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Noradrenergic activation of the basolateral amygdala (BLA) by emotional arousal enhances different forms of recognition memory via functional interactions with the insular cortex (IC). Human neuroimaging studies have revealed that the anterior IC (aIC), as part of the salience network, is dynamically regulated during arousing situations. Emotional stimulation first rapidly increases aIC activity but suppresses it in a delayed fashion. Here, we investigated in male Sprague-Dawley rats whether the BLA influence on recognition memory is associated with an increase or suppression of aIC activity during the postlearning consolidation period. We first employed anterograde and retrograde viral tracing and found that the BLA sends dense monosynaptic projections to the aIC. Memory-enhancing norepinephrine administration into the BLA following an object training experience suppressed aIC activity 1 h later, as determined by a reduced expression of the phosphorylated form of the transcription factor cAMP response element-binding (pCREB) protein and neuronal activity marker c-Fos. In contrast, the number of perisomatic γ-aminobutyric acid (GABA)ergic inhibitory synapses per pCREB-positive neuron was significantly increased, suggesting a dynamic up-regulation of GABAergic tone. In support of this possibility, pharmacological inhibition of aIC activity with a GABAergic agonist during consolidation enhanced object recognition memory. Norepinephrine administration into the BLA did not affect neuronal activity within the posterior IC, which receives sparse innervation from the BLA. The evidence that noradrenergic activation of the BLA enhances the consolidation of object recognition memory via a mechanism involving a suppression of aIC activity provides insight into the broader brain network dynamics underlying emotional regulation of memory.

Background: Stimulation of gamma-aminobutyric acid (GABA) activity through GABA receptor agonists is the basic mechanism of many anticonvulsant drugs. Nevertheless, many of these GABergic drugs have adverse cognitive effects. We previously found that GABAB receptors (GABABRs) in the insula regulate operant associative memory in healthy rats. The present study aimed at investigating the effects of GABABR modulation in the insula on operant associative memory in epileptic rats, along with the underlying mechanisms. Methods: The lithium-pilocarpine model of temporal lobe epilepsy (TLE) was established in male Sprague--Dawley rats. A 22-gauge stainless-steel guide cannula was surgically implanted into the granular insula cortex of the epileptic rats. Baclofen (125 ng/ml, 1 ml), CGP35348 (12.5 mg/ml, 1 ml), or saline (1 ml) was slowly infused through the guide cannula. The Intellicage automated behavioral testing system was used to evaluate operant associative memory of the epileptic rats, including non-spatial operant tasks (basic nosepoke learning and skilled nosepoke learning) and spatial operant tasks (chamber position learning). The expression of the GABABR subunits GB1 and GB2 in the insula was examined by immunofluorescence and Western blotting. Results: The Intellicage tests demonstrated that baclofen significantly impaired basic nosepoke learning, skilled nosepoke learning and chamber position learning of the epileptic rats, while CGP35348 boosted these functions. Immunofluorescence staining revealed that GB1 and GB2 were expressed in the insula of the epileptic rats, and Western blotting analysis showed that baclofen enhanced while CGP35348 inhibited the expression of these subunits. Conclusion: GABABRs in the insula bidirectionally regulate both spatial and non-spatial operant associative memory of epileptic rats. Effects of GABABRs on cognition should be taken into account when evaluating new possible treatments for people with epilepsy. [ABSTRACT FROM AUTHOR]

GABAA receptors are a major contributor to fast inhibitory neurotransmission in the brain. The receptors are activated upon binding the transmitter GABA or allosteric agonists including a number of GABAergic anesthetics and neurosteroids. Functional receptors can be formed by various combinations of the nineteen GABAA subunits cloned to date. GABAA receptors containing the ε subunit exhibit a significant degree of constitutive activity and have been suggested to be unresponsive to allosteric agents. In this study, we have characterized the functional properties of the rat α1β2ε GABAA receptor. We confirm that the α1β2ε receptor exhibits a higher level of constitutive activity than typical of GABAA receptors and show that it is inefficaciously activated by the transmitter and the allosteric agonists propofol, pentobarbital, and allopregnanolone. Manipulations intended to alter ε subunit expression and receptor stoichiometry were largely without effect on receptor properties including sensitivity to GABA and allosteric agonists. Surprisingly, amino acid substitutions at the conserved 9' and 6' positions in the second transmembrane (TM2) domain in the ε subunit did not elicit the expected functional effects of increased constitutive activity and resistance to the channel blocker picrotoxin, respectively. We tested the accessibility of TM2 residues mutated to cysteine using the cysteine-modifying reagent 4-(hydroxymercuri)benzoic acid and found a unique pattern of water-accessible residues in the ε subunit. [ABSTRACT FROM AUTHOR]

During status epilepticus (SE), synaptic γ-aminobutyric acid A receptors (GABAA Rs) become internalized and inactive, whereas spare N-methyl-d-aspartate receptors (NMDARs) assemble, move to the membrane, and become synaptically active. When treatment of SE is delayed, the number of synaptic GABAA Rs is drastically reduced, and a GABAA agonist cannot fully restore inhibition. We used a combination of low-dose diazepam (to stimulate the remaining GABAA Rs), ketamine (to mitigate the effect of the NMDAR increase), and valproate (to enhance inhibition at a nonbenzodiazepine site) to treat seizures in a model of severe cholinergic SE. High doses of diazepam failed to stop electrographic SE, showing that benzodiazepine pharmacoresistance had developed. The diazepam-ketamine-valproate combination was far more effective in stopping SE than triple-dose monotherapy using the same individual drugs. Isobolograms showed that this drug combination's therapeutic actions were synergistic, with positive cooperativity between drugs, whereas drug toxicity was simply additive, without positive or negative cooperativity. As a result, the therapeutic index was improved by this drug combination compared to monotherapy. These results suggest that synergistic drug combinations that target receptor changes can control benzodiazepine-refractory SE.

