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While the trigeminal ganglion is often considered a passive conduit of sensory transmission, neurons and satellite glial cells (SGCs) within it can release neurotransmitters and express neuroreceptors. Some trigeminal ganglion neurons contain the neurotransmitter γ-aminobutyric acid (GABA) and express GABA receptors. There is behavioral evidence that increased GABA levels in the trigeminal ganglion decreases nociception, while a loss of GABA receptors results in hyperalgesia, although the neural mechanisms for this remain to be investigated. In this study, the expression of GABA receptors by trigeminal ganglion neurons that innervate rat labial skin and masseter muscle was compared using immunohistochemistry. The effect of intraganglionic administration of GABA receptor agonists was investigated by single unit recording of trigeminal brainstem and ganglion neuron responses to stimulation of the labial skin and/or masseter muscle in anesthetized rats. The mean frequency of expression of GABAA and GABAB receptors by masseter and labial skin ganglion neurons was 62.5% and 92.7%, and 55.4% and 20.3%, respectively. The expression of both GABA receptors was significantly greater in skin ganglion neurons. Masticatory muscle evoked brainstem trigeminal neuron responses were significantly attenuated by intraganglionic injection of muscimol (GABAA) but not baclofen (GABAB). The mechanical sensitivity of slow and fast conducting masticatory muscle afferent fibers was decreased and increased, respectively, by intraganglionic injection of both muscimol and baclofen. Activation of GABAA receptors may exert a gating effect on sensory transmission through the trigeminal ganglion by decreasing putative nociceptive input and enhancing innocuous sensory input.

The signal transmission in the nervous system operates through a sensitive balance between excitatory (E) inputs and inhibitory (I) responses. Imbalances in this system contribute to the development of pathologies such as seizures. In Caenorhabditis elegans, the locomotor circuit operates via the coordinated activity of cholinergic excitatory (E) and GABAergic inhibitory (I) transmission. Changes in E/I inputs can cause uncontrolled electrical discharges, mimicking the physiology of seizures. Molecules derived from 1,3,4-oxadiazole have been found to exhibit diverse biological activities, including anticonvulsant effect. In this work, we study the activity of the compound 2-[(4-methoxyphenylselenyl)methylthio]-5-phenyl-1,3,4-oxadiazole (MPMT-OX) in the GABAergic and cholinergic systems. We demonstrate that MPMT-OX reduced the locomotor activity of C. elegans with a normal balance between the E/I systems and increased the resistance to paralysis in worms exposed to pentylenetetrazol and aldicarb. MPMT-OX increased seizure resistance and assisted in the recovery of locomotor activity in worms with deletions in the genes unc-46, which regulates the transport of GABA into vesicles, and unc-49, which encodes the GABAA receptor. C. elegans with deletions in the unc-25 and unc-47 genes did not respond to treatment. Therefore, we suggest that the compound MPMT-OX upregulates GABAergic signaling in a manner dependent on the unc-25 gene, which is responsible for GABA synthesis, and unc-47, which encodes the vesicular GABA transporter.

The key question to be answered in the present study was whether the medial septal GABAergic receptors are engaged in the pharmacological profile of the hippocampal formation (HPC) theta rhythm induced by vagal nerve stimulation (VNS). It was demonstrated that the medial septal microinfusion of GABAA and GABAB agonists (muscimol and baclofen) resulted in a progressive reduction of the power of VNS-induced theta. The medial septal microinfusion of GABAA and GABAB antagonists (bicuculline and 2-OH saclofen) resulted in the generation of VNS-induced theta with increased power. The effect of the combined medial septal microinfusion of GABAA agonist and antagonist and GABAB agonist and antagonist on VNS-induced theta rhythm was also evaluated: in the presence of GABAA,B antagonists the effect of agonists predominated. In separate experiments, GABAA and GABAB antagonists were administrated in anesthetized rats pretreated with i.v. administration of atropine sulfate. Atropine was found to abolish spontaneous theta and VNS-induced theta examined in the presence of bicuculline or 2-OH saclofen. The present data provide evidence that the medial septal GABAA and GABAB receptors are involved in the central mechanisms responsible for modulation of VNS-induced HPC theta oscillations. Furthermore, the results of the present study also demonstrate that, in fact, both GABAergic and cholinergic involvement is necessary for the appearance of VNS-induced theta.

Taste neophobia is a feeding system defense mechanism that limits consumption of an unknown, and therefore potentially dangerous, edible until the post-ingestive consequences are experienced. We found that transient pharmacological inhibition (induced with the GABA agonists baclofen and muscimol) of the gustatory thalamus (GT; Experiment 1), but not medial amygdala (MeA; Experiment 2), during exposure to a novel saccharin solution attenuated taste neophobia. In Experiment 3 we found that inhibition of MeA neurons (induced with the chemogenetic receptor hM4DGi) enhanced the expression of taste neophobia whereas excitation of MeA neurons (with hM3DGq) had no influence of taste neophobia. Overall, these results refine the temporal involvement of the GT in the occurrence of taste neophobia and support the hypothesis that neuronal excitation in the GT is necessary for taste neophobia. Conversely, we show that chemogenetically, but not pharmacologically, inhibiting MeA neurons is sufficient to exaggerate the expression of taste neophobia.

The olfactory circuitry in mice involves a well-characterized, vertical receptor type-specific organization, but the localized inhibitory effect from granule cells on action potentials that propagate laterally in secondary dendrites of mitral cell remains open to debate. To understand the functional dynamics of the lateral (horizontal) circuits, we analyzed odor-induced signaling using transgenic mice expressing a genetically encoded Ca2+ indicator specifically in mitral/tufted and some juxtaglomerular cells. Optical imaging of the dorsal olfactory bulb (dOB) revealed specific patterns of glomerular activation in response to odor presentation or direct electric stimulation of the olfactory nerve (ON). Application of a mixture of ionotropic and metabotropic glutamate receptor antagonists onto the exposed dOB completely abolished the responses to direct stimulation of the ON as well as discrete odor-evoked glomerular responses patterns, while a spatially more widespread response component increased and expanded into previously nonresponsive regions. To test whether the widespread odor response component represented signal propagation along mitral cell secondary dendrites, an NMDA receptor antagonist alone was applied to the dOB and was found to also increase and expand odor-evoked response patterns. Finally, with dOB excitatory synaptic transmission completely blocked, application of 1 mM muscimol (a GABAA receptor agonist) to a circumscribed volume in the deep external plexiform layer (EPL) induced an odor non-responsive area. These results indicate that odor stimulation can activate olfactory reciprocal synapses and control lateral interactions among olfactory glomerular modules along a wide range of mitral cell secondary dendrites by modulating the inhibitory effect from granule cells.

The flexible implementation of active and passive strategies to avoid danger is critical to survival. Conversely, the inappropriate allocation of these behaviors may underlie pathological avoidance in neuropsychiatric illnesses. The present study investigated whether these two poles of avoidance may be differentially regulated by subdivsions of the nucleus accumbens, the core (NAcC) and shell (NAcS), which are known to bi-directionally control flexible action selection during reward-seeking. In so doing, we developed a novel cued active/inhibitory avoidance task conducted in operant chambers that entailed presentations of two distinct, 15 s auditory cues. One cue indicated that impending foot-shocks could be avoided by pressing a lever (active avoidance), whereas another cue signaled that shocks could be avoided by withholding presses (inhibitory avoidance). In well-trained rats, pharmacological inactivation of either the NAcC or NAcS impaired active avoidance. In contrast, inhibitory avoidance was disrupted by inactivation of the NAcS, but not NAcC, reflecting a deficit in response-inhibition that manifested as more inhibitory avoidance failures and lever-presses, as well as increased locomotion. Foot-shock sensitivity was unaffected by inactivation of either subregion. In a subsequent experiment, treatment with the monoamine releaser d -amphetamine (1 mg/kg) did not affect active avoidance, but disinhibited lever pressing during inhibitory avoidance trials. These results provide novel insight into the ventral striatal and monoaminergic regulation of flexible response allocation and inhibition that facilitates avoidance behavior and highlight the importance of different subregions of the NAc in action selection during aversively-motivated behaviors.

