GABAB Receptor Antagonist Research Articles

GABAB receptor ligands do not modify conditioned fear responses in BALB/c mice

The GABAB receptor has been well characterised as a substrate of unconditioned anxiety behaviour. Indeed, the anxiolytic effects of positive modulators of the GABAB receptor have been demonstrated across a range of behavioural tests of innate anxiety, whereas GABAB receptor deficient mice have an elevated anxiety phenotype. However, the role of the GABAB receptor in regulating conditioned anxiety behaviour; an important facet of the preclinical study of anxiety disorders such as post-traumatic stress disorder is less well understood. In vitro data suggests that the GABAB receptor plays an important role in regulating the neural circuitry that underpins conditioned fear learning and extinction, but whether these effects translate into alterations in conditioned anxiety behaviour has not been widely investigated. This represents a crucial deficit in the preclinical characterisation of these drugs as putative anxiolytic agents. Using the highly anxious mouse strain, BALB/c, and an auditory fear conditioning protocol, we sought to characterise the GABAB receptor positive modulator GS39783 and GABAB receptor antagonist CGP52432, two compounds not previously evaluated for their effects on conditioned fear. Neither GS39783 nor CGP52432 altered freezing behaviour irrespective of whether drugs were administered before the acquisition, recall or extinction training sessions. These findings suggest limitations to the potential role of GABAB receptor active drugs as clinical agents in the treatment of anxiety.

Co-application of the GABAB receptor agonist, baclofen, and the mGlu receptor agonist, L-CCG-I, facilitates [3H]GABA release from rat cortical nerve endings

Interaction between different transmitter receptor systems is an emerging feature of neurotransmission at central synapses. G protein-coupled receptors' ability to form dimers or larger hetero-oligomers probably serves to facilitate the integration of diverse signals within the cell. We found that, in nerve terminals isolated from the cerebral cortices of rats, co-application of the GABAB agonist, baclofen, and of the non-selective mGlu agonist, L-CCG-I, potentiates the basal and depolarization-evoked release of [(3)H]GABA via a mechanism that involves mobilization of intracellular Ca(2+) ions. The effect of L-CCG-I+baclofen was abolished by the phospholipase C inhibitor U73122, reduced by Xestospongin C (an IP3 receptor blocker), and blocked by 2-APB, an IP3 receptor antagonist. Pretreatment of the synaptosomes with the lipid-soluble Ca(2+) chelator BAPTA-AM also inhibited the effects of L-CCG-I+baclofen. Subtype-selective non-competitive group I mGlu receptor antagonists, MPEP and CPCCOEt, had no effect on the release enhancement produced by baclofen+L-CCG-I. The enhancement was reversed by the GABAB receptor antagonist, CGP54626, and by the group I/group II mGlu receptor antagonist (R,S)-MCPG. The GABA release-enhancing effects of L-CCG-I+baclofen in our model might reflect the presence on cortical nerve endings of GABAB/group I mGlu receptor heteromers with pharmacological properties distinct from those of the component receptors. Activation of these heteromeric receptors might modify the function of the GABAB receptor in such a way that it facilitates GABAergic transmission, an effect that might be useful under conditions of excessive glutamatergic activity.

Cortical Inhibition Reduces Information Redundancy at Presentation of Communication Sounds in the Primary Auditory Cortex

In all sensory modalities, intracortical inhibition shapes the functional properties of cortical neurons but also influences the responses to natural stimuli. Studies performed in various species have revealed that auditory cortex neurons respond to conspecific vocalizations by temporal spike patterns displaying a high trial-to-trial reliability, which might result from precise timing between excitation and inhibition. Studying the guinea pig auditory cortex, we show that partial blockage of GABAA receptors by gabazine (GBZ) application (10 μm, a concentration that promotes expansion of cortical receptive fields) increased the evoked firing rate and the spike-timing reliability during presentation of communication sounds (conspecific and heterospecific vocalizations), whereas GABAB receptor antagonists [10 μm saclofen; 10-50 μm CGP55845 (p-3-aminopropyl-p-diethoxymethyl phosphoric acid)] had nonsignificant effects. Computing mutual information (MI) from the responses to vocalizations using either the evoked firing rate or the temporal spike patterns revealed that GBZ application increased the MI derived from the activity of single cortical site but did not change the MI derived from population activity. In addition, quantification of information redundancy showed that GBZ significantly increased redundancy at the population level. This result suggests that a potential role of intracortical inhibition is to reduce information redundancy during the processing of natural stimuli.