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, the tissue of chronically epileptic rats was collected within 3h of seizure occurrence (≤3h group) or at least 24h after seizure occurrence (≥24h 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 ≤3h group. A biochemical analysis showed that although animals with more frequent/recent seizures (≤3h group) had similar levels of GABAAR at the plasma membrane they showed deficits in inhibitory neurotransmission. By contrast, the tissue obtained from animals experiencing infrequent seizures (≥24h 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.

Administration of pediatric anesthetics with N-methyl D-aspartate (NMDA)-receptor antagonist and/or γ-aminobutyric acid (GABA) agonist activities may result in neuronal degeneration and/or neuronal cell death in neonatal rats. Evaluating pediatric drug candidates for this potential neurotoxicity is often part of overall preclinical new drug development strategy. This specialized assessment may require dosing neonatal rats at postnatal day 7 at the peak of the brain growth spurt and evaluating brain tissue 24 to 48 hours following dosing. The need to identify methods to aid in the accurate and reproducible detection of lesions associated with this type of neurotoxic profile is paramount for meeting the changing needs of neuropathology assessment and addressing emerging challenges in the neuroscience field. We document the use of Fluoro-Jade B (FJB) staining, to be used in conjunction with standard hematoxylin and eosin staining, to detect acute neurodegeneration and neuronal cell death that can be caused by some NMDA-receptor antagonists and/or GABA agonists in the neonatal rat brain. The FJB staining is simple, specific, and sensitive and can be performed on brain specimens from the same cohort of animals utilized for standard neurotoxicity assessment, thus satisfying animal welfare recommendations with no effect on achievement of scientific and regulatory goals. [ABSTRACT FROM AUTHOR]

Use of the highly toxic and easily prepared rodenticide tetramethylenedisulfotetramine (TETS) was banned after thousands of accidental or intentional human poisonings, but it is of continued concern as a chemical threat agent. TETS is a noncompetitive blocker of the GABA type A receptor (GABAAR), but its molecular interaction has not been directly established for lack of a suitable radioligand to localize the binding site. We synthesized [(14)C]TETS (14 mCi/mmol, radiochemical purity >99%) by reacting sulfamide with H(14)CHO and s-trioxane then completion of the sequential cyclization with excess HCHO. The outstanding radiocarbon sensitivity of accelerator mass spectrometry (AMS) allowed the use of [(14)C]TETS in neuroreceptor binding studies with rat brain membranes in comparison with the standard GABAAR radioligand 4'-ethynyl-4-n-[(3)H]propylbicycloorthobenzoate ([(3)H]EBOB) (46 Ci/mmol), illustrating the use of AMS for characterizing the binding sites of high-affinity (14)C radioligands. Fourteen noncompetitive antagonists of widely diverse chemotypes assayed at 1 or 10 µM inhibited [(14)C]TETS and [(3)H]EBOB binding to a similar extent (r(2) = 0.71). Molecular dynamics simulations of these 14 toxicants in the pore region of the α1β2γ2 GABAAR predict unique and significant polar interactions for TETS with α1T1' and γ2S2', which are not observed for EBOB or the GABAergic insecticides. Several GABAAR modulators similarly inhibited [(14)C]TETS and [(3)H]EBOB binding, including midazolam, flurazepam, avermectin Ba1, baclofen, isoguvacine, and propofol, at 1 or 10 μM, providing an in vitro system for recognizing candidate antidotes.

For several decades, studies conducted to evaluate the efficacy of RS(±)-Baclofen in the treatment of alcohol dependence yielded contrasting results. Human and animal studies recently questioned the use of the racemic drug in patients since a potential important role of the different enantiomers has been revealed with an efficacy thought to reside with the active R(+)-enantiomer. Here we conducted experiments in the postdependent rat model of alcohol dependence to compare the efficacy of R(+)-Baclofen or S(-)-Baclofen to that of RS(±)-Baclofen on ethanol intake, seeking, and relapse. R(+)-Baclofen was more effective than RS(±)-Baclofen in reducing ethanol intake and seeking during acute withdrawal and during relapse after abstinence. We also used an original population approach in order to identify drug responders. We found a significant proportion of responders to S(-)-Baclofen and RS(±)-Baclofen, displaying an increase in ethanol intake, and this increasing effect on alcohol intake was not seen in the R(+)-Baclofen group. At an intermediate dose of R(+)-Baclofen, devoid of any motor side effects, we identified a very large proportion of responders (75%) with a large decrease in ethanol intake (90% decrease). Finally, the response to RS(±)-Baclofen on ethanol intake was correlated to plasma level of Baclofen. R(+)-Baclofen and RS(±)-Baclofen were effective in reducing sucrose intake. Our study has important clinical implication since it suggests that the wide variability in the therapeutic responses of patients to RS(±)-Baclofen may come from the sensitivity to the R(+)-Baclofen but also to the one of the S(-)-Baclofen that can promote an increase in ethanol intake. [ABSTRACT FROM AUTHOR]