Somatosensory over-reactivity is common among patients with autism spectrum disorders (ASDs) and is hypothesized to contribute to core ASD behaviors. However, effective treatments for sensory over-reactivity and ASDs are lacking. We found distinct somatosensory neuron pathophysiological mechanisms underlie tactile abnormalities in different ASD mouse models and contribute to some ASD-related behaviors. Developmental loss of ASD-associated genes Shank3 or Mecp2 in peripheral mechanosensory neurons leads to region-specific brain abnormalities, revealing links between developmental somatosensory over-reactivity and the genesis of aberrant behaviors. Moreover, acute treatment with a peripherally restricted GABAA receptor agonist that acts directly on mechanosensory neurons reduced tactile over-reactivity in six distinct ASD models. Chronic treatment of Mecp2 and Shank3 mutant mice improved body condition, some brain abnormalities, anxiety-like behaviors, and some social impairments but not memory impairments, motor deficits, or overgrooming. Our findings reveal a potential therapeutic strategy targeting peripheral mechanosensory neurons to treat tactile over-reactivity and select ASD-related behaviors. Treatment with a peripherally restricted GABAA receptor agonist in multiple distinct autism spectrum disorder mouse models reveals a potential therapeutic strategy for select ASD-related behaviors., National Institutes of Health (U.S.) (Grants F31 MH098641A, R01 MH097104), National Institute of Mental Health (U.S.) (Conte Center Grant P50 MH094271)

Both the lateral septum (LS) and anterior hypothalamus (AHA) regulate behavioural defense. We tested whether those two interconnected structures act in serial in that regard. Infusions of the GABAA agonist muscimol into one side of the LS and the contralateral (but not ipsilateral) AHA suppressed rats' burying in the shock-probe test whereas none of our muscimol infusion approaches altered their open-arm avoidance in the elevated plus-maze. These results suggest that the LS-AHA circuit serves a specialized role in defensive responses towards discrete, localizable threat stimuli but not towards potential threats.

Objective Suppression of the gamma-aminobutyric acid (GABA)ergic activity of the zona incerta (ZI) reportedly plays a role in neuropathic pain after spinal cord injury (SCI). A reduction in GABAergic signaling in the ZI of a thoracic hemisection-SCI rat model has been suggested, but not clearly demonstrated. Accordingly, our objective was to investigate whether GABAergic signals influence SCI-induced neuropathic pain. Methods In vivo, we recorded and compared single-unit, neuronal activity between hemisection-SCI and sham-operated rat models. Furthermore, we analyzed neuronal activity in both models following treatment with either a GABAA receptor agonist (muscimol) or antagonist (bicuculline). Results Rats that underwent hemisection SCI exhibited reduced hindpaw withdrawal thresholds, latencies, and decreased ZI neuronal activity compared with sham-operated controls. Importantly, muscimol treatment increased, whereas bicuculline decreased, the firing rates of the ZI neurons. The muscimol treated, hemisection-SCI rats also exhibited increased hindpaw withdrawal thresholds and latencies. Conclusions These data provide evidence that neuropathic pain after SCI is caused by decreased GABAergic signaling in the ZI. Furthermore, our data demonstrate that infusion of a GABAergic drug into the ZI could restore its inhibitory action and improve neuropathic pain behaviors.

The central nervous system, once thought to be a site of immunological privilege, has since been found to harbour immunocompetent cells and to communicate with the peripheral nervous system. In the central nervous system (CNS), glial cells display immunological responses to pathological and physiological stimuli through pro- and anti-inflammatory cytokine and chemokine signalling, antigen presentation and the clearing of cellular debris through phagocytosis. While this neuroinflammatory signalling can act to reduce neuronal damage and comprises a key facet of CNS homeostasis, persistent inflammation or auto-antigen-mediated immunoreactivity can induce a positive feedback cycle of neuroinflammation that ultimately results in necrosis of glia and neurons. Persistent neuroinflammation has been recognised as a major pathological component of virtually all neurodegenerative diseases and has also been a focus of research into the pathology underlying psychiatric disorders. Thus, pharmacological strategies to curb the pathological effects of persistent neuroinflammation are of interest for many disorders of the CNS. Accumulating evidence suggests that GABAergic activities are closely bound to immune processes and signals, and thus the GABAergic neurotransmitter system might represent an important therapeutic target in modulating neuroinflammation. Here, we review evidence that inflammation induces changes in the GABA neurotransmitter system in the CNS and that GABAergic signalling exerts a reciprocal influence over neuroinflammatory processes. Together, the data support the hypothesis that the GABA system is a potential therapeutic target in the modulation of central inflammation.

Changes in functional GABAAR signaling in hippocampus have previously been evaluated using pre-clinical animal models of either diffuse brain injury or extreme focal brain injury that precludes measurement of cells located ipsilateral to injury. As a result, there is little information about the status of functional GABAAR signaling in dentate granule cells (DGCs) located ipsilateral to focal brain injury, where significant cellular changes have been documented. We used whole-cell patch-clamp recordings from hippocampal slices to measure changes in GABAARs in dentate granule cells (DGCs) at 1-2, 3-5, and 8-13 weeks after controlled cortical impact (CCI) brain injury. Synaptic and tonic GABAAR currents (ITonicGABA) were measured in DGCs at baseline conditions and during application of the GABAAR agonist 4,5,6,7-tetrahydroisoxazolo[5,4-c]pyridine-3-ol hydrochloride (THIP) to assess in the function of δ subunit-containing GABAARs. DGCs ipsilateral to CCI exhibited no changes in the amplitude of resting ITonicGABA relative to DGCs after sham-injury or contralateral to CCI. In contrast, there was a significant reduction in the THIP-evoked ITonicGABA in DGCs ipsilateral to CCI at both time-points. Tonic GABAergic inhibition of DGCs ipsilateral to injury also exhibited reduced responsiveness to the neurosteroid THDOC. ITonicGABA in DGCs ipsilateral to CCI did not exhibit a change in sensitivity to L655,708, an inverse agonist with selectivity for α5 subunit-containing GABAARs, suggesting a lack of functional change in GABAARs containing this subunit. At the 8-13 week time-point, gene expression of GABAAR subunits expected to contribute to ITonicGABA (i.e., α4, α5 and δ) was not significantly altered by CCI injury in isolated dentate gyrus. Collectively, these results demonstrate enduring functional changes in ITonicGABA in DGCs ipsilateral to focal brain injury that occur independent of altered gene expression.

Stimulation of mu opioid receptors using drugs like morphine can increase eating when injected into multiple brain regions including the lateral septum (LS). The LS has been classically associated with reward, anxiety and fearful behaviors but more recently has also received attention with regard to control of feeding. To investigate the role of LS opioid receptors in feeding, we injected mu, delta, and kappa opioid receptor agonists and a mu specific receptor antagonist directly into the LS of rats. We expected that if feeding is mu receptor specific then only mu receptor agonists would increase feeding. We also hypothesized that mu receptor antagonists would suppress the feeding elicited by mu receptor agonists like morphine. Further, because the LS is densely populated with GABA receptors, we used the GABAA receptor agonist muscimol to assess the effect of inhibition of LS neurons on feeding. Our results show that the mu receptor agonist morphine and the specific mu agonist DAMGO reliably and significantly increase feeding behavior across doses tested, while delta and kappa agonists were ineffective. CTAP, a specific mu receptor antagonist, at low doses unexpectedly increased morphine-elicited feeding but at high doses decreased morphine's effect, consistent with mediation by mu receptors. Finally, muscimol rapidly elicited feeding, suggesting a role for LS GABAA receptors in feeding stimulation. These findings suggest that mu opioid receptors in the LS play complex roles in feeding and that neural inhibition may be a mechanism by which they elicit feeding.

GABA is the most common inhibitory neurotransmitter in both vertebrate and invertebrate nervous systems. In insects, inhibition plays important roles at the neuromuscular junction, in the regulation of central pattern generators, and in the modulation of information in higher brain processing centers. Additionally, increasing our understanding of the functions of GABA is important since GABAA receptors are the targets of several classes of pesticides. To investigate the role of GABA in motor function, honey bee foragers were injected with GABA or with agonists or antagonists specific for either GABAA or GABAB receptors. Compounds that activated either type of GABA receptor decreased activity levels. Bees injected with the GABAA receptor antagonist picrotoxin lost the ability to right themselves, whereas blockade of GABAB receptors led to increases in grooming. Injection with antagonists of either GABAA or GABAB receptors resulted in an increase in extended wing behavior, during which bees kept their wings out at right angles to their body rather than folded along their back. These data suggest that the GABA receptor types play distinct roles in behavior and that GABA may affect behavior at several different levels.

