher than that of sham-operated rats. The local administration of DOI in the BLA also produced three types of responses in two groups of rats. Furthermore, the local administration of DOI excited the interneurons in sham-operated rats, whereas the mean firing rate of the interneurons in the lesioned rats was not affected at the same dose. The excitatory effect of the majority of the interneurons after systemic and local administration of DOI was not reversed by the selective 5-HT(2C) receptor antagonist SB242084, and the inhibitory effect of DOI in all the interneurons examined was reversed by GABA(A) receptor antagonist picrotoxinin. The SNc lesion in rats did not change the density of GAD67/5-HT(2A) receptor co-expressing neurons in the BLA. These results indicate that the SNc lesion changes the firing activity of BLA GABA interneurons. Moreover, DOI regulated the firing activity of the interneurons mainly through activation of 5-HT(2A) receptor, and the lesion led to a decreased response of the interneurons to DOI, which attributes to dysfunction of 5-HT(2A) receptor on these interneurons., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Amyotrophic Lateral Sclerosis (ALS) is a neurodegenerative disorder that affects upper and lower motor neurons. Previous evidence has indicated that excitotoxic cell death in ALS may remarkably depend on Cl(-) ion influx through the GABA(A) receptors. In this study we have analysed the effect of Monocyte Chemoattractant Protein-1 (MCP-1), a chemokine expressed to a higher level in ALS patients, on GABAA receptors in cultured cortical neurons from a genetic model of ALS (G93A) and compared with wild type SOD1 (SOD1) and their corresponding non transgenic littermates (Control). By performing electrophysiological experiments we have observed that, in cortical neurons MCP-1 (2-150 ng/ml) induced an enhancement of GABA-evoked currents that was significantly higher in G93A neurons compared to controls. The effect of MCP-1 was not dependent on the activation of its receptor CCR2, while it was blocked by flumazenil, the antagonist of benzodiazepine sites. Analysis of GABAA receptor subunit composition has indicated an altered subunit expression level in G93A cortical neurons compared to controls. Instead, in cultured spinal neurons MCP-1 induced a significant reduction of GABA-evoked currents, also through the benzodiazepine sites, indicating a region-specific mechanism of action. However, no differences were observed in the current reduction between the three neuronal populations. These findings provide the first evidence that MCP-1, acting on benzodiazepine sites, can modulate the GABA-evoked currents, depending on the subunit composition of GABA(A) receptor. In cortical neurons MCP-1 upmodulates the GABA-evoked current and this effect is exacerbated in the mutated neurons. It is reasonable to assume that the higher Cl(-) influx through GABA(A) receptors in the presence of MCP-1 in mutated cortical neurons may induce an excitotoxicity acceleration. Agents able to block the MCP-1 production may then prove useful for ALS treatment., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Recent electrophysiological evidence suggests that ethanol simultaneously exerts opposite effects on the activity of dopamine (DA) neurons in the ventral tegmental area (VTA) through two parallel mechanisms, one promoting and the other reducing the GABA release onto VTA DA neurons. Here we explore the possible behavioural implications of these findings by investigating the role displayed by acetaldehyde (the main metabolite of ethanol) and the non-metabolized fraction of ethanol in motor activity of rats. We analyse the appearance of motor activation or depression after intra-VTA administration of ethanol in rats subjected to different pharmacological pre-treatments designed to preferentially test either the effects of acetaldehyde or the non-metabolized ethanol. Motor activity was evaluated after intra-VTA administration of 35 nmol of ethanol, an apparently ineffective dose that does not modify the motor activity of animals. Pharmacological pre-treatments were used in order to either increase (cyanamide, 10 mg/kg, ip) or decrease (D-penicillamine, 50 mg/kg, ip and sodium azide, 7 mg/kg, ip) acetaldehyde levels in the VTA. Pre-treatments aimed to augment acetaldehyde, increased