Background: The most currently used general anaesthetics are potent potentiators of γ-aminobutyric acid A (GABAA) receptors and are invariably neurotoxic during the early stages of brain development in preclinical animal models. As causality between GABAA potentiation and anaesthetic-induced developmental neurotoxicity has not been established, the question remains whether GABAergic activity is crucial for promoting/enhancing neurotoxicity. Using the neurosteroid analogue, (3α,5α)-3-hydroxy-13,24-cyclo-18,21-dinorchol-22-en-24-ol (CDNC24), which potentiates recombinant GABAA receptors, we examined whether this potentiation is the driving force in inducing neurotoxicity during development.Methods: The neurotoxic potential of CDNC24 was examined vis-à-vis propofol (2,6-diisopropylphenol) and alphaxalone (5α-pregnan-3α-ol-11,20-dione) at the peak of rat synaptogenesis. In addition to the morphological neurotoxicity studies of the subiculum and medial prefrontal cortex (mPFC), we assessed the extra-, pre-, and postsynaptic effects of these agents on GABAergic neurotransmission in acute subicular slices from rat pups.Results: CDNC24, like alphaxalone and propofol, caused dose-dependent hypnosis in vivo, with a higher therapeutic index. CDNC24 and alphaxalone, unlike propofol, did not cause developmental neuroapoptosis in the subiculum and mPFC. Propofol potentiated post- and extrasynaptic GABAA currents as evidenced by increased spontaneous inhibitory postsynaptic current (sIPSC) decay time and prominent tonic currents, respectively. CDNC24 and alphaxalone had a similar postsynaptic effect, but also displayed a strong presynaptic effect as evidenced by decreased frequency of sIPSCs and induced moderate tonic currents.Conclusions: The lack of neurotoxicity of CDNC24 and alphaxalone may be at least partly related to suppression of presynaptic GABA release in the developing brain. [ABSTRACT FROM AUTHOR]

• We inactivated the basolateral amygdala (BLA) in nulliparous and primiparous rats. • BLA inactivation impaired acquisition of fear conditioning in both groups. • BLA inactivation impaired fear extinction retention in nulliparous rats. • BLA inactivation had no effect on fear extinction retention in primiparous rats. In female rats and humans, reproductive experience (i.e., pregnancy) alters the behavioral, hormonal and molecular substrates of fear extinction. Here, we assessed whether the role of a central neural substrate of fear extinction, the basolateral amygdala (BLA), also changes following reproductive experience. Nulliparous (virgin) and primiparous (one prior pregnancy) female rats received infusions of the GABA A agonist, muscimol, to temporarily inactivate the BLA prior to fear conditioning or extinction training. In follow up experiments, the BLA was also inactivated immediately after extinction training. BLA inactivation impaired the acquisition and expression of conditioned fear in both nulliparous and primiparous rats. In nulliparous rats, BLA inactivation prior to or immediately after extinction training impaired extinction retention. In contrast, in primiparous rats, BLA inactivation prior to or immediately after extinction training did not impair extinction retention, despite suppressing freezing during extinction training. These results suggest that, consistent with past findings in males, the BLA is a central component of the neural circuitry of fear acquisition and its extinction in virgin female rats. However, after pregnancy, female rats no longer depend on the BLA to extinguish fear, despite requiring the BLA to acquire conditioned fear. Given that fear extinction forms the basis of exposure therapy for anxiety disorders in humans, the present findings may have clinical implications. To improve the efficacy of exposure therapy for anxiety disorders, we may need to target different mechanisms in females dependent on their reproductive history. [ABSTRACT FROM AUTHOR]

Drugs of abuse induce widespread synaptic adaptations in the mesolimbic dopamine (DA) neurons. Such drug-induced neuroadaptations may constitute an initial cellular mechanism eventually leading to compulsive drug-seeking behavior. To evaluate the impact of GABAB receptors on addiction-related persistent neuroplasticity, we tested the ability of orthosteric agonist baclofen and two positive allosteric modulators (PAMs) of GABAB receptors to suppress neuroadaptations in the ventral tegmental area (VTA) and reward-related behaviors induced by ethanol and cocaine. A novel compound (S)-1-(5-fluoro-2,3-dihydro-1H-inden-2-yl)-4-methyl-6,7,8,9-tetrahydro-[1,2,4]triazolo[4,3-a]quinazolin-5(4H)-one (ORM-27669) was found to be a GABAB PAM of low efficacy as agonist, whereas the reference compound (R,S)-5,7-di-tert-butyl-3-hydroxy-3-trifluoromethyl-3H-benzofuran-2-one (rac-BHFF) had a different allosteric profile being a more potent PAM in the calcium-based assay and an agonist, coupled with potent PAM activity, in the [35 S] GTPγS binding assay in rat and human recombinant receptors. Using autoradiography, the high-efficacy rac-BHFF and the low-efficacy ORM-27669 potentiated the effects of baclofen on [35 S] GTPγS binding with identical brain regional distribution. Treatment of mice with baclofen, rac-BHFF, or ORM-27669 failed to induce glutamate receptor neuroplasticity in the VTA DA neurons. Pretreatment with rac-BHFF at non-sedative doses effectively reversed both ethanol- and cocaine-induced plasticity and attenuated cocaine i.v. self-administration and ethanol drinking. Pretreatment with ORM-27669 only reversed ethanol-induced neuroplasticity and attenuated ethanol drinking but had no effects on cocaine-induced neuroplasticity or self-administration. These findings encourage further investigation of GABAB receptor PAMs with different efficacies in addiction models to develop novel treatment strategies for drug addiction. [ABSTRACT FROM AUTHOR]