GET73 increases rat extracellular hippocampal CA1 GABA levels through a possible involvement of local mGlu5 receptor

N-[(4-trifluoromethyl) benzyl] 4-methoxybutyramide (GET73) is a newly synthesized compound displaying anti-alcohol and anxiolytic properties. In light of the importance of the hippocampal CA1 subregion in alcohol addiction and anxiety-like behaviors-this in vivo microdialysis study characterized the effect of GET73 on extracellular GABA levels in the hippocampal CA1 region of the freely moving rat-including a possible role for mGlu5 receptor in mediating this effect. Both intraperitoneal administration (2-10 mg/kg) and local intra-hippocampal CA1 perfusion with GET73 (50-1000 nM) were associated with a transient, step-wise increase in dialysate hippocampal CA1 GABA levels. The GET73 (10 mg/kg)-induced increase in GABA levels was not affected by intra-CA1 perfusion with either the GABA reuptake inhibitor SKF89976A (0.5 mM) or by local GABAA (bicuculline; 1μM) and GABAB (CGP35348; 500 μM) receptor antagonists. On the contrary, the GET73-induced increase in GABA levels was partially counteracted by the intra-CA1 perfusion with the mGlu5 receptor negative allosteric modulator MPEP (300 µM). Interestingly, GET73 at the lowest (2 mg/kg) dose tested, by itself ineffective, fully counteracted the increase in GABA levels induced by the mGlu5 receptor agonist CHPG (1000 µM). Taken together, these findings suggest that the GET73-induced increase in hippocampal CA1 GABA levels operates independently of local GABA reuptake and/or GABAA or GABAB receptors. Furthermore, the present data lead to hypothesize a possible interaction between GET73 and mGluR5-mediated regulation of hippocampal CA1 GABA transmission, an effect which may be relevant to the ability of GET73 to reduce alcohol intake in an alcohol-preferring rat strain.

Acupuncture suppresses reinstatement of morphine-seeking behavior induced by a complex cue in rats

Morphine causes physical and psychological dependence for individuals after repeated-use. Above all, our previous study showed that acupuncture attenuated reinstatement of morphine-seeking behavior induced by pharmacological cue. In this study, we investigated whether acupuncture could suppress the reinstatement of morphine-seeking behavior induced by the combination of environmental and pharmacological cues and the possible neuronal involvement. Male Sprague-Dawley rats were trained to self-administer morphine (1.0mg/kg) for 3 weeks. Following the withdrawal phase (7 days), the effects of acupuncture on reinstatement of morphine-seeking behavior were investigated. For the investigation of neuronal involvement, the GABAA receptor antagonist bicuculline and the GABAB receptor antagonist SCH 50911 were pre-treated. Morphine-seeking behavior induced by combination of re-exposure to the operant chamber and morphine injection was suppressed perfectly by acupuncture at SI5, but not at the control acupoint LI5 and this effect was blocked by pre-treatment with the GABA receptor antagonists. This study suggests that acupuncture at SI5 can be considered as a predominant therapy for the reinstatement of morphine-seeking behavior in humans.