Objective Since benzodiazepines (BZPs) became clinically available for the treatment of status epilepticus (SE) in children, the incidence of neurological sequelae has increased. However, the cause–effect relationship is poorly understood. In this paper, we examined the effect of BZPs on an inflammation-induced SE (iSE) animal model. Method Inflammation was induced by injecting poly(I:C) (pIC 10 mg/kg, postnatal day 12–14), seizure was induced by injecting pilocarpine hydrochloride (PILO 200 mg/kg, postnatal day 15) into C57BL/6J mice, and the pIC + PILO mice were used as the iSE model (miSE). The GABA-A receptor agonist midazolam (MDL 0.5 mg/kg) was used to inhibit seizures. Sequelae were evaluated by performing behavior and immunohistochemical analyses in the chronic phase. Result The exploratory activity of mice in the miSE plus MDL group increased significantly, indicating that hyperactivity was newly induced by MDL in miSE mice. The contextual fear memory of the miSE mice was also significantly increased and that of miSE treated with MDL returned to the normal level. The parvalbumin-positive GABA neurons were decreased in number by pIC + PILO which was rescued by MDL. Apoptosis marker ssDNA-positive cells were increased by pIC + PILO which could not be rescued by MDL. Therefore, we propose that BZP-dependent therapy for SE needs to be rethought from the perspective of using other treatment approaches.

The central nucleus of the amygdala (CeA) as the main output of amygdala plays an important role in memory processes. In this study we first evaluated the effects of intra-CeA administrations of different doses of a cannabinoid CB1 agonist, WIN55, 212-2, GABA(A) receptor agonist and antagonist, muscimol and bicuculline, alone on memory retention using passive avoidance learning (PAL) test in rats. Then we examined the effects of GABA(A) receptor agents on the responses induced by intra-CeA microinjection of different doses of WIN55, 212-2. We found that administration of WIN55, 212-2 (0.05, 0.1, 0.2 and 0.4μg/rat) immediately after training impaired memory retrieval in a dose-dependent fashion. Although pre-test intra-CeA administration of muscimol (125, 250 and 500ng/rat) alone had no effect on the step-through latency, its co-administration (125ng/rat) with different doses of WIN55, 212-2 potentiated the amnesic effects of any doses of WIN55, 212-2. The results also showed that pre-test intra-CeA administration of bicuculline (200, 400 and 800ng/rat) alone had no significant effect, but at dose of 200ng/rat disrupted post-training WIN55, 212-2-induced amnesia in the retention test. Furthermore, the additional effect of muscimol (125ng/rat) on memory impairment induced by WIN55, 212-2 (0.1μg/rat) was prevented by intra-CeA co-injection of bicuculline (200ng/rat). We indicated that stimulating or blocking GAGA(A) receptors in the CeA by muscimol and bicuculline interfere with WIN55, 212-2-induced deficits in memory retention in a PAL task and therefore suggests an interaction between cannabinergic and GABAergic systems of the CeA in memory process.

Animal models of substance abuse have established a role for the caudal granular insular cortex (CGIC) in drug taking behaviour for several addictive substances, yet nothing has thus far been reported for alcohol. The current research was undertaken to examine the involvement of the CGIC in a rat model of alcohol self-administration. We investigated the inactivating effects of local infusions of a γ-aminobutyric acid agonist mixture (baclofen/muscimol) into the CGIC on alcohol self-administration under a fixed ratio-3 (FR-3). This inactivation of the CGIC decreased operant responding for alcohol along with a corresponding decrease in oral alcohol intake. Our results demonstrate the involvement of the CGIC in alcohol taking behaviour and suggest future studies examine the differential involvement of the various subregions of the insular cortex in various aspects of alcohol consumption.

SummaryTo study rapid sensorimotor remapping, we developed a method to train rats in a behavior in which subjects are cued, on each trial, to apply a sensorimotor association to orient either toward a visual target (“Pro”) or away from it, toward its reverse (“Anti”). Multiple behavioral asymmetries suggested that Anti behavior is cognitively demanding while Pro is easier to learn and perform. This is consistent with a prominent hypothesis in the primate literature that Anti requires prefrontal cortex (PFC), whereas Pro could be mediated by midbrain superior colliculus (SC). Pharmacological inactivation of rat medial PFC supported its expected role in Anti. Remarkably, bilateral SC inactivation substantially impaired Anti while leaving Pro essentially intact. Moreover, SC inactivation eliminated the performance cost of switching from Anti to Pro tasks. Our results establish a rodent model of single-trial sensorimotor remapping and suggest a critical role for SC in the cognitively demanding Anti task.

Our previous work has shown that cerebellar interposed nucleus (IN) modulates immune function. Herein, we reveal mechanism underlying the immunomodulation. Treatment of bilateral cerebellar IN of rats with 3-mercaptopropionic acid (3-MP), a glutamic acid decarboxylase antagonist that reduces γ-aminobutyric acid (GABA) synthesis, enhanced cellular and humoral immune responses to bovine serum albumin, whereas injection of vigabatrin, a GABA-transaminase inhibitor that inhibits GABA degradation, in bilateral cerebellar IN attenuated the immune responses. The 3-MP or vigabatrin administrations in the cerebellar IN decreased or increased hypothalamic GABA content and lymphoid tissues' norepinephrine content, respectively, but did not alter adrenocortical or thyroid hormone levels in serum. In addition, a direct GABAergic projection from cerebellar IN to hypothalamus was found. These findings suggest that GABAergic neurons in cerebellar IN regulate immune system via hypothalamic and sympathetic pathways.

In this study, we analyzed the impact that spontaneous seizures might have on the plasma membrane expression, composition and function of GABAA receptors (GABAARs). For this, tissue of chronically epileptic rats was collected within 3 hours of seizure occurrence (≤3 hours group) or at least 24 hours after seizure occurrence (≥24 hours group). A retrospective analysis of seizure frequency revealed that selecting animals on the bases of seizure proximity also grouped animals in terms of overall seizure burden with a higher seizure burden observed in the ≤3 hours group. A biochemical analysis showed that although animals with more frequent/recent seizures (≤3 hours group) had similar levels of GABAAR at the plasma membrane they showed deficits in inhibitory neurotransmission. In contrast, tissue obtained from animals experiencing infrequent seizures (≥24 hours group) had increased plasma membrane levels of GABAAR and showed no deficit in inhibitory function. Together, our findings offer an initial insight into the molecular changes that might help to explain how alterations in GABAAR function can be associated with differential seizure burden. Our findings also suggest that increased plasma membrane levels of GABAAR might act as a compensatory mechanism to more effectively maintain inhibitory function, repress hyperexcitability and reduce seizure burden. This study is an initial step towards a fuller characterization of the molecular events that trigger alterations in GABAergic neurotransmission during chronic epilepsy.

A range of medications target different aspects of the GABA system; understanding their effects is important to inform further drug development. Effects on the waking EEG comparing these mechanisms have not been reported; in this study we compare the effects on resting MEG spectra of the benzodiazepine receptor agonist zolpidem, the delta sub-unit selective agonist gaboxadol (also known as THIP) and the GABA reuptake inhibitor tiagabine. These were two randomised, single-blind, placebo-controlled, crossover studies in healthy volunteers, one using zolpidem 10 mg, gaboxadol 15 mg and placebo, and the other tiagabine 15 mg and placebo. Whole head MEG recordings and individual MEG spectra were divided into frequency bands. Baseline spectra were subtracted from each post-intervention spectra and then differences between intervention and placebo compared. After zolpidem there were significant increases in beta frequencies and reduction in alpha frequency power; after gaboxadol and tiagabine there were significant increases in power at all frequencies up to beta. Enhancement of tonic inhibition via extrasynaptic receptors by gaboxadol gives rise to a very different MEG signature from the synaptic action of zolpidem. Tiagabine theoretically can affect both types of receptor; from these MEG results it is likely that the latter is the more prominent effect here.This article is part of the Special Issue entitled ‘GABAergic Signaling in Health and Disease’.

a b s t r a c t Abnormal enhancement of polysynaptic brainstem reflexes has been previously reported in patients with spinal cord injury (SCI). We aimed to investigate trigemino-cervical reflex (TCR) in SCI since it may reflect alterations in the connections of trigeminal proprioceptive system and cervical motoneurons. Consecutive 14 patients with SCI and 16 healthy subjects were included in this study. All patients were in the chronic phase. TCR was recorded over sternocleidomastoid (SCM) and splenius capitis (SC) muscles by stimulation of infraorbital nerve. We measured onset latency, amplitudes and durations of responses and compared between groups. We obtained stable responses over both muscles after one sided stimulation in healthy volunteers whereas probability of TCR was decreased in patients over both SCM (78.6% vs. 100%, p = 0.050) and SC (71.4% vs. 100%, p = 0.022). The absence of TCR was related to use of oral baclofen (≥50 mg/day). However, when present, responses of SCI group had higher amplitudes and were more persistent. We demonstrated that TCR probability was similar to healthy subjects in SCI patients who used no or low dose oral baclofen. But it had higher amplitudes and longer durations. It was not obtained in only two patients who used oral baclofen more than 50 mg/day.