Diabetic neuropathy is an incapacitating complication in diabetic patients. The cellular and molecular mechanisms involved in this pathology are poorly understood. Previous studies have suggested that the loss of spinal GABAergic inhibition participate in painful diabetic neuropathy. However, the role of extrasynaptic α5 subunit-containing GABAA (α5GABAA) receptors in this process is not known. The purpose of this study was to investigate the role of α5GABAA receptors in diabetes-induced tactile allodynia, loss of rate-dependent depression (RDD) of the Hoffmann reflex (HR), and modulation of primary afferent excitability. Intraperitoneal administration of streptozotocin induced tactile allodynia. Intrathecal injection of α5GABAA receptor inverse agonist, L-655,708, produced tactile allodynia in naive rats, whereas it reduced allodynia in diabetic rats. In healthy rats, electrical stimulation of the tibial nerve at 5 Hz induced RDD of the HR, although intrathecal treatment with L-655,708 (15 nmol) abolished RDD of the HR. Streptozotocin induced the loss of RDD of the HR, while intrathecal L-655,708 (15 nmol) restored RDD of the HR. L-655,708 (15 nmol) increased tonic excitability of the primary afferents without affecting the phasic excitability produced by the primary afferent depolarization. α5GABAA receptors were immunolocalized in superficial laminae of the dorsal horn and L4 to L6 dorsal root ganglion. Streptozotocin increased mean fluorescence intensity and percentage of neurons expressing α5GABAA receptors in dorsal horn and L4 to L6 dorsal root ganglia in 10-week diabetic rats. Our results suggest that spinal α5GABAA receptors modulate the HR, play an antinociceptive and pronociceptive role in healthy and diabetic rats, respectively, and are tonically active in primary afferents. [ABSTRACT FROM AUTHOR]

Highlights • Aripiprazole elevated NMDA binding in male and female juvenile rats. • Risperidone and aripiprazole increased GABA A binding receptor in juvenile rats. • Olanzapine had no effects on NMDA and GABA A receptor bindings. • Early antipsychotic treatment had brain regional specific effects. • Early antipsychotic treatment caused some gender-dependent changes. Abstract Antipsychotics were developed to treat schizophrenia in adults; however they have been increasingly prescribed in children and adolescents. The NMDA and GABA A receptors are involved in neurodevelopment and the pathophysiology of various mental disorders in children and adolescents. Male and female juvenile rats were treated orally with risperidone (0.3 mg/kg, 3 times/day), aripiprazole (1 mg/kg), olanzapine (1 mg/kg) or vehicle (control), starting from postnatal day (PD) 23 (±1 day) for 3 weeks (corresponding to the childhood-adolescent period in humans). Quantitative autoradiography was used to detect the binding density of [3H]MK-801 (an NMDA receptor antagonist) and [3H]muscimol (a selective GABA A receptor agonist). Aripiprazole elevated the [3H]MK801 binding levels in the NAcC of male rats, and the NAcS and CPu of female rats. Risperidone increased [3H]MK801 levels in the CPu of female rats, and the NAcS of male rats. Aripiprazole upregulated [3H]muscimol binding levels in the CPu and NAcC of male rats, while it elevated the [3H]muscimol levels in the PFC of female rats, compared to controls. These results suggest that early treatment with these antipsychotics modulates NMDA and GABA A neurotransmission in juveniles, which may play a role in their clinical efficacy in the control of mental disorders in children and adolescents. [ABSTRACT FROM AUTHOR]

Objective: In this study, we aimed to evaluate age-dependent cognitive and behavioral changes, neuronal damage, and the amount of gamma-aminobutyric acid-A (GABA-A) alpha 1 in immature, mature, and adolescant rats after status epilepticus (SE). Materials and Methods: SE was induced in immature (17 days), adolescant (45 days), and mature (150 days) rats using pentylenetetrazole (PTZ). After SE, adolescant and mature rats underwent open field test and Morris water maze test. After behavioral tests, the animals were sacrificed and we performed histologic investigations on the immature, adolescant, and mature rats to assess neuronal cell damage (caspase and calpain activity) and amount of GABA-A alpha 1 receptor and compared them with a control group. Results: There were no statistically significant differences between the control and experimental groups in behavioral tests in the early stage after SE. Calpainmediated neuronal damage was observed in mature rats with necrotic morphology after SE, but this was not observed in adolescant and immature rats. Caspasemediated neuronal damage was observed in immature rats with apoptotic morphology after SE. The amount of GABA-A alpha 1 receptor was decreased in the three experimental groups compared with the control groups. The decrease in GABA-A alpha 1 receptor amount was highest in the mature experimental group. The amount of GABA-A alpha 1 receptor in the hippocampus decreased to level higher than in the cortex. Conclusion: This study show that there is no negative impact on learning and behavioral functions in the early stage after PTZ-induced SE, but histologically led to necrosis dominant calpain and caspase-mediated neuronal damage, calpain-mediated cell necrosis is seen particularly in the mature group and caspasedependent apoptotic morphology observed in immature rats.The decreased of GABA-A alpha 1 receptor is highest in the adults. Our study supports that SEinduced cell damage is more pronounced with increased age, and calpain-mediated cell damage that can clearly be observed in the mature group. Long-term follow-up studies are needed to understand the long-term effect of SE-dependent neuronal damage on cognition and behavior. [ABSTRACT FROM AUTHOR]