Baclofen into the lateral parabrachial nucleus induces hypertonic sodium chloride intake during cell dehydration

BackgroundActivation of GABAB receptors with baclofen into the lateral parabrachial nucleus (LPBN) induces ingestion of water and 0.3 M NaCl in fluid replete rats. However, up to now, no study has investigated the effects of baclofen injected alone or combined with GABAB receptor antagonist into the LPBN on water and 0.3 M NaCl intake in rats with increased plasma osmolarity (rats treated with an intragastric load of 2 M NaCl). Male Wistar rats with stainless steel cannulas implanted bilaterally into the LPBN were used.ResultsIn fluid replete rats, baclofen (0.5 nmol/0.2 μl), bilaterally injected into the LPBN, induced ingestion of 0.3 M NaCl (14.3 ± 4.1 vs. saline: 0.2 ± 0.2 ml/210 min) and water (7.1 ± 2.9 vs. saline: 0.6 ± 0.5 ml/210 min). In cell-dehydrated rats, bilateral injections of baclofen (0.5 and 1.0 nmol/0.2 μl) into the LPBN induced an increase of 0.3 M NaCl intake (15.6 ± 5.7 and 21.5 ± 3.5 ml/210 min, respectively, vs. saline: 1.7 ± 0.8 ml/210 min) and an early inhibition of water intake (3.5 ± 1.4 and 6.7 ± 2.1 ml/150 min, respectively, vs. saline: 9.2 ± 1.4 ml/150 min). The pretreatment of the LPBN with 2-hydroxysaclofen (GABAB antagonist, 5 nmol/0.2 μl) potentiated the effect of baclofen on 0.3 M NaCl intake in the first 90 min of test and did not modify the inhibition of water intake induced by baclofen in cell-dehydrated rats. Baclofen injected into the LPBN did not affect blood pressure and heart rate.ConclusionsThus, injection of baclofen into the LPBN in cell-dehydrated rats induced ingestion of 0.3 M NaCl and inhibition of water intake, suggesting that even in a hyperosmotic situation, the blockade of LPBN inhibitory mechanisms with baclofen is enough to drive rats to drink hypertonic NaCl, an effect independent of changes in blood pressure.

Effect of GABA Receptor Agonists or Antagonists Injected Spinally on the Blood Glucose Level in Mice

The possible roles of gamma-amino butyric acid (GABA) receptors located in the spinal cord for the regulation of the blood glucose level were studied in ICR mice. We found in the present study that intrathecal (i.t.) injection with baclofen (a GABAB receptor agonist; 1-10 μg/5 μl) or bicuculline (a GABAA receptor antagonist; 1-10 μg/5 μl) caused an elevation of the blood glucose level in a dose-dependent manner. The hyperglycemic effect induced by baclofen was more pronounced than that induced by bicuculline. However, muscimol (a GABAA receptor agonist; 1-5 μg/5 μl) or phaclofen (a GABAB receptor antagonist; 5-10 μg/5 μl) administered i.t. did not affect the blood glucose level. Baclofen-induced elevation of the blood glucose was dose-dependently attenuated by phaclofen. Furthermore, i.t. pretreatment with pertussis toxin (PTX; 0.05 or 0.1 μg/5 μl) for 6 days dose-dependently reduced the hyperglycemic effect induced by baclofen. Our results suggest that GABAB receptors located in the spinal cord play important roles for the elevation of the blood glucose level. Spinally located PTX-sensitive G-proteins appear to be involved in hyperglycemic effect induced by baclofen. Furthermore, inactivation of GABAA receptors located in the spinal cord appears to be responsible for tonic up-regulation of the blood glucose level.

GABAB receptor‐mediated frequency‐dependent and circadian changes in synaptic plasticity modulate retinal input to the suprachiasmatic nucleus