We have previously shown that GABA protects pancreatic islet cells against apoptosis and exerts anti-inflammatory effects. Notably, GABA inhibited the activation of NF-κB in both islet cells and lymphocytes. NF-κB activation is detrimental to beta cells by promoting apoptosis. However, the mechanisms by which GABA mediates these effects are unknown. Because the above-mentioned effects mimic the activity of sirtuin 1 (SIRT1) in beta cells, we investigated whether it is involved. SIRT1 is an NAD(+)-dependent deacetylase that enhances insulin secretion, and counteracts inflammatory signals in beta cells. We found that the incubation of a clonal beta-cell line (rat INS-1) with GABA increased the expression of SIRT1, as did GABA receptor agonists acting on either type A or B receptors. NAD(+) (an essential cofactor of SIRT1) was also increased. GABA augmented SIRT1 enzymatic activity, which resulted in deacetylation of the p65 component of NF-κB, and this is known to interfere with the activation this pathway. GABA increased insulin production and reduced drug-induced apoptosis, and these actions were reversed by SIRT1 inhibitors. We examined whether SIRT1 is similarly induced in newly isolated human islet cells. Indeed, GABA increased both NAD(+) and SIRT1 (but not sirtuins 2, 3 and 6). It protected human islet cells against spontaneous apoptosis in culture, and this was negated by a SIRT1 inhibitor. Thus, our findings suggest that major beneficial effects of GABA on beta cells are due to increased SIRT1 and NAD(+), and point to a new pathway for diabetes therapy.

The endogenous neuroactive steroid allopregnanolone (ALLO) has previously been shown to induce reinstatement of ethanol seeking in rodents. ALLO is a positive allosteric modulator at both synaptic and extrasynaptic GABA A receptors. The contribution of each class of GABA A receptors in mediating reinstatement of ethanol seeking is unknown. The first aim of the present study was to determine whether ganaxolone (GAN), a longer-acting synthetic analog of ALLO, also promotes reinstatement of ethanol seeking. The second aim was to examine whether preferentially activating extrasynaptic GABA A receptors with the selective agonist gaboxadol (THIP) was sufficient to reinstate responding for ethanol in mice. Male C57BL/6J mice were trained to lever press for access to a 10% ethanol (v/v) solution (10E), using a sucrose-fading procedure. Following extinction of the lever-pressing behavior, systemic THIP (0, 4 and 6 mg/kg) and GAN (0, 10, and 15 mg/kg) were tested for their ability to reinstate ethanol-appropriate responding in the absence of 10E access. GAN significantly increased lever pressing on the previously active lever, while THIP did not alter lever-pressing behavior. The results of this study suggest that direct activation of extrasynaptic GABA A receptors at the GABA site is not sufficient to induce ethanol seeking in the reinstatement procedure. Future studies are necessary to elucidate the mechanisms and brain areas by which differences in the pharmacological activity of GAN and THIP at the GABA A receptor contribute to the dissimilarity in their effect on the reinstatement of ethanol seeking. Nonetheless, based on the increased use of these drugs in clinical trials across multiple disease states, the effects of GAN or THIP on alcohol seeking may be an important consideration if these drugs are to be used clinically in a population with a co-occurring alcohol use disorder.

Background γ -Aminobutyric acid exists throughout the body, and the brain γ -aminobutyric acid receptor (GABAR) regulation reduces oxidative stress (OS). Effects of GABAR regulation in the liver are unknown. Ischemia or reperfusion injury after orthotopic liver transplantation (OLT) or shear stress after split OLT (SOLT) with a small-for-size graft causes OS-induced graft damage. Here, the strategic potential of graft pretreatment in vivo and ex vivo by GABAR regulation was investigated. Materials and methods Recipient rats were divided into seven groups according to the graft pretreatments and graft types: (1) laparotomy, (2) OLT, (3) GABAR regulation in vivo and OLT, (4) GABAR regulation ex vivo and OLT, (5) SOLT, (6) GABAR regulation in vivo and SOLT, and (7) GABAR regulation ex vivo and SOLT. Survival study, biochemical assays, histopathologic or immunohistologic assessments, and Western blotting were performed at 6 h after OLT or SOLT. Results Graft pretreatment in vivo prolonged survival after SOLT. Histopathologic and biochemical profiles verified that graft pretreatment in vivo reduced graft damage after OLT or SOLT. Immunohistologically, graft pretreatment in vivo prevented apoptotic inductions after OLT or SOLT. The 4-hydroxynonenal confirmed the OS after OLT or SOLT, and graft pretreatment in vivo improved the OS. Graft pretreatment in vivo decreased ataxia-telangiectasia–mutated kinase and H2AX after OLT or SOLT. Graft pretreatment in vivo increased phosphatidylinositol 3 kinase and Akt after SOLT. In contrast, GABAR regulation ex vivo did not work. Conclusions Graft pretreatment in vivo , not ex vivo , prevented the ischemia or reperfusion injury–mediated OS after OLT or SOLT via the ataxia-telangiectasia–mutated kinase/H2AX pathway and the shear stress–mediated OS after SOLT with small-for-size graft via the phosphatidylinositol 3 kinase/Akt pathway.

Fragile X syndrome (FXS) is one of the most prevalent and well-studied monogenetic causes of intellectual disability and autism and, although rare, its high penetrance makes it a desirable model for the study of neurodevelopmental disorders more generally. Indeed recent studies suggest that there is functional convergence of a number of genes that are implicated in intellectual disability and autism indicating that an understanding of the cellular and biochemical dysfunction that occurs in monogenic forms of these disorders are likely to reveal common targets for therapeutic intervention. Fundamental research into FXS has provided a wealth of information about how the loss of function of the fragile X mental retardation protein results in biochemical, anatomical and physiological dysfunction leading to the discovery of interventions that correct many of the core pathological phenotypes associated with animal models of FXS. Most promisingly such strategies have led to development of drugs that are now in clinical trials. This review highlights how progress in understanding disorders such as FXS has led to a new era in which targeted molecular treatment towards neurodevelopmental disorders is becoming a reality. This article is part of the Special Issue entitled 'Neurodevelopmental Disorders'.

SummaryCocaine addiction is characterized by long-lasting vulnerability to relapse arising because neutral environmental stimuli become associated with drug use and then act as cues that induce relapse. It is not known how cues elicit cocaine seeking, and why cocaine seeking is more difficult to regulate than seeking a natural reward. We found that cocaine-associated cues initiate cocaine seeking by inducing a rapid, transient increase in dendritic spine size and synaptic strength in the nucleus accumbens. These changes required neural activity in the prefrontal cortex. This is not the case when identical cues were associated with obtaining sucrose, which did not elicit changes in spine size or synaptic strength. The marked cue-induced synaptic changes in the accumbens were correlated with the intensity of cocaine, but not sucrose seeking, and may explain the difficulty addicts experience in managing relapse to cocaine use.