Background: The high rate of relapse to drug use remains a central challenge to treating drug addiction. In human and rat models of addiction, environmental stimuli in contexts associated with previous drug use can provoke a relapse of drug seeking. Pre-clinical studies have used the ABA renewal procedure to study context-induced reinstatement of drug seeking. In the current study, we studied the role of the orbitofrontal cortex (OFC) in context-induced reinstatement to alcohol.Methods: We trained male and female rats to self-administer alcohol in context A, extinguished drug-reinforced responding in a distinct context B, and assessed context-induced reinstatement in context A or B (control group). Next, we determined the effect of context-induced renewal of alcohol-seeking behavior on the expression of Fos (a neuronal activity marker) in the OFC. Finally, we determined the effect of reversible inactivation by GABAa and GABAb receptor agonists (i.e., muscimol and baclofen, respectively) in the OFC.Results and Conclusions: There were no differences between male and female rats in context-induced reinstatement of alcohol-seeking behavior. Re-exposure to Context A, but not Context B, reinstated alcohol-seeking behavior and increased expression of the neural activity marker Fos in the OFC. Reversible inactivation of the OFC with muscimol and baclofen attenuated context-induced reinstatement. Our data indicated that the OFC mediates context-induced reinstatement of alcohol-seeking behavior. [ABSTRACT FROM AUTHOR]

Approximately 7% to 10% of patients develop a chronic pain syndrome after stroke. This chronic pain condition is called central poststroke pain (CPSP). Recent studies have observed an abnormal increase in the secretion of brain-derived neurotrophic factor (BDNF) in spinal cord tissue after spinal cord injury. An animal model of CPSP was established by an intrathalamus injection of collagenase. Mechanical and thermal allodynia was induced after lesions of the thalamic ventral basal complex in rats. Four weeks after the injection, the number of neurons decreased, the number of astrocytes, microglia, and P2X4 receptors increased, and BDNF mRNA expression increased in the brain lesion area. Nociceptive activity in the medial thalamus (MT) and the coherence coefficient of spontaneous field potential oscillations in the anterior cingulate cortex were enhanced in CPSP animals, and these enhancements were blocked by an acute injection of TrkB-Fc and TrkB antagonist Tat Cyclotraxin-B. Instead of being inhibited by the γ-aminobutyric acid (GABA) system in normal rats, multiunit activity in the MT was enhanced after a microinjection of muscimol, a GABAA receptor agonist, in CPSP animals. After CPSP, BDNF expression was enhanced in the MT, whereas the expression of GABAA channels and the cotransporter KCC2 decreased in the same area. These findings suggest that neuronal plasticity in the MT that was induced by BDNF overexpression after the thalamic lesion was a key factor in CPSP. [ABSTRACT FROM AUTHOR]

The rostral ventromedial medulla (RVM) has a well-documented role in pain modulation and exerts antinociceptive and pronociceptive influences mediated by 2 distinct classes of neurons, OFF-cells and ON-cells. OFF-cells are defined by a sudden pause in firing in response to nociceptive inputs, whereas ON-cells are characterized by a "burst" of activity. Although these reflex-related changes in ON- and OFF-cell firing are critical to their pain-modulating function, the pathways mediating these responses have not been identified. The present experiments were designed to test the hypothesis that nociceptive input to the RVM is relayed through the parabrachial complex (PB). In electrophysiological studies, ON- and OFF-cells were recorded in the RVM of lightly anesthetized male rats before and after an infusion of lidocaine or muscimol into PB. The ON-cell burst and OFF-cell pause evoked by noxious heat or mechanical probing were substantially attenuated by inactivation of the lateral, but not medial, parabrachial area. Retrograde tracing studies showed that neurons projecting to the RVM were scattered throughout PB. Few of these neurons expressed calcitonin gene-related peptide, suggesting that the RVM projection from PB is distinct from that to the amygdala. These data show that a substantial component of "bottom-up" nociceptive drive to RVM pain-modulating neurons is relayed through the PB. While the PB is well known as an important relay for ascending nociceptive information, its functional connection with the RVM allows the spinoparabrachial pathway to access descending control systems as part of a recurrent circuit. [ABSTRACT FROM AUTHOR]

Strategies drawn at understanding the functional attributes of specific neural circuits often necessitate electrical stimulation and pharmacological manipulation at the same anatomical site. We describe a simple, inexpensive and reliable method to fabricate a bipolar electrode-cannula assembly for delivery of electric pulses and administration of neuroactive agents at the same site in the rat brain. The assembly consisting of a guide cannula, dummy cannula, internal cannula and bipolar electrode was fabricated using syringe needles, wires and simple electronic components. To test the usefulness of the device, it was implanted on the skull of a rat specifically targeting the posterior ventral tegmental area (pVTA). The rat was conditioned to press the lever in intracranial self-stimulation (ICSS) protocol in an operant chamber. The number of lever presses in a 30 min task was monitored. Intra-pVTA administration with bicuculline (GABA A receptor antagonist) increased the lever press activity, while muscimol (GABA A receptor agonist) had opposite effect. The results confirm that the group of neurons responding to the electrical stimulation probably receive GABAergic inputs. The device is light in weight, costs less than a dollar and can be fabricated from readily available components. It can serve a useful purpose in electrically stimulating any given target in the brain - before, during or after pharmacological manipulation at the same locus and may find application in neuropharmacological and neurobehavioral studies. • We developed a simple and inexpensive method to fabricate electrode-cannula assembly. • The assembly can be used to electrically stimulate and inject neuroactive agents at the same neuroanatomical site. • The usefulness of the device was tested in an intracranial self-stimulation (ICSS) protocol. • In ICSS protocol, GABA A receptors antagonist facilitated self-rewarding activity, but agonist had opposite effect. • The device may find application in neuropharmacology and neurobehavioral studies. [ABSTRACT FROM AUTHOR]