motor activity of rats. Otherwise, pre-treatments intended to decrease local acetaldehyde levels evoked significant reductions in motor activity that were prevented by the local blockade (bicuculline, 17.5 pmol) of the GABAA receptors. Our findings suggest that the brain-generated acetaldehyde is involved in the stimulant effects of ethanol, whereas the non-biotransformed fraction of ethanol, acting through the GABAA receptors, would account for the depressant effects. The present behavioural findings suggest that ethanol dually modulates the activity of DA neurons., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Progesterone is neuroprotective after spinal cord injury, however its mechanism of action remains unexplored. Here we used organotypic spinal cord slice cultures from 3 weeks-old mice to evaluate the mechanisms of neuroprotection by progesterone and its 5α-reduced metabolites. In vitro spinal cord injury, using a weight drop model, induced a decrease in the number of motoneurons. This was correlated with an increase in the number of dying cells (PI⁺ cells) and in LDH release. Addition of 10 μM of progesterone, 5α-dihydroprogesterone (5α-DHP) or allopregnanolone (3α, 5α-tetrahydroprogesterone) to the medium at the time of injury rescued the spinal cord slices from the effects of damage. Progesterone prevented membrane cell damage, motoneuron loss and cell death. These effects were not due to its bioconversion to 5α-DHP nor to allopregnanolone, as supported by the finasteride, an inhibitor of 5α-reductase enzymes, and by the absence of 5α-reduced progesterone metabolites in the slices analyzed by gas chromatography-mass spectrometry. The neuroprotective effects of progesterone required PR as they could not be observed in slices from homozygous knockout PR(-/-) mice. Allopregnanolone treatment was also neuroprotective. Its effects were not due to its bioconversion back to 5α-DHP, which can activate gene transcription via PR, because they were still observed in slices from knockout PR(-/-) mice. Allopregnanolone effects involved GABA(A) receptors, as they were inhibited by the selective GABA(A) receptor antagonist Gabazine, in both PR(+/+) and PR(-/-) mice. Altogether, these findings identify both PR and GABA(A) receptors as important targets for neuroprotection by progestagens after spinal cord injury., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Previous studies demonstrated that the Group III mGlu receptor-selective orthosteric agonist, LSP1-2111 produced anxiolytic- but not antidepressant-like effects upon peripheral administration. Herein, we report the pharmacological actions of Lu AF21934, a novel, selective, and brain-penetrant positive allosteric modulator (PAM) of the mGlu(4) receptor in the stress-induced hyperthermia (SIH), four-plate, marble-burying and Vogel's conflict tests. In all models, except Vogel's conflict test, a dose-dependent anxiolytic-like effect was seen. The anti-hyperthermic effect of Lu AF21934 (5 mg/kg) in the SIH test was inhibited by the benzodiazepine receptor antagonist flumazenil (10 mg/kg) and was not serotonin-dependent, as it persisted in serotonin-deficient mice and upon blockade of either 5-HT(1A) receptors by WAY100635, or 5-HT(2A/2C) receptors by ritanserin. These results suggest that the GABAergic system, but not the serotonergic system, is involved in the mechanism of the anxiolytic-like phenotype of Lu AF21934 in rodents. Lu AF21934 did not produce antidepressant-like effects in the tail suspension test (TST) in mice; however, it decreased the basal locomotor activity of mice that were not habituated to activity cages. This article is part of a Special Issue entitled 'Metabotropic Glutamate Receptors'., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