Alcohol use disorder is a compulsive behavior driven by motivational systems and by a poor control of consummatory behavior. The entopeduncular nucleus (EP) seems to be involved in the regulation of executive mechanisms, hence, in the expression of behavior. Endocannabinoids (eCB) are involved in alcohol intake mechanisms. The eCB receptor name cannabinoid receptor 1 (CB1R) is expressed in the EP in GABAergic terminals. The role of the eCB system (eCBs) of the EP in the modulation of alcohol seeking and intake behavior is unknown. Therefore, we decided to investigate the role of the eCBs and its interaction with GABA transmission in rat EP, in the regulation of alcohol intake behavior. Rats were submitted to a 10-day period of moderate alcohol (10% in tap water) ingestion. No tap water was available. On day 11, either anandamide (AEA, CB1 receptor agonist), AM251 (CB1R inverse agonist), baclofen (BAC, GABAB receptor agonist), or CGP35348 (GABAB receptor antagonist) was administered into the EP. One bottle of water and one of alcohol (10% in water) were available ad libitum for the following 24 h, and consumption was quantified at the end of this period. Results show that administration of AEA into the EP decreased alcohol consumption while AM251 and BAC administered independently increased alcohol consumption. AEA prevented the increase induced by AM251 or BAC. Likewise, CGP35348 prevented alcohol ingestion induced by AM251. These data suggest that eCBs dysfunction in the EP may be playing a crucial role in the abuse and dependence of alcohol and other drugs.

It has been previously reported that the GABA(B) receptor agonist baclofen decreases food intake after oral administration and fat intake after intraperitoneal administration. The aim of the study was to investigate the effects of baclofen (1-4 mg/ kg) administered orally (Experiment 1) on food intake in non-deprived rats (n=6) and intraperitoneally (Experiment 2) on fat intake in non-deprived rats (n=8) that were naïve to baclofen (1st set of trials) and in the same group of rats after they were sub-chronically exposed to baclofen (2nd set of trials). The results from Experiment 1 show that baclofen had no effects on food intake during the 1st set of trials, but the 2 and 4 mg/kg doses significantly increased food consumption during the 2nd set of trials. Baclofen produced sedation during the 1st set of trials, but tolerance occurred to this effect and was not apparent during the 2nd set of trials. These observations suggest that the motor effects may have competed with the hyperphagic effects of baclofen during the 1st set of trials. The data from Experiment 2 show that baclofen had no effects on fat intake during either the 1st or 2nd set of trials. The results of the study thus indicate that orally administrated baclofen increases food intake and intraperitoneal administration has no effect on fat intake in non-deprived rats under the conditions used in this study. These findings may have important implications for research on the use of baclofen in studies concerned with ingestive behaviours.

δ-subunit containing extrasynaptic GABA(A) receptors are potential targets for modifying neuronal activity in a range of brain disorders. With the aim of gaining more insight in synaptic and extrasynaptic inhibition, we used a new positive modulator, AA29504, of δ-subunit containing GABA(A) receptors in mouse neurons in vitro and in vivo. Whole-cell patch-clamp recordings were carried out in the dentate gyrus in mouse brain slices. In granule cells, AA29504 (1 μM) caused a 4.2-fold potentiation of a tonic current induced by THIP (1 μM), while interneurons showed a potentiation of 2.6-fold. Moreover, AA29504 (1 μM) increased the amplitude and prolonged the decay of miniature inhibitory postsynaptic currents (mIPSCs) in granule cells, and this effect was abolished by Zn²⁺ (15 μM). AA29504 (1 μM) also induced a small tonic current (12.7 ± 3.2 pA) per se, and when evaluated in a nominally GABA-free environment using Ca²⁺ imaging in cultured neurons, AA29504 showed GABA(A) receptor agonism in the absence of agonist. Finally, AA29504 exerted dose-dependent stress-reducing and anxiolytic effects in mice in vivo. We propose that AA29504 potentiates δ-containing GABA(A) receptors to enhance tonic inhibition, and possibly recruits perisynaptic δ-containing receptors to participate in synaptic phasic inhibition in dentate gyrus.

The Erwinia ligand-gated ion channel (ELIC) is a bacterial homologue of vertebrate Cys-loop ligand-gated ion channels. It is activated by GABA, and this property, combined with its structural similarity to GABAA and other Cys-loop receptors, makes it potentially an excellent model to probe their structure and function. Here we characterise the pharmacological profile of ELIC, examining the effects of compounds that could activate or inhibit the receptor. We confirm that a range of amino acids and classic GABAA receptor agonists do not elicit responses in ELIC, and we show the receptor can be at least partially activated by 5-aminovaleric acid and γ-hydroxybutyric acid, which are weak agonists. A range of GABAA receptor non-competitive antagonists inhibit GABA-elicited ELIC responses including α-endosulfan (IC50 = 17 μM), dieldrin (IC50 = 66 μM), and picrotoxinin (IC50 = 96 μM) which were the most potent. Docking suggested possible interactions at the 2′ and 6′ pore-lining residues, and mutagenesis of these residues supports this hypothesis for α-endosulfan. A selection of compounds that act at Cys-loop and other receptors also showed some efficacy at blocking ELIC responses, but most were of low potency (IC50 > 100 μM). Overall our data show that a number of compounds can inhibit ELIC, but it has limited pharmacological similarity to GLIC and to Cys-loop receptors., Highlights ► ELIC is structurally and functionally similar to Cys-loop receptors. ► ELIC can be activated by GABA but not other GABAA receptor agonists. ► ELIC responses are blocked by some compounds that block the channel of GABA-activated and other Cys-loop receptors. ► The potency of the compounds that block ELIC is lower than in Cys-loop receptors and in GLIC. ► ELIC pharmacology is distinct from that of related receptors.

Background Preclinical and clinical evidence show that the GABA B agonist, baclofen is a promising treatment for addictive disorders; however, until recently its mechanism of action in the human brain was unknown. In previous work we utilized a laboratory model that included a medication versus placebo regimen to examine baclofen's actions on brain circuitry. Perfusion fMRI [measure of cerebral blood flow (CBF)] data acquired ‘at rest’ before and on the last day of the 21-day medication regimen showed that baclofen diminished CBF bilaterally in the VS, insula and medial orbitofrontal cortex (mOFC). In the present study, we hypothesized that a single dose of baclofen would have effects similar to repeated dosing. Methods To test our hypothesis, in a crossover design, CBF data were acquired using pseudo continuous arterial spin labeled ( p CASL) perfusion fMRI. Subjects were either un-medicated or were administered a 20 mg dose of baclofen approximately 110 min prior to scanning. Results Acute baclofen diminished mOFC, amygdala, and ventral anterior insula CBF without causing sedation (family-wise error corrected at p = 0.001). Conclusions Results demonstrate that similar to repeated dosing, an acute dose of baclofen blunts the ‘limbic’ substrate that is hyper-responsive to drugs and drug cues. Smokers often manage their craving and can remain abstinent for extended periods after quitting, however the risk of eventual relapse approaches 90%. Given that chronic medication may not be a practical solution to the long-term risk of relapse, acute baclofen may be useful on an ‘as-needed’ basis to block craving during ‘at risk’ situations.

While our previous work suggests that the midazolam-induced memory impairment results from the inhibition of new association formation, little is known about the neural correlates underlying these effects beyond the effects of GABA agonists on the brain. We used arterial spin-labeling perfusion MRI to measure cerebral blood flow changes associated with the effects of midazolam on ability to learn arbitrary word-pairs. Using a double-blind, within-subject cross-over design, subjects studied word-pairs for a later cued-recall test while they were scanned. Lists of different word-pairs were studied both before and after an injection of either saline or midazolam. As expected, recall was severely impaired under midazolam. The contrast of MRI signal before and after midazolam administration revealed a decrease in CBF in the left dorsolateral prefrontal cortex (DLPFC), left cingulate gyrus and left posterior cingulate gyrus/precuneus. These effects were observed even after controlling for any effect of injection. A strong correlation between the midazolam-induced changes in neural activity and memory performance was found in the left DLPFC. These findings provide converging evidence that this region plays a critical role in the formation of new associations and that low functioning of this region is associated with anterograde amnesia.