Light is the most important environmental signal that entrains the circadian clock located in the hypothalamic suprachiasmatic nucleus (SCN). The retinohypothalamic tract (RHT) was stimulated to simulate the light intensity-dependent discharges of intrinsically photosensitive retinal ganglion cells projecting axons to the hypothalamus. EPSCs were evoked by paired-pulse stimulation or by application of stimulus trains, and recorded from SCN neurons in rat brain slices. Initial release probability (Pr) and synaptic plasticity changes depended on the strength of GABAB receptor (GABABR)-mediated presynaptic inhibition and could be different at the same GABABR agonist concentration. Facilitation caused by frequency-dependent relief of GABABR-mediated inhibition was observed when the initial Pr was decreased to less than 15% of control during strong activation of presynaptic GABAB receptors by (±)baclofen (10 μm), GABA (2 mm) or by GABA uptake inhibitor nipecotic acid (5 mm). In contrast, short-term synaptic depression appeared during baclofen (10 μm) application when initial Pr was greater than 30% of control. Block of 4-aminopyridine-sensitive K(+) currents increased the amplitude and time constant of decay of evoked EPSCs (eEPSCs), and decreased the GABABR-mediated presynaptic inhibition. The GABAB receptor antagonist CGP55845 (3 μm) increased the eEPSCs amplitude 30% throughout the light-dark cycle. During light and dark phases the RHT inputs to 55% and 33% of recorded neurons, respectively, were under GABAB inhibitory control indicating that the tonic inhibition induced by local changes of endogenous GABA concentration contributes to the circadian variation of RHT transmitter release. We conclude that GABABR-mediated presynaptic inhibition decreased with increasing frequency and broadening of presynaptic action potentials, and depended on the sensitivity of RHT terminals to GABABR agonists, and diurnal changes of the extracellular GABA concentration around RHT axon terminals in the SCN.

Activation of GABAA receptors in the medial prefrontal cortex produces an anxiolytic-like response.

There has been increasing evidence that the γ-aminobutyric acid (GABA)ergic system is involved in the neurobiology of anxiety. The present study aimed to investigate the role of GABAergic systems in the modulation of anxiety in the medial prefrontal cortex (mPFC) of rats using the elevated plus maze test. Rats were anaesthetised with a mixture of ketamine and xylazine, and then special cannulae were inserted stereotaxically into the mPFC. After 5-7 days of recovery, the effects of intra-mPFC administration of GABAergic agents were studied. Bilateral injection of the GABAA receptor agonist muscimol (0.25, 0.5 and 1 μg/rat) produces an anxiolytic-like effect, shown by significant increases in the percentage of open-arm time (%OAT) and percentage of open-arm entries (%OAE). Intra-mPFC administration of the GABAA receptor antagonist bicuculline (0.25, 0.5 and 1 μg/rat) produces significant anxiogenic-like behaviour. However, intra-mPFC injection of the GABAB receptor agonist baclofen (0.05, 0.1 and 0.2 μg/rat) and the GABAB receptor antagonist CGP35348 (5, 10 and 15 μg/rat) did not alter %OAT and %OAE significantly. The results of the present study demonstrate that the GABAergic system of the mPFC modulates anxiety-related behaviours of rats through GABAA receptors.

The Antinociceptive Effect of SNAP5114, a Gamma-Aminobutyric Acid Transporter-3 Inhibitor, in Rat Experimental Pain Models

Gamma-aminobutyric acid (GABA) is the primary inhibitory neurotransmitter in the mammalian central nervous system. GABAergic transmission has an important role in regulating nociception at the spinal dorsal horn. It is terminated by rapid uptake of the neurotransmitter from the synaptic cleft into neurons and glial cells, via specific GABA transporters (GATs). Among the 4 GATs, GAT-3 has the greatest expression in central nervous system regions closely associated with nociceptive transmission, including the spinal cord. In this study, we examined the antinociceptive effect of intrathecal administration of a selective GAT-3 inhibitor, SNAP5114, on acute, inflammatory, and neuropathic pain in experimental models. Male Sprague-Dawley rats were used to assess thermal, mechanical, and chemical nociception in the tail flick and hotplate tests, the paw pressure test, and the formalin test. A rotarod test was performed to assess motor function. Chronic constriction injury to the sciatic nerve was induced in the rats. The electronic von Frey test and the plantar test were then performed to assess mechanical allodynia and thermal hyperalgesia. SNAP5114 (10, 50, 100, or 200 μg) was administered intrathecally to examine antinociceptive activity. To confirm whether the action of SNAP5114 was mediated by GABAergic transmission, the GABAA receptor antagonist bicuculline (0.3 μg) or the GABAB receptor antagonist CGP35348 (30 μg) was administered intrathecally before 200 μg of SNAP5114 in the tail flick test, the formalin test, and the electronic von Frey test. Spinally applied SNAP5114 in normal rats dose-dependently prolonged withdrawal latencies in the tail flick test and suppressed the late-phase response in the formalin test. SNAP5114 did not affect motor performance. In the chronic constriction injury rats, SNAP5114 inhibited mechanical allodynia dose-dependently. The antinociceptive action of SNAP5114 was partially reversed by bicuculline or CGP35348 at doses at which the antagonist alone did not affect baseline behavioral responses. These results suggest that SNAP5114 exerts antinociceptive effects by activating GABAA and GABAB receptors in the spinal cord. The GAT-3 inhibitor may prove useful in treatment of various painful conditions.