We have previously demonstrated that the basolateral amygdala (BLA) is a key component of a neural circuit mediating memory formation for emotionally relevant stimuli in an ethologically-based model of conditioned fear, termed conditioned defeat (CD). In this model, subjects are socially defeated by a larger, more aggressive hamster. Upon subsequent exposure to a smaller, non-aggressive intruder, the defeated animal will show high levels of submissive behaviors and fail to defend its territory. Here we examined whether the medial prefrontal cortex (mPFC), an area with extensive connections with the amygdala, is also a component of this circuit. Temporary inactivation of the mPFC using muscimol, a GABA(A) receptor agonist, significantly enhanced the acquisition but not expression of CD, while blockade of GABA(A) receptors in the mPFC using bicuculline, a GABA(A) antagonist, impaired acquisition of CD. Given these findings, we next sought to test whether plasticity related to the defeat experience occurs in the mPFC. We infused anisomycin, a protein synthesis inhibitor, in the mPFC but this treatment did not alter the acquisition of CD. In our final experiment, we demonstrated that bicuculline failed to alter the acquisition of CD. Together, these results demonstrate for the first time that while the mPFC is both necessary and sufficient for the acquisition of CD, it does not appear to mediate plasticity related to the defeat experience. In contrast, while plasticity underlying CD does appear to occur in the BLA, GABAergic receptor inhibition in the BLA is not sufficient to enhance CD. This article is part of a Special Issue entitled 'Post-Traumatic Stress Disorder'., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

Recent research suggests that adaptations elicited by drugs of abuse share common features with traditional learning models, and that drugs of abuse cause long-term changes in behavior by altering synaptic function and plasticity. In this study, endocannabinoid (eCB) signaling in the dorsolateral striatum, a brain region vital for habit formation, was evaluated in acutely isolated brain slices from ethanol (EtOH)-consuming rats and control rats. EtOH-consuming rats had free access to a 20% EtOH solution for three 24 hour sessions a week during seven weeks and consumed an average of 3.4 g/kg per session. eCB-mediated long-lasting disinhibition (DLL) of population spike (PS) amplitude induced by moderate frequency stimulation was impaired in EtOH-consuming rats, and was not restored by the muscarinic receptor antagonist scopolamine (10 μM). The lack of DLL could be linked to a reduced GABA(A) receptor tone, since bicuculline-mediated disinhibition of striatal output was significantly reduced in slices from EtOH-consuming rats. However, eCB signaling induced by high frequency stimulation (HFS) was also impaired in slices from EtOH-consuming rats and isolated control rats. Activation of presynaptic cannabinoid 1 receptors (CB1R) with WIN55,212-2 (250 nM, 1 μM) significantly modulated PS amplitude in slices from age-matched control rats while slices from EtOH-consuming rats remained unaffected, indicating that eCB signaling is inhibited at a level that is downstream from CB1R activation. Intermittent alcohol intake for seven weeks might thus be sufficient to modulate a presynaptic mechanism that needs to be synergized with CB1R activation for induction of long-term depression (LTD). In conclusion, alcohol consumption inhibits striatal eCB signaling in a way that could be of importance for understanding the neurological underpinnings of addictive behavior., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

The frequency and duration of glutamatergic inputs to the striatum are strong determinants of the net effect of retrograde endocannabinoid (eCB) signaling, and key factors in determining if long-term depression (LTD) has a net disinhibitory or inhibitory action in striatum. Low to moderate frequency stimulation in the dorsolateral striatum elevates eCB levels to an extent that primarily depresses transmitter release at inhibitory synapses, leading to a long-lasting disinhibition (DLL) of synaptic output. The aim of this study was to further characterize the basic features of endocannabinoid-mediated DLL of striatal output induced by moderate frequency stimulation (5 Hz, 60 s). DLL was inhibited in slices treated with the group 1 metabotropic glutamate receptor (mGluR) antagonists MPEP (40 μM) and CPCCOEt (40 μM), the dopamine D2 receptor antagonist sulpiride (5 μM), the L-type calcium channel blocker nifedipine (20 μM), the nicotinic receptor antagonist mecamylamine (10 μM), the muscarinic agonist oxotremorine sesquifumarate (10 μM), and strychnine (0.1 μM). Strychnine did not block DLL induced by WIN55,212-2 (250 nM), showing that glycine receptor-mediated modulation of eCB signaling occurs upstream from CB(1)R activation. Scopolamine (10 μM) restored DLL in strychnine-treated slices, suggesting that inhibition of glycine receptors on cholinergic interneurons could modulate eCB signaling by enhancing muscarinic receptor activation and reducing the opening of L-type calcium channels in response to depolarization. These data suggests that similar activation points are required for stimulation-induced DLL as for LTD at excitatory striatal synapses, and that cholinergic interneurons are key modulators of stimulation-induced eCB signaling in the striatum., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