Ethnopharmacological relevance Montanoa tomentosa also named Cihuapatli is a native plant of Mexico that has been used in traditional medicine for the last five centuries mainly as a remedy for reproductive impairments. However, there are reports indicating that this plant was also consumed by Mexican ancient people for its relaxing properties. In order to corroborate this information, the present study was conducted to evaluate the effect of Montanoa tomentosa lyophilisate (MT) on rat׳s anxiety-like behavior and to analyze its mechanism of action. Materials and methods The anxiolytic-like action of MT (1.5, 3.0, 6.0 and 12.0 mg/kg) was investigated in male Wistar rats tested in three animal models of anxiety: the burying behavior, the elevated plus maze and the hole-board tests. As a positive control, the anti-anxiety effects of different doses of the selective GABA A receptor agonist muscimol were also analyzed. In order to evaluate the participation of the GABA A and oxytocin receptors in the anxiolytic-like actions of MT , the GABA A receptors blockers picrotoxin (0.25 and 0.50 mg/kg), bicuculline (2.0 mg/kg) and flumazenil (5.00 and 10.0 mg/kg), the neurosteroid inhibitor finasteride (50.0 and 100 mg/kg) and the oxytocin receptor antagonist atosiban (0.25 µg) were used. Finally, to evaluate general activity, and motor coordination, the open field and rota-rod tests were used. Results MT at 3.0 mg/kg showed anxiolytic-like effects in the three anxiety paradigms without affecting reactivity, general motor activity or motor coordination; however, at higher doses sedative effects were observed. Picrotoxin (0.25 and 0.50 mg/kg), flumazenil (10.0 mg/kg) and finasteride (100 mg/kg) antagonized the anxiolytic-like actions of MT in the burying behavior test. In the plus maze and hole-board tests bicuculline (2.0 mg/kg) blocked the effects of the plant as well. Atosiban (0.25 µg) did not antagonize the anxiolytic-like actions of MT. Conclusions The results corroborate the anxiolytic-like actions of Montanoa tomentosa and suggest that this effect is mediated through GABA A receptors but not oxytocin receptors. [ABSTRACT FROM AUTHOR]

This study aimed to investigate the effect of baclofen, a γ-aminobutyric acid B (GABAB) receptor agonist, on the expression of p-CREB and NR2B in the spinal dorsal horn of rats with diabetic neuropathic pain (DNP). The DNP rats, which were successfully induced with streptozocin, were distributed among 3 groups that were treated with saline (D1 group), baclofen (D2 group), or CGP55845 + baclofen (D3 group) continuously for 4 days. The rats induced with saline and subsequently treated with saline were used as controls (C group). The times for the paw withdrawal threshold and thermal withdrawal latency of the D1 group were lower than those for the C group, and were significantly increased after baclofen treatment, but not when GABA receptor was pre-blocked with CGP55845 (D3 group). Increased protein expression levels of NR2B and p-CREB and mRNA levels of NR2B were found in the D1 group when compared with the controls. Baclofen treatment significantly suppressed their expression, bringing it close to the levels of controls. However, in the D3 group, the expression of p-CREB and NR2B were still significantly higher than that of the controls. Activation of GABAB receptor by baclofen attenuates diabetic neuropathic pain, which may partly be accomplished via down-regulating the expression of p-CREB and NR2B. [ABSTRACT FROM AUTHOR]

BACKGROUND: Propofol facilitates [gamma]-aminobutyric acid-mediated inhibitory synaptic transmission. In the cerebral cortex, [gamma]-aminobutyric acidergic interneurons target both excitatory pyramidal cells (Pyr) and fast-spiking (FS) and non-FS interneurons. Therefore, the propofol-induced facilitation of inhibitory transmission results in a change in the balance of excitatory and inhibitory inputs to Pyr. However, it is still unknown how propofol modulates [gamma]-aminobutyric acidergic synaptic transmission in each combination of Pyr and interneurons. METHODS: The authors examined whether propofol differentially regulates inhibitory postsynaptic currents (IPSCs) depending on the presynaptic and postsynaptic cell subtypes using multiple whole cell patch clamp recording from [gamma]-aminobutyric acidergic interneurons and Pyr in rat insular cortex. RESULTS: Propofol (10 [mu]M) consistently prolonged decay kinetics of unitary IPSCs (uIPSCs) in all types of inhibitory connections without changing paired-pulse ratio of the second to first uIPSC amplitude or failure rate. The FS-->Pyr connections exhibited greater enhancement of uIPSC charge transfer (2.2 ± 0.5 pC, n = 36) compared with that of FS-->FS/non-FS connections (0.9 ± 0.2 pC, n = 37), whereas the enhancement of charge transfer in non-FS-->Pyr (0.3 ± 0.1 pC, n = 15) and non-FS-->FS/non-FS connections (0.2 ± 0.1 pC, n = 36) was smaller to those in FS-->Pyr/FS/non-FS. Electrical synapses between FS pairs were not affected by propofol. CONCLUSIONS: The principal inhibitory connections (FS-->Pyr) are the most sensitive to propofol-induced facilitation of uIPSCs, which is likely mediated by postsynaptic mechanisms. This preferential uIPSC enhancement in FS-->Pyr connections may result in suppressed neural activities of projection neurons, which in turn reduces excitatory outputs from cortical local circuits. [ABSTRACT FROM AUTHOR]