Postsynaptic GABABReceptors Enhance Extrasynaptic GABAAReceptor Function in Dentate Gyrus Granule Cells

Ambient GABA in the brain tonically activates extrasynaptic GABA(A) receptors, and activity-dependent changes in ambient GABA concentration can also activate GABA(B) receptors. To investigate an interaction between postsynaptic GABA(B) and GABA(A) receptors, we recorded GABA(A) currents elicited by exogenous GABA (10 μm) from dentate gyrus granule cells (DGGCs) in adult rat hippocampal slices. The GABA(B) receptor agonist baclofen (20 μm) enhanced GABA(A) currents. This enhancement was blocked by the GABA(B) receptor antagonist CGP 55845 and intracellular solutions containing the GTP analog GDP-β-s, indicating that baclofen was acting on postsynaptic GABA(B) receptors. Modulation of GABA(A) currents by postsynaptic GABA(B) receptors was not observed in CA1 pyramidal cells or layer 2/3 cortical pyramidal neurons. Baclofen reduced the frequency of spontaneous inhibitory postsynaptic currents (sIPSCs) but did not alter sIPSC amplitude or kinetics. Thus, GABA(A) receptors activated at synapses were not modulated by postsynaptic GABA(B) receptors. In contrast, tonic GABA currents and currents activated by the GABA(A) receptor δ subunit-selective agonist THIP (10 μm) were potentiated by baclofen. Our data indicate that postsynaptic GABA(B) receptors enhance the function of extrasynaptic GABA(A) receptors, including δ subunit-containing receptors that mediate tonic inhibition in DGGCs. The modulation of GABA(A) receptor function by postsynaptic GABA(B) receptors is a newly identified mechanism that will influence the inhibitory tone of DGGCs when GABA(B) and GABA(A) receptors are both activated.

GABAB receptors as a common target for hypothermia and spike and wave seizures: Intersecting mechanisms of thermoregulation and absence epilepsy