The understanding of signaling cascades involved in the induction, promotion, and progression of cancer, although advanced in recent years, is still incomplete. Tracing the imbalance of the impaired, physiologically-essential cellular signaling that drives the neoplastic process is a complex issue. This review discusses the role of the regulator of the fight or flight response, the beta-adrenergic signaling cascade, as a mediator of cancer growth and progression in in vitro and in vivo cancer models. We review a series of experiments from our own laboratory and those of others examining the contribution of this signaling network to lung and other human malignancies and thereby identifying potential targets for chemotherapeutic interventions. The stimulation of the β-adrenergic receptor by lifestyle and environmental factors, as well as a preexisting risk for neoplasm, activates downstream effector molecules (adenylyl cyclase/cAMP/PKA/CREB) concomitant to the transactivation of related pathways (EGFR) that lead to pro-oncogenic signaling; this β-adrenergic pathway thereby encourages cancer growth by evasion of apoptosis, invasion, angiogenesis, and metastasis. GABAergic signaling acts as an antagonist to the β-adrenergic cascade by intercepting adenylyl cyclase activation, and thereby neutralizing the pro-oncogenic effects of β-adrenergic stimulation. The regulation of cancer cell growth by neurobiological signals expands the possibilities for pharmacological interventions in cancer therapy.

Peripherally restricted analgesics are desirable to avoid central nervous system (CNS) side effects of opioids. Nonsteroidal anti-inflammatory drugs produce peripheral analgesia but have significant toxicity. GABA(B) receptors represent peripheral targets for analgesia but selective GABA(B) agonists like baclofen cross the blood-brain barrier. Recently, we found that the CNS-impermeant amino acid, isovaline, produces analgesia without apparent CNS effects. On observing that isovaline has GABA(B) activity in brain slices, we examined the hypothesis that isovaline produces peripheral analgesia mediated by GABA(B) receptors. We compared the peripheral analgesic and CNS effect profiles of isovaline, baclofen, and GABA (a CNS-impermeant, unselective GABA(B) agonist). All three amino acids attenuated allodynia induced by prostaglandin E2 injection into the mouse hindpaw and tested with von Frey filaments. The antiallodynic actions of isovaline, baclofen, and GABA were blocked by the GABA(B) antagonist, CGP52432, and potentiated by the GABA(B) modulator, CGP7930. We measured Behavioural Hyperactivity Scores and temperature change as indicators of GABAergic action in the CNS. ED(95) doses of isovaline and GABA produced no CNS effects while baclofen produced substantial sedation and hypothermia. In a mouse model of osteoarthritis, isovaline restored performance during forced exercise to baseline values. Immunohistochemical staining of cutaneous layers of the analgesic test site demonstrated co-localization of GABA(B1) and GABA(B2) receptor subunits on fine nerve endings and keratinocytes. Isovaline represents a new class of peripherally restricted analgesics without CNS effects, mediated by cutaneous GABA(B) receptors.

Despite evidence that high-affinity GABA(A) receptor subunit mRNA and protein are present in dorsal root ganglia (DRG), low-affinity currents dominate those detected in acutely dissociated DRG neurons in vitro. This observation raises the possibility that high-affinity receptors are normally trafficked out of the DRG toward central and peripheral terminals. We therefore hypothesized that with time in culture, there would be an increase in high-affinity GABA(A) currents in DRG neurons. To test this hypothesis, we studied dissociated DRG neurons 2 h (acute) and 24 h (cultured) after plating with whole-cell patch-clamp techniques, Western blot, and semiquantitative reverse transcriptase polymerase chain reaction (sqRT-PCR) analysis. GABA(A) current density increases dramatically with time in culture in association with the emergence of two persistent currents with EC50's of 0.25±0.01 μM and 3.2±0.02 μM for GABA activation. In a subpopulation of neurons, there was also an increase in the potency of GABA activation of the transient current from an EC50 of 78.16±10.1 μM to 9.56±1.3 μM with time in culture. A fraction of the high-affinity current was potentiated by δ-subunit agonist 4,5,6,7-tetrahydroisoxazolo[5,4-c]pyridine-3-ol (THIP). δ-subunit immunoreactivity was largely restricted to the cytosolic fraction in acute, but the membrane fraction in cultured, DRG neurons, with no detectable change in δ-subunit mRNA. However, the emergence of a high-affinity current blocked by THIP and insensitive to bicuculline was detected in a subpopulation of cultured neurons as well in association with an increase in ρ2- and ρ3-subunit mRNA in cultured DRG neurons. Our results suggest that high-affinity δ-subunit-containing GABA(A) receptors are normally trafficked out of the DRG where they are targeted to peripheral and central processes. They also highlight that the interpretation of data obtained from cultured DRG neurons should be made with caution.

It has been proposed that the medial prefrontal cortex (mPFC) is not necessary for delay eyeblink conditioning (DEC). Here, we investigated the involvement of the mPFC in DEC with a soft or loud tone as the conditioned stimulus (CS) by using electrolytic lesions or muscimol inactivation of guinea pig mPFC. Interestingly, when a soft tone was used as a CS, electrolytic lesions of the mPFC significantly retarded acquisition of the conditioned response (CR), and muscimol infusions into mPFC distinctly inhibited the acquisition and expression of CR, but had no significant effect on consolidation of well-learned CR. In contrast, both electrolytic lesions and muscimol inactivation of mPFC produced no significant deficits in the CR when a loud tone was used as the CS, or in the unconditioned response (UR) when a soft or loud tone was used as the CS. These results demonstrate that the mPFC is essential for the DEC with the soft tone CS but not for the DEC with the loud tone CS.

In the regulation of behavior, the role of GABA neurons has been extensively studied in the circuit of fear, where GABA interneurons play key parts in the acquisition, storage and extinction of fear. Therapeutically, modulators of α(2)/α(3) GABA(A) receptors, such as TPA023, have shown clinical proof of concept as novel anxiolytics, which are superior to classical benzodiazepines by their lack of sedation and much reduced or absent dependence liability. In view of the finding that anxiety disorders and major depression share a GABAergic deficit as a common pathophysiology, the GABA hypothesis of depression has found increasing support. It holds that α(2)/α(3) GABA(A) receptor modulators may serve as novel antidepressants. Initial clinical evidence for this view comes from the significantly enhanced antidepressant therapeutic response when eszopicole, an anxiolytic/hypnotic acting preferentially on α(2)/α(3) and α(1) GABA(A) receptors, was coadministered with an antidepressant. This effect persisted even when sleep items were not considered. These initial results warrant efforts to profile selective α(2)/α(3) GABA(A) receptor modulators, such as TPA023, as novel antidepressants. In addition, GABA(B) receptor antagonists may serve as potential antidepressants. This article is part of a Special Issue entitled 'Anxiety and Depression'.

Primary lens epithelial cell (LEC) cultures derived from newborn (P0) and one-month-old (P30) mouse lenses were used to study GABA (gamma-aminobutyric acid) signaling expression and its effect on the intracellular Ca2+ ([Ca2+]i) level. We have found that these cultures express specific cellular markers for lens epithelial and fiber cells, all components of the functional GABA signaling pathway and GABA, thus recapitulating the developmental program of the ocular lens. Activation of both GABA-A and GABA-B receptors (GABAAR and GABABR) with the specific agonists muscimol and baclofen, respectively induces [Ca2+]i transients that could be blocked by the specific antagonists bicuculline and CGP55845 and were dependent on extracellular Ca2+. Bicuculline did not change the GABA-evoked Ca2+ responses in Ca2-containing buffers, but suppressed them significantly in Ca2+-free buffers suggesting the two receptors couple to convergent Ca2+ mobilization mechanisms with different extracellular Ca2+ sensitivity. Prolonged activation of GABABR induced wave propagation of the Ca2+ signal and persistent oscillations. The number of cells reacting to GABA or GABA + bicuculline in P30 mouse LEC cultures expressing predominantly the synaptic type GABAAR did not differ significantly from the number of reacting cells in P0 mouse LEC cultures. The GABA-induced Ca2+ transients in P30 (but not P0) mouse LEC could be entirely suppressed by co-application of bicuculline and CGP55845. The GABA-mediated Ca2+ signaling may be involved in a variety of Ca2+-dependent cellular processes during lens growth and epithelial cell differentiation.