Overcoming specific fears and subsequent anxiety can be greatly enhanced by the presence of familiar social partners, but the neural circuitry that controls this phenomenon remains unclear. To overcome this, the social interaction (SI) habituation test was developed in this lab to systematically investigate the effects of social familiarity on anxiety-like behavior in rats. Here, we show that social familiarity selectively reduced anxiety-like behaviors induced by an ethological anxiogenic stimulus. The anxiolytic effect of social familiarity could be elicited over multiple training sessions and was specific to both the presence of the anxiogenic stimulus and the familiar social partner. In addition, socially familiar conspecifics served as a safety signal, as anxiety-like responses returned in the absence of the familiar partner. The expression of the social familiarity-induced anxiolysis (SFiA) appears dependent on the prefrontal cortex (PFC), an area associated with cortical regulation of fear and anxiety behaviors. Inhibition of the PFC, with bilateral injections of the GABAA agonist muscimol, selectively blocked the expression of SFiA while having no effect on SI with a novel partner. Finally, the effect of D-cycloserine, a cognitive enhancer that clinically enhances behavioral treatments for anxiety, was investigated with SFiA. D-cycloserine, when paired with familiarity training sessions, selectively enhanced the rate at which SFiA was acquired. Collectively, these outcomes suggest that the PFC has a pivotal role in SFiA, a complex behavior involving the integration of social cues of familiarity with contextual and emotional information to regulate anxiety-like behavior. [ABSTRACT FROM AUTHOR]

Background Administration of the GABAB receptor agonist, baclofen, and positive allosteric modulator, GS39783, has been repeatedly reported to suppress multiple alcohol-related behaviors, including operant oral alcohol self-administration, in rats. This study was designed to compare the effect of baclofen and GS39783 on alcohol self-administration in 3 lines of selectively bred, alcohol-preferring rats: Indiana alcohol-preferring ( P), Sardinian alcohol-preferring ( sP), and Alko Alcohol ( AA). Methods Rats of each line were initially trained to respond on a lever, on a fixed ratio ( FR) 4 ( FR4) schedule of reinforcement, to orally self-administer alcohol (15%, v/v) in daily 30-minute sessions. Once responding reached stable levels, rats were exposed to a sequence of experiments testing baclofen (0, 1, 1.7, and 3 mg/kg; i.p.) and GS39783 (0, 25, 50, and 100 mg/kg; i.g.) on FR4 and progressive ratio ( PR) schedules of reinforcement. Finally, to assess the specificity of baclofen and GS39783 action, rats were slightly food-deprived and trained to lever-respond for food pellets. Results The rank of order of the reinforcing and motivational properties of alcohol was P> sP> AA rats. Under both FR and PR schedules of reinforcement, the rank of order of potency and efficacy of baclofen and GS39783 in suppressing alcohol self-administration was P> sP> AA rats. Only the highest dose of baclofen reduced lever-responding for food pellets; this effect was common to all 3 rat lines. Conversely, no dose of GS39783 altered lever-responding for food in any rat line. Conclusions These results suggest that: (i) the strength of the reinforcing and motivational properties of alcohol differ among P, sP, and AA rats; (ii) the reinforcing and motivational properties of alcohol in P, sP, and AA rats are differentially sensitive to treatment with baclofen and GS39783; (iii) the heterogeneity in sensitivity to baclofen and GS39783 of alcohol self-administration in P, sP, and AA rats may resemble the differential effectiveness of pharmacotherapies among the different typologies of human alcoholics; and (iv) the GABAB receptor is part of the neural substrate mediating the reinforcing and motivational properties of alcohol. [ABSTRACT FROM AUTHOR]

Accumulated evidence has suggested that potentiation of cortical GABAergic inhibitory neurotransmission may be a key mechanism in the treatment of schizophrenia. However, the downstream molecular mechanisms related to GABA potentiation remain unexplored. Recent studies have suggested that dopamine D2 receptor antagonists, which are used in the clinical treatment of schizophrenia, modulate protein kinase B (Akt)/glycogen synthase kinase (GSK)-3 signaling. Here we report that activation of GABAB receptors significantly inhibits Akt/GSK-3 signaling in a β-arrestin-dependent pathway. Agonist stimulation of GABAB receptors enhances the phosphorylation of Akt (Thr-308) and enhances the phosphorylation of GSK-3α (Ser-21)/β (Ser-9) in both HEK-293T cells expressing GABAB receptors and rat hippocampal slices. Furthermore, knocking down the expression of β-arrestin2 using siRNA abolishes the GABAB receptor-mediated modulation of GSK-3 signaling. Our data may help to identify potentially novel targets through which GABAB receptor agents may exert therapeutic effects in the treatment of schizophrenia. [ABSTRACT FROM AUTHOR]