In the current study the link among the γ-hydroxybutyrate (GHB)/pentylenetetrazole (PTZ)-induced absence-like seizures and concomitant decreases in the core temperature, as well as electroencephalographic (EEG) activity during rewarming from deep hypothermia produced by a drug-free protocol were investigated. During the rewarming period after deep cooling, most Wistar rats suffered from bilaterally synchronous spike and waves with no or mild behavioral correlates. Spike and wave seizures were temperature-dependent and were initially registered when body temperature (Tb) reached 25–27°C, but mostly during the mild hypothermia of 0.3–1.3°C (Tb of 36.3–37.3°C). In chemical absence models, spike and wave discharges were also closely accompanied by mild systemic hypothermia, as both PTZ- and GHB-induced temperature decreases ranged from about 1–1.4°C respectively, together with EEG markers of absence activity. Thus, throughout the different experimental designs, the occurrence of spike and wave discharges was always related to a mild (0.3–1.4°C) decrease of Tb. Benzodiazepine diazepam as the GABAA-positive allosteric modulator and CGP 62349 as the selective antagonist of GABAB receptors were used to determine if their well-known anticonvulsant properties also affect hypothermia elicited by these drugs. Finally, during the course of spontaneous rewarming from deep hypothermia, another selective GABAB-blocking agent, CGP 35348, was used to elucidate if GABAB inhibitory system could be critically implicated in the generation of hypothermia-dependent spike and waves. Diazepam prevented both the PTZ-induced hypothermia and electrographic absence seizures, but these two beneficial effects did not occur in the GHB model. Even though diazepam delayed GHB-induced maximal temperature decrease, the GHB effects remained highly significant. The GABAB antagonist CGP 62349 completely prevented hypothermia as well as absence seizures in both chemical models. Likewise, spike and wave discharges, registered during the spontaneous rewarming from deep hypothermia, were completely prevented by CGP 35348. These findings show that systemic hypothermia should definitely be regarded as a marker of GABAB receptor activation. Moreover, the results of this study clearly show that initial mild temperature decrease should be considered as strong absence-provoking factor. Hypothermia-induced nonconvulsive seizures also highlight the importance of continuous EEG monitoring in children undergoing therapeutic hypothermia after cardiac arrest. Since every change in peripheral or systemic temperature ultimately must be perceived by preoptic region of the anterior hypothalamus as the primary thermoregulatory and sleep-inducing center, the preoptic thermosensitive neurons in general and warm-sensitive neurons in particular, simply have to be regarded as the most probable candidate for connected thermoregulatory and absence generating mechanisms. Therefore, additional studies are needed to confirm their potential role in the generation and propagation of absence seizures.

GABAergic mechanisms for shaping transient visual responses in the mouse superior colliculus

An object that suddenly appears in the visual field should be quickly detected and responded to because it could be beneficial or harmful. The superficial layer of the superior colliculus (sSC) is a brain structure capable of such functions, as sSC neurons exhibit sharp transient spike discharges with short latency in response to the appearance of a visual stimulus. However, how transient activity is generated in the sSC is poorly understood. Here, we show that inhibitory inputs actively shape transient activity in the sSC. Juxtacellular recordings from anesthetized mice demonstrate that almost all types of sSC neurons, which were identified by post hoc histochemistry, show transient spike discharges, i.e., ON activity, immediately after visual stimulus onset. ON activity was followed by a pause before the visual stimulus was turned off. To determine whether the pause reflected the absence of excitatory drive or inhibitory conductance, we injected depolarizing currents juxtasomally, which enabled us to observe inhibition as decreased discharges. The pause was observed even under this condition, suggesting that inhibitory input caused the pause. We further found that local application of a mixture of GABAA and GABAB receptor antagonists additively diminished the pause. These results indicate that GABAergic inputs produce transient ON responses by attenuating excitatory activity through the cooperative activation of GABAA and GABAB receptors, allowing sSC neurons to act as a saliency detector.

Endocannabinoid/GABA interactions in the entopeduncular nucleus modulates alcohol intake in rats

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.

A Fluorescent Sensor for GABA and Synthetic GABAB Receptor Ligands

While γ-aminobutyric acid (GABA) is the main inhibitory neurotransmitter, suitable tools to measure its concentration in living cells with high spatiotemporal resolution are missing. Herein, we describe the first ratiometric fluorescent sensor for GABA, dubbed GABA-Snifit, which senses GABA with high specificity and spatiotemporal resolution on the surface of living mammalian cells. GABA-Snifit is a semisynthetic fusion protein containing the GABA(B) receptor, SNAP- and CLIP-tag, a synthetic fluorophore and a fluorescent GABA(B) receptor antagonist. When assembled on cell surfaces, GABA-Snifit displays a GABA-dependent fluorescence emission spectrum in the range of 500-700 nm that permits sensing micromolar to millimolar GABA concentrations. The ratiometric change of the sensor on living cells is 1.8. Furthermore, GABA-Snifit can be utilized to quantify the relative binding affinities of GABA(B) receptor agonists, antagonists and the effect of allosteric modulators. These properties make GABA-Snifit a valuable tool to investigate the role of GABA and GABA(B) in biological systems.