Ivermectin (IVM) is an antiparasitic drug, widely used in domestic animals. In mammals, IVM act as a GABA agonist. This neurotransmitter has an important role in the regulation of sexual behavior. Thus, this study sought to investigate the effects of various medically relevant doses IVM on the sexual behavior of male rats. In particular, we also wished to examine if previous sexual experience modulated responses to IVM. In the first experiment, the sexual behavior of inexperienced male rats was analyzed after they received 0.2, 0.6, 1.0 or 2.0 mg/kg IVM, 15 min prior to behavioral testing. In the second experiment, the effects of four previous sexual experiences on IVM treated rats (1.0 or 2.0 mg/kg, 15 min prior to the 5th session) were assessed. The standard therapeutic dose (0.2 mg/kg) did not impair the sexual behavior of inexperienced male rats. At a more concentrated dose (0.6 mg/kg), which is still within the therapeutic range, the appetitive phase of sexual behavior of inexperienced male rats was impaired. Likewise, 1.0 mg/kg impaired the appetitive phase. Previous sexual experience blocked almost entirely this sexual impairment, suggesting that previous sexual experience exerts a positive effect in attenuating the sexual impairment produced by IVM treatment. Therefore, the standard therapeutic dose of IVM can be used without producing side effects on sexual behavior. Use of more concentrated therapeutic doses is not recommended during reproductive periods, unless the animals have had previous sexual experience.

In order to clarify the mechanisms involved in the susceptibility to GABAA antagonists-induced seizures in morphine dependent rats, we investigated how GABAA agonists modulate this vulnerability. Seizures were induced to animals by infusion of GABAA antagonists: pentylenetetrazole (PTZ), picrotoxin (PIC) and bicuculline (BIC). GABAA agonists, muscimol (MUS) and 4,5,6,7-tetrahydroisoxazolo [5,4-c]pyridin-3-ol (THIP), were administered intravenous (i.v.) before antagonists. Morphine-dependence significantly decreased the PTZ threshold dose (19.16 ± 1.89 versus 25.74 ± 1.25 mg/kg) while, it had no effect on PIC induced seizures. BIC doses for both threshold and tonic–clonic seizures induction were significantly lower in morphine dependent rats (0.10 ± 0.01 and 0.12 ± 0.02 versus 0.25 ± 0.02 and 0.39 ± 0.07 mg/kg respectively). In morphine-dependence, although pre-treatment with MUS significantly increased the required dose of PTZ for seizures threshold, THIP significantly decreased the required dose of PTZ for tonic–clonic convulsion. Moreover, MUS pretreatment completely recovered the effect of morphine dependency on BIC seizure activity. The results suggest that the capability of GABAA agonists on modulation of propensity to seizures induced by different antagonists in morphine-dependence is dissimilar. Therefore, it seems that long-term morphine alters some properties of GABA system so that the responsive rate of GABAA receptors not only to its antagonists, but also to its agonists will change differently.

Rett syndrome is a severe neurodevelopmental disease caused by mutations of the transcriptional repressor methyl-CpG-binding protein 2 (MeCP2) that induce complex, disabling symptoms, including breathing symptoms. Males of Mecp2-deficient mice (Mecp2(-/y)) normally breathe at birth but develop first altered breathing regulations, thereafter erratic breathing with severe apnoeas, aggravating until respiratory distress and premature death. Mecp2(-/y) mice also develop early GABA deficits. To examine whether GABA deficits contributed to breathing defects of Mecp2(-/y) mice, mice were subjected to acute administration of Midazolam, a benzodiazepine of clinical use known to enhance GABA effects. For the first time, we showed that Midazolam abolished, although transiently, the breathing defects of Mecp2(-/y) mice, confirming a crucial role of GABA deficits in their breathing defects.

Deep brain stimulation at high frequencies (HFS) is currently studied in the treatment of therapy-refractory obsessive-compulsive disorder (OCD). The diversity of targeted brain areas and the discrepancy in demonstrating beneficial effects, highlight the need for better mapping of brain regions in which HFS may yield anti-compulsive effects. This goal may be achieved by investigating the effects of HFS in appropriate animal models of OCD. The present study tested the effect of bilateral HFS or pharmacological inactivation (as induced by intracerebral administration of the GABA-agonist muscimol) of both the Globus pallidus (GP; rodent equivalent to human GP externus) and the Nucleus entopeduncularis (EP; rodent equivalent to human GP internus) on checking behaviour in the quinpirole rat model of OCD. We demonstrate that HFS of the GP does not and HFS of the EP only partially reduces OCD-like behaviour in rats. In contrast, pharmacological inactivation of both GP and EP significantly reduces OCD-like behaviour in the model. These data contrast previously derived data on the effectiveness of HFS of the subthalamic nucleus, nucleus accumbens, GP and EP in the same and other rat models of OCD. We conclude that (i) although GP and EP play an important role in the pathophysiology of OCD, these areas may not represent first choice target structures for HFS, (ii) the effectiveness of HFS may depend on different subtypes of OCD, represented in different animal models, and (iii) differential net mechanisms may subserve the effectiveness of HFS and pharmacological inactivation.

This work was aimed to identify the action of several ion channel and pump inhibitors as well as nicotinic, GABAergic, purinergic and serotoninergic drugs on the resting membrane potential (RMP) and assess the role of cholinergic and GABAergic sensitivity in earthworm muscle electrogenesis. The nicotinic agonists acetylcholine (ACh), carbacholine (CCh) and nicotine depolarize the RMP at concentrations of 5 μM and higher. The nicotinic antagonists (+)tubocurarine, α-bungarotoxin, muscarinic antagonists atropine and hexamethonium do not remove or prevent the CCh-induced depolarization. Verapamil, tetrodotoxin, removal of Cl(-) and Ca(2+) from the solution also cannot prevent the depolarization by CCh. In a Na(+)-free medium, however, CCh lost this depolarization ability and this indicates that the drug opens the sodium permeable pathway. Serotonin, glutamate, glycine, adenosine triphosphate (ATP) and cis-4-aminocrotonic acid (GABA(C) receptor antagonist) had no effect on the RMP. On the other hand, isoguvacin, γ-aminobutyric acid (GABA) and baclofen (GABA(B) receptor agonist) hyperpolarized the RMP. Ouabain, bicucullin (GABA(A) antagonist) and phaclofen (GABA(B) antagonist), as well as the removal of Cl(-), suppressed the effect of GABA and baclofen. CCh did not enhance the depolarization generated by ouabain but, on the other hand, hindered the hyperpolarizing activity of baclofen both in the absence and presence of atropine and (+)tubocurarine. The long-term application of CCh depolarizes the RMP primarily by inhibiting the Na(+)/K(+)-ATPase. The muscle membrane also contains A and B type GABA binding sites, the activation of which increases the RMP at the expense of increasing the action of ouabain- and Cl(-) -sensitive electrogenic pumps.

We previously showed that nasal pressure support ventilation (nPSV) can lead to active inspiratory laryngeal narrowing, which originates from the stimulation of bronchopulmonary receptors. Among the three major types of bronchopulmonary receptors, which are variably stimulated by lung distension, C-fiber endings are remarkable, given that their stimulation can also trigger laryngeal closure. Taking advantage of our lamb model with blocked C-fibers, we aimed to assess whether bronchopulmonary C-fiber endings are involved in the active inspiratory laryngeal narrowing during nPSV. Nine lambs were surgically instrumented to assess states of alertness, electrical activity of a glottal constrictor (EaTA), respiratory movements and arterial blood gases. Forty-eight hours later, two polysomnographic recordings were performed during nPSV 15/4 cmH2O, before and after C-fiber blockade. During nPSV, blockade of C-fibers did not prevent inspiratory EaTA (present for 74 ± 41% of respiratory cycles vs. 64 ± 35%, p = 0.9). We conclude that active inspiratory laryngeal narrowing during nPSV does not originate from bronchopulmonary C-fiber endings.

We studied the hypothesis that some of the spinal mechanisms that are involved in neuropathic hypersensitivity play a role in hypersensitivity induced by REM sleep deprivation (REMSD). Rats with a chronic intrathecal (i.t.) catheter had REMSD of 48 h duration that induced hypersensitivity to mechanical stimulation. After REMSD, the animals were treated i.t. with carbenoxolone (a gap junction decoupler), bumetanide (a blocker of Na + –K + –2Cl − cotransporter 1 or NKCC1), muscimol (a GABA A receptor agonist), or pretreated intraperitoneally with minocycline (an inhibitor of microglia activation). Previously, all these treatments attenuated neuropathic hypersensitivity. Following REMSD, carbenoxolone, bumetanide and muscimol had a strong antihypersensitivity effect, whereas pretreatment with minocycline failed to prevent development of hypersensitivity. The results suggest that among spinal pain facilitatory mechanisms that are common to REMSD and neuropathy are NKCC1 blocker- and gap junction decoupler-reversible mechanisms. Moreover, there is a net pain inhibitory effect by spinal administration of an exogenous GABA A receptor agonist following REMSD as shown earlier in neuropathy. In contrast, activation of spinal microglia may not be as important for the development of hypersensitivity induced by REMSD as following nerve injury.