Epileptiform discharges recorded in the 4-aminopyridine (4-AP) in vitro epilepsy model are mediated by glutamatergic and GABAergic signaling. Using a 60-channel perforated multi-electrode array (pMEA) on corticohippocampal slices from 2 to 3 week old mice we recorded interictal- and ictal-like events. When glutamatergic transmission was blocked, interictal-like events no longer initiated in the hilus or CA3/CA1 pyramidal layers but originated from the dentate gyrus granule and molecular layers. Furthermore, frequencies of interictal-like events were reduced and durations were increased in these regions while cortical discharges were completely blocked. Following GABA(A) receptor blockade interictal-like events no longer propagated to the dentate gyrus while their frequency in CA3 increased; in addition, ictal-like cortical events became shorter while increasing in frequency. Lastly, drugs that affect tonic and synaptic GABAergic conductance modulated the frequency, duration, initiation and propagation of interictal-like events. These findings confirm and expand on previous studies indicating that multiple synaptic mechanisms contribute to synchronize neuronal network activity in forebrain structures. This article is part of a Special Issue entitled 'Trends in neuropharmacology: in memory of Erminio Costa'., (Copyright © 2010 Elsevier Ltd. All rights reserved.)

Seldom does a single individual have such a profound effect on the development of a scientific discipline as Erminio Costa had on neuropharmacology. During nearly sixty years of research, Costa and his collaborators helped established many of the basic principles of the pharmacodynamic actions of psychotherapeutics. His contributions range from defining basic neurochemical, physiological and behavioral properties of neurotransmitters and their receptors, to the development of novel theories for drug discovery. Outlined in this report is a portion of his work relating to the involvement of monoamines and GABA in mediating the symptoms of neuropsychiatric disorders and as targets for drug therapies. These studies were selected for review because of their influence on my own work and as an illustration of his logical and insightful approach to research and his clever use of techniques and technologies. Given the significance of his work, the legions of scientist who collaborated with him, and those inspired by his reports, his research will continue to have an impact as long as there is a search for new therapeutics to alleviate the pain and suffering associated with neurological and psychiatric disorders. This article is part of a Special Issue entitled 'Trends in neuropharmacology: in memory of Erminio Costa'., (Copyright © 2010 Elsevier Ltd. All rights reserved.)

Intense noxious stimuli impair GABAergic inhibition in spinal dorsal horn, which has been proposed as a critical contributor to pathological pain. However, how the reduced inhibition exacerbates the transfer of nociceptive information at excitatory glutamatergic synapses is still poorly understood. The present study demonstrated that one of the striking consequences of GABAergic disinhibition was to enhance the function of N-methyl-D-aspartate subtype glutamate receptors (NMDARs), a well-characterized player in central sensitization. We found that intrathecal application of bicuculline, a GABA(A) receptor antagonist, to remove the inhibition readily elicited mechanical allodynia in naive mice, which could be dose-dependently attenuated by NMDARs antagonist D-APV. Biochemical analysis demonstrated that bicuculline did not affect the total expression levels of the obligatory NMDARs subunit NR1 and the regulatory subunit NR2A and NR2B. However, bicuculline promoted NR1 phosphorylation at Serine 897 (NR1-S897) by