Neurogliaform cells of amygdala: a source of slow phasic inhibition in the basolateral complex

Synaptic inhibition in the amygdala actively participates in processing emotional information. To improve the understanding of interneurons in amygdala networks it is necessary to characterize the GABAergic cell types, their connectivity and physiological roles. We used a mouse line expressing a green fluorescent protein (GFP) under the neuropeptide Y (NPY) promoter. Paired recordings between presynaptic NPY-GFP-expressing (+) cells and postsynaptic principal neurons (PNs) of the basolateral amygdala (BLA) were performed. The NPY-GFP+ neurons displayed small somata and short dendrites embedded in a cloud of highly arborized axon, suggesting a neurogliaform cell (NGFC) type. We discovered that a NPY-GFP+ cell evoked a GABA(A) receptor-mediated slow inhibitory postsynaptic current (IPSC) in a PN and an autaptic IPSC. The slow kinetics of these IPSCs was likely caused by the low concentration and spillover of extracellular GABA. We also report that NGFCs of the BLA fired action potentials phase-locked to hippocampal theta oscillations in anaesthetized rats. When this firing was re-played in NPY+-NGFCs in vitro, it evoked a transient depression of the IPSCs. Presynaptic GABA(B) receptors and functional depletion of synaptic vesicles determined this short-term plasticity. Synaptic contacts made by recorded NGFCs showed close appositions, and rarely identifiable classical synaptic structures. Thus, we report here a novel interneuron type of the amygdala that generates volume transmission of GABA. The peculiar functional mode of NGFCs makes them unique amongst all GABAergic cell types of the amygdala identified so far.

Participation of GABAA, GABAB receptors and neurosteroids in toluene-induced hypothermia: Evidence of concentration-dependent differences in the mechanism of action

Toluene is a misused substance that modifies γ-aminobutyric acid (GABA) release and shares behavioral and molecular effects with GABAA and GABAB receptor agonists. GABAergic compounds are involved in thermoregulation processes and volatile substance users have reported that one of the reasons to inhale is to avoid feeling cold. At present, no studies have analyzed the effects of inhalants on body temperature and the mechanism of action involved. Thus, the main purpose of this study was to evaluate the effects of a (60min) acute toluene inhalation (2000, 4000 and 6000ppm) in core temperature. In addition, we tried to prevent the changes of temperature induced by toluene with the specific GABAA receptor blockers picrotoxin (0.01–0.1mg/kg), bicuculline (0.1–0.3mg/kg), and flumazenil (3–30mg/kg); the GABAB receptor antagonist phaclofen (10–30mg/kg) and the neurosteroid synthesis inhibitor finasteride (10–30mg/kg). Results show that toluene reduced core temperature in mice in a concentration-dependent manner. The hypothermia produced by 4000ppm toluene was prevented by picrotoxin, bicuculline, phaclofen and finasteride but not by flumazenil. In contrast none of these antagonists tested blocked the effects of 6000ppm toluene. In conclusion, toluene decreases core temperature, GABA receptors and neurosteroids participate in toluene’s action at 4000ppm; but other mechanisms of action are involved in the hypothermic effects of 6000ppm toluene.

Gamma-Aminobutyric Acid Binds to GABAB Receptor to Inhibit Cholangiocarcinoma Cells Growth via the JAK/STAT3 Pathway