The GABAB agonists block the rewarding properties of opiates. However, the role of GABAB receptors in the discriminative properties of these drugs has received little attention. In this line, the present study was performed to investigate the effects of acute (Experiment 1) and chronic (Experiment 2) pretreatment with baclofen on the discriminative stimulus effects of morphine. Rats were trained to discriminate morphine (5 mg/kg i.p.) from saline under a two-lever fixed-ratio schedule of food reinforcement. Experiment 1: a morphine generalization curve was obtained under standard conditions in control and baclofen (1 and 2 mg/kg i.p.) pretreated animals. Acute baclofen pretreatment attenuated morphine-lever responding and response rate at both doses. Experiment 2: the animals were randomly divided in two groups and a morphine generalization curve was obtained in daily consecutive test sessions before (TEST1) and after (TEST2) chronic saline (Group I) or chronic baclofen (2 mg/kg) (Group II) administration. As expected, chronic saline pretreatment was ineffective, while chronic baclofen pretreatment attenuated the stimulus properties of morphine, without modifying the rate-decreasing effect of the drug.The data support a role for GABAergic neurotransmission in the discriminative effects of opiates and demonstrate that a short-term treatment with baclofen is useful for decreasing the sensitivity to narcotic cue.

Cholinergic neurons of the pontine laterodorsal tegmentum (LDT) play a critical role in regulation of behavioral state. Therefore, elucidation of mechanisms that control their activity is vital for understanding of how switching between wakefulness, sleep and anesthetic states is effectuated. In vivo studies suggest that GABAergic mechanisms within the pons play a critical role in behavioral state switching. However, the postsynaptic, electrophysiological actions of GABA on LDT neurons, as well as the identity of GABA receptors present in the LDT mediating these actions is virtually unexplored. Therefore, we studied the actions of GABA agonists and antagonists on cholinergic LDT cells by performing patch clamp recordings in mouse brain slices. Under conditions where detection of Cl(-) -mediated events was optimized, GABA induced gabazine (GZ)-sensitive inward currents in the majority of LDT neurons. Post-synaptic location of GABA(A) receptors was demonstrated by persistence of muscimol-induced inward currents in TTX and low Ca(2+) solutions. THIP, a selective GABA(A) receptor agonist with a preference for δ-subunit containing GABA(A) receptors, induced inward currents, suggesting the existence of extrasynaptic GABA(A) receptors. LDT cells also possess GABA(B) receptors as baclofen-activated a TTX- and low Ca(2+)-resistant outward current that was attenuated by the GABA(B) antagonists CGP 55845 and saclofen. The tertiapin sensitivity of baclofen-induced outward currents suggests that a G(IRK) mediated this effect. Further, outward currents were never additive with those induced by application of carbachol, suggesting that they were mediated by activation of GABA(B) receptors linked to the same G(IRK) activated in these cells by muscarinic receptor stimulation. Activation of GABA(B) receptors inhibited Ca(2+) increases induced by a depolarizing voltage step shown previously to activate VOCCs in cholinergic LDT neurons. Baclofen-mediated reductions in depolarization-induced Ca(2+) were unaltered by prior emptying of intracellular Ca(2+) stores, but were abolished by low extracellular Ca(2+) and pre-application of nifedipine, indicating that activation of GABA(B) receptors inhibits influx of Ca(2+) involving L-type Ca(2+) channels. Presence of GABA(C) receptors is suggested by the induction of inward current by (E)-4- amino-2-butenoic acid (TACA) and its inhibition by 1,2,5,6-tetrahydropyridine-4-ylmethylphosphinic (TPMPA), a relatively selective agonist and antagonist, respectively, of GABA(C) receptors. All of these GABA-mediated actions were found to occur in histochemically-identified cholinergic neurons. Taken together, these data indicate for the first time that cholinergic neurons of the LDT exhibit functional GABA(A, B and C) receptors, including extrasynaptically located GABA(A) receptors, which may be tonically activated by synaptic overflow of GABA. Accordingly, the activity of cholinergic LDT neurons is likely to be significantly affected by GABAergic tone within the nucleus, and so, demonstrated effects of GABA on behavioral state may be mediated, in part, via direct actions on cholinergic neurons in the LDT.

5-HT7 receptors in the dorsal raphe nucleus (DRN) influence circadian rhythms, sleep, and serotonin release. Because interactions between 5-HT7 receptors and glutamatergic and GABAergic neurons have been demonstrated previously, the current studies tested the hypothesis that GABAergic and/or glutamatergic neurons mediate phase shifts induced by activation of DRN 5-HT7 receptors. Hamsters were fitted with guide cannulae aimed at the DRN, housed in cages with running wheels, and exposed to 14 h light (L):10 h dark (D). In Experiment 1, hamsters received DRN pretreatment with muscimol (87.6 picomoles) or vehicle before DRN 8-OH-DPAT (6 picomoles) microinjections at ZT6. After exposure to constant darkness (10 days), phase shifts were calculated and animals were re-exposed to 14L:10D. The procedure was repeated to give each animal the alternate pretreatment. In Experiment 2, hamsters received DRN pretreatment with NMDA (20 picomoles) or vehicle before 8-OH-DPAT at ZT 6. Other experiments tested the effects of single DRN microinjections of muscimol, bicuculline (136 picomoles), NMDA, MK-801 (10 picomoles) or vehicle. Phase shifts (mean±S.E.M., h) in muscimol/8-OH-DPAT-microinjected hamsters (1.02±0.30) were not different (P=0.11) from those in vehicle/8-OH-DPAT-microinjected hamsters (1.34±0.30), while those in NMDA/8-OH-DPAT-microinjected hamsters (0.67±0.17) were smaller (P

Potential mechanisms of Passiflora incarnata extracts and the effect of extraction methods on ingredients and biological effects were explored. Using the same batch of plant material, total flavonoid yields as measured by high-performance liquid chromatography coupled to diode array detection (HPLC-DAD) increased substantially with hot versus cold extraction methods. Whole Passiflora extract induced prominent, dose-dependent direct GABA(A) currents in hippocampal slices, but the expected modulation of synaptic GABA(A) currents was not seen. GABA was found to be a prominent ingredient of Passiflora extract, and GABA currents were absent when amino acids were removed from the extract. Five different extracts, prepared from a single batch of Passiflora incarnata, were administered to CF-1 mice for 1 week in their drinking water prior to evaluation of their behavioral effects. Anticonvulsant effects against PTZ-induced seizures were seen in mice that received 2 of the 5 Passiflora extracts. Instead of the anxiolytic effects described by others, anxiogenic effects in the elevated plus maze were seen in mice receiving any of the 5 Passiflora extracts.

Articles on pharmacological interventions in disorders of consciousness after brain damage are increasingly appearing in the medical literature. This hypothesis links disorders of consciousness to the depletion of oxygen reliant neurotransmitters based in two biochemical axes, the amino acid axis (glutamate/GABA) and the monoamine axis (dopamine/noradrenalin and serotonin). After a brain injury, an immediate response inside the brain constitutes a surge of amino acids such as glutamate, GABA and others. Glutamate is excitatory and GABA inhibitory. The inhibitory response dominates and the brain becomes suppressed, leading to a loss in consciousness which reduces oxygen requirements. In time, GABA depletes after increased usage and leakage from the brain into the blood. If it cannot be restored sufficiently in some parts of the brain, a secondary response in these regions occurs which makes GABA receptors oversensitive to GABA, so that decreased GABA levels can maintain their suppressive effect. This occurs in prolonged disorders of consciousness as in the Vegetative State, which can be broken by agents such as zolpidem in some patients. In addition to amino acids, monoamine systems such as dopamine are essential to cognition and motor function. Their depletion or the suppression of brain regions in which they function, are proposed as contributors to disorders of consciousness. Reports of successful arousals after amino acid and monoamine based therapies supports the hypothesis that depletion of these brain chemicals may play a fundamental role in disorders of consciousness.