cAMP-dependent protein kinase (PKA). This PKA-mediated phosphorylation incorporated NR1 along with NR2B into synapses. When PKA inhibitor H-89 was intrathecally applied, it totally eliminated bicuculline-induced NMDARs phosphorylation, synaptic redistribution as well as pain sensitization. Importantly, the reduced inhibition also operated to enhance NMDARs functions after peripheral inflammation, because spinal injection of diazepam to rescue the inhibition in inflamed mice greatly depressed PKA phosphorylation of NR1-S897, reduced the synaptic concentration of NR1/NR2B and meanwhile, alleviated the inflammatory pain. These data suggested that removal of GABAergic inhibition allowed for PKA-mediated NMDARs phosphorylation and synaptic accumulation, thus exaggerating NMDARs-dependent nociceptive transmission and behavioral sensitization., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

If the pregnant and lactating female rats are exposed to environmental levels of bisphenol-A (BPA), their male offspring will display hyperactivity and attention-deficit. In patients with attention-deficit/hyperactivity disorder (ADHD), the size of the amygdala is reported to be reduced. This study examined functional alterations in the basolateral amygdala (BLA) of the postnatal 28-day-old male offspring exposed perinatally to BPA (BPA-rats). We specifically focused on the synaptic properties of GABAergic/dopaminergic systems in the BLA. A single electrical stimulation of the capsule fibers evoked multispike responses with an enhanced primary population spikes (1st-PS) in the BPA-rats. A single train of high-frequency stimulation of the fibers induced NMDA receptor (NMDAR) dependent long-term potentiation (LTP) in BPA-rats, but not in control rats. Also, paired-pulse inhibition (PPI, GABA-dependent) in control rats was reversed to paired-pulse facilitation (PPF) in BPA-rats. Perfusion of slices obtained from BPA-rats with the GABA(A) receptor (GABA(A)R) agonist muscimol blocked the multispike responses and LTP, and recovered PPI. By contrast, the dopamine D1 receptor antagonist SCH23390 abolished LTP and attenuated the increased amplitude of 1st-PS in BPA-rats. Conversely, blockade of GABA(A)R by bicuculline could produce the multispike responses and PPF in BLA in control rats. Furthermore, in BLA the infusion of SCH23390, muscimol or the NMDAR blocker MK801 ameliorated the hyperactivity and improved the deficits in attention. These findings suggest that the perinatal exposure to BPA causes GABAergic disinhibition and dopaminergic enhancement, leading to an abnormal cortical-BLA synaptic transmission and plasticity, which may be responsible for the hyperactivity and attention-deficit in BPA-rats. This article is part of a Special Issue entitled 'Synaptic Plasticity & Interneurons'., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

Our earlier studies have demonstrated that the non-selective group III mGlu receptor agonist, ACPT-I, produced anxiolytic rather than antidepressant-like actions after its peripheral administration. Here, we describe the effects of LSP1-2111 ((2S)-2-amino-4-[hydroxy[hydroxy(4-hydroxy-3-methoxy-5-nitro-phenyl)methyl]phosphoryl]butanoic acid), a novel orthosteric, preferential agonist of the mGlu4 receptor, a member of the group III mGlu receptors family, in the stress-induced hyperthermia (SIH) and elevated plus-maze (EPM) tests in mice. In both tests an anxiolytic-like effect was clearly seen in doses of 2 and 5 mg/kg, i.p. The compound did not produce antidepressant-like effects in the tail suspension test (TST) or in the forced swim test (FST) in mice. The potential anxiolytic effect of LSP1-2111 (5 mg/kg) in the SIH test was inhibited by the benzodiazepine receptor antagonist flumazenil (given i.p., 10 mg/kg), and by a 5-HT(1A) receptor antagonist N-{2-[4-(2-methoxyphenyl)-1-piperazinyl]ethyl}-N-(2-pyridynyl)cyclohexane-carboxamide (WAY100635) (0.1 mg/kg, s.c.). At the same time, ritanserin (0.5 mg/kg i.p.), the 5-HT(2A/C) receptor antagonist, did not change the anxiolytic-like effects of LSP1-2111. Moreover, the compound was not effective in 5-HT depleted animals. The results of these studies indicate that the GABAergic and serotonergic systems are involved in the potential anxiolytic action of LSP1-2111., (Copyright © 2010 Elsevier Ltd. All rights reserved.)