Gamma-aminobutyric acid (GABA) has been reported to inhibit the growth of cholangiocarcinoma QBC939 cells, but the mechanisms are still not fully understood. To explore the mechanisms of the anti-cancer effect of GABA on QBC939 cells. An initial immunohistochemistry study of the expressions of GABA receptors in cholangiocarcinoma tissues was followed by the culture and treatment of QBC939 cells for 48 h with GABA, GABA + bicuculine (GABAA receptor antagonist), GABA + phaclofen (GABAB receptor antagonist), and GABA + AG490 (Janus Kinase inhibitor). MTT and Annexin V-FITC/PI binding assays were used to determine the proliferation and apoptosis of the QBC939 cells. The expression of the signal transducer and activator of transcription 3 (STAT3) and phosphorylated STAT3 (Tyr705) [p-STAT3 (Tyr705)] was evaluated by the western blot assay. The effect of GABA on the growth of QBC939 xenograft tumors in athymic nu/nu mice was examined, and p-STAT3 (Tyr705) expression in xenograft tumors was detected by immunohistochemistry. A significant difference was only observed in GABAB receptor expression between cholangiocarcinoma and normal bile tissues. The MTT and Annexin V-FITC/PI assays showed that the antiproliferative and proapoptotic effects of GABA on QBC939 cells could be antagonized by phaclofen and AG490, but not bicuculine. GABA significantly down-regulated p-STAT3 (Tyr705) expression; this action was also antagonized by phaclofen and AG490. GABA also effectively inhibited xenograft tumor growth, and p-STAT3 (Tyr705) expression was significantly decreased in GABA-treated xenograft tumors. GABA may inhibit the growth of cholangiocarcinoma QBC939 cells through the GABAB receptor, and the anti-cancer effects may be partly mediated via the JAK/STAT3 pathway.

Effect of saikosaponin A on maintenance of intravenous morphine self-administration

In this study, we investigated the effects of saikosaponin A (SSA), a major compound of Bupleurum falcatum L., on morphine self-administration behavior. Male Sprague–Dawley rats were trained to self-administer intravenous morphine (0.1mg/kg per injection over 5s) during daily 1-h sessions under a fixed-ratio 1 schedule. Rats were pretreated with SSA (0.25, 0.5, 1.0mg/kg) by intraperitoneal injection 30min prior to the start of the test session. Results demonstrated that pretreatment with SSA reduced morphine-maintained responding dose-dependently. Additionally, SSA inhibition of morphine-reinforced behavior was blocked by the selective GABAB receptor antagonist (2S)(+)-5,5-dimethyl-2-morpholineacetic acid (SCH 50911), but not the selective GABAA receptor antagonist bicuculline. Together, these results suggest that SSA may effectively suppress morphine-reinforced behavior by activating GABAB receptors.

Blockade of the GABAB receptor increases neurogenesis in the ventral but not dorsal adult hippocampus: Relevance to antidepressant action

GABAB receptor antagonists have been shown to have antidepressant-like properties in animal models and thus, could represent a novel approach for the treatment of depression. The neurobiological mechanisms underlying these effects are currently unknown. Adult hippocampal neurogenesis (the birth of new neurons) is thought to play a role in antidepressant drug action. However, the ability of GABAB receptors to modulate the proliferation and survival of newly-born cells in the adult hippocampus remains unexplored. Therefore, we investigated whether the GABAB receptor antagonist, CGP 52432, can induce antidepressant-like behaviour and increase hippocampal neurogenesis in the stress-sensitive mouse strain, BALB/c. Male mice were treated with CGP 52432 either acutely (one injection, 3; 10; 30 mg/kg, i.p.), subchronically (7 days, 3; 10 mg/kg, i.p.) or chronically (21 days, 3; 10 mg/kg, i.p.) and antidepressant-like behaviour was assessed using the forced swim test (FST). The effects of CGP 52432 on the proliferation and survival of newly-born cells in the hippocampus were assessed using BrdU immunohistochemistry. Acute, subchronic and chronic treatment with CGP 52432 induced antidepressant-like behavioural effects in the FST. Moreover, chronic but not acute or subchronic treatment with CGP 52432 increased hippocampal cell proliferation but had no effect on the survival of newly-born cells. This temporal effect is consistent with the time course for the therapeutic action of antidepressants. Interestingly, CGP 52432-induced increases in cell proliferation occurred in the ventral but not in the dorsal hippocampus. This topographical segregation concurs with the hypothesis that the ventral hippocampus is primarily involved in the regulation of stress and emotionality. Taken together, our data suggest that increased hippocampal cell proliferation is a plausible mechanism for the antidepressant-like effects of GABAB receptor antagonists following chronic but not acute treatments. Moreover, altered behavioural effects in the FST does not correlate with changes in neurogenesis.