Abstract: Thickened oil-in-water emulsions are useful model foods in rat studies due to their high acceptance and similarity to foods consumed by humans. Previous work from this laboratory used oil-in-water emulsions thickened with a biopolymer blend containing starch. Intake and effects of baclofen, a GABA-B agonist that decreases fat intake and drug self-administration, were reported, but the contribution of starch was not assessed. In the present study, intake and effects of baclofen were assessed in rats using emulsions prepared with two fat types (32% vegetable shortening, 32% corn oil) and thickened with three biopolymer blends. One biopolymer blend contained starch and the other two did not. Daily 1-h intake of the vegetable shortening emulsion containing starch was significantly greater than the other emulsions. When starch was added to the emulsions originally containing no starch, intake significantly increased. Baclofen generally reduced intake of all emulsions regardless of starch content and stimulated intake of chow. However, effects were more often significant for vegetable shortening emulsions. This report: (1) demonstrates that products used to prepare thickened oil-in-water emulsions have significant effects on rat ingestive behavior, and (2) confirms the ability of baclofen to reduce consumption of fatty foods, while simultaneously stimulating intake of chow. [Copyright &y& Elsevier]

Abstract: 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 2.0mg/kg IVM, 15min prior to behavioral testing. In the second experiment, the effects of four previous sexual experiences on IVM treated rats (1.0 or 2.0mg/kg, 15min prior to the 5th session) were assessed. The standard therapeutic dose (0.2mg/kg) did not impair the sexual behavior of inexperienced male rats. At a more concentrated dose (0.6mg/kg), which is still within the therapeutic range, the appetitive phase of sexual behavior of inexperienced male rats was impaired. Likewise, 1.0mg/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. [Copyright &y& Elsevier]

Neuroactive steroids modulate ethanol intake in several self-administration models with variable effects. The purpose of this work was to examine the effects of the long-acting synthetic GABAergic neurosteroid ganaxolone and the endogenous neurosteroid pregnenolone, a precursor of all GABAergic neuroactive steroids, on the maintenance of ethanol self-administration in an animal model of elevated drinking-the alcohol-preferring (P) rats. P rats were trained to self-administer ethanol (15% v/v) versus water on a concurrent schedule of reinforcement, and the effects of ganaxolone (0 to 30 mg/kg, subcutaneous [SC]) and pregnenolone (0 to 75 mg/kg, intraperitoneal [IP]) were evaluated on the maintenance of ethanol self-administration. After completion of self-administration testing, doses of the neuroactive steroids that altered ethanol self-administration were assessed on spontaneous locomotor activity. Finally, the effect of pregnenolone administration on cerebral cortical levels of the GABAergic neuroactive steroid (3α,5α)-3-hydroxypregnan-20-one (allopregnanolone, 3α,5α-THP) was determined in both ethanol-experienced and ethanol-inexperienced P rats because pregnenolone is a precursor of these steroids. Ganaxolone produced a dose-dependent biphasic effect on ethanol reinforcement, as the lowest dose (1 mg/kg) increased and the highest dose (30 mg/kg) decreased ethanol-reinforced responding. However, the highest ganaxolone dose also produced a nonspecific reduction in locomotor activity. Pregnenolone treatment significantly reduced ethanol self-administration (50 and 75 mg/kg), without altering locomotor activity. Pregnenolone (50 mg/kg) produced a significant increase in cerebral cortical allopregnanolone levels. This increase was observed in the self-administration trained animals, but not in ethanol-naïve P rats. These results indicate that pregnenolone dose-dependently reduces operant ethanol self-administration in P rats without locomotor impairment, suggesting that it may have potential as a novel therapeutic for reducing chronic alcohol drinking in individuals that abuse alcohol. [ABSTRACT FROM AUTHOR]

Background and Purpose: Baclofen (a GABA(B) receptor agonist) is the most commonly used anti-spasticity agent in clinical practice. While effective when administered spinally or systemically, the development of progressive tolerance represents a serious limitation for its long-term use. The goal of the present study was to characterize the treatment potency after intrathecal or systemic treatment with the selective AMPA receptor antagonist NGX424 on stretch reflex activity (SRA) and background muscle activity (BMA) in rats with developed baclofen tolerance.Experimental Approach: Animals were exposed to 10 min of spinal ischaemia to induce an increase in BMA and SRA. Selected animals were implanted with an intrathecal PE-5 catheter and infused intrathecally with baclofen (1 µg·h⁻¹ ) for 14 days. Before and after baclofen infusion, changes in BMA and SRA were measured at 2 day intervals. After development of baclofen tolerance, the animals were injected intrathecally (1 µg) or subcutaneously (3, 6 or 12 mg·kg⁻¹) with NGX424, and changes in BMA and SRA were measured.Key Results: Intrathecal or systemic delivery of NGX424 significantly suppressed the BMA and SRA in baclofen-tolerant animals. This effect was dose dependent. The magnitude of BMA and SRA suppression seen after 1 µg (intrathecal) or 12 mg·kg ⁻¹ (s.c.) of NGX424 injection was similar to that seen during the first 5 days of baclofen infusion. CONCLUSIONS AND IMPLICATIONS These data demonstrate that the use of NGX424 can represent an effective therapy to modulate chronic spasticity in patients who are refractory or tolerant to baclofen treatment. [ABSTRACT FROM AUTHOR]